Toxins

Article

19 pages, 3160 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Differential Antivenom and Small-Molecule Inhibition of Novel Coagulotoxic Variations in Atropoides, Cerrophidion, Metlapilcoatlus, and Porthidium American Viperid Snake Venoms

by Lee Jones, Nicholas J. Youngman, Edgar Neri-Castro, Alid Guadarrama-Martínez, Matthew R. Lewin, Rebecca Carter, Nathaniel Frank and Bryan G. Fry

Toxins 2022, 14(8), 511; https://doi.org/10.3390/toxins14080511 - 26 Jul 2022

Cited by 8 | Viewed by 3426

Abstract

Within Neotropical pit-vipers, the Mexican/Central-American clade consisting of Atropoides, Cerrophidion, Metlapilcoatlus, and Porthidium is a wide-ranging, morphologically and ecologically diverse group of snakes. Despite their prevalence, little is known of the functional aspects of their venoms. This study aimed to [...] Read more.

Within Neotropical pit-vipers, the Mexican/Central-American clade consisting of Atropoides, Cerrophidion, Metlapilcoatlus, and Porthidium is a wide-ranging, morphologically and ecologically diverse group of snakes. Despite their prevalence, little is known of the functional aspects of their venoms. This study aimed to fill the knowledge gap regarding coagulotoxic effects and to examine the potential of different therapeutic approaches. As a general trait, the venoms were shown to be anticoagulant but were underpinned by diverse biochemical actions. Pseudo-procoagulant activity (i.e., thrombin-like), characterized by the direct cleavage of fibrinogen to form weak fibrin clots, was evident for Atropoides picadoi, Cerrophidiontzotzilorum, Metlapilcoatlus mexicanus, M. nummifer, M. occiduus, M. olmec, and Porthidium porrasi. In contrast, other venoms cleaved fibrinogen in a destructive (non-clotting) manner, with C. godmani and C. wilsoni being the most potent. In addition to actions on fibrinogen, clotting enzymes were also inhibited. FXa was only weakly inhibited by most species, but Cerrophidion godmani and C. wilsoni were extremely strong in their inhibitory action. Other clotting enzymes were more widely inhibited by diverse species spanning the full taxonomical range, but in each case, there were species that had these traits notably amplified relatively to the others. C. godmani and C. wilsoni were the most potent amongst those that inhibited the formation of the prothrombinase complex and were also amongst the most potent inhibitors of Factor XIa. While most species displayed only low levels of thrombin inhibition, Porthidium dunni potently inhibited this clotting factor. The regional polyvalent antivenom produced by Instituto Picado Clodomiro was tested and was shown to be effective against the diverse anticoagulant pathophysiological effects. In contrast to the anticoagulant activities of the other species, Porthidium volcanicum was uniquely procoagulant through the activation of Factor VII and Factor XII. This viperid species is the first snake outside of the Oxyuranus/Pseudonaja elapid snake clade to be shown to activate FVII and the first snake venom of any kind to activate FXII. Interestingly, while small-molecule metalloprotease inhibitors prinomastat and marimastat demonstrated the ability to prevent the procoagulant toxicity of P. volcanicum, neither ICP antivenom nor inhibitor DMPS showed this effect. The extreme variation among the snakes here studied underscores how venom is a dynamic trait and how this can shape clinical outcomes and influence evolving treatment strategies. Full article

(This article belongs to the Section Animal Venoms)

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13 pages, 328 KiB

Open AccessEditor’s ChoiceArticle

Deterministic and Probabilistic Dietary Exposure Assessment to Deoxynivalenol in Spain and the Catalonia Region

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

Toxins 2022, 14(7), 506; https://doi.org/10.3390/toxins14070506 - 20 Jul 2022

Cited by 6 | Viewed by 2313

Abstract

Deoxynivalenol (DON) remains one of the most concerning mycotoxins produced by the Fusarium genus due to the wide occurrence in highly consumed cereal-based food and its associated toxicological effects. Previous studies conducted in Spain and other European countries suggested that some vulnerable groups [...] Read more.

Deoxynivalenol (DON) remains one of the most concerning mycotoxins produced by the Fusarium genus due to the wide occurrence in highly consumed cereal-based food and its associated toxicological effects. Previous studies conducted in Spain and other European countries suggested that some vulnerable groups such as children could be exceeding the tolerable daily intakes. Thus, the aim of this study was to conduct a comprehensive and updated dietary exposure assessment study in Spain, with a specific analysis in the region of Catalonia. Cereal-based food samples collected during 2019 were analysed using liquid chromatography coupled to tandem mass spectrometry for multi-mycotoxin detection including DON and its main metabolites and derivatives. Consumption data were gathered from the nation-wide food surveys ENALIA and ENALIA2 conducted in Spain, and a specific survey conducted in Catalonia. The data were combined using deterministic and semi-parametric probabilistic methods. The results showed that DON was widely present in cereal-based food highly consumed in Spain and the Catalonia region. Exposure to DON among the adult population was globally low; however, among infants aged 3–9 years, it resulted in the median of 192 ng/kg body weight/day and the 95th percentiles of 604 ng/kg body weight/day, that would exceed the most conservative safety threshold for infants. Bread and pasta were the main contributing foodstuffs to the global exposure to DON, even among infants; thus, those foods should be considered a priority for food control or to develop strategies to reduce the exposure. In any case, further toxicological and epidemiological studies are required in order to refine the safety thresholds accounting for the sensitivity of the infant population. Full article

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

32 pages, 4441 KiB

Open AccessEditor’s ChoiceArticle

Cocktails of Mycotoxins, Phytoestrogens, and Other Secondary Metabolites in Diets of Dairy Cows in Austria: Inferences from Diet Composition and Geo-Climatic Factors

by Felipe Penagos-Tabares, Ratchaneewan Khiaosa-ard, Marlene Schmidt, Eva-Maria Bartl, Johanna Kehrer, Veronika Nagl, Johannes Faas, Michael Sulyok, Rudolf Krska and Qendrim Zebeli

Toxins 2022, 14(7), 493; https://doi.org/10.3390/toxins14070493 - 15 Jul 2022

Cited by 10 | Viewed by 3706

Abstract

Dairy production is a pivotal economic sector of Austrian and European agriculture. Dietary toxins and endocrine disruptors of natural origin such as mycotoxins and phytoestrogens can affect animal health, reproduction, and productivity. This study characterized the profile of a wide spectrum of fungal, [...] Read more.

