Toxins

Article

21 pages, 2525 KiB

Open AccessEditor’s ChoiceArticle

A Novel Cytotoxic Mechanism for Triple-Negative Breast Cancer Cells Induced by the Type II Heat-Labile Enterotoxin LT-IIc through Ganglioside Ligation

by Natalie D. King-Lyons, Aryana S. Bhati, John C. Hu, Lorrie M. Mandell, Gautam N. Shenoy, Hugh J. Willison and Terry D. Connell

Toxins 2024, 16(7), 311; https://doi.org/10.3390/toxins16070311 - 11 Jul 2024

Viewed by 1028

Abstract

Triple-negative breast cancer (TNBC), which constitutes 10–20 percent of all breast cancers, is aggressive, has high metastatic potential, and carries a poor prognosis due to limited treatment options. LT-IIc, a member of the type II subfamily of ADP-ribosylating—heat-labile enterotoxins that bind to a [...] Read more.

Triple-negative breast cancer (TNBC), which constitutes 10–20 percent of all breast cancers, is aggressive, has high metastatic potential, and carries a poor prognosis due to limited treatment options. LT-IIc, a member of the type II subfamily of ADP-ribosylating—heat-labile enterotoxins that bind to a distinctive set of cell-surface ganglioside receptors—is cytotoxic toward TNBC cell lines, but has no cytotoxic activity for non-transformed breast epithelial cells. Here, primary TNBC cells, isolated from resected human tumors, showed an enhanced cytotoxic response specifically toward LT-IIc, in contrast to other enterotoxins that were tested. MDA-MB-231 cells, a model for TNBC, were used to evaluate potential mechanisms of cytotoxicity by LT-IIc, which induced elevated intracellular cAMP and stimulated the cAMP response element-binding protein (CREB) signaling pathway. To dissect the role of ADP-ribosylation, cAMP induction, and ganglioside ligation in the cytotoxic response, MDA-MB-231 cells were exposed to wild-type LT-IIc, the recombinant B-pentamer of LT-IIc that lacks the ADP-ribosylating A polypeptide, or mutants of LT-IIc with an enzymatically inactivated A1-domain. These experiments revealed that the ADP-ribosyltransferase activity of LT-IIc was nonessential for inducing the lethality of MDA-MB-231 cells. In contrast, a mutant LT-IIc with an altered ganglioside binding activity failed to trigger a cytotoxic response in MDA-MB-231 cells. Furthermore, the pharmacological inhibition of ganglioside expression protected MDA-MB-231 cells from the cytotoxic effects of LT-IIc. These data establish that ganglioside ligation, but not the induction of cAMP production nor ADP-ribosyltransferase activity, is essential to initiating the LT-IIc-dependent cell death of MDA-MB-231 cells. These experiments unveiled previously unknown properties of LT-IIc and gangliosides in signal transduction, offering the potential for the targeted treatment of TNBC, an option that is desperately needed. Full article

(This article belongs to the Section Bacterial Toxins)

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20 pages, 4165 KiB

Open AccessEditor’s ChoiceArticle

Bioremediation of Aflatoxin B₁ by Meyerozyma guilliermondii AF01 in Peanut Meal via Solid-State Fermentation

by Wan Zhang, Changpo Sun, Wei Wang and Zhongjie Zhang

Toxins 2024, 16(7), 305; https://doi.org/10.3390/toxins16070305 - 4 Jul 2024

Viewed by 949

Abstract

The use of microorganisms to manage aflatoxin contamination is a gentle and effective approach. The aim of this study was to test the removal of AFB₁ from AFB₁-contaminated peanut meal by a strain of Meyerozyma guilliermondii AF01 screened by the [...] Read more.

The use of microorganisms to manage aflatoxin contamination is a gentle and effective approach. The aim of this study was to test the removal of AFB₁ from AFB₁-contaminated peanut meal by a strain of Meyerozyma guilliermondii AF01 screened by the authors and to optimize the conditions of the biocontrol. A regression model with the removal ratio of AFB₁ as the response value was established by means of single-factor and response surface experiments. It was determined that the optimal conditions for the removal of AFB₁ from peanut meal by AF01 were 75 h at 29 °C under the natural pH, with an inoculum of 5.5%; the removal ratio of AFB₁ reached 69.31%. The results of simulating solid-state fermentation in production using shallow pans and fermentation bags showed that the removal ratio of AFB₁ was 68.85% and 70.31% in the scaled-up experiments, respectively. This indicated that AF01 had strong adaptability to the environment with facultative anaerobic fermentation detoxification ability. The removal ratio of AFB₁ showed a positive correlation with the growth of AF01, and there were no significant changes in the appearance and quality of the peanut meal after fermentation. This indicated that AF01 had the potential to be used in practical production. Full article

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10 pages, 538 KiB

Open AccessEditor’s ChoiceArticle

Enhancing Botulinum Toxin Injection Precision: The Efficacy of a Single Cadaveric Ultrasound Training Intervention for Improved Anatomical Localization

by Camille Heslot, Omar Khan, Alexis Schnitzler, Chloe Haldane, Romain David and Rajiv Reebye

Toxins 2024, 16(7), 304; https://doi.org/10.3390/toxins16070304 - 2 Jul 2024

Viewed by 902

Abstract

Ultrasound guidance can enhance existing landmark-based injection methods, even through a brief and single exposure during a cadaveric training course. A total of twelve participants were enrolled in this training program, comprising nine physical medicine and rehabilitation specialists, one pediatrician, and two physician [...] Read more.

Ultrasound guidance can enhance existing landmark-based injection methods, even through a brief and single exposure during a cadaveric training course. A total of twelve participants were enrolled in this training program, comprising nine physical medicine and rehabilitation specialists, one pediatrician, and two physician assistants. For each participant, one upper-limb muscle and one lower-limb muscle were randomly chosen from the preselected muscle group. Subsequently, participants were tasked with injecting both of their chosen cadaveric muscles with 1 mL of acrylic paint using a manual needle palpation technique, relying solely on their knowledge of anatomic landmarks. Participants then underwent a personalized, one-to-one ultrasound teaching session, lasting approximately five minutes, conducted by two highly experienced instructors. Following this instructive phase, participants were tasked with a second round of injections, targeting the same two muscles in the lower and upper limbs. However, this time, the injections were performed using anatomical landmarks and ultrasound guidance. To facilitate differentiation from the initial injections, a distinct color of acrylic paint was employed. When employing the anatomical landmark-based approach, the overall success rate for injections was 67%, with 16 out of 24 targeted muscles accurately injected. With the incorporation of ultrasound guidance, the success rate was 92%, precisely targeting 22 out of the 24 muscles under examination. There was an improvement in injection accuracy achievable through the integration of ultrasound guidance, even with minimal training exposure. Our single cadaveric ultra-sound training program contributes valuable insights to the utilization of ultrasound for anatomy training to help optimize the targeting of BoNT-A. Full article

(This article belongs to the Special Issue Uses of Botulinum Toxin Injection in Medicine)

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24 pages, 4815 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

The Clot Thickens: Differential Coagulotoxic and Cardiotoxic Activities of Anguimorpha Lizard Venoms

by James Dobson, Abhinandan Chowdhury, Jeremie Tai-A-Pin, Harold van der Ploeg, Amber Gillett and Bryan G. Fry

Toxins 2024, 16(6), 283; https://doi.org/10.3390/toxins16060283 - 20 Jun 2024

Cited by 1 | Viewed by 1377

Abstract

Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards have long been assumed to be for defensive purposes, there is increasing evidence of toxic activities more useful [...] Read more.

Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards have long been assumed to be for defensive purposes, there is increasing evidence of toxic activities more useful for predation than defence (such as paralytic neurotoxicity). This study aimed to ascertain the effects of Heloderma, Lanthanotus, and Varanus lizard venoms on the coagulation and cardiovascular systems. Anticoagulant toxicity was demonstrated for the Varanus species studied, with the venoms prolonging clotting times in human and bird plasma due to the destructive cleavage of fibrinogen. In contrast, thromboelastographic analyses on human and bird plasmas in this study demonstrated a procoagulant bioactivity for Heloderma venoms. A previous study on Heloderma venom using factor-depleted plasmas as a proxy model suggested a procoagulant factor was present that activated either Factor XI or Factor XII, but could not ascertain the precise target. Our activation studies using purified zymogens confirmed FXII activation. Comparisons of neonate and adult H. exasperatum, revealed the neonates to be more potent in the ability to activate FXII, being more similar to the venom of the smaller species H. suspectum than the adult H. exasperatum. This suggests potent FXII activation a basal trait in the genus, present in the small bodied last common ancestor. This also indicates an ontogenetic difference in prey preferences in the larger Heloderma species paralleing the change in venom biochemistry. In addition, as birds lack Factor XII, the ability to clot avian plasma suggested an additional procoagulant site of action, which was revealed to be the activation of Factor VII, with H. horridum being the most potent. This study also examined the effects upon the cardiovascular system, including the liberation of kinins from kininogen, which contributes to hypotension induction. This form of toxicity was previously described for Heloderma venoms, and was revealed in this study was to also be a pathophysiological effect of Lanthanotus and Varanus venoms. This suggests that this toxic activity was present in the venom of the last common ancestor of the anguimorph lizards, which is consistent with kallikrein enzymes being a shared toxin trait. This study therefore uncovered novel actions of anguimorph lizard venoms, not only contributing to the evolutionary biology body of knowledge but also revealing novel activities to mine for drug design lead compounds. Full article

(This article belongs to the Special Issue Venom Components Acting on the Hemostatic System: Structural and Mechanistic Insights)

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15 pages, 3020 KiB

Open AccessEditor’s ChoiceArticle

The Possible Role of Mycotoxins in the Pathogenesis of Endometrial Cancer

by Márkó Unicsovics, Zsófia Molnár, Miklós Mézes, Katalin Posta, György Nagyéri, Szabolcs Várbíró, Nándor Ács, Levente Sára and Zsuzsanna Szőke

Toxins 2024, 16(6), 236; https://doi.org/10.3390/toxins16060236 - 23 May 2024

Viewed by 1772

Abstract

Endometrial cancer is one of the most common cancer types among women. Many factors can contribute to the development of this disease, including environmental factors and, thus, eating habits. Our study aims to determine the levels of various mycotoxins and their metabolites in [...] Read more.

Endometrial cancer is one of the most common cancer types among women. Many factors can contribute to the development of this disease, including environmental factors and, thus, eating habits. Our study aims to determine the levels of various mycotoxins and their metabolites in the blood serum and endometrial tissue samples of participants with previously proven endometrial cancer and to find possible contributions to cancer development. In the cohort clinical trial, 52 participants aged between 44 and 86 were studied. The participants were divided into two groups: patients or matched controls. All patients had previously histologically diagnosed endometrial cancer. The cancer patients were divided into low-grade endometrioid and low- plus high-grade endometrioid groups. Controls had no history of endometrial malignancy or premalignancy. Blood serum and endometrial tissue samples were obtained from all study patients. We compared the concentrations of total Aflatoxins (Afs), Deoxynivalenol (DON), Ochratoxin-A (OTA), T2-toxin and HT2 toxin (T2/HT2 toxin), Zearalenone (ZEN), alpha-Zearalenol (α-ZOL), and Fumonisin B1 (FB1) in the serum and endometrium between the different study groups. As a result, we can see a significant correlation between the higher levels of Afs and zearalenone and the presence of endometrial cancer. In the case of Afs, DON, OTA, T2/HT2 toxins, ZEN, and alpha-ZOL, we measured higher endometrial concentrations than in serum. Considering the effect of mycotoxins and eating habits on cancer development, our results might lead to further research exploring the relationship between certain mycotoxins and endometrium cancer. Full article

(This article belongs to the Special Issue Advances in Contamination, Detection and Risk Assessment of Mycotoxins)

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10 pages, 4613 KiB

Open AccessEditor’s ChoiceArticle

Borrelia burgdorferi 0755, a Novel Cytotoxin with Unknown Function in Lyme Disease

by Sam T. Donta

Toxins 2024, 16(6), 233; https://doi.org/10.3390/toxins16060233 - 21 May 2024

Viewed by 2020

Abstract

The pathophysiology of Lyme disease, especially in its persistent form, remains to be determined. As many of the neurologic symptoms are similar to those seen in other toxin-associated disorders, a hypothesis was generated that B. burgdorferi, the causative agent of Lyme disease, [...] Read more.

The pathophysiology of Lyme disease, especially in its persistent form, remains to be determined. As many of the neurologic symptoms are similar to those seen in other toxin-associated disorders, a hypothesis was generated that B. burgdorferi, the causative agent of Lyme disease, may produce a neurotoxin to account for some of the symptoms. Using primers against known conserved bacterial toxin groups, and PCR technology, a candidate neurotoxin was discovered. The purified protein was temporarily named BbTox, and was subsequently found to be identical to BB0755, a protein deduced from the genome sequence of B. burgdorferi that has been annotated as a Z ribonuclease. BbTox has cytotoxic activity against cells of neural origin in tissue culture. Its toxic activity appears to be directed against cytoskeletal elements, similar to that seen with toxins of Clostridioides difficile and Clostridioides botulinum, but differing from that of cholera and E. coli toxins, and other toxins. It remains to be determined whether BbTox has direct cytotoxic effects on neural or glial cells in vivo, or its activity is primarily that of a ribonuclease analogous to other bacterial ribonucleases that are involved in antibiotic tolerance remains to be determined. Full article

(This article belongs to the Section Bacterial Toxins)

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15 pages, 4425 KiB

Open AccessEditor’s ChoiceArticle

Specificity of DNA ADP-Ribosylation Reversal by NADARs

by Bara Cihlova, Yang Lu, Andreja Mikoč, Marion Schuller and Ivan Ahel

Toxins 2024, 16(5), 208; https://doi.org/10.3390/toxins16050208 - 28 Apr 2024

Cited by 1 | Viewed by 1825

Abstract

Recent discoveries establish DNA and RNA as bona fide substrates for ADP-ribosylation. NADAR (“NAD- and ADP-ribose”-associated) enzymes reverse guanine ADP-ribosylation and serve as antitoxins in the DarT-NADAR operon. Although NADARs are widespread across prokaryotes, eukaryotes, and viruses, their specificity and broader physiological roles [...] Read more.

Recent discoveries establish DNA and RNA as bona fide substrates for ADP-ribosylation. NADAR (“NAD- and ADP-ribose”-associated) enzymes reverse guanine ADP-ribosylation and serve as antitoxins in the DarT-NADAR operon. Although NADARs are widespread across prokaryotes, eukaryotes, and viruses, their specificity and broader physiological roles remain poorly understood. Using phylogenetic and biochemical analyses, we further explore de-ADP-ribosylation activity and antitoxin functions of NADAR domains. We demonstrate that different subfamilies of NADAR proteins from representative E. coli strains and an E. coli-infecting phage retain biochemical activity while displaying specificity in providing protection from toxic guanine ADP-ribosylation in cells. Furthermore, we identify a myxobacterial enzyme within the YbiA subfamily that functions as an antitoxin for its associated DarT-unrelated ART toxin, which we termed YarT, thus presenting a hitherto uncharacterised ART-YbiA toxin–antitoxin pair. Our studies contribute to the burgeoning field of DNA ADP-ribosylation, supporting its physiological relevance within and beyond bacterial toxin–antitoxin systems. Notably, the specificity and confinement of NADARs to non-mammals infer their potential as highly specific targets for antimicrobial drugs with minimal off-target effects. Full article

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17 pages, 10327 KiB

Open AccessEditor’s ChoiceArticle

Edodin: A New Type of Toxin from Shiitake Mushroom (Lentinula edodes) That Inactivates Mammalian Ribosomes

by Lucía Citores, Sara Ragucci, Claudia C. Gay, Rosita Russo, Angela Chambery, Antimo Di Maro, Rosario Iglesias and José M. Ferreras

Toxins 2024, 16(4), 185; https://doi.org/10.3390/toxins16040185 - 10 Apr 2024

Viewed by 1902

Abstract

Ribosome-inactivating proteins (RIPs) are a group of proteins with rRNA N-glycosylase activity that irreversibly inhibit protein synthesis and consequently cause cell death. Recently, an RIP called ledodin has been found in shiitake; it is cytotoxic, strongly inhibits protein synthesis, and shows rRNA N-glycosylase [...] Read more.

Ribosome-inactivating proteins (RIPs) are a group of proteins with rRNA N-glycosylase activity that irreversibly inhibit protein synthesis and consequently cause cell death. Recently, an RIP called ledodin has been found in shiitake; it is cytotoxic, strongly inhibits protein synthesis, and shows rRNA N-glycosylase activity. In this work, we isolated and characterized a 50 kDa cytotoxic protein from shiitake that we named edodin. Edodin inhibits protein synthesis in a mammalian cell-free system, but not in insect-, yeast-, and bacteria-derived systems. It exhibits rRNA N-glycosylase and DNA-nicking activities, which relate it to plant RIPs. It was also shown to be toxic to HeLa and COLO 320 cells. Its structure is not related to other RIPs found in plants, bacteria, or fungi, but, instead, it presents the characteristic structure of the fold type I of pyridoxal phosphate-dependent enzymes. Homologous sequences have been found in other fungi of the class Agaricomycetes; thus, edodin could be a new type of toxin present in many fungi, some of them edible, which makes them of great interest in health, both for their involvement in food safety and for their potential biomedical and biotechnological applications. Full article

(This article belongs to the Special Issue Biological Activities of Ribosome Inactivating Proteins II)

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11 pages, 4216 KiB

Open AccessEditor’s ChoiceArticle

High-Voltage Toxin’Roll: Electrostatic Charge Repulsion as a Dynamic Venom Resistance Trait in Pythonid Snakes

by Uthpala Chandrasekara, Emilie M. Broussard, Darin R. Rokyta and Bryan G. Fry

Toxins 2024, 16(4), 176; https://doi.org/10.3390/toxins16040176 - 4 Apr 2024

Viewed by 1984

Abstract

The evolutionary interplay between predator and prey has significantly shaped the development of snake venom, a critical adaptation for subduing prey. This arms race has spurred the diversification of the components of venom and the corresponding emergence of resistance mechanisms in the prey [...] Read more.

The evolutionary interplay between predator and prey has significantly shaped the development of snake venom, a critical adaptation for subduing prey. This arms race has spurred the diversification of the components of venom and the corresponding emergence of resistance mechanisms in the prey and predators of venomous snakes. Our study investigates the molecular basis of venom resistance in pythons, focusing on electrostatic charge repulsion as a defense against α-neurotoxins binding to the alpha-1 subunit of the postsynaptic nicotinic acetylcholine receptor. Through phylogenetic and bioactivity analyses of orthosteric site sequences from various python species, we explore the prevalence and evolution of amino acid substitutions that confer resistance by electrostatic repulsion, which initially evolved in response to predatory pressure by Naja (cobra) species (which occurs across Africa and Asia). The small African species Python regius retains the two resistance-conferring lysines (positions 189 and 191) of the ancestral Python genus, conferring resistance to sympatric Naja venoms. This differed from the giant African species Python sebae, which has secondarily lost one of these lysines, potentially due to its rapid growth out of the prey size range of sympatric Naja species. In contrast, the two Asian species Python brongersmai (small) and Python bivittatus (giant) share an identical orthosteric site, which exhibits the highest degree of resistance, attributed to three lysine residues in the orthosteric sites. One of these lysines (at orthosteric position 195) evolved in the last common ancestor of these two species, which may reflect an adaptive response to increased predation pressures from the sympatric α-neurotoxic snake-eating genus Ophiophagus (King Cobras) in Asia. All these terrestrial Python species, however, were less neurotoxin-susceptible than pythons in other genera which have evolved under different predatory pressure as: the Asian species Malayopython reticulatus which is arboreal as neonates and juveniles before rapidly reaching sizes as terrestrial adults too large for sympatric Ophiophagus species to consider as prey; and the terrestrial Australian species Aspidites melanocephalus which occupies a niche, devoid of selection pressure from α-neurotoxic predatory snakes. Our findings underline the importance of positive selection in the evolution of venom resistance and suggest a complex evolutionary history involving both conserved traits and secondary evolution. This study enhances our understanding of the molecular adaptations that enable pythons to survive in environments laden with venomous threats and offers insights into the ongoing co-evolution between venomous snakes and their prey. Full article

(This article belongs to the Section Animal Venoms)

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20 pages, 912 KiB

Open AccessEditor’s ChoiceArticle

Paradoxical Exception to Island Tameness: Increased Defensiveness in an Insular Population of Rattlesnakes

by William K. Hayes, Carl E. Person, Gerad A. Fox, Julie L. King, Erick Briggs and Eric C. K. Gren

Toxins 2024, 16(3), 157; https://doi.org/10.3390/toxins16030157 - 18 Mar 2024

Cited by 1 | Viewed by 2965

Abstract

Island tameness results largely from a lack of natural predators. Because some insular rattlesnake populations lack functional rattles, presumably the consequence of relaxed selection from reduced predation, we hypothesized that the Santa Catalina Island, California, USA, population of the southern Pacific rattlesnake ( [...] Read more.

Island tameness results largely from a lack of natural predators. Because some insular rattlesnake populations lack functional rattles, presumably the consequence of relaxed selection from reduced predation, we hypothesized that the Santa Catalina Island, California, USA, population of the southern Pacific rattlesnake (Crotalus helleri, which possesses a functional rattle), would exhibit a decrement in defensive behavior relative to their mainland counterparts. Contrary to our prediction, rattlesnakes from the island not only lacked tameness compared to mainland snakes, but instead exhibited measurably greater levels of defensiveness. Island snakes attempted to bite 4.7 times more frequently as we endeavored to secure them by hand, and required 2.1-fold more time to be pinned and captured. When induced to bite a beaker after being grasped, the island snakes also delivered 2.1-fold greater quantities of venom when controlling for body size. The additional venom resulted from 2.1-fold larger pulses of venom ejected from the fangs. We found no effects of duration in captivity (2–36 months), which suggests an absence of long-term habituation of antipredator behaviors. Breeding bird surveys and Christmas bird counts indicated reduced population densities of avian predators on Catalina compared to the mainland. However, historical estimates confirmed that populations of foxes and introduced mammalian predators (cats and pigs) and antagonists (herbivorous ungulates) substantially exceeded those on the mainland in recent centuries, and therefore best explain the paradoxically exaggerated defensive behaviors exhibited by Catalina’s rattlesnakes. These findings augment our understanding of anthropogenic effects on the behaviors of island animals and underscore how these effects can negatively affect human safety. Full article

(This article belongs to the Section Animal Venoms)

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15 pages, 1034 KiB

Open AccessEditor’s ChoiceArticle

Association of Urinary Mycotoxins with Sperm Quality: A Case-Control Study in Southern Nigeria

by Friday Ebhodaghe Okonofua, Lorretta Favour Chizomam Ntoimo, Emmanuel Iyayi Unuabonah, Titus Afred Makudali Msagati, Oladiran Ayodeji, Michael Aziken, Akhere Omonkhua, Victor Ohenhen, Celestina Olafusi and Moses O. Alfred

Toxins 2024, 16(3), 119; https://doi.org/10.3390/toxins16030119 - 29 Feb 2024

Cited by 2 | Viewed by 2438

Abstract

The objective of this study was to determine the association between mycotoxins and the quality of spermatozoa in Nigeria. We designed a prospective case-control study involving 136 men diagnosed with reduced sperm count and quality in five infertility clinics in southwest Nigeria and [...] Read more.

The objective of this study was to determine the association between mycotoxins and the quality of spermatozoa in Nigeria. We designed a prospective case-control study involving 136 men diagnosed with reduced sperm count and quality in five infertility clinics in southwest Nigeria and 154 normal fertile controls. Sperm analysis was conducted in accordance with the recommendations of the World Health OrganizationWHO, while Liquid Chromatography–Mass Spectrometry was used to assay three metabolites of mycotoxins (zearalenone, ochratoxin A, and deoxyvinelol) in the urine samples of cases and controls. The data were analysed with descriptive statistics and non-parametric linear regression. The results showed no overall significant difference in levels of these metabolites between the cases and control groups. In contrast, higher levels of zearalenone and ochratoxin A significantly decreased sperm motility in the cases. Similarly, an increase in the level of ochratoxin A decreased sperm morphology in the unadjusted model in the cases. We conclude that exposure to mycotoxins reduces the quality of spermatozoa (motility and morphology) in Nigerian men but may have no effect on sperm count. Efforts to reduce the exposure of men to mycotoxins are important interventions to improve sperm quality and reduce the prevalence of male infertility in the country. Full article

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13 pages, 3878 KiB

Open AccessEditor’s ChoiceArticle

Bee Venom Stimulates Growth Factor Release from Adipose-Derived Stem Cells to Promote Hair Growth

by Jung Hyun Kim, Tae Yoon Kim, Bonhyuk Goo and Yeoncheol Park

Toxins 2024, 16(2), 84; https://doi.org/10.3390/toxins16020084 - 4 Feb 2024

Cited by 2 | Viewed by 2766

Abstract

Limited evidence suggests that stimulating adipose-derived stem cells (ASCs) indirectly promotes hair growth. We examined whether bee venom (BV) activated ASCs and whether BV-induced hair growth was facilitated by enhanced growth factor release by ASCs. The induction of the telogen-to-anagen phase was studied [...] Read more.

Limited evidence suggests that stimulating adipose-derived stem cells (ASCs) indirectly promotes hair growth. We examined whether bee venom (BV) activated ASCs and whether BV-induced hair growth was facilitated by enhanced growth factor release by ASCs. The induction of the telogen-to-anagen phase was studied in mice. The underlying mechanism was investigated using organ cultures of mouse vibrissa hair follicles. When BV-treated ASCs were injected subcutaneously into mice, the telogen-to-anagen transition was accelerated and, by day 14, the hair weight increased. Quantitative polymerase chain reaction (qPCR) revealed that BV influenced the expression of several molecules, including growth factors, chemokines, channels, transcription factors, and enzymes. Western blot analysis was employed to verify the protein expression levels of extracellular-signal-regulated kinase (ERK) and phospho-ERK. Both the Boyden chamber experiment and scratch assay confirmed the upregulation of cell migration by BV. Additionally, ASCs secreted higher levels of growth factors after exposure to BV. Following BV therapy, the gene expression levels of alkaline phosphatase (ALP), fibroblast growth factor (FGF)-1 and 6, endothelial cell growth factor, and platelet-derived growth factor (PDGF)-C were upregulated. The findings of this study suggest that bee venom can potentially be utilized as an ASC-preconditioning agent for hair regeneration. Full article

(This article belongs to the Section Animal Venoms)

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14 pages, 1745 KiB

Open AccessEditor’s ChoiceArticle

TDP-43 and Alzheimer’s Disease Pathology in the Brain of a Harbor Porpoise Exposed to the Cyanobacterial Toxin BMAA

by Susanna P. Garamszegi, Daniel J. Brzostowicki, Thomas M. Coyne, Regina T. Vontell and David A. Davis

Toxins 2024, 16(1), 42; https://doi.org/10.3390/toxins16010042 - 12 Jan 2024

Cited by 6 | Viewed by 2583

Abstract

Cetaceans are well-regarded as sentinels for toxin exposure. Emerging studies suggest that cetaceans can also develop neuropathological changes associated with neurodegenerative disease. The occurrence of neuropathology makes cetaceans an ideal species for examining the impact of marine toxins on the brain across the [...] Read more.