Dairy production is a pivotal economic sector of Austrian and European agriculture. Dietary toxins and endocrine disruptors of natural origin such as mycotoxins and phytoestrogens can affect animal health, reproduction, and productivity. This study characterized the profile of a wide spectrum of fungal, plant, and unspecific secondary metabolites, including regulated, emerging, and modified mycotoxins, phytoestrogens, and cyanogenic glucosides, in complete diets of lactating cows from 100 Austrian dairy farms. To achieve this, a validated multi-metabolite liquid chromatography/electrospray ionization–tandem mass spectrometric (LC/ESI–MS/MS) method was employed, detecting 155 of >800 tested metabolites. Additionally, the most influential dietary and geo-climatic factors related to the dietary mycotoxin contamination of Austrian dairy cattle were recognized. We evidenced that the diets of Austrian dairy cows presented ubiquitous contamination with mixtures of mycotoxins and phytoestrogens. Metabolites derived from Fusarium spp. presented the highest concentrations, were the most recurrent, and had the highest diversity among the detected fungal compounds. Zearalenone, deoxynivalenol, and fumonisin B1 were the most frequently occurring mycotoxins considered in the EU legislation, with detection frequencies >70%. Among the investigated dietary factors, inclusion of maize silage (MS) and straw in the diets was the most influential factor in contamination with Fusarium-derived and other fungal toxins and metabolites, and temperature was the most influential among the geo-climatic factors. Full article

(This article belongs to the Special Issue Mycotoxins in Food and Feed: Detection and Identification)

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19 pages, 6329 KiB

Open AccessEditor’s ChoiceArticle

Improved Therapy of B-Cell Non-Hodgkin Lymphoma by Obinutuzumab-Dianthin Conjugates in Combination with the Endosomal Escape Enhancer SO1861

by Hossein Panjideh, Nicole Niesler, Alexander Weng and Hendrik Fuchs

Toxins 2022, 14(7), 478; https://doi.org/10.3390/toxins14070478 - 13 Jul 2022

Cited by 5 | Viewed by 2725

Abstract

Immunotoxins do not only bind to cancer-specific receptors to mediate the elimination of tumor cells through the innate immune system, but also increase target cytotoxicity by the intrinsic toxin activity. The plant glycoside SO1861 was previously reported to enhance the endolysosomal escape of [...] Read more.

Immunotoxins do not only bind to cancer-specific receptors to mediate the elimination of tumor cells through the innate immune system, but also increase target cytotoxicity by the intrinsic toxin activity. The plant glycoside SO1861 was previously reported to enhance the endolysosomal escape of antibody-toxin conjugates in non-hematopoietic cells, thus increasing their cytotoxicity manifold. Here we tested this technology for the first time in a lymphoma in vivo model. First, the therapeutic CD20 antibody obinutuzumab was chemically conjugated to the ribosome-inactivating protein dianthin. The cytotoxicity of obinutuzumab-dianthin (ObiDi) was evaluated on human B-lymphocyte Burkitt’s lymphoma Raji cells and compared to human T-cell leukemia off-target Jurkat cells. When tested in combination with SO1861, the cytotoxicity for target cells was 131-fold greater than for off-target cells. In vivo imaging in a xenograft model of B-cell lymphoma in mice revealed that ObiDi/SO1861 efficiently prevents tumor growth (51.4% response rate) compared to the monotherapy with ObiDi (25.9%) and non-conjugated obinutuzumab (20.7%). The reduction of tumor volume and overall survival was also improved. Taken together, our results substantially contribute to the development of a combination therapy with SO1861 as a platform technology to enhance the efficacy of therapeutic antibody-toxin conjugates in lymphoma and leukemia. Full article

(This article belongs to the Special Issue Immunotoxin and beyond—Past, Present and Future Perspectives)

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15 pages, 4453 KiB

Open AccessEditor’s ChoiceArticle

RIPpore: A Novel Host-Derived Method for the Identification of Ricin Intoxication through Oxford Nanopore Direct RNA Sequencing

by Yan Ryan, Abbie Harrison, Hannah Trivett, Catherine Hartley, Jonathan David, Graeme C. Clark and Julian A. Hiscox

Toxins 2022, 14(7), 470; https://doi.org/10.3390/toxins14070470 - 9 Jul 2022

Cited by 1 | Viewed by 2234

Abstract

Ricin is a toxin which enters cells and depurinates an adenine base in the sarcin-ricin loop in the large ribosomal subunit, leading to the inhibition of protein translation and cell death. We postulated that this depurination event could be detected using Oxford Nanopore [...] Read more.

Ricin is a toxin which enters cells and depurinates an adenine base in the sarcin-ricin loop in the large ribosomal subunit, leading to the inhibition of protein translation and cell death. We postulated that this depurination event could be detected using Oxford Nanopore Technologies (ONT) direct RNA sequencing, detecting a change in charge in the ricin loop. In this study, A549 cells were exposed to ricin for 2–24 h in order to induce depurination. In addition, a novel software tool was developed termed RIPpore that could quantify the adenine modification of ribosomal RNA induced by ricin upon respiratory epithelial cells. We provided demonstrable evidence for the first time that this base change detected is specific to RIP activity using a neutralising antibody against ricin. We believe this represents the first detection of depurination in RNA achieved using ONT sequencers. Collectively, this work highlights the potential for ONT and direct RNA sequencing to detect and quantify depurination events caused by ribosome-inactivating proteins such as ricin. RIPpore could have utility in the evaluation of new treatments and/or in the diagnosis of exposure to ricin. Full article

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

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13 pages, 3208 KiB

Open AccessEditor’s ChoiceArticle

Cytotoxic and Hemolytic Activities of Extracts of the Fish Parasite Dinoflagellate Amyloodinium ocellatum

by Márcio Moreira, Lucía Soliño, Cátia L. Marques, Vincent Laizé, Pedro Pousão-Ferreira, Pedro Reis Costa and Florbela Soares

Toxins 2022, 14(7), 467; https://doi.org/10.3390/toxins14070467 - 8 Jul 2022

Cited by 6 | Viewed by 4643

Abstract

The dinoflagellate Amyloodinium ocellatum is the etiological agent of a parasitic disease named amyloodiniosis. Mortalities of diseased fish are usually attributed to anoxia, osmoregulatory impairment, or opportunistic bacterial infections. Nevertheless, the phylogenetic proximity of A. ocellatum to a group of toxin-producing dinoflagellates from [...] Read more.