Cetaceans are well-regarded as sentinels for toxin exposure. Emerging studies suggest that cetaceans can also develop neuropathological changes associated with neurodegenerative disease. The occurrence of neuropathology makes cetaceans an ideal species for examining the impact of marine toxins on the brain across the lifespan. Here, we describe TAR DNA-binding protein 43 (TDP-43) proteinopathy and Alzheimer’s disease (AD) neuropathological changes in a beached harbor porpoise (Phocoena phocoena) that was exposed to a toxin produced by cyanobacteria called β-N-methylamino-L-alanine (BMAA). We found pathogenic TDP-43 cytoplasmic inclusions in neurons throughout the cerebral cortex, midbrain and brainstem. P62/sequestosome-1, responsible for the autophagy of misfolded proteins, was observed in the amygdala, hippocampus and frontal cortex. Genes implicated in AD and TDP-43 neuropathology such as APP and TARDBP were expressed in the brain. AD neuropathological changes such as amyloid-β plaques, neurofibrillary tangles, granulovacuolar degeneration and Hirano bodies were present in the hippocampus. These findings further support the development of progressive neurodegenerative disease in cetaceans and a potential causative link to cyanobacterial toxins. Climate change, nutrient pollution and industrial waste are increasing the frequency of harmful cyanobacterial blooms. Cyanotoxins like BMAA that are associated with neurodegenerative disease pose an increasing public health risk. Full article

(This article belongs to the Section Marine and Freshwater Toxins)

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16 pages, 767 KiB

Open AccessEditor’s ChoiceArticle

A Randomized, Double-Blind, Active Control, Multicenter, Phase 3 Study to Evaluate the Efficacy and Safety of Liztox^® versus Botox^® in Post-Stroke Upper Limb Spasticity

by Dong Hyun Ye, Min Ho Chun, Yoon Ghil Park, Nam-Jong Paik, Shi-Uk Lee, Seung Don Yoo and Deog Young Kim

Toxins 2023, 15(12), 697; https://doi.org/10.3390/toxins15120697 - 12 Dec 2023

Cited by 1 | Viewed by 2173

Abstract

Botulinum toxin type A (BTX-A) injection is a commonly used therapeutic intervention for upper limb spasticity in stroke patients. This study was designed as a randomized, active-drug-controlled, double-blind, multicenter, phase 3 clinical trial to evaluate the safety and efficacy of Liztox^® in [...] Read more.

Botulinum toxin type A (BTX-A) injection is a commonly used therapeutic intervention for upper limb spasticity in stroke patients. This study was designed as a randomized, active-drug-controlled, double-blind, multicenter, phase 3 clinical trial to evaluate the safety and efficacy of Liztox^® in comparison to onabotulinum toxin A (Botox^®) for individuals with post-stroke upper limb spasticity. The primary outcome was the alteration in wrist flexor muscle tone from the initial assessment to the fourth week, evaluated using the modified Ashworth scale (MAS). Secondary outcomes included MAS score changes for the wrist at weeks 8 and 12 from baseline; MAS score changes for finger and elbow flexors; and changes in the Disability Assessment Scale (DAS), Subject’s Global Assessment (SGA), the Investigator’s Global Assessment (IGA), and Caregiver Burden Scale (CBS) at weeks 4, 8, and 12 from baseline. The MAS score for wrist flexor spasticity decreased by −1.14 ± 0.59 in the Liztox^® group and −1.22 ± 0.59 in the Botox^® group from baseline to week 4. The difference [97.5% confidence interval (CI)] between the test and control groups was 0.08 [−∞, 0.26], confirming the non-inferiority of the test group compared to the control group. Furthermore, there were consistent improvements in the IGA, SGA, and CBS scores across all assessment intervals, with no statistically significant variances detected between the two groups. No safety-related concerns were reported during the study. In conclusion, Liztox^® injection proved to be a secure and efficacious intervention for managing upper extremity spasticity in post-stroke patients. Full article

(This article belongs to the Special Issue Botulinum Toxins Injections for Hypertonic Limbs—Is It So Effective?)

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17 pages, 1783 KiB

Open AccessEditor’s ChoiceArticle

Comparative Analysis of Secondary Metabolites Produced by Ascochyta fabae under In Vitro Conditions and Their Phytotoxicity on the Primary Host, Vicia faba, and Related Legume Crops

by Eleonora Barilli, Pierluigi Reveglia, Francisco J. Agudo-Jurado, Vanessa Cañete García, Alessio Cimmino, Antonio Evidente and Diego Rubiales

Toxins 2023, 15(12), 693; https://doi.org/10.3390/toxins15120693 - 9 Dec 2023

Viewed by 2270

Abstract

Ascochyta blight, caused by Ascochyta fabae, poses a significant threat to faba bean and other legumes worldwide. Necrotic lesions on stems, leaves, and pods characterize the disease. Given the economic impact of this pathogen and the potential involvement of secondary metabolites in [...] Read more.

Ascochyta blight, caused by Ascochyta fabae, poses a significant threat to faba bean and other legumes worldwide. Necrotic lesions on stems, leaves, and pods characterize the disease. Given the economic impact of this pathogen and the potential involvement of secondary metabolites in symptom development, a study was conducted to investigate the fungus’s ability to produce bioactive metabolites that might contribute to its pathogenicity. For this investigation, the fungus was cultured in three substrates (Czapek-Dox, PDB, and rice). The produced metabolites were analyzed by NMR and LC-HRMS methods, resulting in the dereplication of seven metabolites, which varied with the cultural substrates. Ascochlorin, ascofuranol, and (R)-mevalonolactone were isolated from the Czapek-Dox extract; ascosalipyrone, benzoic acid, and tyrosol from the PDB extract; and ascosalitoxin and ascosalipyrone from the rice extract. The phytotoxicity of the pure metabolites was assessed at different concentrations on their primary hosts and related legumes. The fungal exudates displayed varying degrees of phytotoxicity, with the Czapek-Dox medium’s exudate exhibiting the highest activity across almost all legumes tested. The species belonging to the genus Vicia spp. were the most susceptible, with faba bean being susceptible to all metabolites, at least at the highest concentration tested, as expected. In particular, ascosalitoxin and benzoic acid were the most phytotoxic in the tested condition and, as a consequence, expected to play an important role on necrosis’s appearance. Full article

(This article belongs to the Special Issue Biological Activities and Potential Applications of Phytotoxins)

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17 pages, 1087 KiB

Open AccessEditor’s ChoiceArticle

Turmeric Powder Counteracts Oxidative Stress and Reduces AFB1 Content in the Liver of Broilers Exposed to the EU Maximum Levels of the Mycotoxin

by Neenu Amminikutty, Veronica Spalenza, Watanya Jarriyawattanachaikul, Paola Badino, Maria Teresa Capucchio, Elena Colombino, Achille Schiavone, Donato Greco, Vito D’Ascanio, Giuseppina Avantaggiato, Sihem Dabbou, Carlo Nebbia and Flavia Girolami

Toxins 2023, 15(12), 687; https://doi.org/10.3390/toxins15120687 - 7 Dec 2023

Cited by 2 | Viewed by 2123

Abstract

The most frequent adverse effects of AFB1 in chicken are low performance, the depression of the immune system, and a reduced quality of both eggs and meat, leading to economic losses. Since oxidative stress plays a major role in AFB1 toxicity, natural products [...] Read more.

The most frequent adverse effects of AFB1 in chicken are low performance, the depression of the immune system, and a reduced quality of both eggs and meat, leading to economic losses. Since oxidative stress plays a major role in AFB1 toxicity, natural products are increasingly being used as an alternative to mineral binders to tackle AFB1 toxicosis in farm animals. In this study, an in vivo trial was performed by exposing broilers for 10 days to AFB1 at dietary concentrations approaching the maximum limits set by the EU (0.02 mg/kg feed) in the presence or absence of turmeric powder (TP) (included in the feed at 400 mg/kg). The aims were to evaluate (i) the effects of AFB1 on lipid peroxidation, antioxidant parameters, histology, and the expression of drug transporters and biotransformation enzymes in the liver; (ii) the hepatic accumulation of AFB1 and its main metabolites (assessed using an in-house-validated HPLC-FLD method); (iii) the possible modulation of the above parameters elicited by TP. Broilers exposed to AFB1 alone displayed a significant increase in lipid peroxidation in the liver, which was completely reverted by the concomitant administration of TP. Although no changes in glutathione levels and antioxidant enzyme activities were detected in any treatment group, AFB1 significantly upregulated and downregulated the mRNA expression of CYP2A6 and Nrf2, respectively. TP counteracted such negative effects and increased the hepatic gene expression of selected antioxidant enzymes (i.e., CAT and SOD2) and drug transporters (i.e., ABCG2), which were further enhanced in combination with AFB1. Moreover, both AFB1 and TP increased the mRNA levels of ABCC2 and ABCG2 in the duodenum. The latter changes might be implicated in the decrease in hepatic AFB1 to undetectable levels (<LOD) in the TP supplemented group. Overall, our findings further support the use of TP as an effective feeding strategy to prevent AFB1-related adverse effects in broilers. Full article

(This article belongs to the Special Issue Mycotoxins in Farm Animals: Health Disorders and Preventive Strategies)

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16 pages, 2712 KiB

Open AccessEditor’s ChoiceCommunication

ADDovenom: Thermostable Protein-Based ADDomer Nanoparticles as New Therapeutics for Snakebite Envenoming

by Stefanie K. Menzies, Raquel Arinto-Garcia, Fernanda Gobbi Amorim, Iara Aimê Cardoso, Camille Abada, Thomas Crasset, Fabien Durbesson, Rebecca J. Edge, Priscila El-Kazzi, Sophie Hall, Damien Redureau, Richard Stenner, Johara Boldrini-França, Huan Sun, António Roldão, Paula M. Alves, Robert A. Harrison, Renaud Vincentelli, Imre Berger, Loïc Quinton, Nicholas R. Casewell and Christiane Schaffitzel Show full author list Hide full author list

Toxins 2023, 15(12), 673; https://doi.org/10.3390/toxins15120673 - 28 Nov 2023

Cited by 1 | Viewed by 2458

Abstract

Snakebite envenoming can be a life-threatening medical emergency that requires prompt medical intervention to neutralise the effects of venom toxins. Each year up to 138,000 people die from snakebites and threefold more victims suffer life-altering disabilities. The current treatment of snakebite relies solely [...] Read more.

Snakebite envenoming can be a life-threatening medical emergency that requires prompt medical intervention to neutralise the effects of venom toxins. Each year up to 138,000 people die from snakebites and threefold more victims suffer life-altering disabilities. The current treatment of snakebite relies solely on antivenom—polyclonal antibodies isolated from the plasma of hyperimmunised animals—which is associated with numerous deficiencies. The ADDovenom project seeks to deliver a novel snakebite therapy, through the use of an innovative protein-based scaffold as a next-generation antivenom. The ADDomer is a megadalton-sized, thermostable synthetic nanoparticle derived from the adenovirus penton base protein; it has 60 high-avidity binding sites to neutralise venom toxins. Here, we outline our experimental strategies to achieve this goal using state-of-the-art protein engineering, expression technology and mass spectrometry, as well as in vitro and in vivo venom neutralisation assays. We anticipate that the approaches described here will produce antivenom with unparalleled efficacy, safety and affordability. Full article

(This article belongs to the Section Animal Venoms)

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20 pages, 10566 KiB

Open AccessEditor’s ChoiceArticle

An Assessment of Potential Threats to Human Health from Algae Blooms in the Indian River Lagoon (USA) 2018–2021: Unique Patterns of Cytotoxicity Associated with Toxins

by Esther A. Guzmán, Tara A. Peterson, Priscilla L. Winder, Kirstie T. Francis, Malcolm McFarland, Jill C. Roberts, Jennifer Sandle and Amy E. Wright

Toxins 2023, 15(11), 664; https://doi.org/10.3390/toxins15110664 - 17 Nov 2023

Viewed by 3938

Abstract

The Indian River Lagoon (IRL), a 156-mile-long estuary located on the eastern coast of Florida, experiences phytoplankton bloom events due to increased seasonal temperatures coupled with anthropogenic impacts. This study aimed to gather data on the toxicity to human cells and to identify [...] Read more.

The Indian River Lagoon (IRL), a 156-mile-long estuary located on the eastern coast of Florida, experiences phytoplankton bloom events due to increased seasonal temperatures coupled with anthropogenic impacts. This study aimed to gather data on the toxicity to human cells and to identify secondary metabolites found in water samples collected in the IRL. Water samples from 20 sites of the IRL were collected during the wet and dry seasons over a three-year period. A panel of cell lines was used to test cytotoxicity. Hemagglutination, hemolysis, and inhibition of protein phosphatase 2A (PP2A) were also measured. Cytotoxic blooms were seen both in the south (Microcystis) and the north (Pyrodinium) of the IRL. Each toxin induced a consistent pattern of cytotoxicity in the panel of human cell lines assayed. During blooms, cytotoxicity due to a single type of toxin is obvious from this pattern. In the absence of blooms, the cytotoxicity seen reflected either a mixture of toxins or it was caused by an unidentified toxin. These observations suggest that other toxins with the potential to be harmful to human health may be present in the IRL. Moreover, the presence of toxins in the IRL is not always associated with blooms of known toxin-producing organisms. Full article

(This article belongs to the Section Marine and Freshwater Toxins)

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11 pages, 1922 KiB

Open AccessEditor’s ChoiceArticle

Synergistic Effects of Essential Oils and Organic Acids against Aspergillus flavus Contamination in Poultry Feed

by Tim Satterlee, Callie Megan McDonough, Scott E. Gold, Chongxiao Chen, Anthony E. Glenn and Anthony Pokoo-Aikins

Toxins 2023, 15(11), 635; https://doi.org/10.3390/toxins15110635 - 31 Oct 2023

Cited by 2 | Viewed by 2050

Abstract

Organic acids and essential oils are commonly used in the poultry industry as antimicrobials and for their beneficial effects on gut health, growth performance, and meat quality. A common postharvest storage fungal colonist, Aspergillus flavus, contaminates corn, the primary component of poultry [...] Read more.

Organic acids and essential oils are commonly used in the poultry industry as antimicrobials and for their beneficial effects on gut health, growth performance, and meat quality. A common postharvest storage fungal colonist, Aspergillus flavus, contaminates corn, the primary component of poultry feed, with the highly detrimental mycotoxin, aflatoxin. Aflatoxin adversely affects poultry feed intake, feed conversion efficiency, weight gain, egg production, fertility, hatchability, and poultry meat yield. Both organic acids and essential oils have been reported to inhibit the growth of A. flavus. Thus, we evaluated if the inhibitory synergy between combined essential oils (cinnamon, lemongrass, and oregano) and organic acids (acetic, butyric, and propionic) prevents A. flavus growth. The study confirmed that these compounds inhibit the growth of A. flavus and that synergistic interactions do occur between some of them. Overall, cinnamon oil was shown to have the highest synergy with all the organic acids tested, requiring 1000 µL/L air of cinnamon oil and 888 mg/kg of butyric acid to fully suppress A. flavus growth on corn kernels. With the strong synergism demonstrated, combining certain essential oils and organic acids offers a potentially effective natural method for controlling postharvest aflatoxin contamination in poultry feed. Full article

(This article belongs to the Section Mycotoxins)

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15 pages, 2959 KiB

Open AccessEditor’s ChoiceArticle

Ensemble Machine Learning of Gradient Boosting (XGBoost, LightGBM, CatBoost) and Attention-Based CNN-LSTM for Harmful Algal Blooms Forecasting

by Jung Min Ahn, Jungwook Kim and Kyunghyun Kim

Toxins 2023, 15(10), 608; https://doi.org/10.3390/toxins15100608 - 10 Oct 2023

Cited by 19 | Viewed by 3579

Abstract

Harmful algal blooms (HABs) are a serious threat to ecosystems and human health. The accurate prediction of HABs is crucial for their proactive preparation and management. While mechanism-based numerical modeling, such as the Environmental Fluid Dynamics Code (EFDC), has been widely used in [...] Read more.

Harmful algal blooms (HABs) are a serious threat to ecosystems and human health. The accurate prediction of HABs is crucial for their proactive preparation and management. While mechanism-based numerical modeling, such as the Environmental Fluid Dynamics Code (EFDC), has been widely used in the past, the recent development of machine learning technology with data-based processing capabilities has opened up new possibilities for HABs prediction. In this study, we developed and evaluated two types of machine learning-based models for HABs prediction: Gradient Boosting models (XGBoost, LightGBM, CatBoost) and attention-based CNN-LSTM models. We used Bayesian optimization techniques for hyperparameter tuning, and applied bagging and stacking ensemble techniques to obtain the final prediction results. The final prediction result was derived by applying the optimal hyperparameter and bagging and stacking ensemble techniques, and the applicability of prediction to HABs was evaluated. When predicting HABs with an ensemble technique, it is judged that the overall prediction performance can be improved by complementing the advantages of each model and averaging errors such as overfitting of individual models. Our study highlights the potential of machine learning-based models for HABs prediction and emphasizes the need to incorporate the latest technology into this important field. Full article

(This article belongs to the Section Marine and Freshwater Toxins)

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17 pages, 1968 KiB

Open AccessEditor’s ChoiceArticle

Discovery of an Insect Neuroactive Helix Ring Peptide from Ant Venom

by Valentine Barassé, Laurence Jouvensal, Guillaume Boy, Arnaud Billet, Steven Ascoët, Benjamin Lefranc, Jérôme Leprince, Alain Dejean, Virginie Lacotte, Isabelle Rahioui, Catherine Sivignon, Karen Gaget, Mélanie Ribeiro Lopes, Federica Calevro, Pedro Da Silva, Karine Loth, Françoise Paquet, Michel Treilhou, Elsa Bonnafé and Axel Touchard

Toxins 2023, 15(10), 600; https://doi.org/10.3390/toxins15100600 - 5 Oct 2023

Cited by 2 | Viewed by 2225

Abstract

Ants are among the most abundant terrestrial invertebrate predators on Earth. To overwhelm their prey, they employ several remarkable behavioral, physiological, and biochemical innovations, including an effective paralytic venom. Ant venoms are thus cocktails of toxins finely tuned to disrupt the physiological systems [...] Read more.

Ants are among the most abundant terrestrial invertebrate predators on Earth. To overwhelm their prey, they employ several remarkable behavioral, physiological, and biochemical innovations, including an effective paralytic venom. Ant venoms are thus cocktails of toxins finely tuned to disrupt the physiological systems of insect prey. They have received little attention yet hold great promise for the discovery of novel insecticidal molecules. To identify insect-neurotoxins from ant venoms, we screened the paralytic activity on blowflies of nine synthetic peptides previously characterized in the venom of Tetramorium bicarinatum. We selected peptide U₁₁, a 34-amino acid peptide, for further insecticidal, structural, and pharmacological experiments. Insecticidal assays revealed that U₁₁ is one of the most paralytic peptides ever reported from ant venoms against blowflies and is also capable of paralyzing honeybees. An NMR spectroscopy of U₁₁ uncovered a unique scaffold, featuring a compact triangular ring helix structure stabilized by a single disulfide bond. Pharmacological assays using Drosophila S2 cells demonstrated that U₁₁ is not cytotoxic, but suggest that it may modulate potassium conductance, which structural data seem to corroborate and will be confirmed in a future extended pharmacological investigation. The results described in this paper demonstrate that ant venom is a promising reservoir for the discovery of neuroactive insecticidal peptides. Full article

(This article belongs to the Special Issue Ant Venom)

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17 pages, 1558 KiB

Open AccessEditor’s ChoiceArticle

Use of Yeast Cell Wall Extract for Growing Pigs Consuming Feed Contaminated with Mycotoxins below or above Regulatory Guidelines: A Meta-Analysis with Meta-Regression

by Alexandra C. Weaver, Daniel M. Weaver, Nicholas Adams and Alexandros Yiannikouris

Toxins 2023, 15(10), 596; https://doi.org/10.3390/toxins15100596 - 3 Oct 2023

Cited by 2 | Viewed by 2821

Abstract

Using a random-effects meta-analysis, the performance of growing pigs under a mycotoxin challenge (MT) with or without supplementation of yeast cell wall extract (YCWE, Mycosorb^®, Alltech Inc.) was evaluated. Both MT and YCWE were also compared to animal controls not receiving [...] Read more.

Using a random-effects meta-analysis, the performance of growing pigs under a mycotoxin challenge (MT) with or without supplementation of yeast cell wall extract (YCWE, Mycosorb^®, Alltech Inc.) was evaluated. Both MT and YCWE were also compared to animal controls not receiving mycotoxins (CTRL). Meta-regression was used to further explore the impacts of MT at/below (category 1) or above (category 2) global regulatory guidelines. Following the screening, 23 suitable references (30 mycotoxin treatments) were used. Overall, MT lowered average daily gain (ADG, p < 0.001) and average daily feed intake (ADFI, p < 0.0001) from CTRL by −84 and −165 g, respectively. Inclusion of YCWE during mycotoxin challenges (YCWE+MT, average 2.1 kg/ton) tended to result in greater ADG (+17 g, p = 0.068) compared to MT treatments. The gain-to-feed ratio (G:F) was not impacted by MT or YCWE+MT. Further investigation by meta-regression revealed that pigs fed MT in category 1 had lower ADG (−78.5 g, p < 0.001) versus CTRL, while YCWE+MT had higher ADG (+48 g, p < 0.001) over MT and was similar to CTRL. The ADFI was not impacted, although YCWE+MT had ADFI values similar to the CTRL. In category 2, ADG and ADFI of pigs fed MT were lower than CTRL (−85.1 and −166 g, respectively, p < 0.0001), with a tendency for YCWE+MT to result in higher ADFI (+25.3 g, p = 0.062). In summary, the inclusion of YCWE provided benefits to performance during common mycotoxin challenge levels (at or below regulatory guidelines). Full article

(This article belongs to the Special Issue Current Research on Mycotoxins in Food and Feed: From Detection and Unravelling of Toxicity to Control)

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14 pages, 3931 KiB

Open AccessEditor’s ChoiceArticle

Presence, Co-Occurrence, and Daily Intake Estimates of Aflatoxins and Fumonisins in Maize Consumed in Food-Insecure Regions of Western Honduras

by Luis Sabillón, Jackeline Alvarado, Alejandra Leiva, Rodrigo Mendoza, Raúl Espinal, John F. Leslie and Andréia Bianchini

Toxins 2023, 15(9), 559; https://doi.org/10.3390/toxins15090559 - 7 Sep 2023

Cited by 1 | Viewed by 1784

Abstract

Foodborne mycotoxins are a significant food safety risk in developing countries. Our objective was to determine the occurrence of and exposure levels to aflatoxins (AFs) and fumonisins (FBs) in maize intended for human and animal consumption in food-insecure regions of western Honduras. Total [...] Read more.

Foodborne mycotoxins are a significant food safety risk in developing countries. Our objective was to determine the occurrence of and exposure levels to aflatoxins (AFs) and fumonisins (FBs) in maize intended for human and animal consumption in food-insecure regions of western Honduras. Total AFs and FBs were quantified with a monoclonal antibody-based affinity spectrofluorimetric method. FBs were detected in 614/631 samples of maize destined for human consumption at 0.3 to 41 mg/kg (mean, 2.7 mg/kg). Of the 614 positive samples, 147 had FB levels exceeding the U.S. Food and Drug Administration (FDA) advisory threshold of 4.0 mg/kg. AFs were detected in 109/631 samples of maize for human consumption with concentrations between 1.0 and 490 µg/kg (mean, 10 µg/kg). AF levels in 34 samples exceeded the FDA regulatory limit (i.e., 20 µg/kg). The average probable daily intake of AFs in western Honduras ranged from 0 to 260 ng/kg body weight/day, and for FBs, the average probable daily intake ranged from 17 to 53 μg/kg body weight/day. AFs and FBs co-occurred in 106/631 samples with 60 samples containing both toxins at levels greater than the FDA regulatory levels. Samples of maize intended for animal feed had significantly higher AF (mean, 22 µg/kg) and FB (mean, 7.6 mg/kg) contamination levels than those observed in samples destined for human consumption. Thus, the maize supply chain in western Honduras is contaminated with mycotoxins at levels that pose health risks to both humans and livestock. More effective mycotoxin surveillance and implementation of effective mitigation strategies are needed to reduce mycotoxin contamination and exposure. Full article

(This article belongs to the Special Issue Occurrence and Determination of Mycotoxins)

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19 pages, 4401 KiB

Open AccessEditor’s ChoiceArticle

Identification of Acid Hydrolysis Metabolites of the Pimelea Toxin Simplexin for Targeted UPLC-MS/MS Analysis

by Zhi Hung Loh, Natasha L. Hungerford, Diane Ouwerkerk, Athol V. Klieve and Mary T. Fletcher

Toxins 2023, 15(9), 551; https://doi.org/10.3390/toxins15090551 - 5 Sep 2023

Cited by 1 | Viewed by 1843

Abstract

Pimelea poisoning of cattle is a unique Australian toxic condition caused by the daphnane orthoester simplexin present in native Pimelea pasture plants. Rumen microorganisms have been proposed to metabolise simplexin by enzymatic reactions, likely at the orthoester and epoxide moieties of simplexin, but [...] Read more.

Pimelea poisoning of cattle is a unique Australian toxic condition caused by the daphnane orthoester simplexin present in native Pimelea pasture plants. Rumen microorganisms have been proposed to metabolise simplexin by enzymatic reactions, likely at the orthoester and epoxide moieties of simplexin, but a metabolic pathway has not been confirmed. This study aimed to investigate this metabolic pathway through the analysis of putative simplexin metabolites. Purified simplexin was hydrolysed with aqueous hydrochloric acid and sulfuric acid to produce target metabolites for UPLC-MS/MS analysis of fermentation fluid samples, bacterial isolate samples, and other biological samples. UPLC-MS/MS analysis identified predicted hydrolysed products from both acid hydrolysis procedures with MS breakdown of these putative products sharing high-resolution accurate mass (HRAM) fragmentation ions with simplexin. However, targeted UPLC-MS/MS analysis of the biological samples failed to detect the H₂SO₄ degradation products, suggesting that the rumen microorganisms were unable to produce similar simplexin degradation products at detectable levels, or that metabolites, once formed, were further metabolised. Overall, in vitro acid hydrolysis was able to hydrolyse simplexin at the orthoester and epoxide functionalities, but targeted UPLC-MS/MS analysis of biological samples did not detect any of the identified simplexin hydrolysis products. Full article

(This article belongs to the Special Issue Toxic Compounds from Poisonous Plants: Identification and Analytical Methods)

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16 pages, 14537 KiB

Open AccessEditor’s ChoiceArticle

Intramuscular Bleeding and Formation of Microthrombi during Skeletal Muscle Damage Caused by a Snake Venom Metalloprotease and a Cardiotoxin

by Medha Sonavane, José R. Almeida, Elanchezhian Rajan, Harry F. Williams, Felix Townsend, Elizabeth Cornish, Robert D. Mitchell, Ketan Patel and Sakthivel Vaiyapuri

Toxins 2023, 15(9), 530; https://doi.org/10.3390/toxins15090530 - 28 Aug 2023

Cited by 1 | Viewed by 2748

Abstract

The interactions between specific snake venom toxins and muscle constituents are the major cause of severe muscle damage that often result in amputations and subsequent socioeconomic ramifications for snakebite victims and/or their families. Therefore, improving our understanding of venom-induced muscle damage and determining [...] Read more.

The interactions between specific snake venom toxins and muscle constituents are the major cause of severe muscle damage that often result in amputations and subsequent socioeconomic ramifications for snakebite victims and/or their families. Therefore, improving our understanding of venom-induced muscle damage and determining the underlying mechanisms of muscle degeneration/regeneration following snakebites is critical to developing better strategies to tackle this issue. Here, we analysed intramuscular bleeding and thrombosis in muscle injuries induced by two different snake venom toxins (CAMP—Crotalus atrox metalloprotease (a PIII metalloprotease from the venom of this snake) and a three-finger toxin (CTX, a cardiotoxin from the venom of Naja pallida)). Classically, these toxins represent diverse scenarios characterised by persistent muscle damage (CAMP) and successful regeneration (CTX) following acute damage, as normally observed in envenomation by most vipers and some elapid snakes of Asian, Australasian, and African origin, respectively. Our immunohistochemical analysis confirmed that both CAMP and CTX induced extensive muscle destruction on day 5, although the effects of CTX were reversed over time. We identified the presence of fibrinogen and P-selectin exposure inside the damaged muscle sections, suggesting signs of bleeding and the formation of platelet aggregates/microthrombi in tissues, respectively. Intriguingly, CAMP causes integrin shedding but does not affect any blood clotting parameters, whereas CTX significantly extends the clotting time and has no impact on integrin shedding. The rates of fibrinogen clearance and reduction in microthrombi were greater in CTX-treated muscle compared to CAMP-treated muscle. Together, these findings reveal novel aspects of venom-induced muscle damage and highlight the relevance of haemostatic events such as bleeding and thrombosis for muscle regeneration and provide useful mechanistic insights for developing better therapeutic interventions. Full article

(This article belongs to the Special Issue Pre-clinical and Clinical Management of Snakebite Envenomation)

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17 pages, 7408 KiB

Open AccessEditor’s ChoiceArticle

A Multi-Layer-Controlled Strategy for Cloning and Expression of Toxin Genes in Escherichia coli

by Jessie Vandierendonck, Yana Girardin, Pieter De Bruyn, Henri De Greve and Remy Loris

Toxins 2023, 15(8), 508; https://doi.org/10.3390/toxins15080508 - 18 Aug 2023

Cited by 3 | Viewed by 2462

Abstract

Molecular cloning and controlled expression remain challenging when the target gene encodes a protein that is toxic to the host. We developed a set of multi-layer control systems to enable cloning of genes encoding proteins known to be highly toxic in Escherichia coli [...] Read more.

Molecular cloning and controlled expression remain challenging when the target gene encodes a protein that is toxic to the host. We developed a set of multi-layer control systems to enable cloning of genes encoding proteins known to be highly toxic in Escherichia coli and other bacteria. The different multi-layer control systems combine a promoter–operator system on a transcriptional level with a riboswitch for translational control. Additionally, replicational control is ensured by using a strain that reduces the plasmid copy number. The use of weaker promoters (such as P_BAD or PfdeA) in combination with the effective theophylline riboswitch is essential for cloning genes that encode notoriously toxic proteins that directly target translation and transcription. Controlled overexpression is possible, allowing the system to be used for evaluating in vivo effects of the toxin. Systems with a stronger promoter can be used for successful overexpression and purification of the desired protein but are limited to toxins that are more moderate and do not interfere with their own production. Full article

(This article belongs to the Section Bacterial Toxins)

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16 pages, 2621 KiB

Open AccessEditor’s ChoiceArticle

A Metalloproteinase Cocktail from the Venom of Protobothrops flavoviridis Cleaves Amyloid Beta Peptides at the α-Cleavage Site

by Eugene Futai, Hajime Kawasaki, Shinichi Sato, Khadija Daoudi, Masafumi Hidaka, Taisuke Tomita and Tomohisa Ogawa

Toxins 2023, 15(8), 500; https://doi.org/10.3390/toxins15080500 - 12 Aug 2023

Cited by 2 | Viewed by 2886

Abstract

A disintegrin and metalloproteinase (ADAM) family proteins are a major class of membrane-anchored multidomain proteinases that are responsible for the shedding of cell surface protein ectodomains, including amyloid precursor protein (APP). Human ADAM 9, 10, and 17 proteolyze APPs and produce non-amyloid-genic p3 [...] Read more.

A disintegrin and metalloproteinase (ADAM) family proteins are a major class of membrane-anchored multidomain proteinases that are responsible for the shedding of cell surface protein ectodomains, including amyloid precursor protein (APP). Human ADAM 9, 10, and 17 proteolyze APPs and produce non-amyloid-genic p3 peptides, instead of neurotoxic amyloid-β peptides (Aβs; Aβ40 and Aβ42), which form fibrils and accumulate in the brain of patients with Alzheimer’s disease (AD). The ADAM family is closely related to snake venom metalloproteinases (SVMPs), which are derived from ancestral ADAMs but act as soluble proteinases. To test the therapeutic potential of SVMPs, we purified SVMPs from Protobothrops flavoviridis venom using metal ion affinity and pooled into a cocktail. Thus, 9 out of 11 SVMPs in the P. flavoviridis genome were identified in the cocktail. SVMPs inhibited Aβ secretion when added to human cell culture medium without affecting APP proteolysis. SVMPs degraded synthetic Aβ40 and Aβ42 peptides at the same cleavage site (α-site of APP) as ADAM9, 10, and 17. SVMPs did not degrade Aβ fibrils but interfered with their formation, assessed using thioflavin-T. Thus, SVMPs have therapeutic potential for AD as an Aβ-degrading protease, and the finding adds to the discovery of bioactive peptides from venoms as novel therapeutics. Full article

(This article belongs to the Special Issue Potential Therapeutic Applications of Animal Venoms and Toxins)

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17 pages, 2440 KiB

Open AccessEditor’s ChoiceArticle

Characterization of Particle-Size-Based Homogeneity and Mycotoxin Distribution Using Laser Diffraction Particle Size Analysis

by Kai Zhang, Ivy Tran and Steven Tan

Toxins 2023, 15(7), 450; https://doi.org/10.3390/toxins15070450 - 6 Jul 2023

Cited by 2 | Viewed by 2696

Abstract

Sample homogeneity dictates whether analyzing a test portion of an entire sample can provide representative information about incurred mycotoxins. In this study, we evaluated particle-size-distribution-based homogeneity of laboratory mycotoxin samples using laser diffraction particle size analysis and International Organization for Standardization (ISO) Guide [...] Read more.

Sample homogeneity dictates whether analyzing a test portion of an entire sample can provide representative information about incurred mycotoxins. In this study, we evaluated particle-size-distribution-based homogeneity of laboratory mycotoxin samples using laser diffraction particle size analysis and International Organization for Standardization (ISO) Guide 35: 2017. Incurred whole corn, compound feed, peanut butter, and wheat flour (500 g each) were comminuted using wet, cryogenic, or dry milling. We used a sample dividing (riffling) device to obtain representative subsamples (25 g each) and developed a laser diffraction particle size analysis procedure by optimizing key parameters such as the refractive index, absorption, and stirring rate. The homogeneity of the particle size distribution within laboratory subsamples was characterized using the optimized laser diffraction procedure. An assessment of homogeneity was also performed for individual mycotoxins in each incurred matrix sample following the procedure described in ISO Guide 35. The concentrations of the incurred mycotoxins were determined using liquid chromatography–mass spectrometry (LC-MS). Within- and between-subsample variances of incurred aflatoxin B1 in peanut butter; deoxynivalenol in corn, compound feed, and wheat flour; and fumonisins in compound feed corroborated that when the particle size measurements were less than 850 µm, mycotoxins concentrations were consistent across independent test portions, which was confirmed using an analysis of variance (F-test). This study highlights the benefits of laser diffraction particle size analysis and suggests its use as a test procedure to evaluate homogeneity in new sample commodities. Full article

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23 pages, 11155 KiB

Open AccessEditor’s ChoiceArticle

Cyanotoxins Increase Cytotoxicity and Promote Nonalcoholic Fatty Liver Disease Progression by Enhancing Cell Steatosis

by Suryakant Niture, Sashi Gadi, Qi Qi, Leslimar Rios-Colon, Sabin Khatiwada, Vandana, Reshan A. Fernando, Keith E. Levine and Deepak Kumar

Toxins 2023, 15(7), 411; https://doi.org/10.3390/toxins15070411 - 25 Jun 2023

Cited by 6 | Viewed by 2290

Abstract

Freshwater prokaryotic cyanobacteria within harmful algal blooms produce cyanotoxins which are considered major pollutants in the aquatic system. Direct exposure to cyanotoxins through inhalation, skin contact, or ingestion of contaminated drinking water can target the liver and may cause hepatotoxicity. In the current [...] Read more.

Freshwater prokaryotic cyanobacteria within harmful algal blooms produce cyanotoxins which are considered major pollutants in the aquatic system. Direct exposure to cyanotoxins through inhalation, skin contact, or ingestion of contaminated drinking water can target the liver and may cause hepatotoxicity. In the current study, we investigated the effect of low concentrations of cyanotoxins on cytotoxicity, inflammation, modulation of unfolded protein response (UPR), steatosis, and fibrosis signaling in human hepatocytes and liver cell models. Exposure to low concentrations of microcystin-LR (MC-LR), microcystin-RR (MC-RR), nodularin (NOD), and cylindrospermopsin (CYN) in human bipotent progenitor cell line HepaRG and hepatocellular carcinoma (HCC) cell lines HepG2 and SK-Hep1 resulted in increased cell toxicity. MC-LR, NOD, and CYN differentially regulated inflammatory signaling, activated UPR signaling and lipogenic gene expression, and induced cellular steatosis and fibrotic signaling in HCC cells. MC-LR, NOD, and CYN also regulated AKT/mTOR signaling and inhibited autophagy. Chronic exposure to MC-LR, NOD, and CYN upregulated the expression of lipogenic and fibrosis biomarkers. Moreover, RNA sequencing (RNA seq) data suggested that exposure of human hepatocytes, HepaRG, and HCC HepG2 cells to MC-LR and CYN modulated expression levels of several genes that regulate non-alcoholic fatty liver disease (NAFLD). Our data suggest that low concentrations of cyanotoxins can cause hepatotoxicity and cell steatosis and promote NAFLD progression. Full article

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31 pages, 7206 KiB

Open AccessEditor’s ChoiceArticle

Diversity of Mycotoxins and Other Secondary Metabolites Recovered from Blood Oranges Infected by Colletotrichum, Alternaria, and Penicillium Species

by Ermes Ivan Rovetto, Carlos Luz, Federico La Spada, Giuseppe Meca, Mario Riolo and Santa Olga Cacciola

Toxins 2023, 15(7), 407; https://doi.org/10.3390/toxins15070407 - 21 Jun 2023

Cited by 10 | Viewed by 2427

Abstract

This study identified secondary metabolites produced by Alternaria alternata, Colletotrichum gloeosporioides, and Penicillium digitatum in fruits of two blood orange cultivars before harvest. Analysis was performed by UHPLC–Q-TOF-MS. Three types of fruits were selected, asymptomatic, symptomatic showing necrotic lesions caused by [...] Read more.

This study identified secondary metabolites produced by Alternaria alternata, Colletotrichum gloeosporioides, and Penicillium digitatum in fruits of two blood orange cultivars before harvest. Analysis was performed by UHPLC–Q-TOF-MS. Three types of fruits were selected, asymptomatic, symptomatic showing necrotic lesions caused by hail, and mummified. Extracts from peel and juice were analyzed separately. Penicillium digitatum was the prevalent species recovered from mummified and hail-injured fruits. Among 47 secondary metabolites identified, 16, 18, and 13 were of A. alternata, C. gloeosporioides, and P. digitatum, respectively. Consistently with isolations, indicating the presence of these fungi also in asymptomatic fruits, the metabolic profiles of the peel of hail-injured and asymptomatic fruits did not differ substantially. Major differences were found in the profiles of juice from hail-injured and mummified fruits, such as a significant higher presence of 5,4-dihydroxy-3,7,8-trimethoxy-6C-methylflavone and Atrovenetin, particularly in the juice of mummified fruits of the Tarocco Lempso cultivar. Moreover, the mycotoxins patulin and Rubratoxin B were detected exclusively in mummified fruits. Patulin was detected in both the juice and peel, with a higher relative abundance in the juice, while Rubratoxin B was detected only in the juice. These findings provide basic information for evaluating and preventing the risk of contamination by mycotoxins in the citrus fresh fruit supply chain and juice industry. Full article

(This article belongs to the Section Mycotoxins)

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23 pages, 5583 KiB

Open AccessEditor’s ChoiceArticle

The Clostridium botulinum C2 Toxin Subunit C2IIa Delivers Enzymes with Positively Charged N-Termini into the Cytosol of Target Cells

by Sebastian Heber, Joscha Borho, Nicole Stadler, Fanny Wondany, Irina König, Jens Michaelis, Panagiotis Papatheodorou, Holger Barth and Maximilian Fellermann

Toxins 2023, 15(6), 390; https://doi.org/10.3390/toxins15060390 - 9 Jun 2023

Cited by 4 | Viewed by 2233

Abstract

The binary Clostridium (C.) botulinum C2 toxin consists of two non-linked proteins. The proteolytically activated binding/transport subunit C2IIa forms barrel-shaped homoheptamers, which bind to cell surface receptors, mediate endocytosis, and translocate the enzyme subunit C2I into the cytosol of target cells. [...] Read more.

The binary Clostridium (C.) botulinum C2 toxin consists of two non-linked proteins. The proteolytically activated binding/transport subunit C2IIa forms barrel-shaped homoheptamers, which bind to cell surface receptors, mediate endocytosis, and translocate the enzyme subunit C2I into the cytosol of target cells. Here, we investigate whether C2IIa can be harnessed as a transporter for proteins/enzymes fused to polycationic tags, as earlier demonstrated for the related anthrax toxin transport subunit PA₆₃. To test C2IIa-mediated transport in cultured cells, reporter enzymes are generated by fusing different polycationic tags to the N- or C-terminus of other bacterial toxins’ catalytic A subunits. C2IIa as well as PA₆₃ deliver N-terminally polyhistidine-tagged proteins more efficiently compared to C-terminally tagged ones. However, in contrast to PA₆₃, C2IIa does not efficiently deliver polylysine-tagged proteins into the cytosol of target cells. Moreover, untagged enzymes with a native cationic N-terminus are efficiently transported by both C2IIa and PA₆₃. In conclusion, the C2IIa-transporter serves as a transport system for enzymes that harbor positively charged amino acids at their N-terminus. The charge distribution at the N-terminus of cargo proteins and their ability to unfold in the endosome and subsequently refold in the cytosol determine transport feasibility and efficiency. Full article

(This article belongs to the Special Issue Intracellular Transport of Toxins: Insights into Mechanisms of Cytotoxicity and Applications in Cell Biology and Medicine)

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15 pages, 7662 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Domperidone Protects Cells from Intoxication with Clostridioides difficile Toxins by Inhibiting Hsp70-Assisted Membrane Translocation

by Maria Braune-Yan, Jinfang Jia, Mary Wahba, Johannes Schmid, Panagiotis Papatheodorou, Holger Barth and Katharina Ernst

Toxins 2023, 15(6), 384; https://doi.org/10.3390/toxins15060384 - 7 Jun 2023

Cited by 2 | Viewed by 2007

Abstract

Clostridioides difficile infections cause severe symptoms ranging from diarrhea to pseudomembranous colitis due to the secretion of AB-toxins, TcdA and TcdB. Both toxins are taken up into cells through receptor-mediated endocytosis, autoproteolytic processing and translocation of their enzyme domains from acidified endosomes into [...] Read more.

Clostridioides difficile infections cause severe symptoms ranging from diarrhea to pseudomembranous colitis due to the secretion of AB-toxins, TcdA and TcdB. Both toxins are taken up into cells through receptor-mediated endocytosis, autoproteolytic processing and translocation of their enzyme domains from acidified endosomes into the cytosol. The enzyme domains glucosylate small GTPases such as Rac1, thereby inhibiting processes such as actin cytoskeleton regulation. Here, we demonstrate that specific pharmacological inhibition of Hsp70 activity protected cells from TcdB intoxication. In particular, the established inhibitor VER-155008 and the antiemetic drug domperidone, which was found to be an Hsp70 inhibitor, reduced the number of cells with TcdB-induced intoxication morphology in HeLa, Vero and intestinal CaCo-2 cells. These drugs also decreased the intracellular glucosylation of Rac1 by TcdB. Domperidone did not inhibit TcdB binding to cells or enzymatic activity but did prevent membrane translocation of TcdB’s glucosyltransferase domain into the cytosol. Domperidone also protected cells from intoxication with TcdA as well as CDT toxin produced by hypervirulent strains of Clostridioides difficile. Our results reveal Hsp70 requirement as a new aspect of the cellular uptake mechanism of TcdB and identified Hsp70 as a novel drug target for potential therapeutic strategies required to combat severe Clostridioides difficile infections. Full article

(This article belongs to the Special Issue Toxin-Host Interaction of Clostridium Toxins)

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22 pages, 2662 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

One Size Fits All—Venomics of the Iberian Adder (Vipera seoanei, Lataste 1878) Reveals Low Levels of Venom Variation across Its Distributional Range

by Ignazio Avella, Maik Damm, Inês Freitas, Wolfgang Wüster, Nahla Lucchini, Óscar Zuazo, Roderich D. Süssmuth and Fernando Martínez-Freiría

Toxins 2023, 15(6), 371; https://doi.org/10.3390/toxins15060371 - 1 Jun 2023

Cited by 6 | Viewed by 3039

Abstract

European vipers (genus Vipera) are medically important snakes displaying considerable venom variation, occurring at different levels in this group. The presence of intraspecific venom variation, however, remains understudied in several Vipera species. Vipera seoanei is a venomous snake endemic to the northern [...] Read more.

European vipers (genus Vipera) are medically important snakes displaying considerable venom variation, occurring at different levels in this group. The presence of intraspecific venom variation, however, remains understudied in several Vipera species. Vipera seoanei is a venomous snake endemic to the northern Iberian Peninsula and south-western France, presenting notable phenotypic variation and inhabiting several diverse habitats across its range. We analysed the venoms of 49 adult specimens of V. seoanei from 20 localities across the species’ Iberian distribution. We used a pool of all individual venoms to generate a V. seoanei venom reference proteome, produced SDS-PAGE profiles of all venom samples, and visualised patterns of variation using NMDS. By applying linear regression, we then assessed presence and nature of venom variation between localities, and investigated the effect of 14 predictors (biological, eco-geographic, genetic) on its occurrence. The venom comprised at least 12 different toxin families, of which five (i.e., PLA₂, svSP, DI, snaclec, svMP) accounted for about 75% of the whole proteome. The comparative analyses of the SDS-PAGE venom profiles showed them to be remarkably similar across the sampled localities, suggesting low geographic variability. The regression analyses suggested significant effects of biological and habitat predictors on the little variation we detected across the analysed V. seoanei venoms. Other factors were also significantly associated with the presence/absence of individual bands in the SDS-PAGE profiles. The low levels of venom variability we detected within V. seoanei might be the result of a recent population expansion, or of processes other than directional positive selection. Full article

(This article belongs to the Section Animal Venoms)

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15 pages, 1860 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Next-Generation Sequencing for Venomics: Application of Multi-Enzymatic Limited Digestion for Inventorying the Snake Venom Arsenal

by Fernanda Gobbi Amorim, Damien Redureau, Thomas Crasset, Lou Freuville, Dominique Baiwir, Gabriel Mazzucchelli, Stefanie K. Menzies, Nicholas R. Casewell and Loïc Quinton

Toxins 2023, 15(6), 357; https://doi.org/10.3390/toxins15060357 - 25 May 2023

Cited by 4 | Viewed by 2585

Abstract

To improve the characterization of snake venom protein profiles, we report the application of a new generation of proteomic methodology to deeply characterize complex protein mixtures. The new approach, combining a synergic multi-enzymatic and a time-limited digestion (MELD), is a versatile and straightforward [...] Read more.

To improve the characterization of snake venom protein profiles, we report the application of a new generation of proteomic methodology to deeply characterize complex protein mixtures. The new approach, combining a synergic multi-enzymatic and a time-limited digestion (MELD), is a versatile and straightforward protocol previously developed by our group. The higher number of overlapping peptides generated during MELD increases the quality of downstream peptide sequencing and of protein identification. In this context, this work aims at applying the MELD strategy to a venomics purpose for the first time, and especially for the characterization of snake venoms. We used four venoms as the test models for this proof of concept: two Elapidae (Dendroaspis polylepis and Naja naja) and two Viperidae (Bitis arietans and Echis ocellatus). Each venom was reduced and alkylated before being submitted to two different protocols: the classical bottom-up proteomics strategy including a digestion step with trypsin only, or MELD, which combines the activities of trypsin, Glu-C and chymotrypsin with a limited digestion approach. The resulting samples were then injected on an M-Class chromatographic system, and hyphenated to a Q-Exactive Mass Spectrometer. Toxins and protein identification were performed by Peaks Studio X+. The results show that MELD considerably improves the number of sequenced (de novo) peptides and identified peptides from protein databases, leading to the unambiguous identification of a greater number of toxins and proteins. For each venom, MELD was successful, not only in terms of the identification of the major toxins (increasing of sequence coverage), but also concerning the less abundant cellular components (identification of new groups of proteins). In light of these results, MELD represents a credible methodology to be applied as the next generation of proteomics approaches dedicated to venomic analysis. It may open new perspectives for the sequencing and inventorying of the venom arsenal and should expand global knowledge about venom composition. Full article

(This article belongs to the Special Issue Omics Approaches to Study Toxins)

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34 pages, 3313 KiB

Open AccessEditor’s ChoiceArticle

MicotoXilico: An Interactive Database to Predict Mutagenicity, Genotoxicity, and Carcinogenicity of Mycotoxins

by Josefa Tolosa, Eva Serrano Candelas, José Luis Vallés Pardo, Addel Goya, Salvador Moncho, Rafael Gozalbes and Martina Palomino Schätzlein

Toxins 2023, 15(6), 355; https://doi.org/10.3390/toxins15060355 - 24 May 2023

Cited by 4 | Viewed by 2675

Abstract

Mycotoxins are secondary metabolites produced by certain filamentous fungi. They are common contaminants found in a wide variety of food matrices, thus representing a threat to public health, as they can be carcinogenic, mutagenic, or teratogenic, among other toxic effects. Several hundreds of [...] Read more.

Mycotoxins are secondary metabolites produced by certain filamentous fungi. They are common contaminants found in a wide variety of food matrices, thus representing a threat to public health, as they can be carcinogenic, mutagenic, or teratogenic, among other toxic effects. Several hundreds of mycotoxins have been reported, but only a few of them are regulated, due to the lack of data regarding their toxicity and mechanisms of action. Thus, a more comprehensive evaluation of the toxicity of mycotoxins found in foodstuffs is required. In silico toxicology approaches, such as Quantitative Structure–Activity Relationship (QSAR) models, can be used to rapidly assess chemical hazards by predicting different toxicological endpoints. In this work, for the first time, a comprehensive database containing 4360 mycotoxins classified in 170 categories was constructed. Then, specific robust QSAR models for the prediction of mutagenicity, genotoxicity, and carcinogenicity were generated, showing good accuracy, precision, sensitivity, and specificity. It must be highlighted that the developed QSAR models are compliant with the OECD regulatory criteria, and they can be used for regulatory purposes. Finally, all data were integrated into a web server that allows the exploration of the mycotoxin database and toxicity prediction. In conclusion, the developed tool is a valuable resource for scientists, industry, and regulatory agencies to screen the mutagenicity, genotoxicity, and carcinogenicity of non-regulated mycotoxins. Full article

(This article belongs to the Special Issue Mycotoxins and Fungal Toxins: Current Status and Future Perspectives)

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13 pages, 2421 KiB

Open AccessEditor’s ChoiceArticle

Bioactives Overproduction through Operational Strategies in the Ichthyotoxic Microalga Heterosigma akashiwo Culture

by Adrián Macías-de la Rosa, Miguel Ángel González-Cardoso, María del Carmen Cerón-García, Lorenzo López-Rosales, Juan José Gallardo-Rodríguez, Sergio Seoane, Asterio Sánchez-Mirón and Francisco García-Camacho

Toxins 2023, 15(5), 349; https://doi.org/10.3390/toxins15050349 - 20 May 2023

Cited by 2 | Viewed by 1920

Abstract

The red tide-forming microalga Heterosigma akashiwo has been associated with massive events of fish deaths, both wild and cultured. Culture conditions are responsible for the synthesis or accumulation of some metabolites with different interesting bioactivities. H. akashiwo LC269919 strain was grown in a [...] Read more.

The red tide-forming microalga Heterosigma akashiwo has been associated with massive events of fish deaths, both wild and cultured. Culture conditions are responsible for the synthesis or accumulation of some metabolites with different interesting bioactivities. H. akashiwo LC269919 strain was grown in a 10 L bubble column photobioreactor artificially illuminated with multi-coloured LED lights. Growth and production of exopolysaccharides, polyunsaturated fatty acids (PUFAs), and carotenoids were evaluated under different culture modes (batch, fed-batch, semicontinuous, and continuous) at two irradiance levels (300 and 700 µE·s⁻¹·m⁻²). Continuous mode at the dilution rate of 0.2·day⁻¹ and 700 µE·s⁻¹·m⁻² provided the highest production of biomass, PUFAs (132.6 and 2.3 mg·L⁻¹·day⁻¹), and maximum fucoxanthin productivity (0.16 mg·L⁻¹·day⁻¹). The fed-batch mode accumulated exopolysaccharides in a concentration (1.02 g·L⁻¹) 10-fold over the batch mode. An extraction process based on a sequential gradient partition with water and four water-immiscible organic solvents allowed the isolation of bioactive fucoxanthin from methanolic extracts of H. akashiwo. Metabolites present in H. akashiwo, fucoxanthin and polar lipids (i.e., eicosapentaenoic acid (EPA)), or probably such as phytosterol (β-Sitosterol) from other microalgae, were responsible for the antitumor activity obtained. Full article

(This article belongs to the Section Marine and Freshwater Toxins)

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16 pages, 2106 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

A Three-Monoclonal Antibody Combination Potently Neutralizes BoNT/G Toxin in Mice

by Yongfeng Fan, Jianlong Lou, Christina C. Tam, Weihua Wen, Fraser Conrad, Priscila Leal da Silva Alves, Luisa W. Cheng, Consuelo Garcia-Rodriguez, Shauna Farr-Jones and James D. Marks

Toxins 2023, 15(5), 316; https://doi.org/10.3390/toxins15050316 - 30 Apr 2023

Cited by 5 | Viewed by 2073

Abstract

Equine-derived antitoxin (BAT^®) is the only treatment for botulism from botulinum neurotoxin serotype G (BoNT/G). BAT^® is a foreign protein with potentially severe adverse effects and is not renewable. To develop a safe, more potent, and renewable antitoxin, humanized monoclonal [...] Read more.

Equine-derived antitoxin (BAT^®) is the only treatment for botulism from botulinum neurotoxin serotype G (BoNT/G). BAT^® is a foreign protein with potentially severe adverse effects and is not renewable. To develop a safe, more potent, and renewable antitoxin, humanized monoclonal antibodies (mAbs) were generated. Yeast displayed single chain Fv (scFv) libraries were prepared from mice immunized with BoNT/G and BoNT/G domains and screened with BoNT/G using fluorescence-activated cell sorting (FACS). Fourteen scFv-binding BoNT/G were isolated with K_D values ranging from 3.86 nM to 103 nM (median K_D 20.9 nM). Five mAb-binding non-overlapping epitopes were humanized and affinity matured to create antibodies hu6G6.2, hu6G7.2, hu6G9.1, hu6G10, and hu6G11.2, with IgG K_D values ranging from 51 pM to 8 pM. Three IgG combinations completely protected mice challenged with 10,000 LD₅₀s of BoNT/G at a total mAb dose of 6.25 μg per mouse. The mAb combinations have the potential for use in the diagnosis and treatment of botulism due to serotype G and, along with antibody combinations to BoNT/A, B, C, D, E, and F, provide the basis for a fully recombinant heptavalent botulinum antitoxin to replace the legacy equine product. Full article

(This article belongs to the Special Issue Human Antibody Engineering for Prevention and Treatment of Botulism)

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10 pages, 1805 KiB

Open AccessEditor’s ChoiceArticle

The Co-Occurrence of T-2 Toxin, Deoxynivalenol, and Fumonisin B1 Activated the Glutathione Redox System in the EU-Limiting Doses in Laying Hens

by Szabina Kulcsár, Benjámin Kövesi, Krisztián Balogh, Erika Zándoki, Zsolt Ancsin, Márta Erdélyi and Miklós Mézes

Toxins 2023, 15(5), 305; https://doi.org/10.3390/toxins15050305 - 23 Apr 2023

Cited by 9 | Viewed by 1947

Abstract

Different mycotoxins in feed lead to combined exposure, increasing adverse effects on animal health. Trichothecene mycotoxins have been associated with inducing oxidative stress, which is neutralized by the glutathione system within the antioxidant defense, depending on the dose and duration of exposure. T-2 [...] Read more.