The dinoflagellate Amyloodinium ocellatum is the etiological agent of a parasitic disease named amyloodiniosis. Mortalities of diseased fish are usually attributed to anoxia, osmoregulatory impairment, or opportunistic bacterial infections. Nevertheless, the phylogenetic proximity of A. ocellatum to a group of toxin-producing dinoflagellates from Pfiesteria, Parvodinium and Paulsenella genera suggests that it may produce toxin-like compounds, adding a new dimension to the possible cause of mortalities in A. ocellatum outbreaks. To address this question, extracts prepared from different life stages of the parasite were tested in vitro for cytotoxic effects using two cell lines derived from branchial arches (ABSa15) and the caudal fin (CFSa1) of the gilthead seabream (Sparus aurata), and for hemolytic effects using erythrocytes purified from the blood of gilthead seabream juveniles. Cytotoxicity and a strong hemolytic effect, similar to those observed for Karlodinium toxins, were observed for the less polar extracts of the parasitic stage (trophont). A similar trend was observed for the less polar extracts of the infective stage (dinospores), although cell viability was only affected in the ABSa15 line. These results suggest that A. ocellatum produces tissue-specific toxic compounds that may have a role in the attachment of the dinospores’ and trophonts’ feeding process. Full article

(This article belongs to the Special Issue Effects of Harmful Algal Blooms on Aquatic Organisms)

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22 pages, 2510 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Exploring the Utility of ssDNA Aptamers Directed against Snake Venom Toxins as New Therapeutics for Snakebite Envenoming

by Nessrin Alomran, Raja Chinnappan, Jaffer Alsolaiss, Nicholas R. Casewell and Mohammed Zourob

Toxins 2022, 14(7), 469; https://doi.org/10.3390/toxins14070469 - 8 Jul 2022

Cited by 11 | Viewed by 2739

Abstract

Snakebite is a neglected tropical disease that causes considerable death and disability in the tropical world. Although snakebite can cause a variety of pathologies in victims, haemotoxic effects are particularly common and are typically characterised by haemorrhage and/or venom-induced consumption coagulopathy. Antivenoms are [...] Read more.

Snakebite is a neglected tropical disease that causes considerable death and disability in the tropical world. Although snakebite can cause a variety of pathologies in victims, haemotoxic effects are particularly common and are typically characterised by haemorrhage and/or venom-induced consumption coagulopathy. Antivenoms are the mainstay therapy for treating the toxic effects of snakebite, but despite saving thousands of lives annually, these therapies are associated with limited cross-snake species efficacy due to venom variation, which ultimately restricts their therapeutic utility to particular geographical regions. In this study, we sought to explore the potential of ssDNA aptamers as toxin-specific inhibitory alternatives to antibodies. As a proof of principle model, we selected snake venom serine protease toxins, which are responsible for contributing to venom-induced coagulopathy following snakebite envenoming, as our target. Using SELEX technology, we selected ssDNA aptamers against recombinantly expressed versions of the fibrinogenolytic SVSPs ancrod from the venom of C. rhodostoma and batroxobin from B. atrox. From the resulting pool of specific ssDNA aptamers directed against each target, we identified candidates that exhibited low nanomolar binding affinities to their targets. Downstream aptamer-linked immobilised sorbent assay, fibrinogenolysis, and coagulation profiling experiments demonstrated that the candidate aptamers were able to recognise native and recombinant SVSP toxins and inhibit the toxin- and venom-induced prolongation of plasma clotting times and the consumption of fibrinogen, with inhibitory potencies highly comparable to commercial polyvalent antivenoms. Our findings demonstrate that rationally selected toxin-specific aptamers can exhibit broad in vitro cross-reactivity against toxin isoforms found in different snake venoms and are capable of inhibiting toxins in pathologically relevant in vitro and ex vivo models of venom activity. These data highlight the potential utility of ssDNA aptamers as novel toxin-inhibiting therapeutics of value for tackling snakebite envenoming. Full article

(This article belongs to the Section Animal Venoms)

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13 pages, 2280 KiB

Open AccessEditor’s ChoiceArticle

Development of a Monoclonal scFv against Cytotoxin to Neutralize Cytolytic Activity Induced by Naja atra Venom on Myoblast C2C12 Cells

by Chien-Chun Liu, Cho-Ju Wu, Tsai-Ying Chou, Geng-Wang Liaw, Yung-Chin Hsiao, Lichieh-Julie Chu, Chi-Hsin Lee, Po-Jung Wang, Cheng-Hsien Hsieh, Chun-Kuei Chen and Jau-Song Yu

Toxins 2022, 14(7), 459; https://doi.org/10.3390/toxins14070459 - 4 Jul 2022

Cited by 8 | Viewed by 2182

Abstract

The Taiwanese cobra, Naja atra, is a clinically significant species of snake observed in the wild in Taiwan. Victims bitten by N. atra usually experience severe pain and local tissue necrosis. Although antivenom is available for treatment of cobra envenomation, its neutralization [...] Read more.