Different mycotoxins in feed lead to combined exposure, increasing adverse effects on animal health. Trichothecene mycotoxins have been associated with inducing oxidative stress, which is neutralized by the glutathione system within the antioxidant defense, depending on the dose and duration of exposure. T-2 toxin, deoxynivalenol (DON), and fumonisin B1 (FB1) are commonly found in feed commodities simultaneously. In the present study, the intracellular biochemical and gene expression changes were investigated in the case of multi-mycotoxin exposure, focusing on certain elements of the glutathione redox system. In a short-term feeding trial, an in vivo study was performed with low (EU-proposed) doses: T-2/HT-2 toxin: 0.25 mg; DON/2-AcDON/15-AcDON.: 5 mg; FB1: 20 mg/kg feed, and high doses (twice the low dose) in laying hens. The multi-mycotoxin exposure affected the glutathione system; GSH concentration and GPx activity was higher in the liver in the low-dose group on day 1 compared to the control. Furthermore, the gene expression of antioxidant enzymes increased significantly on day 1 in both exposure levels compared to the control. The results suggest that when EU-limiting doses are applied, individual mycotoxins may have a synergistic effect in the induction of oxidative stress. Full article

(This article belongs to the Section Mycotoxins)

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15 pages, 3169 KiB

Open AccessEditor’s ChoiceArticle

Immunomodulatory Effects of Cylindrospermopsin in Human T Cells and Monocytes

by Antonio Casas-Rodríguez, Óscar Cebadero-Dominguez, María Puerto, Ana María Cameán and Angeles Jos

Toxins 2023, 15(4), 301; https://doi.org/10.3390/toxins15040301 - 20 Apr 2023

Cited by 3 | Viewed by 1909

Abstract

Cylindrospermopsin (CYN) is a cyanotoxin with an increasing occurrence, and therefore it is important to elucidate its toxicity profile. CYN has been classified as a cytotoxin, although the scientific literature has already revealed that it affects a wide range of organs and systems. [...] Read more.

Cylindrospermopsin (CYN) is a cyanotoxin with an increasing occurrence, and therefore it is important to elucidate its toxicity profile. CYN has been classified as a cytotoxin, although the scientific literature has already revealed that it affects a wide range of organs and systems. However, research on its potential immunotoxicity is still limited. Thus, this study aimed to evaluate the impact of CYN on two human cell lines representative of the immune system: THP-1 (monocytes) and Jurkat (lymphocytes). CYN reduced cell viability, leading to mean effective concentrations (EC50 24 h) of 6.00 ± 1.04 µM and 5.20 ± 1.20 µM for THP-1 and Jurkat cells, respectively, and induced cell death mainly by apoptosis in both experimental models. Moreover, CYN decreased the differentiation of monocytes to macrophages after 48 h of exposure. In addition, an up-regulation of the mRNA expression of different cytokines, such as interleukin (IL) 2, IL-8, tumor necrosis factor-alpha (TNF-α) and interferon-gamma (INF-γ), was also observed mainly after 24 h exposure in both cell lines. However, only an increase in TNF-α in THP-1 supernatants was observed by ELISA. Overall, these results suggest the immunomodulatory activity of CYN in vitro. Therefore, further research is required to evaluate the impact of CYN on the human immune system. Full article

(This article belongs to the Special Issue Toxicology Research on Cyanotoxins)

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22 pages, 3174 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Application of an Extracellular Matrix-Mimicking Fluorescent Polymer for the Detection of Proteolytic Venom Toxins

by Eric Wachtel, Matyas A. Bittenbinder, Bas van de Velde, Julien Slagboom, Axel de Monts de Savasse, Luis L. Alonso, Nicholas R. Casewell, Freek J. Vonk and Jeroen Kool

Toxins 2023, 15(4), 294; https://doi.org/10.3390/toxins15040294 - 18 Apr 2023

Cited by 1 | Viewed by 2167

Abstract

The cytotoxicity caused by snake venoms is a serious medical problem that greatly contributes to the morbidity observed in snakebite patients. The cytotoxic components found in snake venoms belong to a variety of toxin classes and may cause cytotoxic effects by targeting a [...] Read more.

The cytotoxicity caused by snake venoms is a serious medical problem that greatly contributes to the morbidity observed in snakebite patients. The cytotoxic components found in snake venoms belong to a variety of toxin classes and may cause cytotoxic effects by targeting a range of molecular structures, including cellular membranes, the extracellular matrix (ECM) and the cytoskeleton. Here, we present a high-throughput assay (384-well plate) that monitors ECM degradation by snake venom toxins via the application of fluorescent versions of model ECM substrates, specifically gelatin and collagen type I. Both crude venoms and fractionated toxins of a selection of medically relevant viperid and elapid species, separated via size-exclusion chromatography, were studied using the self-quenching, fluorescently labelled ECM–polymer substrates. The viperid venoms showed significantly higher proteolytic degradation when compared to elapid venoms, although the venoms with higher snake venom metalloproteinase content did not necessarily exhibit stronger substrate degradation than those with a lower one. Gelatin was generally more readily cleaved than collagen type I. In the viperid venoms, which were subjected to fractionation by SEC, two (B. jararaca and C. rhodostoma, respectively) or three (E. ocellatus) active proteases were identified. Therefore, the assay allows the study of proteolytic activity towards the ECM in vitro for crude and fractionated venoms. Full article

(This article belongs to the Special Issue Animal Venoms: Proteomics, Biochemical Activities and Application)

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12 pages, 1938 KiB

Open AccessEditor’s ChoiceArticle

Quantitative and Qualitative Pain Evaluation in Response to OnabotulinumtoxinA for Chronic Migraine: An Observational Real-Life Study

by Claudia Altamura, Nicoletta Brunelli, Giovanna Viticchi, Sergio Salvemini, Gianluca Cecchi, Marilena Marcosano, Luisa Fofi, Mauro Silvestrini and Fabrizio Vernieri

Toxins 2023, 15(4), 284; https://doi.org/10.3390/toxins15040284 - 15 Apr 2023

Cited by 6 | Viewed by 1797

Abstract

(1) Background: Randomized controlled trials and real-life studies demonstrated the efficacy of OnabotulinumtoxinA (OBT-A) for CM prevention. However, no studies specifically addressed its effect on pain’s quantitative intensity and qualitative characteristics. (2) Methods: This is an ambispective study: a post-hoc retrospective analysis of [...] Read more.

(1) Background: Randomized controlled trials and real-life studies demonstrated the efficacy of OnabotulinumtoxinA (OBT-A) for CM prevention. However, no studies specifically addressed its effect on pain’s quantitative intensity and qualitative characteristics. (2) Methods: This is an ambispective study: a post-hoc retrospective analysis of real-life prospectively collected data from two Italian headache centers on CM patients treated with OBT-A over one year (i.e., Cy1-4). The primary endpoint was the changes in pain intensity (Numeric Rating Scale, NRS; the Present Pain Intensity (PPI) scale, the 6-point Behavioral Rating Scale (BRS-6)) and quality scale (the short-form McGill Pain Questionnaire (SF-MPQ)) scores. We also assessed the relationship between changes in intensity and quality of pain and disability scale (MIDAS; HIT-6) scores, monthly headache days (MHDs), and monthly acute medication intake (MAMI) (3) Results: We retrieved 152 cases (51.5 years SD 11.3, 80.3% females). From baseline to Cy-4, MHDs, MAMI, NRS, PPI, and BRS-6 scores decreased (consistently p < 0.001). Only the throbbing (p = 0.004), splitting (p = 0.018), and sickening (p = 0.017) qualities of pain collected in the SF-MPQ were reduced. Score variations in MIDAS related to those in PPI scales (p = 0.035), in the BRS-6 (p = 0.001), and in the NRS (p = 0.003). Similarly, HIT-6 score changes related to PPI score modifications (p = 0.027), in BRS-6 (p = 0.001) and NRS (p = 0.006). Conversely, MAMI variation was not associated with qualitative or quantitative pain score modifications except BRS-6 (p = 0.018). (4) Conclusions: Our study shows that OBT-A alleviates migraine by reducing its impact on multiple aspects, such as frequency, disability, and pain intensity. The beneficial effect on pain intensity seems specific to pain characteristics related to C-fiber transmission and is associated with a reduction in migraine-related disability. Full article

(This article belongs to the Special Issue Botulinum Toxin and Migraine: Goals and Perspectives)

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12 pages, 329 KiB

Open AccessEditor’s ChoiceArticle

Proton-Pump Inhibitors and Serum Concentrations of Uremic Toxins in Patients with Chronic Kidney Disease

by Carolla El Chamieh, Islam Amine Larabi, Solène M. Laville, Christian Jacquelinet, Christian Combe, Denis Fouque, Maurice Laville, Luc Frimat, Roberto Pecoits-Filho, Céline Lange, Bénédicte Stengel, Natalia Alencar De Pinho, Jean-Claude Alvarez, Ziad A. Massy and Sophie Liabeuf

Toxins 2023, 15(4), 276; https://doi.org/10.3390/toxins15040276 - 7 Apr 2023

Cited by 2 | Viewed by 2607

Abstract

Use of proton-pump inhibitors (PPIs) is common in patients with chronic kidney disease (CKD). PPIs and many uremic toxins (UTs) are eliminated by the kidney’s tubular organic anion transporter system. In a cross-sectional study, we sought to evaluate the association between PPI prescription [...] Read more.

Use of proton-pump inhibitors (PPIs) is common in patients with chronic kidney disease (CKD). PPIs and many uremic toxins (UTs) are eliminated by the kidney’s tubular organic anion transporter system. In a cross-sectional study, we sought to evaluate the association between PPI prescription and serum concentrations of various UTs. We studied a randomly selected sub-group of participants in the CKD-REIN cohort (adult patients with a confirmed diagnosis of CKD and estimated glomerular filtration rate (eGFR) < 60 mL/min/1.73 m²) with available frozen samples collected at baseline. PPI prescription was recorded at baseline. Serum concentrations of 10 UTs were measured using a validated liquid chromatography tandem mass spectrometry technique. Multiple linear regression was performed, with the log UT concentration as the dependent variable. Of the 680 included patients (median age: 68 years; median eGFR: 32 mL/min/1.73 m²), 31% had PPI prescriptions at baseline. Patients using PPIs had higher levels of certain UTs in comparison to other patients, including total and free indoxyl sulfate (IS), total and free p-cresylsulfate, total and free p-cresylglucuronide (PCG), phenylacetylglutamine (PAG), free kynurenine, and free hippuric acid. After adjustment for baseline co-morbidities, number of co-prescribed drugs, and laboratory data, including eGFR, associations between PPI prescription and elevated serum concentrations of free and total IS, free and total PCG, and PAG remained significant. Our results indicate that PPI prescription is independently associated with serum UT retention. These findings are interesting to better understand the factors that may modulate serum UT concentration in CKD patients, however, they will need to be confirmed by longitudinal studies. Full article

(This article belongs to the Section Uremic Toxins)

17 pages, 2908 KiB

Open AccessEditor’s ChoiceArticle

Proteomics of Vespa velutina nigrithorax Venom Sac Queens and Workers: A Quantitative SWATH-MS Analysis

by Manuela Alonso-Sampedro, Xesús Feás, Susana Belén Bravo, María Pilar Chantada-Vázquez and Carmen Vidal

Toxins 2023, 15(4), 266; https://doi.org/10.3390/toxins15040266 - 3 Apr 2023

Cited by 4 | Viewed by 3869

Abstract

Health risks caused by stings from Vespa velutina nigrithorax (VV), also known as the yellow-legged Asian hornet, have become a public concern, but little is known about its venom composition. This study presents the proteome profile of the VV’s venom sac (VS) based on [...] Read more.

Health risks caused by stings from Vespa velutina nigrithorax (VV), also known as the yellow-legged Asian hornet, have become a public concern, but little is known about its venom composition. This study presents the proteome profile of the VV’s venom sac (VS) based on Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH-MS). The study also performed proteomic quantitative analysis and examined the biological pathways and molecular functions of the proteins in the VS of VV gynes (i.e., future queens [SQ]) and workers [SW]. The total protein content per VS was significantly higher in the SW than in the SQ (274 ± 54 µg/sac vs. 175 ± 22 µg/sac; p = 0.02). We quantified a total of 228 proteins in the VS, belonging to 7 different classes: Insecta (n = 191); Amphibia and Reptilia (n = 20); Bacilli, γ-Proteobacteria and Pisoniviricetes (n = 12); and Arachnida (n = 5). Among the 228 identified proteins, 66 showed significant differential expression between SQ and SW. The potential allergens hyaluronidase A, venom antigen 5 and phospholipase A1 were significantly downregulated in the SQ venom. Full article

(This article belongs to the Special Issue Advances in Venom Immunology and Allergy)

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13 pages, 11408 KiB

Open AccessEditor’s ChoiceArticle

In Vitro High-Throughput Genotoxicity Testing Using γH2AX Biomarker, Microscopy and Reproducible Automatic Image Analysis in ImageJ—A Pilot Study with Valinomycin

by Bára Křížkovská, Martin Schätz, Jan Lipov, Jitka Viktorová and Eva Jablonská

Toxins 2023, 15(4), 263; https://doi.org/10.3390/toxins15040263 - 1 Apr 2023

Cited by 1 | Viewed by 1732

Abstract

(1) Background: The detection of DNA double-strand breaks in vitro using the phosphorylated histone biomarker (γH2AX) is an increasingly popular method of measuring in vitro genotoxicity, as it is sensitive, specific and suitable for high-throughput analysis. The γH2AX response is either detected by [...] Read more.

(1) Background: The detection of DNA double-strand breaks in vitro using the phosphorylated histone biomarker (γH2AX) is an increasingly popular method of measuring in vitro genotoxicity, as it is sensitive, specific and suitable for high-throughput analysis. The γH2AX response is either detected by flow cytometry or microscopy, the latter being more accessible. However, authors sparsely publish details, data, and workflows from overall fluorescence intensity quantification, which hinders the reproducibility. (2) Methods: We used valinomycin as a model genotoxin, two cell lines (HeLa and CHO-K1) and a commercial kit for γH2AX immunofluorescence detection. Bioimage analysis was performed using the open-source software ImageJ. Mean fluorescent values were measured using segmented nuclei from the DAPI channel and the results were expressed as the area-scaled relative fold change in γH2AX fluorescence over the control. Cytotoxicity is expressed as the relative area of the nuclei. We present the workflows, data, and scripts on GitHub. (3) Results: The outputs obtained by an introduced method are in accordance with expected results, i.e., valinomycin was genotoxic and cytotoxic to both cell lines used after 24 h of incubation. (4) Conclusions: The overall fluorescence intensity of γH2AX obtained from bioimage analysis appears to be a promising alternative to flow cytometry. Workflow, data, and script sharing are crucial for further improvement of the bioimage analysis methods. Full article

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20 pages, 1295 KiB

Open AccessEditor’s ChoiceArticle

Model of the Origin of a Ciguatoxic Grouper (Plectropomus leopardus)

by Michael J. Holmes and Richard J. Lewis

Toxins 2023, 15(3), 230; https://doi.org/10.3390/toxins15030230 - 21 Mar 2023

Cited by 5 | Viewed by 1987

Abstract

Published data were used to model the transfer of ciguatoxins (CTX) across three trophic levels of a marine food chain on the Great Barrier Reef (GBR), Australia, to produce a mildly toxic common coral trout (Plectropomus leopardus), one of the most [...] Read more.

Published data were used to model the transfer of ciguatoxins (CTX) across three trophic levels of a marine food chain on the Great Barrier Reef (GBR), Australia, to produce a mildly toxic common coral trout (Plectropomus leopardus), one of the most targeted food fishes on the GBR. Our model generated a 1.6 kg grouper with a flesh concentration of 0.1 µg/kg of Pacific-ciguatoxin-1 (P-CTX-1 = CTX1B) from 1.1 to 4.3 µg of P-CTX-1 equivalents (eq.) entering the food chain from 0.7 to 2.7 million benthic dinoflagellates (Gambierdiscus sp.) producing 1.6 pg/cell of the P-CTX-1 precursor, P-CTX-4B (CTX4B). We simulated the food chain transfer of ciguatoxins via surgeonfishes by modelling Ctenochaetus striatus feeding on turf algae. A C. striatus feeding on ≥1000 Gambierdiscus/cm² of turf algae accumulates sufficient toxin in <2 days that when preyed on, produces a 1.6 kg common coral trout with a flesh concentration of 0.1 µg/kg P-CTX-1. Our model shows that even transient blooms of highly ciguatoxic Gambierdiscus can generate ciguateric fishes. In contrast, sparse cell densities of ≤10 Gambierdiscus/cm² are unlikely to pose a significant risk, at least in areas where the P-CTX-1 family of ciguatoxins predominate. The ciguatera risk from intermediate Gambierdiscus densities (~100 cells/cm²) is more difficult to assess, as it requires feeding times for surgeonfish (~4–14 days) that overlap with turnover rates of turf algae that are grazed by herbivorous fishes, at least in regions such as the GBR, where stocks of herbivorous fishes are not impacted by fishing. We use our model to explore how the duration of ciguatoxic Gambierdiscus blooms, the type of ciguatoxins they produce, and fish feeding behaviours can produce differences in relative toxicities between trophic levels. Our simple model indicates thresholds for the design of risk and mitigation strategies for ciguatera and the variables that can be manipulated to explore alternate scenarios for the accumulation and transfer of P-CTX-1 analogues through marine food chains and, potentially, for other ciguatoxins in other regions, as more data become available. Full article

(This article belongs to the Special Issue Ciguatoxins 2022–2023)

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11 pages, 1604 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Artificial Substrates Coupled with qPCR (AS-qPCR) Assay for the Detection of the Toxic Benthopelagic Dinoflagellate Vulcanodinium rugosum

by Aurélien Bouquet, Christine Felix, Estelle Masseret, Coralie Reymond, Eric Abadie, Mohamed Laabir and Jean Luc Rolland

Toxins 2023, 15(3), 217; https://doi.org/10.3390/toxins15030217 - 11 Mar 2023

Cited by 2 | Viewed by 1793

Abstract

Vulcanodinium rugosum is an emerging benthopelagic neuro-toxic dinoflagellate species responsible for seasonal Pinnatoxins and Portimines contaminations of shellfish and marine animals. This species is challenging to detect in the environment, as it is present in low abundance and difficult to be identified using [...] Read more.

Vulcanodinium rugosum is an emerging benthopelagic neuro-toxic dinoflagellate species responsible for seasonal Pinnatoxins and Portimines contaminations of shellfish and marine animals. This species is challenging to detect in the environment, as it is present in low abundance and difficult to be identified using light microscopy. In this work, we developed a method using artificial substrates coupled with qPCR (AS-qPCR) to detect V. rugosum in a marine environment. This sensitive, specific and easy-to-standardize alternative to current techniques does not require specialized expertise in taxonomy. After determining the limits and specificity of the qPCR, we searched for the presence of V. rugosum in four French Mediterranean lagoons using artificial substrates collected every two weeks for one year. The AS-qPCR method revealed its occurrences in summer 2021 in every studied lagoon and detected cells in more samples than light microscopy. As V. rugosum development induces shellfish contamination even at low microalga densities, the AS-qPCR method is accurate and relevant for monitoring V. rugosum in a marine environment. Full article

(This article belongs to the Special Issue Advances in Marine Toxins: Characterization, Analysis and Surveillance)

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13 pages, 1822 KiB

Open AccessEditor’s ChoiceArticle

Dispersive Magnetic Solid-Phase Extraction as a Novelty Sample Treatment for the Determination of the Main Aflatoxins in Paprika

by María García-Nicolás, Natalia Arroyo-Manzanares and Pilar Viñas

Toxins 2023, 15(2), 160; https://doi.org/10.3390/toxins15020160 - 15 Feb 2023

Cited by 5 | Viewed by 2430

Abstract

Dispersive magnetic solid-phase extraction (DMSPE) technique is proposed as a new sensitive and effective sample treatment method for the determination of aflatoxins in paprika samples. DMSPE was followed by ultrahigh-performance liquid chromatography and high-resolution mass spectrometry detection (UHPLC-HRMS) using a non-targeted acquisition mode [...] Read more.

Dispersive magnetic solid-phase extraction (DMSPE) technique is proposed as a new sensitive and effective sample treatment method for the determination of aflatoxins in paprika samples. DMSPE was followed by ultrahigh-performance liquid chromatography and high-resolution mass spectrometry detection (UHPLC-HRMS) using a non-targeted acquisition mode for the detection of main aflatoxins (aflatoxin G1, G2, B1 and B2) and derivatives. DMSPE was based on the use of magnetic nanocomposite coated with polypyrrole (PPy) polymer and the main experimental parameters influencing the extraction efficiency in adsorption and desorption steps have been studied and optimized. Analyses were performed using 250 µL magnetic PPy nanocomposite into the sample solution, adsorbing the analytes in 30 min and desorbing them with ethyl acetate (2 mL) in 15 min. The method has been validated, obtaining quantification limits between 3.5 and 4.7 µg kg⁻¹ and recoveries between 89.5–97.7%. The high recovery rate, wide detection range and the use for the first time of the reusable Fe₃O₄@PPy nanomaterial in suspension for solid food matrices, guarantee the usefulness of the method developed for adequate control of aflatoxins levels in paprika. The proposed methodology was applied for the analysis of 31 samples (conventional and organic) revealing the absence of aflatoxins in the samples. Full article

(This article belongs to the Collection Aflatoxins)

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0 pages, 3854 KiB

Open AccessEditor’s ChoiceArticle

Mixtures of Mycotoxins, Phytoestrogens, and Other Secondary Metabolites in Whole-Plant Corn Silages and Total Mixed Rations of Dairy Farms in Central and Northern Mexico

by Felipe Penagos-Tabares, Michael Sulyok, Juan-Ignacio Artavia, Samanta-Irais Flores-Quiroz, César Garzón-Pérez, Ezequías Castillo-Lopez, Luis Zavala, Juan-David Orozco, Johannes Faas, Rudolf Krska and Qendrim Zebeli

Toxins 2023, 15(2), 153; https://doi.org/10.3390/toxins15020153 - 13 Feb 2023

Cited by 6 | Viewed by 2654

Abstract

Mycotoxins and endocrine disruptors such as phytoestrogens can affect cattle health, reproduction, and productivity. Most studies of mycotoxins in dairy feeds in Mexico and worldwide have been focused on a few (regulated) mycotoxins. In contrast, less known fungal toxins, phytoestrogens, and other metabolites [...] Read more.

Mycotoxins and endocrine disruptors such as phytoestrogens can affect cattle health, reproduction, and productivity. Most studies of mycotoxins in dairy feeds in Mexico and worldwide have been focused on a few (regulated) mycotoxins. In contrast, less known fungal toxins, phytoestrogens, and other metabolites have been neglected and underestimated. This study analyzed a broad spectrum (>800) of mycotoxins, phytoestrogens, and fungal, plant, and unspecific secondary metabolites in whole-plant corn silages (WPCSs) and total mixed rations (TMRs) collected from 19 Mexican dairy farms. A validated multi-metabolite liquid chromatography/electrospray ionization–tandem mass spectrometric (LC/ESI–MS/MS) method was used. Our results revealed 125 of >800 tested (potentially toxic) secondary metabolites. WPCSs/TMRs in Mexico presented ubiquitous contamination with mycotoxins, phytoestrogens, and other metabolites. The average number of mycotoxins per TMR was 24, ranging from 9 to 31. Fusarium-derived secondary metabolites showed the highest frequencies, concentrations, and diversity among the detected fungal compounds. The most frequently detected mycotoxins in TMRs were zearalenone (ZEN) (100%), fumonisin B1 (FB1) (84%), and deoxynivalenol (84%). Aflatoxin B1 (AFB1) and ochratoxin A (OTA), previously reported in Mexico, were not detected. All TMR samples tested positive for phytoestrogens. Among the investigated dietary ingredients, corn stover, sorghum silage, and concentrate proportions were the most correlated with levels of total mycotoxins, fumonisins (Fs), and ergot alkaloids, respectively. Full article

(This article belongs to the Special Issue Detection, Control and Contamination of Mycotoxins (Volume II))

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15 pages, 636 KiB

Open AccessEditor’s ChoiceArticle

Advantages of Multiplexing Ability of the Orbitrap Mass Analyzer in the Multi-Mycotoxin Analysis

by Dávid Rakk, József Kukolya, Biljana D. Škrbić, Csaba Vágvölgyi, Mónika Varga and András Szekeres

Toxins 2023, 15(2), 134; https://doi.org/10.3390/toxins15020134 - 7 Feb 2023

Cited by 5 | Viewed by 2005

Abstract

In routine measurements, the length of the analysis time and nfumber of samples analysed during a time unit are crucial parameters, which are especially important for the food analysis, particularly in the case of mycotoxin determinations. High-resolution equipment, including time-of-flight or Orbitrap analyzators, [...] Read more.

In routine measurements, the length of the analysis time and nfumber of samples analysed during a time unit are crucial parameters, which are especially important for the food analysis, particularly in the case of mycotoxin determinations. High-resolution equipment, including time-of-flight or Orbitrap analyzators, can provide stable instrumental background for high-throughput analyses. In this report, a short, 1 min MS-based multi-mycotoxin method was developed with the application of a short column as a reduced chromatographic separation, taking advantages of the multiplexing and high-resolution capability of the QExactive Orbitrap MS possessing sub-1 ppm mass accuracy. The performance of the method was evaluated regarding selectivity, LOD, LOQ, linearity, matrix effect, and recovery, and compared to a UHPLC-MS/MS method. The final multiplexing method was able to quantify 11 mycotoxins in defined ranges (aflatoxins (corn, 2.8–600 μg/kg; wheat, 1.5–350 μg/kg), deoxynivalenol (corn, 640–9600 μg/kg; wheat, 128–3500 μg/kg), fumonisins (corn, 20–1500 μg/kg; wheat, 30–3500 μg/kg), HT-2 (corn, 64–5200 μg/kg; wheat, 61–3500 μg/kg), T-2 (corn, 10–800 μg/kg; wheat, 4–250 μg/kg), ochratoxin (corn, 4.7–600 μg/kg; wheat, 1–1000 μg/kg), zearalenone (corn, 64–4800 μg/kg; wheat, 4–500 μg/kg)) within one minute in corn and wheat matrices at the MRL levels stated by the European Union. Full article

(This article belongs to the Special Issue Rapid Detection of Mycotoxin Contamination 2.0)

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20 pages, 4386 KiB

Open AccessEditor’s ChoiceArticle

Effect of Plasma-Activated Water Bubbles on Fusarium graminearum, Deoxynivalenol, and Germination of Naturally Infected Barley during Steeping

by Ehsan Feizollahi, Urmila Basu, Rudolph Fredua-Agyeman, Brasathe Jeganathan, Lusine Tonoyan, Stephen E. Strelkov, Thava Vasanthan, Arno G. Siraki and M. S. Roopesh

Toxins 2023, 15(2), 124; https://doi.org/10.3390/toxins15020124 - 3 Feb 2023

Cited by 7 | Viewed by 3303

Abstract

Contamination of barley by deoxynivalenol (DON), a mycotoxin produced by Fusarium graminearum, causes considerable financial loss to the grain and malting industries. In this study, two atmospheric cold plasma (ACP) reactors were used to produce plasma-activated water (PAW) bubbles. The potential of [...] Read more.

Contamination of barley by deoxynivalenol (DON), a mycotoxin produced by Fusarium graminearum, causes considerable financial loss to the grain and malting industries. In this study, two atmospheric cold plasma (ACP) reactors were used to produce plasma-activated water (PAW) bubbles. The potential of PAW bubbles for the steeping of naturally infected barley (NIB) during the malting process was investigated. The PAW bubbles produced by treating water for 30 min using a bubble spark discharge (BSD) at low temperature resulted in the greatest concentration of oxygen-nitrogen reactive species (RONS). This treatment resulted in 57.3% DON degradation compared with 36.9% in the control sample; however, the same treatment reduced germination significantly (p < 0.05). Direct BSD ACP treatment for 20 min at low temperature and indirect treatment for 30 min increased the percentage of germinated rootlets of the seedlings compared with the control. Considering both the DON reduction and germination improvement of barley seeds, continuous jet ACP treatment for 30 min performed better than the other treatments used in this study. At higher temperature of PAW bubbles, the concentration of RONS was significantly (p < 0.05) reduced. Based on quantitative polymerase chain reaction (qPCR) analysis and fungal culture tests, the PAW bubble treatment did not significantly reduce infection of NIB. Nonetheless, this study provides useful information for the malting industry for PAW treatment optimization and its use in barley steeping for DON reduction and germination improvement. Full article

(This article belongs to the Section Mycotoxins)

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17 pages, 2702 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Brown Spider Venom Phospholipase-D Activity upon Different Lipid Substrates

by Daniele Chaves-Moreira, Luiza Helena Gremski, Fábio Rogério de Moraes, Larissa Vuitika, Ana Carolina Martins Wille, Jorge Enrique Hernández González, Olga Meiri Chaim, Andrea Senff-Ribeiro, Raghuvir Krishnaswamy Arni and Silvio Sanches Veiga

Toxins 2023, 15(2), 109; https://doi.org/10.3390/toxins15020109 - 27 Jan 2023

Cited by 6 | Viewed by 2242

Abstract

Brown spider envenomation results in dermonecrosis, characterized by an intense inflammatory reaction. The principal toxins of brown spider venoms are phospholipase-D isoforms, which interact with different cellular membrane components, degrade phospholipids, and generate bioactive mediators leading to harmful effects. The Loxosceles intermedia phospholipase [...] Read more.

Brown spider envenomation results in dermonecrosis, characterized by an intense inflammatory reaction. The principal toxins of brown spider venoms are phospholipase-D isoforms, which interact with different cellular membrane components, degrade phospholipids, and generate bioactive mediators leading to harmful effects. The Loxosceles intermedia phospholipase D, LiRecDT1, possesses a loop that modulates the accessibility to the active site and plays a crucial role in substrate. In vitro and in silico analyses were performed to determine aspects of this enzyme’s substrate preference. Sphingomyelin d18:1/6:0 was the preferred substrate of LiRecDT1 compared to other Sphingomyelins. Lysophosphatidylcholine 16:0/0:0 was preferred among other lysophosphatidylcholines, but much less than Sphingomyelin d18:1/6:0. In contrast, phosphatidylcholine d18:1/16:0 was not cleaved. Thus, the number of carbon atoms in the substrate plays a vital role in determining the optimal activity of this phospholipase-D. The presence of an amide group at C2 plays a key role in recognition and activity. In silico analyses indicated that a subsite containing the aromatic residues Y228 and W230 appears essential for choline recognition by cation-π interactions. These findings may help to explain why different cells, with different phospholipid fatty acid compositions exhibit distinct susceptibilities to brown spider venoms. Full article

(This article belongs to the Special Issue Animal Venom: Challenges and Perspectives in Drug Discovery)

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13 pages, 431 KiB

Open AccessEditor’s ChoiceArticle

The Effects of Incremental Doses of Aflatoxin B₁ on In Vitro Ruminal Nutrient Digestibility and Fermentation Profile of a Lactating Dairy Cow Diet in a Dual-Flow Continuous Culture System

by Felipe Xavier Amaro, Yun Jiang, Kathy Arriola, Matheus R. Pupo, Bruna C. Agustinho, Sarah L. Bennett, James R. Vinyard, Lais Tomaz, Richard R. Lobo, Andres Pech-Cervantes, Jose A. Arce-Cordero, Antonio P. Faciola, Adegbola Tolulope Adesogan and Diwakar Vyas

Toxins 2023, 15(2), 90; https://doi.org/10.3390/toxins15020090 - 18 Jan 2023

Cited by 1 | Viewed by 2222

Abstract

Aflatoxin B₁ (AFB₁) is a mycotoxin known to impair human and animal health. It is also believed to have a deleterious effect on ruminal nutrient digestibility under in vitro batch culture systems. The objective of this study was to evaluate [...] Read more.

Aflatoxin B₁ (AFB₁) is a mycotoxin known to impair human and animal health. It is also believed to have a deleterious effect on ruminal nutrient digestibility under in vitro batch culture systems. The objective of this study was to evaluate the effects of increasing the dose of AFB₁ on ruminal dry matter and nutrient digestibility, fermentation profile, and N flows using a dual-flow continuous culture system fed a diet formulated for lactating dairy cows. Eight fermenter vessels were used in a replicated 4 × 4 Latin square design with 10 d periods (7 d adaptation and 3 d sample collection). Treatments were randomly applied to fermenters on diet DM basis: (1) 0 μg of AFB₁/kg of DM (Control); (2) 50 μg of AFB₁/kg of DM (AF50); (3) 100 μg of AFB₁/kg of DM (AF100); and (4) 150 μg of AFB₁/kg of DM (AF150). Treatments did not affect nutrient digestibility, fermentation, and N flows. Aflatoxin B₁ concentration in ruminal fluid increased with dose but decreased to undetectable levels after 4 h post-dosing. In conclusion, adding incremental doses of AFB₁ did not affect ruminal fermentation, digestibility of nutrients, and N flows in a dual-flow continuous culture system fed diets formulated for lactating dairy cows. Full article

(This article belongs to the Collection Aflatoxins)

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23 pages, 2709 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Bivalve Shellfish Safety in Portugal: Variability of Faecal Levels, Metal Contaminants and Marine Biotoxins during the Last Decade (2011–2020)

by Ana Catarina Braga, Susana Margarida Rodrigues, Helena Maria Lourenço, Pedro Reis Costa and Sónia Pedro

Toxins 2023, 15(2), 91; https://doi.org/10.3390/toxins15020091 - 18 Jan 2023

Cited by 7 | Viewed by 2675

Abstract

Bivalves are a high-value product whose production has markedly increased, reaching 9863 tonnes in Portugal in 2021. Bivalves’ habitats—lagoons, estuaries and coastal waters—are exposed to biological and anthropogenic contaminants, which can bioaccumulate in these organisms and pose a significant public health risk. The [...] Read more.

Bivalves are a high-value product whose production has markedly increased, reaching 9863 tonnes in Portugal in 2021. Bivalves’ habitats—lagoons, estuaries and coastal waters—are exposed to biological and anthropogenic contaminants, which can bioaccumulate in these organisms and pose a significant public health risk. The need to obtain a safe product for human consumption led to the implementation of standardised hygiene regulations for harvesting and marketing bivalve molluscs, resulting in routine monitoring of bivalve production areas for microbial quality, metal contaminants, and marine biotoxins. While excessive levels of biotoxins and metal contamination lead to temporary harvesting bans, high faecal contamination leads to area reclassification and impose post-harvest treatments. In this study, the seasonal and temporal variability of these parameters were analysed using historical data generated by the monitoring programme during the last decade. Moreover, the impact of the monitoring program on bivalve harvesting from 2011 to 2020 was assessed. This program presented a considerable improvement over time, with an increase in the sampling effort and the overall program representativeness. Finally, contamination risk, revising control measures, and defining recommendations for risk mitigation measures are given in the light of ten years’ monitoring. Full article

(This article belongs to the Special Issue Bioactive Toxins in Marine Organism: Detection and Harmful Impacts)

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16 pages, 1050 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Effects of Botulinum Toxin Type A on the Nociceptive and Lemniscal Somatosensory Systems in Chronic Migraine: An Electrophysiological Study

by Gabriele Sebastianelli, Francesco Casillo, Antonio Di Renzo, Chiara Abagnale, Ettore Cioffi, Vincenzo Parisi, Cherubino Di Lorenzo, Mariano Serrao, Francesco Pierelli, Jean Schoenen and Gianluca Coppola

Toxins 2023, 15(1), 76; https://doi.org/10.3390/toxins15010076 - 14 Jan 2023

Cited by 7 | Viewed by 3762

Abstract

(1) Background: OnabotulinumtoxinA (BoNT-A) is a commonly used prophylactic treatment for chronic migraine (CM). Although randomized placebo studies have shown its clinical efficacy, the mechanisms by which it exerts its therapeutic effect are still incompletely understood and debated. (2) Methods: We studied in [...] Read more.

(1) Background: OnabotulinumtoxinA (BoNT-A) is a commonly used prophylactic treatment for chronic migraine (CM). Although randomized placebo studies have shown its clinical efficacy, the mechanisms by which it exerts its therapeutic effect are still incompletely understood and debated. (2) Methods: We studied in 15 CM patients the cephalic and extracephalic nociceptive and lemniscal sensory systems using electrophysiological techniques before and 1 and 3 months after one session of pericranial BoNT-A injections according to the PREEMPT protocol. We recorded the nociceptive blink reflex (nBR), the trigemino-cervical reflex (nTCR), the pain-related cortical evoked potential (PREP), and the upper limb somatosensory evoked potential (SSEP). (3) Results: Three months after a single session of prophylactic therapy with BoNT-A in CM patients, we found (a) an increase in the homolateral and contralateral nBR AUC, (b) an enhancement of the contralateral nBR AUC habituation slope and the nTCR habituation slope, (c) a decrease in PREP N-P 1st and 2nd amplitude block, and (d) no effect on SSEPs. (4) Conclusions: Our study provides electrophysiological evidence for the ability of a single session of BoNT-A injections to exert a neuromodulatory effect at the level of trigeminal system through a reduction in input from meningeal and other trigeminovascular nociceptors. Moreover, by reducing activity in cortical pain processing areas, BoNT-A restores normal functioning of the descending pain modulation systems. Full article

(This article belongs to the Special Issue Botulinum Toxin and Migraine: Goals and Perspectives)

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18 pages, 6580 KiB

Open AccessEditor’s ChoiceArticle

Highly Evolvable: Investigating Interspecific and Intraspecific Venom Variation in Taipans (Oxyuranus spp.) and Brown Snakes (Pseudonaja spp.)

by Jory van Thiel, Luis L. Alonso, Julien Slagboom, Nathan Dunstan, Roel M. Wouters, Cassandra M. Modahl, Freek J. Vonk, Timothy N. W. Jackson and Jeroen Kool

Toxins 2023, 15(1), 74; https://doi.org/10.3390/toxins15010074 - 13 Jan 2023

Cited by 11 | Viewed by 3305

Abstract

Snake venoms are complex mixtures of toxins that differ on interspecific (between species) and intraspecific (within species) levels. Whether venom variation within a group of closely related species is explained by the presence, absence and/or relative abundances of venom toxins remains largely unknown. [...] Read more.

Snake venoms are complex mixtures of toxins that differ on interspecific (between species) and intraspecific (within species) levels. Whether venom variation within a group of closely related species is explained by the presence, absence and/or relative abundances of venom toxins remains largely unknown. Taipans (Oxyuranus spp.) and brown snakes (Pseudonaja spp.) represent medically relevant species of snakes across the Australasian region and provide an excellent model clade for studying interspecific and intraspecific venom variation. Using liquid chromatography with ultraviolet and mass spectrometry detection, we analyzed a total of 31 venoms covering all species of this monophyletic clade, including widespread localities. Our results reveal major interspecific and intraspecific venom variation in Oxyuranus and Pseudonaja species, partially corresponding with their geographical regions and phylogenetic relationships. This extensive venom variability is generated by a combination of the absence/presence and differential abundance of venom toxins. Our study highlights that venom systems can be highly dynamical on the interspecific and intraspecific levels and underscores that the rapid toxin evolvability potentially causes major impacts on neglected tropical snakebites. Full article

(This article belongs to the Section Animal Venoms)

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17 pages, 2837 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Montane Rattlesnakes in México: Venoms of Crotalus tancitarensis and Related Species within the Crotalus intermedius Group

by Emily R. Grabowsky, Anthony J. Saviola, Javier Alvarado-Díaz, Adrian Quijada Mascareñas, Kirk C. Hansen, John R. Yates III and Stephen P. Mackessy

Toxins 2023, 15(1), 72; https://doi.org/10.3390/toxins15010072 - 13 Jan 2023

Cited by 2 | Viewed by 2760

Abstract

The Crotalus intermedius group is a clade of rattlesnakes consisting of several species adapted to a high elevation habitat, primarily in México. Crotalus tancitarensis was previously classified as C. intermedius, until individuals occurring on Cerro Tancítaro in Michoacán, México, were reevaluated and [...] Read more.

The Crotalus intermedius group is a clade of rattlesnakes consisting of several species adapted to a high elevation habitat, primarily in México. Crotalus tancitarensis was previously classified as C. intermedius, until individuals occurring on Cerro Tancítaro in Michoacán, México, were reevaluated and classified as a new species (C. tancitarensis) based on scale pattern and geographic location. This study aimed to characterize the venom of C. tancitarensis and compare the venom profile to those of other species within the Crotalus intermedius group using gel electrophoresis, biochemical assays, reverse-phase high performance liquid chromatography, mass spectrometry, and lethal toxicity (LD₅₀) assays. Results show that the venom profiles of species within the Crotalus intermedius group are similar, but with distinct differences in phospholipase A₂ (PLA₂), metalloproteinase PI (SVMP PI), and kallikrein-like serine proteinase (SVSP) activity and relative abundance. Proteomic analysis indicated that the highland forms produce venoms with 50–60 protein isoforms and a composition typical of type I rattlesnake venoms (abundant SVMPs, lack of presynaptic PLA₂-based neurotoxins), as well as a diversity of typical Crotalus venom components such as serine proteinases, PLA₂s, C-type lectins, and less abundant toxins (LAAOs, CRiSPs, etc.). The overall venom profile of C. tancitarensis appears most similar to C. transversus, which is consistent with a previous mitochondrial DNA analysis of the Crotalus intermedius group. These rattlesnakes of the Mexican highlands represent a radiation of high elevation specialists, and in spite of divergence of species in these Sky Island habitats, venom composition of species analyzed here has remained relatively conserved. The majority of protein family isoforms are conserved in all members of the clade, and as seen in other more broadly distributed rattlesnake species, differences in their venoms are largely due to relative concentrations of specific components. Full article

(This article belongs to the Section Animal Venoms)

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14 pages, 29465 KiB

Open AccessEditor’s ChoiceArticle

Solanum nigrum Fruit Extract Modulates Immune System Activity of Mealworm Beetle, Tenebrio molitor L.

by Arkadiusz Urbański, Natalia Konopińska, Natalia Bylewska, Radosław Gmyrek, Marta Spochacz-Santoro, Sabino Aurelio Bufo and Zbigniew Adamski

Toxins 2023, 15(1), 68; https://doi.org/10.3390/toxins15010068 - 12 Jan 2023

Cited by 3 | Viewed by 2422

Abstract

Here, we report the first evidence concerning the modulation of insect immune system activity after applying Solanum nigrum fruit extract (EXT). We focused on two main issues: (1) is EXT cytotoxic for Tenebrio molitor haemocytes? and (2) how EXT affects the basic immune [...] Read more.

Here, we report the first evidence concerning the modulation of insect immune system activity after applying Solanum nigrum fruit extract (EXT). We focused on two main issues: (1) is EXT cytotoxic for Tenebrio molitor haemocytes? and (2) how EXT affects the basic immune mechanisms of T. molitor. The results indicate cytotoxic action of 0.01 and 0.1% EXT on beetle haemocytes. Both the injection of EXT and incubating haemocytes with the EXT solution on microscopic slides significantly increased the number of apoptotic cells. However, 24 h after injection of 0.1% EXT cytotoxic effect of the tested extract probably was masked by the increased number of circulating haemocytes. Application of 0.01 and 0.1% EXT led to impairment of the activity of basic immune mechanisms such as phenoloxidase activity and the lysozyme-like antimicrobial activity of T. molitor haemolymph. Moreover, the EXT elicited significant changes in the expression level of selected immune genes. However, some of the immunomodulatory effects of EXT were different in beetles with and without an activated immune system. The obtained results are an essential step toward a complete understanding of the EXT mode of action on the T. molitor physiology and its potential usage in pest control. Full article

(This article belongs to the Special Issue Toxic Secondary Plant Metabolites: Toxicology, Medicine, Pharmacology, Ecotoxicology and Crop Protection)

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16 pages, 1613 KiB

Open AccessEditor’s ChoiceArticle

Sustainable Strategies to Counteract Mycotoxins Contamination and Cowpea Weevil in Chickpea Seeds during Post-Harvest

by Claudia Pisuttu, Samuele Risoli, Lorenzo Moncini, Cristina Nali, Elisa Pellegrini and Sabrina Sarrocco

Toxins 2023, 15(1), 61; https://doi.org/10.3390/toxins15010061 - 11 Jan 2023

Cited by 11 | Viewed by 3436

Abstract

Mycotoxins contamination and pest infestation of foods and feeds represent a pivotal threat for food safety and security worldwide, with crucial implications for human and animal health. Controlled atmosphere could be a sustainable strategy to reduce mycotoxins content and counteract the vitality of [...] Read more.

Mycotoxins contamination and pest infestation of foods and feeds represent a pivotal threat for food safety and security worldwide, with crucial implications for human and animal health. Controlled atmosphere could be a sustainable strategy to reduce mycotoxins content and counteract the vitality of deleterious organisms in foodstuff. Ozone treatment (O₃, 500 ppb for 30, 60 or 90 min) and high nitrogen concentration (N₂, 99% for 21 consecutive days) were tested in the post-harvest management of four batches of Cicer arietinum grains to control the presence of mycotoxigenic fungi and their secondary metabolites, as well as pest (i.e., Callosobruchus maculatus) infestation. At the end of the treatment, O₃ significantly decreased the incidence of Penicillium spp. (by an average of −50%, independently to the time of exposure) and reduced the patulin and aflatoxins content after 30 min (−85 and −100%, respectively). High N₂ concentrations remarkably reduced mycotoxins contamination (by an average of −94%) and induced pest mortality (at 100% after 5 days of exposure). These results confirm the promising potential of O₃ and N₂ in post-harvest conservation strategies, leading to further investigations to evaluate the effects on the qualitative characteristics of grains. Full article

(This article belongs to the Special Issue Novel Strategies for Biodegradation and Detoxification of Mycotoxins)

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15 pages, 1634 KiB

Open AccessEditor’s ChoiceArticle

In Vitro Efficacy of Antivenom and Varespladib in Neutralising Chinese Russell’s Viper (Daboia siamensis) Venom Toxicity

by Mimi Lay, Qing Liang, Geoffrey K. Isbister and Wayne C. Hodgson

Toxins 2023, 15(1), 62; https://doi.org/10.3390/toxins15010062 - 11 Jan 2023

Cited by 9 | Viewed by 2562

Abstract

The venom of the Russell’s viper (Daboia siamensis) contains neurotoxic and myotoxic phospholipase A₂ toxins which can cause irreversible damage to motor nerve terminals. Due to the time delay between envenoming and antivenom administration, antivenoms may have limited efficacy against [...] Read more.

The venom of the Russell’s viper (Daboia siamensis) contains neurotoxic and myotoxic phospholipase A₂ toxins which can cause irreversible damage to motor nerve terminals. Due to the time delay between envenoming and antivenom administration, antivenoms may have limited efficacy against some of these venom components. Hence, there is a need for adjunct treatments to circumvent these limitations. In this study, we examined the efficacy of Chinese D. siamensis antivenom alone, and in combination with a PLA₂ inhibitor, Varespladib, in reversing the in vitro neuromuscular blockade in the chick biventer cervicis nerve-muscle preparation. Pre-synaptic neurotoxicity and myotoxicity were not reversed by the addition of Chinese D. siamensis antivenom 30 or 60 min after venom (10 µg/mL). The prior addition of Varespladib prevented the neurotoxic and myotoxic activity of venom (10 µg/mL) and was also able to prevent further reductions in neuromuscular block and muscle twitches when added 60 min after venom. The addition of the combination of Varespladib and antivenom 60 min after venom failed to produce further improvements than Varespladib alone. This demonstrates that the window of time in which antivenom remains effective is relatively short compared to Varespladib and small-molecule inhibitors may be effective in abrogating some activities of Chinese D. siamensis venom. Full article

(This article belongs to the Special Issue Advanced Research on Animal Venoms in China)

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34 pages, 4265 KiB

Open AccessEditor’s ChoiceArticle

A Feasibility Study into the Production of a Mussel Matrix Reference Material for the Cyanobacterial Toxins Microcystins and Nodularins

by Andrew D. Turner, Daniel G. Beach, Amanda Foss, Ingunn A. Samdal, Kjersti L. E. Løvberg, Julia Waack, Christine Edwards, Linda A. Lawton, Karl J. Dean, Benjamin H. Maskrey and Adam M. Lewis

Toxins 2023, 15(1), 27; https://doi.org/10.3390/toxins15010027 - 30 Dec 2022

Cited by 5 | Viewed by 2386

Abstract

Microcystins and nodularins, produced naturally by certain species of cyanobacteria, have been found to accumulate in aquatic foodstuffs such as fish and shellfish, resulting in a risk to the health of the seafood consumer. Monitoring of toxins in such organisms for risk management [...] Read more.

Microcystins and nodularins, produced naturally by certain species of cyanobacteria, have been found to accumulate in aquatic foodstuffs such as fish and shellfish, resulting in a risk to the health of the seafood consumer. Monitoring of toxins in such organisms for risk management purposes requires the availability of certified matrix reference materials to aid method development, validation and routine quality assurance. This study consequently targeted the preparation of a mussel tissue reference material incurred with a range of microcystin analogues and nodularins. Nine targeted analogues were incorporated into the material as confirmed through liquid chromatography with tandem mass spectrometry (LC-MS/MS), with an additional 15 analogues detected using LC coupled to non-targeted high resolution mass spectrometry (LC-HRMS). Toxins in the reference material and additional source tissues were quantified using LC-MS/MS, two different enzyme-linked immunosorbent assay (ELISA) methods and with an oxidative-cleavage method quantifying 3-methoxy-2-methyl-4-phenylbutyric acid (MMPB). Correlations between the concentrations quantified using the different methods were variable, likely relating to differences in assay cross-reactivities and differences in the abilities of each method to detect bound toxins. A consensus concentration of total soluble toxins determined from the four independent test methods was 2425 ± 575 µg/kg wet weight. A mean 43 ± 9% of bound toxins were present in addition to the freely extractable soluble form (57 ± 9%). The reference material produced was homogenous and stable when stored in the freezer for six months without any post-production stabilization applied. Consequently, a cyanotoxin shellfish reference material has been produced which demonstrates the feasibility of developing certified seafood matrix reference materials for a large range of cyanotoxins and could provide a valuable future resource for cyanotoxin risk monitoring, management and mitigation. Full article

(This article belongs to the Special Issue Cyanotoxins in the Food Chain)

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11 pages, 544 KiB

Open AccessEditor’s ChoiceArticle

Occurrence and Dietary Exposure Assessment to Enniatin B through Consumption of Cereal-Based Products in Spain and the Catalonia Region

by Jose A. Gallardo, Sonia Marín, Antonio J. Ramos, German Cano-Sancho and Vicente Sanchis

Toxins 2023, 15(1), 24; https://doi.org/10.3390/toxins15010024 - 29 Dec 2022

Cited by 7 | Viewed by 2187

Abstract

Enniatin B (ENNB) is a mycotoxin produced by moulds from the Fusarium genera and its toxic effects are still not fully elucidated, hence a safe reference exposure value has not been established yet. ENNB is the most prevalent emerging mycotoxin and is widely [...] Read more.

Enniatin B (ENNB) is a mycotoxin produced by moulds from the Fusarium genera and its toxic effects are still not fully elucidated, hence a safe reference exposure value has not been established yet. ENNB is the most prevalent emerging mycotoxin and is widely found in cereal-based products, nevertheless, there are no comprehensive exposure assessment studies. For that reason, the aim of this study was to characterise the occurrence of ENNB and estimate the exposure of the Spanish and Catalan populations. A total of 347 cereal-based products were collected in 2019 and were analysed using liquid chromatography-tandem mass spectrometry. Consumption data were obtained from the national food consumption surveys (ENALIA) and a regional survey conducted in Catalonia. The global exposure was estimated using deterministic and probabilistic methods. The results showed a high occurrence of close to 100% in all foodstuffs, with a range from 6 to 269 µg/kg, and a strong correlation with the levels of deoxynivalenol. Children aged one–nine years were the most exposed, showing mean estimates in the range 308–324 ng/kg bw/day and 95th percentiles 697–781 ng/kg bw/day. This study stresses the need for further toxicological data to establish reference doses and conclude formal risk assessment, accounting for the co-occurrence with deoxynivalenol. Full article

(This article belongs to the Special Issue Mycotoxin Exposure: Assessment, Corresponding Adverse Health Outcomes and Control Plans)

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18 pages, 2789 KiB

Open AccessEditor’s ChoiceArticle

A Novel Trichothecene Toxin Phenotype Associated with Horizontal Gene Transfer and a Change in Gene Function in Fusarium

by Robert H. Proctor, Guixia Hao, Hye-Seon Kim, Briana K. Whitaker, Imane Laraba, Martha M. Vaughan and Susan P. McCormick

Toxins 2023, 15(1), 12; https://doi.org/10.3390/toxins15010012 - 24 Dec 2022

Cited by 5 | Viewed by 2099

Abstract

Fusarium trichothecenes are among the mycotoxins of most concern to food and feed safety. Production of these mycotoxins and presence of the trichothecene biosynthetic gene (TRI) cluster have been confirmed in only two multispecies lineages of Fusarium: the Fusarium incarnatum [...] Read more.