The Taiwanese cobra, Naja atra, is a clinically significant species of snake observed in the wild in Taiwan. Victims bitten by N. atra usually experience severe pain and local tissue necrosis. Although antivenom is available for treatment of cobra envenomation, its neutralization potency against cobra-induced necrosis is weak, with more than 60% of cobra envenoming patients developing tissue necrosis after antivenom administration. The present study found that cytotoxin (CTX) is a key component of N. atra venom responsible for cytotoxicity against myoblast cells. Anti-CTX IgY was generated in hens, and the spleens of these hens were used to construct libraries for the development of single chain variable fragments (scFv). Two anti-CTX scFv, S1 and 2S7, were selected using phage display technology and biopanning. Both polyclonal IgY and monoclonal scFv S1 reacted specifically with CTX in cobra venom. In a cell model assay, the CTX-induced cytolytic effect was inhibited only by monoclonal scFv S1, not by polyclonal IgY. Moreover, the neutralization potency of scFv S1 was about 3.8 mg/mg, approximately three times higher than that of conventional freeze-dried neurotoxic antivenom (FNAV). Collectively, these results suggest that scFv S1 can effectively neutralize CTX-induced cytotoxicity and, when combined with currently available antivenom, can improve the potency of the latter, thereby preventing tissue damage induced by cobra envenoming. Full article

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

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24 pages, 5625 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Exploring the Utility of Recombinant Snake Venom Serine Protease Toxins as Immunogens for Generating Experimental Snakebite Antivenoms

by Nessrin Alomran, Patricia Blundell, Jaffer Alsolaiss, Edouard Crittenden, Stuart Ainsworth, Charlotte A. Dawson, Rebecca J. Edge, Steven R. Hall, Robert A. Harrison, Mark C. Wilkinson, Stefanie K. Menzies and Nicholas R. Casewell

Toxins 2022, 14(7), 443; https://doi.org/10.3390/toxins14070443 - 29 Jun 2022

Cited by 13 | Viewed by 3472

Abstract

Snakebite is a neglected tropical disease that causes high rates of global mortality and morbidity. Although snakebite can cause a variety of pathologies in victims, haemotoxic effects are particularly common and are typically characterised by haemorrhage and/or venom-induced consumption coagulopathy. Despite polyclonal antibody-based [...] Read more.

Snakebite is a neglected tropical disease that causes high rates of global mortality and morbidity. Although snakebite can cause a variety of pathologies in victims, haemotoxic effects are particularly common and are typically characterised by haemorrhage and/or venom-induced consumption coagulopathy. Despite polyclonal antibody-based antivenoms being the mainstay life-saving therapy for snakebite, they are associated with limited cross-snake species efficacy, as there is often extensive toxin variation between snake venoms, including those used as immunogens for antivenom production. This restricts the therapeutic utility of any antivenom to certain geographical regions. In this study, we explored the feasibility of using recombinantly expressed toxins as immunogens to stimulate focused, pathology-specific, antibodies in order to broadly counteract specific toxins associated with snakebite envenoming. Three snake venom serine proteases (SVSP) toxins, sourced from geographically diverse and medically important viper snake venoms, were successfully expressed in HEK293F mammalian cells and used for murine immunisation. Analyses of the resulting antibody responses revealed that ancrod and RVV-V stimulated the strongest immune responses, and that experimental antivenoms directed against these recombinant SVSP toxins, and a mixture of the three different immunogens, extensively recognised and exhibited immunological binding towards a variety of native snake venoms. While the experimental antivenoms showed some reduction in abnormal clotting parameters stimulated by the toxin immunogens and crude venom, specifically reducing the depletion of fibrinogen levels and prolongation of prothrombin times, fibrinogen degradation experiments revealed that they broadly protected against venom- and toxin-induced fibrinogenolytic functional activities. Overall, our findings further strengthen the case for the use of recombinant venom toxins as supplemental immunogens to stimulate focused and desirable antibody responses capable of neutralising venom-induced pathological effects, and therefore potentially circumventing some of the limitations associated with current snakebite therapies. Full article

(This article belongs to the Section Animal Venoms)

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17 pages, 1955 KiB

Open AccessEditor’s ChoiceArticle

Natural and Induced Tolerance to Hymenoptera Venom: A Single Mechanism?

by Ana Navas, Berta Ruiz-Leon, Pilar Serrano, Manuel Martí, M Luisa Espinazo, Nadine Blanco, Juan Molina, Corona Alonso, Aurora Jurado and Carmen Moreno-Aguilar

Toxins 2022, 14(7), 426; https://doi.org/10.3390/toxins14070426 - 22 Jun 2022

Cited by 6 | Viewed by 2268

Abstract

Inducing tolerance in Hymenoptera-allergic patients, bee venom immunotherapy (BVIT) is a widely accepted method to treat severe allergy to bee stings. In order to increase the existing knowledge on the underlying immunological mechanisms and look for possible biomarkers predictive of efficacy, a group [...] Read more.

Inducing tolerance in Hymenoptera-allergic patients, bee venom immunotherapy (BVIT) is a widely accepted method to treat severe allergy to bee stings. In order to increase the existing knowledge on the underlying immunological mechanisms and look for possible biomarkers predictive of efficacy, a group of 20 bee-venom-allergic patients (AG) were thoroughly examined during their first year of BVIT. In addition, the results of treated patients with those of an untreated group of 20 tolerant beekeepers (TG) who had previously shown a firm suppressor-regulatory profile were compared. Tolerance in AG patients was invariably associated with a significant regulatory response characterised by the expansion of Helios⁻ subpopulation and increased IL-10, specific IgG4 (sIgG4), and kynurenine levels. Although specific IgE (sIgE) levels increased transiently, surprisingly, the T helper type 2 (Th2) population and IL-4 levels rose significantly after one year of immunotherapy. Thus, the picture of two parallel phenomena emerges: a tolerogenic response and an allergenic one. Comparing these results with those obtained from the TG, different immunological mechanisms appear to govern natural and acquired tolerance to immunotherapy. Of particular interest, the kynurenine levels and T regulatory (Treg) Helios⁻ population could be proposed as new biomarkers of response to BVIT. Full article

(This article belongs to the Section Animal Venoms)

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16 pages, 2271 KiB

Open AccessEditor’s ChoiceArticle

Computer-Aided Analysis of West Sub-Saharan Africa Snakes Venom towards the Design of Epitope-Based Poly-Specific Antivenoms

by Albert Ros-Lucas, Pascal Bigey, Jean-Philippe Chippaux, Joaquim Gascón and Julio Alonso-Padilla

Toxins 2022, 14(6), 418; https://doi.org/10.3390/toxins14060418 - 18 Jun 2022

Cited by 4 | Viewed by 2394

Abstract

Snakebite envenomation is a neglected tropical disease that causes over 100,000 deaths each year. The only effective treatment consists of antivenoms derived from animal sera, but these have been deemed with highly variable potency and are usually inaccessible and too costly for victims. [...] Read more.

Snakebite envenomation is a neglected tropical disease that causes over 100,000 deaths each year. The only effective treatment consists of antivenoms derived from animal sera, but these have been deemed with highly variable potency and are usually inaccessible and too costly for victims. The production of antivenoms by venom-independent techniques, such as the immunization with multi-epitope constructs, could circumvent those drawbacks. Herein, we present a knowledge-based pipeline to prioritize potential epitopes of therapeutic relevance from toxins of medically important snakes in West Sub-Saharan Africa. It is mainly based on sequence conservation and protein structural features. The ultimately selected 41 epitopes originate from 11 out of 16 snake species considered of highest medical importance in the region and 3 out of 10 of those considered as secondary medical importance. Echis ocellatus, responsible for the highest casualties in the area, would be covered by 12 different epitopes. Remarkably, this pipeline is versatile and customizable for the analysis of snake venom sequences from any other region of the world. Full article

(This article belongs to the Special Issue Snake Venom-Omics and Next Generation Antivenom)

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11 pages, 3071 KiB

Open AccessEditor’s ChoiceArticle

Investigation of Weather Triggers Preceding Outbreaks of Acute Bovine Liver Disease in Australia

by Eve M. Manthorpe, Grant T. Rawlin, Mark A. Stevenson, Lucy Woolford and Charles G. B. Caraguel

Toxins 2022, 14(6), 414; https://doi.org/10.3390/toxins14060414 - 17 Jun 2022

Viewed by 1591

Abstract

Acute bovine liver disease (ABLD) is a hepatic disease affecting cattle sporadically in southern Australia, characterised histologically by striking periportal hepatocellular necrosis. The cause of ABLD is unknown; however, the seasonality and acute presentation of outbreaks suggest mycotoxin involvement. We described the geographical [...] Read more.

Acute bovine liver disease (ABLD) is a hepatic disease affecting cattle sporadically in southern Australia, characterised histologically by striking periportal hepatocellular necrosis. The cause of ABLD is unknown; however, the seasonality and acute presentation of outbreaks suggest mycotoxin involvement. We described the geographical and seasonal occurrence of ABLD reports from 2010 to 2020 in Victoria, Australia, and explored potential weather triggers preceding 26 outbreaks occurring across 23 properties using a case-crossover design. Outbreaks occurred most frequently in autumn/early winter and in herds located along the southern coastal plain of Victoria, and occasionally within the low-lying regions of the Great Dividing Range. Lactating adult dairy cattle represented the most reported cases. We observed a significant association between an increase in average daily dewpoint in the 15 days preceding an ABLD outbreak, suggesting that dew formation may be a key determinant for this disease. Our findings support the etiology of a potent hepatotoxic agent that requires moisture for proliferation and/or toxin production. Full article

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12 pages, 1344 KiB

Open AccessEditor’s ChoiceArticle

A Pilot Randomized Controlled Trial of Botulinum Toxin Treatment Combined with Robot-Assisted Therapy, Mirror Therapy, or Active Control Treatment in Patients with Spasticity Following Stroke

by Jen-Wen Hung, Chu-Ling Yen, Ku-Chou Chang, Wei-Chi Chiang, I-Ching Chuang, Ya-Ping Pong, Wen-Chi Wu and Ching-Yi Wu

Toxins 2022, 14(6), 415; https://doi.org/10.3390/toxins14060415 - 17 Jun 2022

Cited by 4 | Viewed by 3490

Abstract

Effects of the combined task-oriented trainings with botulinum toxin A (BoNT-A) injection on improving motor functions and reducing spasticity remains unclear. This study aims to investigate effects of 3 task-oriented trainings (robot-assisted therapy (RT), mirror therapy (MT), and active control treatment (AC)) in [...] Read more.

Effects of the combined task-oriented trainings with botulinum toxin A (BoNT-A) injection on improving motor functions and reducing spasticity remains unclear. This study aims to investigate effects of 3 task-oriented trainings (robot-assisted therapy (RT), mirror therapy (MT), and active control treatment (AC)) in patients with stroke after BoNT-A injection. Thirty-seven patients with chronic spastic hemiplegic stroke were randomly assigned to receive RT, MT, or AC following BoNT-A injection over spastic upper extremity muscles. Each session of RT, MT, and AC was 75 min, 3 times weekly, for 8 weeks. Outcome measures were assessed at pretreatment, post-treatment, and 3-month follow-up, involving the Fugl-Meyer Assessment (FMA), Modified Ashworth Scale (MAS), Motor Activity Log (MAL), including amount of use (AOU) and quality of movement (QOM), and arm activity level. All 3 combined treatments improved FMA, MAS, and MAL. The AC induced a greater effect on QOM in MAL at the 3-month follow-up than RT or MT. All 3 combined trainings induced minimal effect on arm activity level. Our findings suggest that for patients with stroke who received BoNT-A injection over spastic UE muscles, the RT, MT, or AC UE training that followed was effective in improving motor functions, reducing spasticity, and enhancing daily function. Full article

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14 pages, 3314 KiB

Open AccessEditor’s ChoiceArticle

Genotoxicity of 12 Mycotoxins by the SOS/umu Test: Comparison of Liver and Kidney S9 Fraction

by Maria Alonso-Jauregui, Elena González-Peñas, Adela López de Cerain and Ariane Vettorazzi

Toxins 2022, 14(6), 400; https://doi.org/10.3390/toxins14060400 - 10 Jun 2022

Cited by 6 | Viewed by 2148

Abstract

Liver S9 fraction is usually employed in mutagenicity/genotoxicity in vitro assays, but some genotoxic compounds may need another type of bioactivation. In the present work, an alternative S9 fraction from the kidneys was used for the genotoxicity assessment of 12 mycotoxins with the [...] Read more.