Fusarium trichothecenes are among the mycotoxins of most concern to food and feed safety. Production of these mycotoxins and presence of the trichothecene biosynthetic gene (TRI) cluster have been confirmed in only two multispecies lineages of Fusarium: the Fusarium incarnatum-equiseti (Incarnatum) and F. sambucinum (Sambucinum) species complexes. Here, we identified and characterized a TRI cluster in a species that has not been formally described and is represented by Fusarium sp. NRRL 66739. This fungus is reported to be a member of a third Fusarium lineage: the F. buharicum species complex. Cultures of NRRL 66739 accumulated only two trichothecenes, 7-hydroxyisotrichodermin and 7-hydroxyisotrichodermol. Although these are not novel trichothecenes, the production profile of NRRL 66739 is novel, because in previous reports 7-hydroxyisotrichodermin and 7-hydroxyisotrichodermol were components of mixtures of 6–8 trichothecenes produced by several Fusarium species in Sambucinum. Heterologous expression analysis indicated that the TRI13 gene in NRRL 66739 confers trichothecene 7-hydroxylation. This contrasts the trichothecene 4-hydroxylation function of TRI13 in other Fusarium species. Phylogenetic analyses suggest that NRRL 66739 acquired the TRI cluster via horizontal gene transfer from a close relative of Incarnatum and Sambucinum. These findings provide insights into evolutionary processes that have shaped the distribution of trichothecene production among Fusarium species and the structural diversity of the toxins. Full article

(This article belongs to the Special Issue Fusarium Mycotoxins: Chemistry, Genetics and Biology)

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18 pages, 2788 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Commercial Antivenoms Exert Broad Paraspecific Immunological Binding and In Vitro Inhibition of Medically Important Bothrops Pit Viper Venoms

by Jaffer Alsolaiss, Nessrin Alomran, Laura Hawkins and Nicholas R. Casewell

Toxins 2023, 15(1), 1; https://doi.org/10.3390/toxins15010001 - 20 Dec 2022

Cited by 8 | Viewed by 1942

Abstract

Snakebite envenoming is a life threatening neglected tropical disease that represents a considerable public health concern in the tropics. Viperid snakes of the genus Bothrops are among those of greatest medical importance in Latin America, and they frequently cause severe systemic haemotoxicity and [...] Read more.

Snakebite envenoming is a life threatening neglected tropical disease that represents a considerable public health concern in the tropics. Viperid snakes of the genus Bothrops are among those of greatest medical importance in Latin America, and they frequently cause severe systemic haemotoxicity and local tissue destructive effects in human victims. Although snakebite antivenoms can be effective therapeutics, their efficacy is undermined by venom toxin variation among snake species. In this study we investigated the extent of paraspecific venom cross-reactivity exhibited by three distinct anti-Bothrops antivenoms (Soro antibotrópico-crotálico, BothroFav and PoliVal-ICP) against seven different Bothrops pit viper venoms from across Latin America. We applied a range of in vitro assays to assess the immunological binding and recognition of venom toxins by the antivenoms and their inhibitory activities against specific venom functionalities. Our findings demonstrated that, despite some variations, the monovalent antivenom BothroFav and the polyvalent antivenoms Soro antibotrópico-crotálico and PoliVap-ICP exhibited extensive immunological recognition of the distinct toxins found in the different Bothrops venoms, with Soro antibotrópico-crotálico generally outperformed by the other two products. In vitro functional assays revealed outcomes largely consistent with the immunological binding data, with PoliVap-ICP and BothroFav exhibiting the greatest inhibitory potencies against procoagulant and fibrinogen-depleting venom activities, though Soro antibotrópico-crotálico exhibited potent inhibition of venom metalloproteinase activities. Overall, our findings demonstrate broad levels of antivenom paraspecificity, with in vitro immunological binding and functional inhibition often highly comparable between venoms used to manufacture the antivenoms and those from related species, even in the case of the monovalent antivenom BothroFav. Our findings suggest that the current clinical utility of these antivenoms could possibly be expanded to other parts of Latin America that currently suffer from a lack of specific snakebite therapies. Full article

(This article belongs to the Section Animal Venoms)

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16 pages, 1756 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Botulism in Spain: Epidemiology and Outcomes of Antitoxin Treatment, 1997–2019

by Marina Peñuelas, María Guerrero-Vadillo, Sylvia Valdezate, María Jesús Zamora, Inmaculada Leon-Gomez, Ángeles Flores-Cuéllar, Gema Carrasco, Oliva Díaz-García and Carmen Varela

Toxins 2023, 15(1), 2; https://doi.org/10.3390/toxins15010002 - 20 Dec 2022

Cited by 4 | Viewed by 3519

Abstract

Background: Botulism is a low incidence but potentially fatal infectious disease caused by neurotoxins produced mainly by Clostridium botulinum. There are different routes of acquisition, food-borne and infant/intestinal being the most frequent presentation, and antitoxin is the treatment of choice in all [...] Read more.

Background: Botulism is a low incidence but potentially fatal infectious disease caused by neurotoxins produced mainly by Clostridium botulinum. There are different routes of acquisition, food-borne and infant/intestinal being the most frequent presentation, and antitoxin is the treatment of choice in all cases. In Spain, botulism is under surveillance, and case reporting is mandatory. Methods: This retrospective study attempts to provide a more complete picture of the epidemiology of botulism in Spain from 1997 to 2019 and an assessment of the treatment, including the relationship between a delay in antitoxin administration and the length of hospitalization using the Cox proportional hazards test and Kruskal–Wallis test, and an approach to the frequency of adverse events, issues for which no previous national data have been published. Results: Eight of the 44 outbreaks were associated with contaminated commercial foods involving ≤7 cases/outbreak; preserved vegetables were the main source of infection, followed by fish products; early antitoxin administration significantly reduces the hospital stay, and adverse reactions to the antitoxin affect around 3% of treated cases. Full article

(This article belongs to the Special Issue Toxins and Infectious Diseases)

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11 pages, 1055 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Targeted Proteomics Analysis of Staphylococcal Superantigenic Toxins in Menstrual Fluid from Women with Menstrual Toxic Shock Syndrome (mTSS)

by Marie Courçon, Cédric Badiou, Mathilde Louwagie, Sibyle Etievant, Michel Jaquinod, Gérard Lina and Virginie Brun

Toxins 2022, 14(12), 886; https://doi.org/10.3390/toxins14120886 - 19 Dec 2022

Viewed by 2352

Abstract

Menstrual toxic shock syndrome (mTSS) is a rare life-threatening febrile illness that occurs in women using intravaginal menstrual protection. It is caused by toxic shock syndrome toxin 1 (TSST-1) produced by Staphylococcus aureus, triggering a sudden onset of rash and hypotension, subsequently [...] Read more.

Menstrual toxic shock syndrome (mTSS) is a rare life-threatening febrile illness that occurs in women using intravaginal menstrual protection. It is caused by toxic shock syndrome toxin 1 (TSST-1) produced by Staphylococcus aureus, triggering a sudden onset of rash and hypotension, subsequently leading to multiple organ failure. Detecting TSST-1 and S. aureus virulence factors in menstrual fluid could accelerate the diagnosis and improve therapeutic management of mTSS. However, menstrual fluid is a highly complex matrix, making detection of bacterial toxins challenging. Here, we present a mass-spectrometry-based proteomics workflow for the targeted, quantitative analysis of four S. aureus superantigenic toxins in menstrual fluids (TSST-1, SEA, SEC, and SED). This method was applied to characterize toxin levels in menstrual fluids collected from patients with mTSS and healthy women. Toxins were detectable in samples from patients with mTSS and one healthy donor at concentrations ranging from 0 to 0.46 µg/mL for TSST-1, and 0 to 1.07 µg/mL for SEC. SEA and SED were never detected in clinical specimens, even though many S. aureus strains were positive for the corresponding genes. The method presented here could be used to explore toxin production in vivo in users of intravaginal devices to improve the diagnosis, understanding, and prevention of mTSS. Full article

(This article belongs to the Special Issue Advances in Toxins and Virulence Analysis of Bacteria)

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14 pages, 2272 KiB

Open AccessEditor’s ChoiceArticle

Candidalysin Is the Hemolytic Factor of Candida albicans

by Selene Mogavero, Sarah Höfs, Alexa N. Lauer, Rita Müller, Sascha Brunke, Stefanie Allert, Franziska Gerwien, Sabrina Groth, Edward Dolk, Duncan Wilson, Thomas Gutsmann and Bernhard Hube

Toxins 2022, 14(12), 874; https://doi.org/10.3390/toxins14120874 - 15 Dec 2022

Cited by 13 | Viewed by 4277

Abstract

Candida albicans produces an important virulence factor, the hypha-associated Ece1-derived secreted peptide toxin candidalysin, which is crucial for the establishment of mucosal and systemic infections. C. albicans has also long been known to be hemolytic, yet the hemolytic factor has not been clearly [...] Read more.

Candida albicans produces an important virulence factor, the hypha-associated Ece1-derived secreted peptide toxin candidalysin, which is crucial for the establishment of mucosal and systemic infections. C. albicans has also long been known to be hemolytic, yet the hemolytic factor has not been clearly identified. Here, we show that candidalysin is the hemolytic factor of C. albicans. Its hemolytic activity is modulated by fragments of another Ece1 peptide, P7. Hemolysis by candidalysin can be neutralized by the purinergic receptor antagonist pyridoxal-phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS). PPADS also affects candidalysin’s ability to intercalate into synthetic membranes. We also describe the neutralization potential of two anti-candidalysin nanobodies, which are promising candidates for future anti-Candida therapy. This work provides evidence that the historically proposed hemolytic factor of C. albicans is in fact candidalysin and sheds more light on the complex roles of this toxin in C. albicans biology and pathogenicity. Full article

(This article belongs to the Section Mycotoxins)

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15 pages, 1722 KiB

Open AccessEditor’s ChoiceArticle

Characterization of NanR Regulation of Sialidase Production, Sporulation and Enterotoxin Production by Clostridium perfringens Type F Strains Carrying a Chromosomal Enterotoxin Gene

by Jihong Li, Eric Mi, Arhat Pradhan and Bruce A. McClane

Toxins 2022, 14(12), 872; https://doi.org/10.3390/toxins14120872 - 13 Dec 2022

Cited by 2 | Viewed by 1711

Abstract

Clostridium perfringens type F food poisoning (FP) strains produce C. perfringens enterotoxin (CPE) to cause a common bacterial food-borne illness in the United States. During FP, CPE is synthesized in the intestines when C. perfringens sporulates. Besides CPE, FP strains also produce sialidases. [...] Read more.

Clostridium perfringens type F food poisoning (FP) strains produce C. perfringens enterotoxin (CPE) to cause a common bacterial food-borne illness in the United States. During FP, CPE is synthesized in the intestines when C. perfringens sporulates. Besides CPE, FP strains also produce sialidases. Most FP strains carry their cpe gene on the chromosome and all surveyed chromosomal cpe (c-cpe) FP strains produce NanH sialidase or both NanJ and NanH sialidases. NanR has been shown previously to regulate sialidase activity in non-FP strains. The current study investigated whether NanR also regulates sialidase activity or influences sporulation and CPE production for c-cpe FP strains SM101 and 01E809. In sporulation medium, the SM101 nanR null mutant showed lower sialidase activity, sporulation, and CPE production than its wild-type parent, while the 01E809 nanR null mutant showed roughly similar sialidase activity, sporulation, and CPE production as its parent. In vegetative medium, the nanR null mutants of both strains produced more spores than their parents while NanR repressed sialidase activity in SM101 but positively regulated sialidase activity in 01E809. These results demonstrate that NanR regulates important virulence functions of c-cpe strains, with this control varying depending on strain and culture conditions. Full article

(This article belongs to the Collection Clostridium difficile/Clostridium sordellii and Clostridium perfringens Toxins)

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16 pages, 2345 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

The Crystal Structure of Bacillus thuringiensis Tpp80Aa1 and Its Interaction with Galactose-Containing Glycolipids

by Hannah L. Best, Lainey J. Williamson, Magdalena Lipka-Lloyd, Helen Waller-Evans, Emyr Lloyd-Evans, Pierre J. Rizkallah and Colin Berry

Toxins 2022, 14(12), 863; https://doi.org/10.3390/toxins14120863 - 8 Dec 2022

Cited by 7 | Viewed by 2963

Abstract

Tpp80Aa1 from Bacillus thuringiensis is a Toxin_10 family protein (Tpp) with reported action against Culex mosquitoes. Here, we demonstrate an expanded target range, showing Tpp80Aa1 is also active against the larvae of Anopheles gambiae and Aedes aegypti mosquitoes. We report the first crystal [...] Read more.

Tpp80Aa1 from Bacillus thuringiensis is a Toxin_10 family protein (Tpp) with reported action against Culex mosquitoes. Here, we demonstrate an expanded target range, showing Tpp80Aa1 is also active against the larvae of Anopheles gambiae and Aedes aegypti mosquitoes. We report the first crystal structure of Tpp80Aa1 at a resolution of 1.8 Å, which shows Tpp80Aa1 consists of two domains: an N-terminal β-trefoil domain resembling a ricin B lectin and a C-terminal putative pore-forming domain sharing structural similarity with the aerolysin family. Similar to other Tpp family members, we observe Tpp80Aa1 binds to the mosquito midgut, specifically the posterior midgut and the gastric caecum. We also identify that Tpp80Aa1 can interact with galactose-containing glycolipids and galactose, and this interaction is critical for exerting full insecticidal action against mosquito target cell lines. Full article

(This article belongs to the Special Issue Bacillus thuringiensis: A Broader View of Its Biocidal Activity)

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17 pages, 1424 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Design of a Diagnostic Immunoassay for Aflatoxin M1 Based on a Plant-Produced Antibody

by Cristina Capodicasa, Erica Bastiani, Thea Serra, Laura Anfossi and Marcello Catellani

Toxins 2022, 14(12), 851; https://doi.org/10.3390/toxins14120851 - 3 Dec 2022

Cited by 1 | Viewed by 2284

Abstract

A new green competitive ELISA for aflatoxin M1 quantification in raw milk was developed. This diagnostic tool is based on an anti AFM1 mAb produced by plant molecular farming in alternative to classical systems. Our assay, showing an IC₅₀ below 25 ng/L, [...] Read more.

A new green competitive ELISA for aflatoxin M1 quantification in raw milk was developed. This diagnostic tool is based on an anti AFM1 mAb produced by plant molecular farming in alternative to classical systems. Our assay, showing an IC₅₀ below 25 ng/L, fits with the requirements of EU legislation limits for AFM1 (50 ng/L). Optimal accuracy was achieved in correspondence of the decision levels (25 and 50 ng/L), and the assay enabled AFM1 quantification in the range 5–110 ng/L, with limit of detection 3 ng/L. Moreover, to evaluate a real applicability in diagnostics, raw milk-spiked samples were analysed, achieving satisfactory recovery rates of AFM1. In conclusion, an efficient and ready-to-use diagnostic assay for the quantification of aflatoxin M1 in milk, based on a plant-produced recombinant mAb, has been successfully developed. Full article

(This article belongs to the Special Issue Advanced Analytical Methodologies for the Determination of Mycotoxins in Food and Environmental Objects)

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25 pages, 6207 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Subchronic Oral Cylindrospermopsin Exposure Alters the Host Gut Microbiome and Is Associated with Progressive Hepatic Inflammation, Stellate Cell Activation, and Mild Fibrosis in a Preclinical Study

by Punnag Saha, Macayla Upright, Dipro Bose, Subhajit Roy, Ayushi Trivedi, Madhura More, Geoff I. Scott, Bryan W. Brooks and Saurabh Chatterjee

Toxins 2022, 14(12), 835; https://doi.org/10.3390/toxins14120835 - 1 Dec 2022

Cited by 6 | Viewed by 3702

Abstract

Epidemiological studies have reported a strong association between liver injury and incidences of hepatocellular carcinoma in sections of humans globally. Several preclinical studies have shown a strong link between cyanotoxin exposure and the development of nonalcoholic steatohepatitis, a precursor of hepatocellular carcinoma. Among [...] Read more.

Epidemiological studies have reported a strong association between liver injury and incidences of hepatocellular carcinoma in sections of humans globally. Several preclinical studies have shown a strong link between cyanotoxin exposure and the development of nonalcoholic steatohepatitis, a precursor of hepatocellular carcinoma. Among the emerging threats from cyanotoxins, new evidence shows cylindrospermopsin release in freshwater lakes. A known hepatotoxin in higher concentrations, we examined the possible role of cylindrospermopsin in causing host gut dysbiosis and its association with liver pathology in a mouse model of toxico-pharmacokinetics and hepatic pathology. The results showed that oral exposure to cylindrospermopsin caused decreased diversity of gut bacteria phyla accompanied by an increased abundance of Clostridioides difficile and decreased abundance of probiotic flora such as Roseburia, Akkermanssia, and Bacteroides thetaiotamicron, a signature most often associated with intestinal and hepatic pathology and underlying gastrointestinal disease. The altered gut dysbiosis was also associated with increased Claudin2 protein in the intestinal lumen, a marker of gut leaching and endotoxemia. The study of liver pathology showed marked liver inflammation, the release of damage-associated molecular patterns, and activation of toll-like receptors, a hallmark of consistent and progressive liver damage. Hepatic pathology was also linked to increased Kupffer cell activation and stellate cell activation, markers of progressive liver damage often linked to the development of liver fibrosis and carcinoma. In conclusion, the present study provides additional evidence of cylindrospermopsin-linked progressive liver pathology that may be very well-linked to gut dysbiosis, though definitive evidence involving this link needs to be studied further. Full article

(This article belongs to the Special Issue Ecology and Toxicology of Cyanobacteria and Cyanotoxins)

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23 pages, 2255 KiB

Open AccessEditor’s ChoiceArticle

The Fast and the Furriest: Investigating the Rate of Selection on Mammalian Toxins

by Leah Lucy Joscelyne Fitzpatrick, Vincent Nijman, Rodrigo Ligabue-Braun and K. Anne-Isola Nekaris

Toxins 2022, 14(12), 842; https://doi.org/10.3390/toxins14120842 - 1 Dec 2022

Cited by 2 | Viewed by 3519

Abstract

The evolution of venom and the selection pressures that act on toxins have been increasingly researched within toxinology in the last two decades, in part due to the exceptionally high rates of diversifying selection observed in animal toxins. In 2015, Sungar and Moran [...] Read more.

The evolution of venom and the selection pressures that act on toxins have been increasingly researched within toxinology in the last two decades, in part due to the exceptionally high rates of diversifying selection observed in animal toxins. In 2015, Sungar and Moran proposed the ‘two-speed’ model of toxin evolution linking evolutionary age of a group to the rates of selection acting on toxins but due to a lack of data, mammals were not included as less than 30 species of venomous mammal have been recorded, represented by elusive species which produce small amounts of venom. Due to advances in genomics and transcriptomics, the availability of toxin sequences from venomous mammals has been increasing. Using branch- and site-specific selection models, we present the rates of both episodic and pervasive selection acting upon venomous mammal toxins as a group for the first time. We identified seven toxin groups present within venomous mammals, representing Chiroptera, Eulipotyphla and Monotremata: KLK1, Plasminogen Activator, Desmallipins, PACAP, CRiSP, Kunitz Domain One and Kunitz Domain Two. All but one group (KLK1) was identified by our results to be evolving under both episodic and pervasive diversifying selection with four toxin groups having sites that were implicated in the fitness of the animal by TreeSAAP (Selection on Amino Acid Properties). Our results suggest that venomous mammal ecology, behaviour or genomic evolution are the main drivers of selection, although evolutionary age may still be a factor. Our conclusion from these results indicates that mammalian toxins are following the two-speed model of selection, evolving predominately under diversifying selection, fitting in with other younger venomous taxa like snakes and cone snails—with high amounts of accumulating mutations, leading to more novel adaptions in their toxins. Full article

(This article belongs to the Special Issue Evolution of Venomous and Poisonous Animals)

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17 pages, 4433 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Comparative Study of Toxic Effects and Pathophysiology of Envenomations Induced by Carybdea brevipedalia (Cnidaria: Cubozoa) and Nemopilema nomurai (Cnidaria: Scyphozoa) Jellyfish Venoms

by Du Hyeon Hwang, Phil-Ok Koh, Ramachandran Loganathan Mohan Prakash, Jinho Chae, Changkeun Kang and Euikyung Kim

Toxins 2022, 14(12), 831; https://doi.org/10.3390/toxins14120831 - 28 Nov 2022

Cited by 1 | Viewed by 2516

Abstract

Jellyfish stings can result in local tissue damage and systemic pathophysiological sequelae. Despite constant occurrences of jellyfish stings in oceans throughout the world, the toxinological assessment of these jellyfish envenomations has not been adequately reported in quantitative as well as in qualitative measurements. [...] Read more.

Jellyfish stings can result in local tissue damage and systemic pathophysiological sequelae. Despite constant occurrences of jellyfish stings in oceans throughout the world, the toxinological assessment of these jellyfish envenomations has not been adequately reported in quantitative as well as in qualitative measurements. Herein, we have examined and compared the in vivo toxic effects and pathophysiologic alterations using experimental animal models for two representative stinging jellyfish classes, i.e., Cubozoa and Scyphozoa. For this study, mice were administered with venom extracts of either Carybdea brevipedalia (Cnidaria: Cubozoa) or Nemopilema nomurai (Cnidaria: Scyphozoa). From the intraperitoneal (IP) administration study, the median lethal doses leading to the deaths of mice 24 h post-treatment after (LD₅₀) for C. brevipedalia venom (CbV) and N. nomurai venom (NnV) were 0.905 and 4.4697 mg/kg, respectively. The acute toxicity (i.e., lethality) of CbV was much higher with a significantly accelerated time to death value compared with those of NnV. The edematogenic activity induced by CbV was considerably (83.57/25 = 3.343-fold) greater than NnV. For the evaluation of their dermal toxicities, the epidermis, dermis, subcutaneous tissues, and skeletal muscles were evaluated toxinologically/histopathologically following the intradermal administration of the venoms. The minimal hemorrhagic doses (MHD) of the venoms were found to be 55.6 and 83.4 μg/mouse for CbV and NnV, respectively. Furthermore, the CbV injection resulted in extensive alterations of mouse dermal tissues, including severe edema, and hemorrhagic/necrotic lesions, with the minimum necrotizing dose (MND) of 95.42 µg/kg body weight. The skin damaging effects of CbV appeared to be considerably greater, compared with those of NnV (MND = 177.99 µg/kg). The present results indicate that the toxicities and pathophysiologic effects of jellyfish venom extracts may vary from species to species. As predicted from the previous reports on these jellyfish envenomations, the crude venom extracts of C. brevipedalia exhibit much more potent toxicity than that of N. nomurai in the present study. These observations may contribute to our understanding of the toxicities of jellyfish venoms, as well as their mode of toxinological actions, which might be helpful for establishing the therapeutic strategies of jellyfish stings. Full article

(This article belongs to the Special Issue Venom-Induced Tissue Damage)

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19 pages, 3422 KiB

Open AccessEditor’s ChoiceArticle

Targeted Sphingolipid Analysis in Heart, Gizzard, and Breast Muscle in Chickens Reveals Possible New Target Organs of Fumonisins

by Philippe Guerre, Caroline Gilleron, Maria Matard-Mann and Pi Nyvall Collén

Toxins 2022, 14(12), 828; https://doi.org/10.3390/toxins14120828 - 24 Nov 2022

Cited by 5 | Viewed by 1878

Abstract

Alteration of sphingolipid synthesis is a key event in fumonisins toxicity, but only limited data have been reported regarding the effects of fumonisins on the sphingolipidome. Recent studies in chickens found that the changes in sphingolipids in liver, kidney, lung, and brain differed [...] Read more.

Alteration of sphingolipid synthesis is a key event in fumonisins toxicity, but only limited data have been reported regarding the effects of fumonisins on the sphingolipidome. Recent studies in chickens found that the changes in sphingolipids in liver, kidney, lung, and brain differed greatly. This study aimed to determine the effects of fumonisins on sphingolipids in heart, gizzard, and breast muscle in chickens fed 20.8 mg FB1 + FB2/kg for 9 days. A significant increase in the sphinganine:sphingosine ratio due to an increase in sphinganine was observed in heart and gizzard. Dihydroceramides and ceramides increased in the hearts of chickens fed fumonisins, but decreased in the gizzard. The dihydrosphingomyelin, sphingomyelin, and glycosylceramide concentrations paralleled those of ceramides, although the effects were less pronounced. In the heart, sphingolipids with fatty acid chain lengths of 20 to 26 carbons were more affected than those with 14–16 carbons; this difference was not observed in the gizzard. Partial least squares-discriminant analysis on sphingolipids in the heart allowed chickens to be divided into two distinct groups according to their diet. The same was the case for the gizzard. Pearson coefficients of correlation among all the sphingolipids assayed revealed strong positive correlations in the hearts of chickens fed fumonisins compared to chickens fed a control diet, as well as compared to gizzard, irrespective of the diet fed. By contrast, no effect of fumonisins was observed on sphingolipids in breast muscle. Full article

(This article belongs to the Special Issue Mechanism of Action of Mycotoxins)

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17 pages, 3488 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Distinct Metabolic States Are Observed in Hypoglycemia Induced in Mice by Ricin Toxin or by Fasting

by Jacob Kempa, Galen O’Shea-Stone, Corinne E. Moss, Tami Peters, Tamera K. Marcotte, Brian Tripet, Brian Eilers, Brian Bothner, Valérie Copié and Seth H. Pincus

Toxins 2022, 14(12), 815; https://doi.org/10.3390/toxins14120815 - 22 Nov 2022

Cited by 1 | Viewed by 2321

Abstract

Hypoglycemia may be induced by a variety of physiologic and pathologic stimuli and can result in life-threatening consequences if untreated. However, hypoglycemia may also play a role in the purported health benefits of intermittent fasting and caloric restriction. Previously, we demonstrated that systemic [...] Read more.

Hypoglycemia may be induced by a variety of physiologic and pathologic stimuli and can result in life-threatening consequences if untreated. However, hypoglycemia may also play a role in the purported health benefits of intermittent fasting and caloric restriction. Previously, we demonstrated that systemic administration of ricin toxin induced fatal hypoglycemia in mice. Here, we examine the metabolic landscape of the hypoglycemic state induced in the liver of mice by two different stimuli: systemic ricin administration and fasting. Each stimulus produced the same decrease in blood glucose and weight loss. The polar metabolome was studied using ¹H NMR, quantifying 59 specific metabolites, and untargeted LC-MS on approximately 5000 features. Results were analyzed by multivariate analyses, using both principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA), to identify global metabolic patterns, and by univariate analyses (ANOVA) to assess individual metabolites. The results demonstrated that while there were some similarities in the responses to the two stimuli including decreased glucose, ADP, and glutathione, they elicited distinct metabolic states. The metabolite showing the greatest difference was O-phosphocholine, elevated in ricin-treated animals and known to be affected by the pro-inflammatory cytokine TNF-α. Another difference was the alternative fuel source utilized, with fasting-induced hypoglycemia primarily ketotic, while the response to ricin-induced hypoglycemia involves protein and amino acid catabolism. Full article

(This article belongs to the Special Issue Advances in Ricin Antitoxins: From Intoxication to Diagnosis and Treatment)

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26 pages, 4283 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Extracellular Vesicles from Bothrops jararaca Venom Are Diverse in Structure and Protein Composition and Interact with Mammalian Cells

by Larissa Gonçalves-Machado, Brunno Renato Farias Verçoza, Fábio César Sousa Nogueira, Rafael Donadélli Melani, Gilberto Barbosa Domont, Silas Pessini Rodrigues, Juliany Cola Fernandes Rodrigues and Russolina Benedeta Zingali

Toxins 2022, 14(11), 806; https://doi.org/10.3390/toxins14110806 - 19 Nov 2022

Cited by 2 | Viewed by 2770

Abstract

Snake venoms are complex cocktails of non-toxic and toxic molecules that work synergistically for the envenoming outcome. Alongside the immediate consequences, chronic manifestations and long-term sequelae can occur. Recently, extracellular vesicles (EVs) were found in snake venom. EVs mediate cellular communication through long [...] Read more.