Liver S9 fraction is usually employed in mutagenicity/genotoxicity in vitro assays, but some genotoxic compounds may need another type of bioactivation. In the present work, an alternative S9 fraction from the kidneys was used for the genotoxicity assessment of 12 mycotoxins with the SOS/umu test. The results were compared with liver S9 fraction, and 2–4 independent experiments were performed with each mycotoxin. The expected results were obtained with positive controls (4-nitroquinoline-N-oxide and 2-aminoanthracene) without metabolic activation or with liver S9, but a potent dose-dependent effect with 4-nitroquinoline-N-oxide and no activity of 2-aminoanthracene with kidney S9 were noticed. Aflatoxin B1 was genotoxic with metabolic activation, the effect being greater with liver S9. Sterigmatocystin was clearly genotoxic with liver S9 but equivocal with kidney S9. Ochratoxin A, zearalenone and fumonisin B1 were negative in all conditions. Trichothecenes were negative, except for nivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, T-2 and HT-2 toxins, which showed equivocal results with kidney S9 because a clear dose-response effect was not observed. Most of the mycotoxins have been assessed with kidney S9 and the SOS/umu test for the first time here. The results with the positive controls and the mycotoxins confirm that the organ used for the S9 fraction preparation has an influence on the genotoxic activity of some compounds. Full article

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

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17 pages, 3418 KiB

Open AccessEditor’s ChoiceArticle

Liza ramada Juveniles after Exposure to the Toxic Dinoflagellate Vulcanodinium rugosum: Effects on Fish Viability, Tissue Contamination and Microalgae Survival after Gut Passage

by Aurélien Bouquet, Marie Anaïs Perdrau, Mohamed Laabir, Elodie Foucault, Nicolas Chomérat, Jean Luc Rolland and Eric Abadie

Toxins 2022, 14(6), 401; https://doi.org/10.3390/toxins14060401 - 10 Jun 2022

Cited by 3 | Viewed by 2750

Abstract

Pinnatoxins (PnTX) and Portimines (Prtn), two toxins produced by the benthic dinoflagellate Vulcanodinium rugosum, are known to be lethal to mice after intraperitoneal or oral administration. They are also known to accumulate in shellfish such as mussels and clams, but their effect [...] Read more.

Pinnatoxins (PnTX) and Portimines (Prtn), two toxins produced by the benthic dinoflagellate Vulcanodinium rugosum, are known to be lethal to mice after intraperitoneal or oral administration. They are also known to accumulate in shellfish such as mussels and clams, but their effect on fish and the upper food chain remains unknown. In this work, juveniles of the fish Liza ramada (Mullet) were exposed to a strain of V. rugosum producing PnTX G and Prtn A. The fishes’ viability and contamination were recorded at times interval. Results showed that L. ramada juveniles were able to feed on V. rugosum and that their tissues could be contaminated by PnTX G and Prtn A without impact on fish viability. Furthermore, the microalgae temporary cysts survived and germinated after fish gut passage. This study showed the potential of L. ramada to transfer PnTX and Prtn toxins to the upper food chain and to disseminate V. rugosum in environment. Full article

(This article belongs to the Special Issue Effects of Harmful Algal Blooms on Aquatic Organisms)

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15 pages, 2571 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

In Vitro Metabolism of Phenylspirodrimanes Derived from the Indoor Fungus Stachybotrys

by Viktoria Lindemann, Annika Jagels, Matthias Behrens, Florian Hübner and Hans-Ulrich Humpf

Toxins 2022, 14(6), 395; https://doi.org/10.3390/toxins14060395 - 8 Jun 2022

Cited by 4 | Viewed by 2099

Abstract

Fungi belonging to the genus Stachybotrys are frequently detected in water-damaged indoor environments, and a potential correlation between emerging health problems of inhabitants of affected housing and the fungi is controversially discussed. Secondary metabolites (i.e., mycotoxins) produced by Stachybotrys, such as the [...] Read more.

Fungi belonging to the genus Stachybotrys are frequently detected in water-damaged indoor environments, and a potential correlation between emerging health problems of inhabitants of affected housing and the fungi is controversially discussed. Secondary metabolites (i.e., mycotoxins) produced by Stachybotrys, such as the highly toxic macrocyclic trichothecenes (MCTs), are of potential concern to human health. The present study, however, focused on the potential effects of the more broadly and abundantly formed group of phenylspirodrimanes (PSDs). The phase I and II metabolism of four structurally different PSDs were investigated in vitro using hepatic models in combination with high-performance liquid chromatography high-resolution mass spectrometry (HPLC-HRMS) analysis. In addition to metabolite detection by HRMS, isolation and structure elucidation by nuclear magnetic resonance spectroscopy (NMR) was part of the conducted study as well. Full article

(This article belongs to the Section Mycotoxins)

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18 pages, 2522 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Mycotoxin Analysis of Grain via Dust Sampling: Review, Recent Advances and the Way Forward: The Contribution of the MycoKey Project

by Biancamaria Ciasca, Sarah De Saeger, Marthe De Boevre, Mareike Reichel, Michelangelo Pascale, Antonio F. Logrieco and Veronica M. T. Lattanzio

Toxins 2022, 14(6), 381; https://doi.org/10.3390/toxins14060381 - 31 May 2022

Cited by 4 | Viewed by 3563

Abstract

The sampling protocols for the official control of the levels of mycotoxins in foodstuffs are very costly and time-consuming. More efforts are needed to implement alternative sampling plans able to support official control, or to adapt the current ones. The aim of the [...] Read more.

The sampling protocols for the official control of the levels of mycotoxins in foodstuffs are very costly and time-consuming. More efforts are needed to implement alternative sampling plans able to support official control, or to adapt the current ones. The aim of the research carried out within the European Horizon 2020 MycoKey project was to evaluate the applicability at industrial scale of the dust sampling approach to detect multiple mycotoxins in grains. To this end, two trials were performed on an EU industrial site: (i) control of the unloading of wheat from train wagons; (ii) control of the unloading of wheat from trucks. In line with previous studies, the MycoKey results indicated that dust sampling and mycotoxin analysis represent a fitness for purpose approach for non–destructive and rapid identification of wheat commodities compliant to the maximum permitted levels. Based on reviewed and newly generated results, this article discusses potential applications and limits of the dust sampling methodology, identifying future research needs. Full article

(This article belongs to the Special Issue Selected Papers from MycoKey - MycoTWIN - ISM international conference)

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11 pages, 1381 KiB

Open AccessEditor’s ChoiceArticle

Botulinum Toxin Type A for Treatment of Forehead Hyperhidrosis: Multicenter Clinical Experience and Review from Literature

by Anna Campanati, Emanuela Martina, Stamatis Gregoriou, George Kontochristopoulos, Matteo Paolinelli, Federico Diotallevi, Giulia Radi, Ivan Bobyr, Barbara Marconi, Giulio Gualdi, Paolo Amerio and Annamaria Offidani

Toxins 2022, 14(6), 372; https://doi.org/10.3390/toxins14060372 - 27 May 2022

Cited by 5 | Viewed by 6727

Abstract

Among the forms of idiopathic hyperhidrosis, those involving the forehead have the greatest impact on patients’ quality of life, as symptoms are not very controllable and are difficult to mask for patients. Although the local injection therapy with Incobotulinum toxin type A (IncoBTX-A [...] Read more.

Among the forms of idiopathic hyperhidrosis, those involving the forehead have the greatest impact on patients’ quality of life, as symptoms are not very controllable and are difficult to mask for patients. Although the local injection therapy with Incobotulinum toxin type A (IncoBTX-A therapy) can be considered a rational treatment, data from the literature describing both efficacy and safety of the treatment over the long term are poor. The aim of this report is to describe the single-center experience of five patients seeking treatment, for forehead hyperhidrosis with IncoBTX-A. To evaluate the benefits, safety profile and duration of anhidrosis, patients were treated following a standardized procedure and then followed until clinical relapse. The amount of sweating was measured by gravimetric testing, the extension of hyperhidrosis area was measured through Minor’s iodine starch test, and response to the treatment was evaluated using the Hyperhidrosis Disease Severity Scale (HDSS) and the Dermatology Life Quality Index (DLQI). In all treated patients, a significant anhidrotic effect was observed 4 weeks after the treatment and lasted for approximately 36 weeks. The reduction in sweat production was associated with significant amelioration of symptoms and quality of life for all treated patients. No serious side effects occurred; one patient complained of a mild transient bilateral ptosis. Although further wider studies are required, our preliminary results seem to encourage the use of IncoBTX-A in forehead hyperhidrosis. Full article

(This article belongs to the Special Issue Toxins in Dermatology)

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18 pages, 2015 KiB

Open AccessEditor’s ChoiceArticle

Venomics of the Central European Myrmicine Ants Myrmica rubra and Myrmica ruginodis

by Sabine Hurka, Karina Brinkrolf, Rabia Özbek, Frank Förster, André Billion, John Heep, Thomas Timm, Günter Lochnit, Andreas Vilcinskas and Tim Lüddecke

Toxins 2022, 14(5), 358; https://doi.org/10.3390/toxins14050358 - 21 May 2022

Cited by 8 | Viewed by 4338

Abstract

Animal venoms are a rich source of novel biomolecules with potential applications in medicine and agriculture. Ants are one of the most species-rich lineages of venomous animals. However, only a fraction of their biodiversity has been studied so far. Here, we investigated the [...] Read more.

Animal venoms are a rich source of novel biomolecules with potential applications in medicine and agriculture. Ants are one of the most species-rich lineages of venomous animals. However, only a fraction of their biodiversity has been studied so far. Here, we investigated the venom components of two myrmicine (subfamily Myrmicinae) ants: Myrmica rubra and Myrmica ruginodis. We applied a venomics workflow based on proteotranscriptomics and found that the venoms of both species are composed of several protein classes, including venom serine proteases, cysteine-rich secretory protein, antigen 5 and pathogenesis-related 1 (CAP) superfamily proteins, Kunitz-type serine protease inhibitors and venom acid phosphatases. Several of these protein classes are known venom allergens, and for the first time we detected phospholipase A1 in the venom of M. ruginodis. We also identified two novel epidermal growth factor (EGF) family toxins in the M. ruginodis venom proteome and an array of additional EGF-like toxins in the venom gland transcriptomes of both species. These are similar to known toxins from the related myrmicine ant, Manica rubida, and the myrmecine (subfamily Myrmeciinae) Australian red bulldog ant Myrmecia gullosa, and are possibly deployed as weapons in defensive scenarios or to subdue prey. Our work suggests that M.rubra and M. ruginodis venoms contain many enzymes and other high-molecular-weight proteins that cause cell damage. Nevertheless, the presence of EGF-like toxins suggests that myrmicine ants have also recruited smaller peptide components into their venom arsenal. Although little is known about the bioactivity and function of EGF-like toxins, their presence in myrmicine and myrmecine ants suggests they play a key role in the venom systems of the superfamily Formicoidea. Our work adds to the emerging picture of ant venoms as a source of novel bioactive molecules and highlights the need to incorporate such taxa in future venom bioprospecting programs. Full article

(This article belongs to the Section Animal Venoms)

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11 pages, 4335 KiB

Open AccessEditor’s ChoiceArticle

Structural Features of Clostridium botulinum Neurotoxin Subtype A2 Cell Binding Domain

by Kyle S. Gregory, Tejaswini B. Mahadeva, Sai Man Liu and K. Ravi Acharya

Toxins 2022, 14(5), 356; https://doi.org/10.3390/toxins14050356 - 19 May 2022

Cited by 6 | Viewed by 4659

Abstract

Botulinum neurotoxins (BoNT) are a group of clostridial toxins that cause the potentially fatal neuroparalytic disease botulism. Although highly toxic, BoNTs are utilized as therapeutics to treat a range of neuromuscular conditions. Several serotypes (BoNT/A-/G, /X) have been identified with vastly differing toxicological [...] Read more.