Snake venoms are complex cocktails of non-toxic and toxic molecules that work synergistically for the envenoming outcome. Alongside the immediate consequences, chronic manifestations and long-term sequelae can occur. Recently, extracellular vesicles (EVs) were found in snake venom. EVs mediate cellular communication through long distances, delivering proteins and nucleic acids that modulate the recipient cell’s function. However, the biological roles of snake venom EVs, including possible cross-organism communication, are still unknown. This knowledge may expand the understanding of envenoming mechanisms. In the present study, we isolated and characterized the EVs from Bothrops jararaca venom (Bj-EVs), giving insights into their biological roles. Fresh venom was submitted to differential centrifugation, resulting in two EV populations with typical morphology and size range. Several conserved EV markers and a subset of venom related EV markers, represented mainly by processing enzymes, were identified by proteomic analysis. The most abundant protein family observed in Bj-EVs was 5’-nucleotidase, known to be immunosuppressive and a low abundant and ubiquitous toxin in snake venoms. Additionally, we demonstrated that mammalian cells efficiently internalize Bj-EVs. The commercial antibothropic antivenom partially recognizes Bj-EVs and inhibits cellular EV uptake. Based on the proteomic results and the in vitro interaction assays using macrophages and muscle cells, we propose that Bj-EVs may be involved not only in venom production and processing but also in host immune modulation and long-term effects of envenoming. Full article

(This article belongs to the Section Animal Venoms)

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25 pages, 2846 KiB

Open AccessEditor’s ChoiceArticle

Lactic Acid Bacteria as Potential Agents for Biocontrol of Aflatoxigenic and Ochratoxigenic Fungi

by Eva María Mateo, Andrea Tarazona, Misericordia Jiménez and Fernando Mateo

Toxins 2022, 14(11), 807; https://doi.org/10.3390/toxins14110807 - 19 Nov 2022

Cited by 8 | Viewed by 2746

Abstract

Aflatoxins (AF) and ochratoxin A (OTA) are fungal metabolites that have carcinogenic, teratogenic, embryotoxic, genotoxic, neurotoxic, and immunosuppressive effects in humans and animals. The increased consumption of plant-based foods and environmental conditions associated with climate change have intensified the risk of mycotoxin intoxication. [...] Read more.

Aflatoxins (AF) and ochratoxin A (OTA) are fungal metabolites that have carcinogenic, teratogenic, embryotoxic, genotoxic, neurotoxic, and immunosuppressive effects in humans and animals. The increased consumption of plant-based foods and environmental conditions associated with climate change have intensified the risk of mycotoxin intoxication. This study aimed to investigate the abilities of eleven selected LAB strains to reduce/inhibit the growth of Aspergillus flavus, Aspergillus parasiticus, Aspergillus carbonarius, Aspergillus niger, Aspergillus welwitschiae, Aspergillus steynii, Aspergillus westerdijkiae, and Penicillium verrucosum and AF and OTA production under different temperature regiments. Data were treated by ANOVA, and machine learning (ML) models able to predict the growth inhibition percentage were built, and their performance was compared. All factors LAB strain, fungal species, and temperature significantly affected fungal growth and mycotoxin production. The fungal growth inhibition range was 0–100%. Overall, the most sensitive fungi to LAB treatments were P. verrucosum and A. steynii, while the least sensitive were A. niger and A. welwitschiae. The LAB strains with the highest antifungal activity were Pediococcus pentosaceus (strains S11sMM and M9MM5b). The reduction range for AF was 19.0% (aflatoxin B1)-60.8% (aflatoxin B2) and for OTA, 7.3–100%, depending on the bacterial and fungal strains and temperatures. The LAB strains with the highest anti-AF activity were the three strains of P. pentosaceus and Leuconostoc mesenteroides ssp. dextranicum (T2MM3), and those with the highest anti-OTA activity were Leuconostoc paracasei ssp. paracasei (3T3R1) and L. mesenteroides ssp. dextranicum (T2MM3). The best ML methods in predicting fungal growth inhibition were multilayer perceptron neural networks, followed by random forest. Due to anti-fungal and anti-mycotoxin capacity, the LABs strains used in this study could be good candidates as biocontrol agents against aflatoxigenic and ochratoxigenic fungi and AFL and OTA accumulation. Full article

(This article belongs to the Special Issue Evaluation and Prevention of Mycotoxin Contamination and Toxicological Effects)

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10 pages, 1989 KiB

Open AccessEditor’s ChoiceArticle

Proteomic Analysis of the Predatory Venom of Conus striatus Reveals Novel and Population-Specific κA-Conotoxin SIVC

by Fabrice Saintmont, Guillaume Cazals, Claudia Bich and Sebastien Dutertre

Toxins 2022, 14(11), 799; https://doi.org/10.3390/toxins14110799 - 17 Nov 2022

Cited by 2 | Viewed by 2021

Abstract

Animal venoms are a rich source of pharmacological compounds with ecological and evolutionary significance, as well as with therapeutic and biotechnological potentials. Among the most promising venomous animals, cone snails produce potent neurotoxic venom to facilitate prey capture and defend against aggressors. Conus [...] Read more.

Animal venoms are a rich source of pharmacological compounds with ecological and evolutionary significance, as well as with therapeutic and biotechnological potentials. Among the most promising venomous animals, cone snails produce potent neurotoxic venom to facilitate prey capture and defend against aggressors. Conus striatus, one of the largest piscivorous species, is widely distributed, from east African coasts to remote Polynesian Islands. In this study, we investigated potential intraspecific differences in venom composition between distinct geographical populations from Mayotte Island (Indian Ocean) and Australia (Pacific Ocean). Significant variations were noted among the most abundant components, namely the κA-conotoxins, which contain three disulfide bridges and complex glycosylations. The amino acid sequence of a novel κA-conotoxin SIVC, including its N-terminal acetylated variant, was deciphered using tandem mass spectrometry (MS/MS). In addition, the glycosylation pattern was found to be consisting of two HexNAc and four Hex for the Mayotte population, which diverge from the previously characterized two HexNAc and three Hex combinations for this species, collected elsewhere. Whereas the biological and ecological roles of these modifications remain to be investigated, population-specific glycosylation patterns provide, for the first time, a new level of intraspecific variations in cone snail venoms. Full article

(This article belongs to the Special Issue Evolution, Genomics and Proteomics of Venom)

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12 pages, 680 KiB

Open AccessEditor’s ChoiceArticle

An Eight-Year Survey on Aflatoxin B1 Indicates High Feed Safety in Animal Feed and Forages in Northern Italy

by Luca Ferrari, Francesca Fumagalli, Nicoletta Rizzi, Elisa Grandi, Serena Vailati, Michele Manoni, Matteo Ottoboni, Federica Cheli and Luciano Pinotti

Toxins 2022, 14(11), 763; https://doi.org/10.3390/toxins14110763 - 4 Nov 2022

Cited by 10 | Viewed by 3466

Abstract

Aflatoxins (AFs) remain the main concern for the agricultural and dairy industries due to their effects on the performances and quality of livestock production. Aflatoxins are always unavoidable and should be monitored. The objective of this paper is to bring to light a [...] Read more.

Aflatoxins (AFs) remain the main concern for the agricultural and dairy industries due to their effects on the performances and quality of livestock production. Aflatoxins are always unavoidable and should be monitored. The objective of this paper is to bring to light a significant volume of data on AF contamination in several animal feed ingredients in Northern Italy. The Regional Breeders Association of Lombardy has been conducting a survey program to monitor mycotoxin contamination in animal feeds, and in this paper, we present data relating to AFB1 contamination. In most cases (95%), the concentrations were low enough to ensure compliance with the European Union’s (EU’s) maximum admitted levels for animal feed ingredients. However, the data show a high variability in AF contamination between different matrices and, within the same matrix, a high variability year over year. High levels of AFs were detected in maize and cotton, especially in the central part of the second decade of this century, i.e., 2015–2018, which has shown a higher risk of AF contamination in feed materials in Northern Italy. Variability due to climate change and the international commodity market affect future prospects to predict the presence of AFs. Supplier monitoring and control and reduced buying of contaminated raw materials, as well as performing analyses of each batch, help reduce AF spread. Full article

(This article belongs to the Special Issue Improved and Innovative Actions for Mycotoxin Management in the Food and Feed Chain 2.0)

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13 pages, 942 KiB

Open AccessEditor’s ChoiceArticle

Toxin Profile of Two Gymnodinium catenatum Strains from Iberian Coastal Waters

by Joana F. Leal, Gabriel Bombo, Hugo Pereira, Bernardo Vicente, Ana Amorim and Maria L. S. Cristiano

Toxins 2022, 14(11), 762; https://doi.org/10.3390/toxins14110762 - 4 Nov 2022

Cited by 5 | Viewed by 2429

Abstract

Gymnodinium catenatum has been the main species responsible for paralytic shellfish poisoning events along the Portuguese coast (Iberian Peninsula), causing bans on bivalve harvesting that result in huge economic losses. This work presents the characterization of two novel isolates of G. catenatum regarding [...] Read more.

Gymnodinium catenatum has been the main species responsible for paralytic shellfish poisoning events along the Portuguese coast (Iberian Peninsula), causing bans on bivalve harvesting that result in huge economic losses. This work presents the characterization of two novel isolates of G. catenatum regarding their growth and toxin profiles. Laboratory growth experiments revealed that, although low growth rates were obtained during cultivation, the cell yields were high compared to those reported in the literature. Evaluation of the toxin profiles, by HPLC-FLD, essentially confirmed the typical composition of toxins of this regional population (Iberian Peninsula), namely, the absence or low representation of the toxins dcNEO, GTX1,4 and NEO and a higher ratio of the toxins C1,2, GTX6 and GTX5. However, the percentage of the identified toxins varied among the strains of this study (under the same isolation, growth, and analysis conditions), and also differed from that of other strains described in the literature. Interestingly, we found a comparatively high abundance of dcSTX in both strains, relative to the other toxins, and an unquantifiable amount of C3,4 toxins. In addition to the geographic relationship between toxin profiles, chemical conversions among toxins may explain some differences encountered in the toxin profiles of G. catenatum strains. Full article

(This article belongs to the Special Issue Animal Poisoning: Toxins from Plants or Feed)

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19 pages, 3279 KiB

Open AccessEditor’s ChoiceArticle

Interaction of Clostridium perfringens Epsilon Toxin with the Plasma Membrane: The Role of Amino Acids Y42, Y43 and H162

by Skye Marshall, Beth McGill, Helen Morcrette, C. Peter Winlove, Catalin Chimerel, Peter G. Petrov and Monika Bokori-Brown

Toxins 2022, 14(11), 757; https://doi.org/10.3390/toxins14110757 - 3 Nov 2022

Cited by 1 | Viewed by 2990

Abstract

Clostridium perfringens epsilon toxin (Etx) is a pore forming toxin that causes enterotoxaemia in ruminants and may be a cause of multiple sclerosis in humans. To date, most in vitro studies of Etx have used the Madin-Darby canine kidney (MDCK) cell line. However, [...] Read more.

Clostridium perfringens epsilon toxin (Etx) is a pore forming toxin that causes enterotoxaemia in ruminants and may be a cause of multiple sclerosis in humans. To date, most in vitro studies of Etx have used the Madin-Darby canine kidney (MDCK) cell line. However, studies using Chinese hamster ovary (CHO) cells engineered to express the putative Etx receptor, myelin and lymphocyte protein (MAL), suggest that amino acids important for Etx activity differ between species. In this study, we investigated the role of amino acids Y42, Y43 and H162, previously identified as important in Etx activity towards MDCK cells, in Etx activity towards CHO-human MAL (CHO-hMAL) cells, human red blood cells (hRBCs) and synthetic bilayers using site-directed mutants of Etx. We show that in CHO-hMAL cells Y42 is critical for Etx binding and not Y43 as in MDCK cells, indicating that surface exposed tyrosine residues in the receptor binding domain of Etx impact efficiency of cell binding to MAL-expressing cells in a species-specific manner. We also show that Etx mutant H162A was unable to lyse CHO-hMAL cells, lysed hRBCs, whilst it was able to form pores in synthetic bilayers, providing evidence of the complexity of Etx pore formation in different lipid environments. Full article

(This article belongs to the Special Issue Pore-Forming Toxin Interactions with the Membrane)

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15 pages, 3603 KiB

Open AccessEditor’s ChoiceArticle

Impact of Stagnation on the Diversity of Cyanobacteria in Drinking Water Treatment Plant Sludge

by Farhad Jalili, Hana Trigui, Juan Francisco Guerra Maldonado, Sarah Dorner, Arash Zamyadi, B. Jesse Shapiro, Yves Terrat, Nathalie Fortin, Sébastien Sauvé and Michèle Prévost

Toxins 2022, 14(11), 749; https://doi.org/10.3390/toxins14110749 - 31 Oct 2022

Cited by 3 | Viewed by 1913

Abstract

Health-related concerns about cyanobacteria-laden sludge of drinking water treatment plants (DWTPs) have been raised in the past few years. Microscopic taxonomy, shotgun metagenomic sequencing, and microcystin (MC) measurement were applied to study the fate of cyanobacteria and cyanotoxins after controlled sludge storage (stagnation) [...] Read more.

Health-related concerns about cyanobacteria-laden sludge of drinking water treatment plants (DWTPs) have been raised in the past few years. Microscopic taxonomy, shotgun metagenomic sequencing, and microcystin (MC) measurement were applied to study the fate of cyanobacteria and cyanotoxins after controlled sludge storage (stagnation) in the dark in a full-scale drinking water treatment plant within 7 to 38 days. For four out of eight dates, cyanobacterial cell growth was observed by total taxonomic cell counts during sludge stagnation. The highest observed cell growth was 96% after 16 days of stagnation. Cell growth was dominated by potential MC producers such as Microcystis, Aphanocapsa, Chroococcus, and Dolichospermum. Shotgun metagenomic sequencing unveiled that stagnation stress shifts the cyanobacterial communities from the stress-sensitive Nostocales (e.g., Dolichospermum) order towards less compromised orders and potential MC producers such as Chroococcales (e.g., Microcystis) and Synechococcales (e.g., Synechococcus). The relative increase of cyanotoxin producers presents a health challenge when the supernatant of the stored sludge is recycled to the head of the DWTP or discharged into the source. These findings emphasize the importance of a strategy to manage cyanobacteria-laden sludge and suggest practical approaches should be adopted to control health/environmental impacts of cyanobacteria and cyanotoxins in sludge. Full article

(This article belongs to the Special Issue Management of Cyanobacteria and Cyanotoxins in Waters)

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14 pages, 1787 KiB

Open AccessEditor’s ChoiceArticle

Ochratoxin A Defective Aspergillus carbonarius Mutants as Potential Biocontrol Agents

by Belén Llobregat, Luis González-Candelas and Ana-Rosa Ballester

Toxins 2022, 14(11), 745; https://doi.org/10.3390/toxins14110745 - 31 Oct 2022

Cited by 6 | Viewed by 2374

Abstract

Aspergillus carbonarius is one of the main species responsible for wine, coffee and cocoa toxin contamination. The main mycotoxin produced by this fungus, ochratoxin A (OTA), is a secondary metabolite categorized as a possible carcinogen because of its significant nephrotoxicity and immunosuppressive effects. [...] Read more.

Aspergillus carbonarius is one of the main species responsible for wine, coffee and cocoa toxin contamination. The main mycotoxin produced by this fungus, ochratoxin A (OTA), is a secondary metabolite categorized as a possible carcinogen because of its significant nephrotoxicity and immunosuppressive effects. A polyketide synthase gene (otaA) encodes the first enzyme in the OTA biosynthetic pathway. It is known that the filamentous fungi, growth, development and production of secondary metabolites are interconnected processes governed by global regulatory factors whose encoding genes are generally located outside the gene clusters involved in the biosynthesis of each secondary metabolite, such as the veA gene, which forms part of the VELVET complex. Different fungal strains compete for nutrients and space when they infect their hosts, and safer non-mycotoxigenic strains may be able to outcompete mycotoxigenic strains during colonization. To determine the possible utility of biopesticides based on the competitive exclusion of mycotoxigenic strains by non-toxigenic ones, we used A. carbonarius ΔotaA and ΔveA knockout mutants. Our results showed that during both in vitro growth and infection of grapes, non-mycotoxigenic strains could outcompete the wild-type strain. Additionally, the introduction of the non-mycotoxigenic strain led to a drastic decrease in OTA during both in vitro growth and infection of grapes. Full article

(This article belongs to the Special Issue Occurrence and Determination of Mycotoxins)

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30 pages, 5095 KiB

Open AccessEditor’s ChoiceArticle

A Combined Bioassay and Nanofractionation Approach to Investigate the Anticoagulant Toxins of Mamba and Cobra Venoms and Their Inhibition by Varespladib

by Arif Arrahman, Taline D. Kazandjian, Kristina B. M. Still, Julien Slagboom, Govert W. Somsen, Freek J. Vonk, Nicholas R. Casewell and Jeroen Kool

Toxins 2022, 14(11), 736; https://doi.org/10.3390/toxins14110736 - 27 Oct 2022

Cited by 6 | Viewed by 2475

Abstract

Envenomation by elapid snakes primarily results in neurotoxic symptoms and, consequently, are the primary focus of therapeutic research concerning such venoms. However, mounting evidence suggests these venoms can additionally cause coagulopathic symptoms, as demonstrated by some Asian elapids and African spitting cobras. This [...] Read more.

Envenomation by elapid snakes primarily results in neurotoxic symptoms and, consequently, are the primary focus of therapeutic research concerning such venoms. However, mounting evidence suggests these venoms can additionally cause coagulopathic symptoms, as demonstrated by some Asian elapids and African spitting cobras. This study sought to investigate the coagulopathic potential of venoms from medically important elapids of the genera Naja (true cobras), Hemachatus (rinkhals), and Dendroaspis (mambas). Crude venoms were bioassayed for coagulant effects using a plasma coagulation assay before RPLC/MS was used to separate and identify venom toxins in parallel with a nanofractionation module. Subsequently, coagulation bioassays were performed on the nanofractionated toxins, along with in-solution tryptic digestion and proteomics analysis. These experiments were then repeated on both crude venoms and on the nanofractionated venom toxins with the addition of either the phospholipase A₂ (PLA₂) inhibitor varespladib or the snake venom metalloproteinase (SVMP) inhibitor marimastat. Our results demonstrate that various African elapid venoms have an anticoagulant effect, and that this activity is significantly reduced for cobra venoms by the addition of varespladib, though this inhibitor had no effect against anticoagulation caused by mamba venoms. Marimastat showed limited capacity to reduce anticoagulation in elapids, affecting only N. haje and H. haemachatus venom at higher doses. Proteomic analysis of nanofractionated toxins revealed that the anticoagulant toxins in cobra venoms were both acidic and basic PLA₂s, while the causative toxins in mamba venoms remain uncertain. This implies that while PLA₂ inhibitors such as varespladib and metalloproteinase inhibitors such as marimastat are viable candidates for novel snakebite treatments, they are not likely to be effective against mamba envenomings. Full article

(This article belongs to the Special Issue New Trends in Identification and Characterization of Venom Components)

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14 pages, 2065 KiB

Open AccessEditor’s ChoiceArticle

A Robust and Sensitive Spectrophotometric Assay for the Enzymatic Activity of Bacterial Adenylate Cyclase Toxins

by Marilyne Davi, Mirko Sadi, Irene Pitard, Alexandre Chenal and Daniel Ladant

Toxins 2022, 14(10), 691; https://doi.org/10.3390/toxins14100691 - 8 Oct 2022

Cited by 1 | Viewed by 2059

Abstract

Various bacterial pathogens are producing toxins that target the cyclic Nucleotide Monophosphate (cNMPs) signaling pathways in order to facilitate host colonization. Among them, several are exhibiting potent nucleotidyl cyclase activities that are activated by eukaryotic factors, such as the adenylate cyclase (AC) toxin, [...] Read more.

Various bacterial pathogens are producing toxins that target the cyclic Nucleotide Monophosphate (cNMPs) signaling pathways in order to facilitate host colonization. Among them, several are exhibiting potent nucleotidyl cyclase activities that are activated by eukaryotic factors, such as the adenylate cyclase (AC) toxin, CyaA, from Bordetella pertussis or the edema factor, EF, from Bacillus anthracis. The characterization of these toxins frequently requires accurate measurements of their enzymatic activity in vitro, in particular for deciphering their structure-to-function relationships by protein engineering and site-directed mutagenesis. Here we describe a simple and robust in vitro assay for AC activity based on the spectrophotometric detection of cyclic AMP (cAMP) after chromatographic separation on aluminum oxide. This assay can accurately detect down to fmol amounts of B. pertussis CyaA and can even be used in complex media, such as cell extracts. The relative advantages and disadvantages of this assay in comparison with other currently available methods are briefly discussed. Full article

(This article belongs to the Special Issue Toxins: Mr Hyde or Dr Jekyll?)

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16 pages, 3208 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Cytokine and Chemokine Retention Profile in COVID-19 Patients with Chronic Kidney Disease

by Paola Ciceri, Valeria Bono, Lorenza Magagnoli, Matteo Sala, Antonella d’Arminio Monforte, Andrea Galassi, Alessandra Barassi, Giulia Marchetti and Mario Cozzolino

Toxins 2022, 14(10), 673; https://doi.org/10.3390/toxins14100673 - 28 Sep 2022

Cited by 8 | Viewed by 2266

Abstract

Chronic kidney disease (CKD) patients are more susceptible to infections compared to the general population. SARS-CoV-2 virus pathology is characterized by a cytokine storm responsible for the systemic inflammation typical of the COVID-19 disease. Since CKD patients have a reduced renal clearance, we [...] Read more.

Chronic kidney disease (CKD) patients are more susceptible to infections compared to the general population. SARS-CoV-2 virus pathology is characterized by a cytokine storm responsible for the systemic inflammation typical of the COVID-19 disease. Since CKD patients have a reduced renal clearance, we decided to investigate whether they accumulate harmful mediators during the COVID-19 disease. We conducted a retrospective study on 77 COVID-19 hospitalized subjects in the acute phase of the illness. Thirteen different cytokines were assessed in plasma collected upon hospitalization. The patients were divided into three groups according to their estimated glomerular filtration rate, eGFR < 30 (n = 23), 30 < eGFR < 60 (n = 33), eGFR > 60 mL/min (n = 21). We found that Tumor Necrosis Factor α and its receptors I and II, Interleukin-7, Leukemia Inhibitory Factor, FAS receptor, Chitinase 3-like I, and the Vascular Endothelial Growth Factor showed an increased accumulation that negatively correlate with eGFR. Moreover, non-survivor patients with an impaired kidney function have significantly more elevated levels of the same mediators. In conclusion, there is a tendency in COVID-19 ESRD patients to accumulate harmful cytokines. The accumulation seems to associate with mortality outcomes and may be due to reduced clearance but also to increased biosynthesis in most severe cases. Full article

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10 pages, 985 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Potassium Bioaccessibility in Uncooked and Cooked Plant Foods: Results from a Static In Vitro Digestion Methodology

by Costanza Ceccanti, Lucia Guidi, Claudia D’Alessandro and Adamasco Cupisti

Toxins 2022, 14(10), 668; https://doi.org/10.3390/toxins14100668 - 26 Sep 2022

Cited by 8 | Viewed by 3364

Abstract

Hyperkalemia is a major concern in chronic kidney disease and in end-stage renal disease, representing a predictor of hospitalization and mortality. To prevent and treat hyperkalemia, dietary management is of great clinical interest. Currently, the growing use of plant-based diets causes an increasing [...] Read more.

Hyperkalemia is a major concern in chronic kidney disease and in end-stage renal disease, representing a predictor of hospitalization and mortality. To prevent and treat hyperkalemia, dietary management is of great clinical interest. Currently, the growing use of plant-based diets causes an increasing concern about potassium load in renal patients. The aim of this study was to assess the bioaccessibility of potassium in vegetables, concerning all aspects of the plants (fruit, flower, root, tuber, leaf and seed) and to what extent different boiling techniques affect potassium content and bioaccessibility of plant-based foods. Bioaccessibility was evaluated by an in vitro digestion methodology, resembling human gastro-intestinal tract. Potassium content was higher in seeds and leaves, despite it not being possible to define a common “rule” according to the type of organ, namely seed, leaf or fruit. Boiling reduced potassium content in all vegetables excluding carrot, zucchini, and cauliflower; boiling starting from cold water contributed to a greater reduction of the potassium content in potato, peas, and beans. Bioaccessibility after in vitro digestion ranged from 12 (peas) to 93% (tomato) regardless of species and organs. Higher bioaccessibility was found in spinach, chicory, zucchini, tomato, kiwi, and cauliflower, and lower bioaccessibility in peas. Potassium from leaf resulted in the highest bioaccessibility after digestion; as a whole potassium bioaccessibility in the fruits and vegetables studied was 67% on average, with differences in relation to the different organs and species. Further, considering the method of boiling to reduce potassium content, these data indicate that the effective potassium load from plant-based foods may be lower than originally expected. This supports the clinical advices to maintain a wide use of plant-based food in the management of renal patients. Full article

(This article belongs to the Special Issue New Strategies for the Reduction of Uremic Toxins (2022))

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24 pages, 5411 KiB

Open AccessEditor’s ChoiceArticle

Longitudinal Metabolomics and Lipidomics Analyses Reveal Alterations Associated with Envenoming by Bothrops asper and Daboia russelii in an Experimental Murine Model

by Nishikant Wase, José María Gutiérrez, Alexandra Rucavado and Jay W. Fox

Toxins 2022, 14(10), 657; https://doi.org/10.3390/toxins14100657 - 23 Sep 2022

Cited by 2 | Viewed by 2137

Abstract

Longitudinal metabolomics and lipidomics analyses were carried out on the blood plasma of mice injected intramuscularly with venoms of the viperid species Bothrops asper or Daboia russelii. Blood samples were collected 1, 3, 6, and 24 h after venom injection, and a [...] Read more.

Longitudinal metabolomics and lipidomics analyses were carried out on the blood plasma of mice injected intramuscularly with venoms of the viperid species Bothrops asper or Daboia russelii. Blood samples were collected 1, 3, 6, and 24 h after venom injection, and a control group of non-envenomed mice was included. Significant perturbations in metabolomics and lipidomics were observed at 1, 3, and 6 h, while values returned close to those of control mice by 24 h, hence reflecting a transient pattern of metabolic disturbance. Both venoms induced significant changes in amino acids, as well as in several purines and pyrimidines, and in some metabolites of the tricarboxylic acid cycle. KEGG analysis of metabolic pathways that showed those with the greatest change included aminoacyl tRNA synthesis and amino acid biosynthesis and metabolism pathways. With regard to lipid metabolism, there was an increase in triglycerides and some acyl carnitines and a concomitant drop in the levels of some phospholipids. In addition, envenomed mice had higher levels of cortisol, heme, and some oxidative stress markers. The overall pattern of metabolic changes in envenomed mice bears similarities with the patterns described in several traumatic injuries, thus underscoring a metabolic response/adaptation to the injurious action of the venoms. Full article

(This article belongs to the Section Animal Venoms)

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16 pages, 3222 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Improved Analytical Approach for Determination of Tropane Alkaloids in Leafy Vegetables Based on µ-QuEChERS Combined with HPLC-MS/MS

by Lorena González-Gómez, Sonia Morante-Zarcero, Jorge A. M. Pereira, José S. Câmara and Isabel Sierra

Toxins 2022, 14(10), 650; https://doi.org/10.3390/toxins14100650 - 20 Sep 2022

Cited by 14 | Viewed by 3251

Abstract

This work presents an optimized methodology based on the miniaturization of the original QuEChERS (μ-QuEChERS) followed by liquid chromatography coupled to mass spectrometry (HPLC-MS/MS) for the determination of tropane alkaloids (TAs), atropine, and scopolamine in leafy vegetable samples. The analytical methodology was successfully [...] Read more.

This work presents an optimized methodology based on the miniaturization of the original QuEChERS (μ-QuEChERS) followed by liquid chromatography coupled to mass spectrometry (HPLC-MS/MS) for the determination of tropane alkaloids (TAs), atropine, and scopolamine in leafy vegetable samples. The analytical methodology was successfully validated, demonstrating quantitation limits (MQL) ≤ 2.3 ng/g, good accuracy, and precision, with recoveries between 90–100% and RSD ≤ 13% for both analytes. The method was applied to the analysis of TA-producing plants (Brugmansia versicolor, Solandra maxima, and Convolvulus arvensis). High concentrations of scopolamine were found in flowers (1771 mg/kg) and leaves (297 mg/kg) of B. versicolor. The highest concentration of atropine was found in flowers of S. maxima (10.4 mg/kg). Commercial mixed leafy vegetables contaminated with B. versicolor and S. maxima were analysed to verify the efficacy of the method, showing recoveries between 82 and 110% for both analytes. Finally, the method was applied to the analysis of eighteen samples of leafy vegetables, finding atropine in three samples of mixed leafy vegetables, with concentrations of 2.7, 3.2, and 3.4 ng/g, and in nine samples with concentrations ≤MQL. In turn, scopolamine was only found in a sample of chopped Swiss chard with a concentration ≤MQL. Full article

(This article belongs to the Section Plant Toxins)

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16 pages, 2298 KiB

Open AccessEditor’s ChoiceArticle

The Origin of Teratogenic Retinoids in Cyanobacteria

by Luděk Sehnal, Marie Smutná, Lucie Bláhová, Pavel Babica, Petra Šplíchalová and Klára Hilscherová

Toxins 2022, 14(9), 636; https://doi.org/10.3390/toxins14090636 - 15 Sep 2022

Cited by 6 | Viewed by 2289

Abstract

Although information about the occurrence and distribution of retinoids in the environment is scarce, cyanobacterial water blooms have been identified as a significant source of these small molecules. Despite the confirmed presence of retinoids in the freshwater blooms dominated by cyanobacteria and their [...] Read more.

Although information about the occurrence and distribution of retinoids in the environment is scarce, cyanobacterial water blooms have been identified as a significant source of these small molecules. Despite the confirmed presence of retinoids in the freshwater blooms dominated by cyanobacteria and their described teratogenic effects, reliable identification of retinoid producers and the mechanism of their biosynthesis is missing. In this study, the cultures of several taxonomically diverse species of axenic cyanobacteria were confirmed as significant producers of retinoid-like compounds. The consequent bioinformatic analysis suggested that the enzymatic background required for the biosynthesis of all-trans retinoic acid from retinal is not present across phylum Cyanobacteria. However, we demonstrated that retinal conversion into other retinoids can be mediated non-enzymatically by free radical oxidation, which leads to the production of retinoids widely detected in cyanobacteria and environmental water blooms, such as all-trans retinoic acid or all-trans 5,6epoxy retinoic acid. Importantly, the production of these metabolites by cyanobacteria in association with the mass development of water blooms can lead to adverse impacts in aquatic ecosystems regarding the described teratogenicity of retinoids. Moreover, our finding that retinal can be non-enzymatically converted into more bioactive retinoids, also in water, and out of the cells, increases the environmental significance of this process. Full article

(This article belongs to the Special Issue Cyanobacterial Toxin and Secondary Metabolite Detection, Fate and Toxicity Assessment)

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18 pages, 5014 KiB

Open AccessEditor’s ChoiceArticle

A Uremic Pig Model for Peritoneal Dialysis

by Joost C. de Vries, Maaike K. van Gelder, Anneke S. Monninkhof, Sabbir Ahmed, Diënty H. M. Hazenbrink, Tri Q. Nguyen, Gèrard A. P. de Kort, Evert-Jan P. A. Vonken, Koen R. D. Vaessen, Jaap A. Joles, Marianne C. Verhaar and Karin G. F. Gerritsen

Toxins 2022, 14(9), 635; https://doi.org/10.3390/toxins14090635 - 14 Sep 2022

Cited by 3 | Viewed by 2555

Abstract

With increasing interest in home dialysis, there is a need for a translational uremic large animal model to evaluate technical innovations in peritoneal dialysis (PD). To this end, we developed a porcine model with kidney failure. Stable chronic kidney injury was induced by [...] Read more.

With increasing interest in home dialysis, there is a need for a translational uremic large animal model to evaluate technical innovations in peritoneal dialysis (PD). To this end, we developed a porcine model with kidney failure. Stable chronic kidney injury was induced by bilateral subtotal renal artery embolization. Before applying PD, temporary aggravation of uremia was induced by administration of gentamicin (10 mg/kg i.v. twice daily for 7 days), to obtain uremic solute levels within the range of those of dialysis patients. Peritoneal transport was assessed using a standard peritoneal permeability assessment (SPA). After embolization, urea and creatinine concentrations transiently increased from 1.6 ± 0.3 to 7.5 ± 1.2 mM and from 103 ± 14 to 338 ± 67 µM, respectively, followed by stabilization within 1–2 weeks to 2.5 ± 1.1 mM and 174 ± 28 µM, respectively. Gentamicin induced temporary acute-on-chronic kidney injury with peak urea and creatinine concentrations of 16.7 ± 5.3 mM and 932 ± 470 µM respectively. PD was successfully applied, although frequently complicated by peritonitis. SPA showed a low transport status (D/P creatinine at 4 h of 0.41 (0.36–0.53)) with a mass transfer area coefficient of 9.6 ± 3.1, 4.6 ± 2.6, 3.4 ± 2.3 mL/min for urea, creatinine, and phosphate respectively. In conclusion, this porcine model with on-demand aggravation of uremia is suitable for PD albeit with peritoneal transport characterized by a low transport status. Full article

(This article belongs to the Special Issue Analysis and Applications of Uremic Animal Models)

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11 pages, 978 KiB

Open AccessEditor’s ChoiceArticle

Bites by Non-Native Reptiles in France: Species, Circumstances and Outcome

by Gaël Le Roux, Guillaume Grenet, Corinne Schmitt, French Poison Control Centers Research Group, Sébastien Larréché and Alexis Descatha

Toxins 2022, 14(8), 570; https://doi.org/10.3390/toxins14080570 - 20 Aug 2022

Cited by 3 | Viewed by 2121

Abstract

We aimed to make an exhaustive assessment of circumstances of bites by exotic reptiles bred in France. A retrospective observational study was conducted in all the reported cases from 2000 to 2020 in French poison control centers (PCCs). Two hundred and eighteen cases [...] Read more.

We aimed to make an exhaustive assessment of circumstances of bites by exotic reptiles bred in France. A retrospective observational study was conducted in all the reported cases from 2000 to 2020 in French poison control centers (PCCs). Two hundred and eighteen cases of bites were recorded. The sex ratio (M/F) of the patients was 1.79 and the mean age of the patients was 29.0 ± 15.8 years. Twenty-two cases (10.1%) occurred during the deep night. One hundred and eighty-six bites (85.7%) occurred in a private context; however, there were more cases of high severity when it occurred in a professional setting (60.0% vs. 11.2%, p < 0.01). The feeding/nursing activity accounted for 54.7% cases. Forty-three species of snake were identified; 28 were considered venomous. There were no deaths among the patients in the study. Most of the cases (85.8%) were of mild severity. All of the patients bitten by a venomous reptile were hospitalized: 10 patients received an antivenom; and 2 required surgery. Bites occurred at home and by a small number of popular non-venomous reptile species (pythons and boas, colubrids). These occurred mainly when handling the animals. The rare envenomations were mainly by Asian and American crotalids, followed by elapids. One-third of them were treated with antivenom when available. Full article

(This article belongs to the Special Issue Snakebite and Clinical Toxinology)

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16 pages, 3023 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Isolation, Characterization and Biological Action of Type-1 Ribosome-Inactivating Proteins from Tissues of Salsola soda L.

by Nicola Landi, Sara Ragucci, Lucía Citores, Angela Clemente, Hafiza Z. F. Hussain, Rosario Iglesias, José M. Ferreras and Antimo Di Maro

Toxins 2022, 14(8), 566; https://doi.org/10.3390/toxins14080566 - 19 Aug 2022

Cited by 11 | Viewed by 2556

Abstract

Ribosome-inactivating proteins (RIPs) are known as RNA N-glycosylases. They depurinate the major rRNA, damaging ribosomes and inhibiting protein synthesis. Here, new single-chain (type-1) RIPs named sodins were isolated from the seeds (five proteins), edible leaves (one protein) and roots (one protein) of Salsola [...] Read more.

Ribosome-inactivating proteins (RIPs) are known as RNA N-glycosylases. They depurinate the major rRNA, damaging ribosomes and inhibiting protein synthesis. Here, new single-chain (type-1) RIPs named sodins were isolated from the seeds (five proteins), edible leaves (one protein) and roots (one protein) of Salsola soda L. Sodins are able to release Endo’s fragment when incubated with rabbit and yeast ribosomes and inhibit protein synthesis in cell-free systems (IC₅₀ = 4.83–79.31 pM). In addition, sodin 5, the major form isolated from seeds, as well as sodin eL and sodin R, isolated from edible leaves and roots, respectively, display polynucleotide:adenosine glycosylase activity and are cytotoxic towards the Hela and COLO 320 cell lines (IC₅₀ = 0.41–1200 nM), inducing apoptosis. The further characterization of sodin 5 reveals that this enzyme shows a secondary structure similar to other type-1 RIPs and a higher melting temperature (Tm = 76.03 ± 0.30 °C) and is non-glycosylated, as other sodins are. Finally, we proved that sodin 5 possesses antifungal activity against Penicillium digitatum. Full article

(This article belongs to the Special Issue Biological Activities of Ribosome-Inactivating Proteins)

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13 pages, 1661 KiB

Open AccessEditor’s ChoiceArticle

Iridoid Glycosides Isolated from Bellardia trixago Identified as Inhibitors of Orobanche cumana Radicle Growth

by Gabriele Soriano, Antonietta Siciliano, Mónica Fernández-Aparicio, Antonio Cala Peralta, Marco Masi, Antonio Moreno-Robles, Marco Guida and Alessio Cimmino

Toxins 2022, 14(8), 559; https://doi.org/10.3390/toxins14080559 - 17 Aug 2022

Cited by 4 | Viewed by 2295

Abstract

Orobanche cumana is an obligate holoparasitic plant with noxious effects in sunflower crops. Bellardia trixago is a facultative hemiparasitic plant that infects ruderal plants without noxious significance in agriculture and is known to produce a wide spectrum of bioactive metabolites. The objective of [...] Read more.

Orobanche cumana is an obligate holoparasitic plant with noxious effects in sunflower crops. Bellardia trixago is a facultative hemiparasitic plant that infects ruderal plants without noxious significance in agriculture and is known to produce a wide spectrum of bioactive metabolites. The objective of this study was to evaluate the allelopathic effects of B. trixago on the growth of O. cumana seedlings. Three different extracts using solvents of increasing polarity (n-hexane, dichloromethane and ethyl acetate) were prepared from the flowers, aerial green organs and roots of two populations, a white-flowered and a yellow-flowered population of B. trixago, both collected in southern Spain. Each extract was studied using allelopathic screenings on O. cumana which resulted in the identification of allelopathic activity of the ethyl acetate extracts against Orobanche radicles. Five iridoid glycosides were isolated together with benzoic acid from the ethyl acetate extract of aerial green organs by bio-guided purification. These compounds were identified as bartsioside, melampyroside, mussaenoside, gardoside methyl ester and aucubin. Among them, melampyroside was found to be the most abundant constituent in the extract (44.3% w/w), as well as the most phytotoxic iridoid on O. cumana radicle, showing a 72.6% inhibition of radicle growth. This activity of melampyroside was significantly high when compared with the inhibitory activity of benzoic acid (25.9%), a phenolic acid with known allelopathic activity against weeds. The ecotoxicological profile of melampyroside was evaluated using organisms representing different trophic levels of the aquatic and terrestrial ecosystems, namely producers (green freshwater algae Raphidocelis subcapitata and macrophyte Lepidium sativum), consumers (water flea Daphnia magna and nematode Caenorhabditis elegans) and decomposers (bacterium Aliivibrio fischeri). The ecotoxicity of melampyroside differed significantly depending on the test organism showing the highest toxicity to daphnia, nematodes and bacteria, and a lower toxicity to algae and macrophytes. The findings of the present study may provide useful information for the generation of green alternatives to synthetic herbicides for the control of O. cumana. Full article

(This article belongs to the Special Issue Biological Activities and Potential Applications of Phytotoxins)

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31 pages, 3760 KiB

Open AccessEditor’s ChoiceArticle

Microcalorimetric Investigations of Reversible Staphylococcal Enterotoxin Unfolding

by Susan C. Berry, Odbert A. Triplett, Li-Rong Yu, Mark E. Hart, Lauren S. Jackson and William H. Tolleson

Toxins 2022, 14(8), 554; https://doi.org/10.3390/toxins14080554 - 15 Aug 2022

Cited by 2 | Viewed by 3024

Abstract

Staphylococcal food poisoning (SFP) is a common food-borne illness often associated with contamination during food handling. The genes for Staphylococcal enterotoxin (SE) isoforms SEA and SEB are frequently detected in human nasal Staphylococcus aureus isolates and these toxins are commonly associated with SFP. [...] Read more.

Staphylococcal food poisoning (SFP) is a common food-borne illness often associated with contamination during food handling. The genes for Staphylococcal enterotoxin (SE) isoforms SEA and SEB are frequently detected in human nasal Staphylococcus aureus isolates and these toxins are commonly associated with SFP. Past studies described the resistance of preformed SE proteins to heat inactivation and their reactivation upon cooling in foods. Full thermodynamic analyses for these processes have not been reported, however. The thermal stabilities of SEA, SEB, and SEH and reversibility of unfolding in simple buffers were investigated at pH 4.5 and pH 6.8 using differential scanning calorimetry (DSC). SEA and SEB unfolding was irreversible at pH 6.8 and at least partially reversible at pH 4.5 while SEH unfolding was irreversible at pH 4.5 and reversible at pH 6.8. Additional studies showed maximum refolding for SEB at pH 3.5–4.0 and diminished refolding at pH 4.5 with increasing ionic strength. SE-stimulated secretion of interferon-gamma by human peripheral blood mononuclear cells was used to assess residual SE biological activity following heat treatments using conditions matching those used for DSC studies. The biological activities of SEB and SEH exhibited greater resistance to heat inactivation than that of SEA. The residual activities of heat-treated SEB and SEH were measurable but diminished further in the presence of reconstituted nonfat dry milk adjusted to pH 4.5 or pH 6.8. To different extents, the pH and ionic strengths typical for foods influenced the thermal stabilities of SEA, SEB, and SEH and their potentials to renature spontaneously after heat treatments. Full article

(This article belongs to the Section Bacterial Toxins)

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16 pages, 2729 KiB

Open AccessEditor’s ChoiceArticle

Modification of Deoxynivalenol by a Fungal Laccase Paired with Redox Mediator TEMPO

by Hina Shanakhat, Susan P. McCormick, Mark Busman, Joseph O. Rich and Matthew G. Bakker

Toxins 2022, 14(8), 548; https://doi.org/10.3390/toxins14080548 - 11 Aug 2022

Cited by 3 | Viewed by 2603

Abstract

Mycotoxins such as deoxynivalenol introduce a health risk to the food supply and are costly to manage or avoid. Technologies for reducing or eliminating the toxicity of deoxynivalenol could be useful in a variety of processes, such as in preserving the value as [...] Read more.

Mycotoxins such as deoxynivalenol introduce a health risk to the food supply and are costly to manage or avoid. Technologies for reducing or eliminating the toxicity of deoxynivalenol could be useful in a variety of processes, such as in preserving the value as animal feed of byproducts of ethanol production. We characterized transformation products of deoxynivalenol that were formed by the combination of a fungal laccase paired with the chemical mediator 2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPO), using chromatography, mass spectrometry, and nuclear magnetic resonance spectroscopy. Alcohol groups at the C3 and C15 positions of deoxynivalenol were oxidized to ketones, and the chemical mediator became covalently linked to the C4 position. Conditions experienced during gas chromatography led to the dissociation of TEMPO, forming 3,15-diketodeoxynivalenol. Understanding the range of possible modifications to deoxynivalenol and other trichothecenes is a necessary step toward effective remediation of contaminated grain. Full article

(This article belongs to the Special Issue Innovative Mycotoxin Detoxification: Discoveries, Mechanisms and Applications)

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12 pages, 2186 KiB

Open AccessEditor’s ChoiceArticle

Application of Aspergillus niger Fumonisin Amine Oxidase (AnFAO) to Detoxify Fumonisin-Contaminated Maize

by Patrick G. Telmer, Megan J. Kelman, Justin B. Renaud, Mark W. Sumarah and Christopher P. Garnham

Toxins 2022, 14(8), 544; https://doi.org/10.3390/toxins14080544 - 9 Aug 2022

Cited by 6 | Viewed by 2308

Abstract

Fumonisin mycotoxins are a family of secondary metabolites produced by Fusarium verticillioides and related species, as well as some strains of Aspergillus niger. Fumonisin contamination of maize is a concern when grown under hot, dry conditions. When present above regulatory levels, there [...] Read more.

Fumonisin mycotoxins are a family of secondary metabolites produced by Fusarium verticillioides and related species, as well as some strains of Aspergillus niger. Fumonisin contamination of maize is a concern when grown under hot, dry conditions. When present above regulatory levels, there can be effects on animal health. New tools to reduce the toxicity of maize and maize products with high concentrations of fumonisin are needed. Recently, we reported an amine oxidase (AnFAO) from a fumonisin-producing Aspergillus niger strain capable of oxidatively deaminating intact fumonisins. In this study, AnFAO was used to reduce intact fumonisin concentrations in milled maize flour, whole kernel maize inoculated with fumonisin-producing Fusarium verticillioides, and dried distillers’ grains with solubles (DDGS). The data showed that milled maize flour incubated with 1 µM AnFAO for 1 h resulted in complete deamination of FB₁ and FB₂. A greater than 90% reduction in FB_1–3 concentrations was observed following a simple washing procedure of whole kernel maize in the presence of 1 µM AnFAO for 1 h. Similarly, a ≥86% reduction in FB_1–3 concentrations was observed in DDGS after 4 h incubation with 1 µM AnFAO. Finally, we engineered the methylotrophic yeast Pichia pastoris to produce functional AnFAO in both a secreted and intracellular form. These results support the further development and application of AnFAO as a promising tool to remediate fumonisin-contaminated maize and maize products. Full article

(This article belongs to the Special Issue Innovative Mycotoxin Detoxification: Discoveries, Mechanisms and Applications)

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24 pages, 3761 KiB

Open AccessEditor’s ChoiceArticle

Diversity of Phospholipases A₂ from Bothrops atrox Snake Venom: Adaptive Advantages for Snakes Compromising Treatments for Snakebite Patients

by Leijiane F. Sousa, Amanda P. Freitas, Bruna L. Cardoso, Tiago H. M. Del-Rei, Vanessa A. Mendes, Daniele P. Oréfice, Marisa M. T. Rocha, Benedito C. Prezoto and Ana M. Moura-da-Silva

Toxins 2022, 14(8), 543; https://doi.org/10.3390/toxins14080543 - 8 Aug 2022

Cited by 2 | Viewed by 3155

Abstract

The evolution of snake venoms resulted in multigene toxin families that code for structurally similar isoforms eventually harboring distinct functions. PLA₂s are dominant toxins in viper venoms, and little is known about the impact of their diversity on human envenomings and [...] Read more.

The evolution of snake venoms resulted in multigene toxin families that code for structurally similar isoforms eventually harboring distinct functions. PLA₂s are dominant toxins in viper venoms, and little is known about the impact of their diversity on human envenomings and neutralization by antivenoms. Here, we show the isolation of three distinct PLA₂s from B. atrox venom. FA1 is a Lys-49 homologue, and FA3 and FA4 are catalytic Asp-49 PLA₂s. FA1 and FA3 are basic myotoxic proteins, while FA4 is an acid non-myotoxic PLA₂. FA3 was the most potent toxin, inducing higher levels of edema, inflammatory nociception, indirect hemolysis, and anticoagulant activity on human, rat, and chicken plasmas. FA4 presented lower anticoagulant activity, and FA1 had only a slight effect on human and rat plasmas. PLA₂s presented differential reactivities with antivenoms, with an emphasis on FA3, which was not recognized or neutralized by the antivenoms used in this study. Our findings reveal the functional and antigenic diversity among PLA₂s from B. atrox venom, highlighting the importance of assessing venom variability for understanding human envenomations and treatment with antivenoms, particularly evident here as the antivenom fails to recognize FA3, the most active multifunctional toxin described. Full article

(This article belongs to the Special Issue Animal Venoms and Their Components: Molecular Mechanisms of Action v 2.0)

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26 pages, 2326 KiB

Open AccessEditor’s ChoiceArticle

Pooled Analysis of Real-World Evidence Supports Anti-CGRP mAbs and OnabotulinumtoxinA Combined Trial in Chronic Migraine

by Damiana Scuteri, Paolo Tonin, Pierluigi Nicotera, Marilù Vulnera, Giuseppina Cristina Altieri, Assunta Tarsitano, Giacinto Bagetta and Maria Tiziana Corasaniti

Toxins 2022, 14(8), 529; https://doi.org/10.3390/toxins14080529 - 1 Aug 2022

Cited by 20 | Viewed by 3980

Abstract

OnabotulinumtoxinA, targeting the CGRP machinery, has been approved for the last two decades for chronic migraine prevention. The recently approved monoclonal antibodies (mAbs) directed towards the calcitonin gene-related peptide (CGRP) pathway open a new age for chronic migraine control. However, some 40% patients [...] Read more.

OnabotulinumtoxinA, targeting the CGRP machinery, has been approved for the last two decades for chronic migraine prevention. The recently approved monoclonal antibodies (mAbs) directed towards the calcitonin gene-related peptide (CGRP) pathway open a new age for chronic migraine control. However, some 40% patients suffering from chronic migraine is still resistant to treatment. The aim of this work is to answer the following PICOS (participants intervention comparator outcome study design) question: Is there evidence of efficacy and safety of the combined administration of anti-CGRP mAbs and onabotulinumtoxinA in chronic migraine? A systematic review and meta-analysis [Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 recommendations] was made up to 19 April 2022. The results are encouraging: the combined treatment proved to afford ≥50% monthly headache days (MHDs)/frequency reduction respect to baseline in up to 58.8% of patients; in comparison, anti-CGRP mAbs reduce MHDs of 1.94 days from baseline and botulinum toxin of 1.86 days. Our study demonstrates for the first time that the combination therapy of onabotulinumtoxinA with anti-CGRP mAbs affords a reduction of 2.67 MHDs with respect to onabotulinumtoxinA alone, with moderate certainty of evidence. Adequately powered, good-quality studies are needed to confirm the response to combination therapy in terms of efficacy and safety. PROSPERO registration: CRD42022313640. Full article

(This article belongs to the Special Issue Botulinum Neurotoxin: Shared/Common Mechanisms in the Treatment of Pain, Spasticity and Movement Disorders)

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14 pages, 2356 KiB

Open AccessEditor’s ChoiceArticle

Measurement of Fumonisins in Maize Using a Portable Mass Spectrometer

by Chris M. Maragos, Kristin Barnett, Luke Morgan, Martha M. Vaughan and Kristal K. Sieve

Toxins 2022, 14(8), 523; https://doi.org/10.3390/toxins14080523 - 30 Jul 2022

Cited by 7 | Viewed by 2485

Abstract

Fumonisins are a group of mycotoxins that routinely contaminate maize. Their presence is monitored at multiple stages from harvest to final product. Immunoassays are routinely used to screen commodities in the field while laboratory-based methods, such as mass spectrometry (MS), are used for [...] Read more.

Fumonisins are a group of mycotoxins that routinely contaminate maize. Their presence is monitored at multiple stages from harvest to final product. Immunoassays are routinely used to screen commodities in the field while laboratory-based methods, such as mass spectrometry (MS), are used for confirmation. The use of a portable mass spectrometer unlocks the potential to conduct confirmatory analyses outside of traditional laboratories. Herein, a portable mass spectrometer was used to measure fumonisins in maize. Samples were extracted with aqueous methanol, cleaned up on an immunoaffinity column, and tested with the portable MS. The limits of detection were 0.15, 0.19, and 0.28 mg/kg maize for fumonisins B₁ (FB₁), FB₂/FB₃, and total fumonisins, respectively. The corresponding limits of quantitation in maize were 0.33, 0.59, and 0.74 mg/kg. Recoveries ranged from 93.6% to 108.6%. However, RSDs ranged from 12.0 to 29.8%. The method was applied to the detection of fumonisins in 64 samples of maize collected as part of the Illinois Department of Agriculture’s monitoring program. Good correlations were observed between the portable MS and a laboratory-based LC-MS method (r² from 0.9132 to 0.9481). Results suggest the portable MS can be applied to the measurement of fumonisins in maize at levels relevant to international regulations. Full article

(This article belongs to the Special Issue Advances towards Novel Rapid and Eco-Friendly Methods for the Analysis of Mycotoxins in Food)

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13 pages, 1343 KiB

Open AccessEditor’s ChoiceArticle

Acer pseudoplatanus: A Potential Risk of Poisoning for Several Herbivore Species

by Benoît Renaud, Caroline-Julia Kruse, Anne-Christine François, Lisa Grund, Carolin Bunert, Lucie Brisson, François Boemer, Gilbert Gault, Barbara Ghislain, Thierry Petitjean, Pascal Gustin and Dominique-Marie Votion

Toxins 2022, 14(8), 512; https://doi.org/10.3390/toxins14080512 - 26 Jul 2022

Cited by 7 | Viewed by 2637

Abstract

Acer pseudoplatanus is a worldwide-distributed tree which contains toxins, among them hypoglycin A (HGA). This toxin is known to be responsible for poisoning in various species, including humans, equids, Père David’s deer and two-humped camels. We hypothesized that any herbivore pasturing with A. [...] Read more.

Acer pseudoplatanus is a worldwide-distributed tree which contains toxins, among them hypoglycin A (HGA). This toxin is known to be responsible for poisoning in various species, including humans, equids, Père David’s deer and two-humped camels. We hypothesized that any herbivore pasturing with A. pseudoplatanus in their vicinity may be at risk for HGA poisoning. To test this hypothesis, we surveyed the HGA exposure from A. pseudoplatanus in species not yet described as being at risk. Animals in zoological parks were the major focus, as they are at high probability to be exposed to A. pseudoplatanus in enclosures. We also searched for a toxic metabolite of HGA (i.e., methylenecyclopropylacetyl-carnitine; MCPA-carnitine) in blood and an alteration of the acylcarnitines profile in HGA-positive animals to document the potential risk of declaring clinical signs. We describe for the first instance cases of HGA poisoning in Bovidae. Two gnus (Connochaetes taurinus taurinus) exposed to A. pseudoplatanus in their enclosure presented severe clinical signs, serum HGA and MCPA-carnitine and a marked modification of the acylcarnitines profile. In this study, even though all herbivores were exposed to A. pseudoplatanus, proximal fermenters species seemed less susceptible to HGA poisoning. Therefore, a ruminal transformation of HGA is hypothesized. Additionally, we suggest a gradual alteration of the fatty acid metabolism in case of HGA poisoning and thus the existence of subclinical cases. Full article

(This article belongs to the Special Issue Animal Poisoning: Toxins from Plants or Feed)

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