Botulinum neurotoxins (BoNT) are a group of clostridial toxins that cause the potentially fatal neuroparalytic disease botulism. Although highly toxic, BoNTs are utilized as therapeutics to treat a range of neuromuscular conditions. Several serotypes (BoNT/A-/G, /X) have been identified with vastly differing toxicological profiles. Each serotype can be further sub-categorised into subtypes due to subtle variations in their protein sequence. These minor changes have been attributed to differences in both the duration of action and potency for BoNT/A subtypes. BoNTs are composed of three domains—a cell-binding domain, a translocation domain, and a catalytic domain. In this paper, we present the crystal structures of the botulinum neurotoxin A2 cell binding domain, both alone and in complex with its receptor ganglioside GD1a at 1.63 and 2.10 Å, respectively. The analysis of these structures reveals a potential redox-dependent Lys-O-Cys bridge close to the ganglioside binding site and a hinge motion between the H_CN and H_CC subdomains. Furthermore, we make a detailed comparison with the previously reported H_C/A2:SV2C structure for a comprehensive structural analysis of H_C/A2 receptor binding. Full article

(This article belongs to the Section Bacterial Toxins)

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21 pages, 1553 KiB

Open AccessEditor’s ChoiceArticle

Carry-Over of Zearalenone and Its Metabolites to Intestinal Tissues and the Expression of CYP1A1 and GSTπ1 in the Colon of Gilts before Puberty

by Magdalena Mróz, Magdalena Gajęcka, Paweł Brzuzan, Sylwia Lisieska-Żołnierczyk, Dawid Leski, Łukasz Zielonka and Maciej T. Gajęcki

Toxins 2022, 14(5), 354; https://doi.org/10.3390/toxins14050354 - 18 May 2022

Cited by 4 | Viewed by 1995

Abstract

The objective of this study was to evaluate whether low doses of zearalenone (ZEN) affect the carry-over of ZEN and its metabolites to intestinal tissues and the expression of CYP1A1 and GSTπ1 in the large intestine. Prepubertal gilts (with a BW of up [...] Read more.

The objective of this study was to evaluate whether low doses of zearalenone (ZEN) affect the carry-over of ZEN and its metabolites to intestinal tissues and the expression of CYP1A1 and GSTπ1 in the large intestine. Prepubertal gilts (with a BW of up to 14.5 kg) were exposed in group ZEN to daily ZEN5 doses of 5 μg/kg BW (n = 15); in group ZEN10, 10 μg/kg BW (n = 15); in group ZEN15, 15 μg/kg BW (n = 15); or were administered a placebo (group C, n = 15) throughout the experiment. After euthanasia, tissues were sampled on exposure days 7, 21, and 42 (D1, D2, and D3, respectively). The results confirmed that the administered ZEN doses (LOAEL, NOAEL, and MABEL) were appropriate to reliably assess the carry-over of ZEN. Based on the observations made during 42 days of exposure to pure ZEN, it can be hypothesized that all mycotoxins (ZEN, α-zearalenol, and β-zearalenol) contribute to a balance between intestinal cells and the expression of selected genes encoding enzymes that participate in biotransformation processes in the large intestine; modulate feminization processes in prepubertal gilts; and elicit flexible, adaptive responses of the macroorganism to mycotoxin exposure at the analyzed doses. Full article

(This article belongs to the Special Issue Influence of Deoxynivalenol and Zearalenone in Feed on Animal Health)

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16 pages, 995 KiB

Open AccessEditor’s ChoiceArticle

Effects of Two Toxin-Producing Harmful Algae, Alexandrium catenella and Dinophysis acuminata (Dinophyceae), on Activity and Mortality of Larval Shellfish

by Sarah K. D. Pease, Michael L. Brosnahan, Marta P. Sanderson and Juliette L. Smith

Toxins 2022, 14(5), 335; https://doi.org/10.3390/toxins14050335 - 10 May 2022

Cited by 14 | Viewed by 3411

Abstract

Harmful algal bloom (HAB) species Alexandrium catenella and Dinophysis acuminata are associated with paralytic shellfish poisoning (PSP) and diarrhetic shellfish poisoning (DSP) in humans, respectively. While PSP and DSP have been studied extensively, less is known about the effects of these HAB species [...] Read more.

Harmful algal bloom (HAB) species Alexandrium catenella and Dinophysis acuminata are associated with paralytic shellfish poisoning (PSP) and diarrhetic shellfish poisoning (DSP) in humans, respectively. While PSP and DSP have been studied extensively, less is known about the effects of these HAB species or their associated toxins on shellfish. This study investigated A. catenella and D. acuminata toxicity in a larval oyster (Crassostrea virginica) bioassay. Larval activity and mortality were examined through 96-h laboratory exposures to live HAB cells (10–1000 cells/mL), cell lysates (1000 cells/mL equivalents), and purified toxins (10,000 cells/mL equivalents). Exposure to 1000 cells/mL live or lysed D. acuminata caused larval mortality (21.9 ± 7.0%, 10.2 ± 4.0%, respectively) while exposure to any tested cell concentration of live A. catenella, but not lysate, caused swimming arrest and/or mortality in >50% of larvae. Exposure to high concentrations of saxitoxin (STX) or okadaic acid (OA), toxins traditionally associated with PSP and DSP, respectively, had no effect on larval activity or mortality. In contrast, pectenotoxin-2 (PTX2) caused rapid larval mortality (49.6 ± 5.8% by 48 h) and completely immobilized larval oysters. The results indicate that the toxic effects of A. catenella and D. acuminata on shellfish are not linked to the primary toxins associated with PSP and DSP in humans, and that PTX2 is acutely toxic to larval oysters. Full article

(This article belongs to the Special Issue Effects of Harmful Algal Blooms on Aquatic Organisms)

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21 pages, 6883 KiB

Open AccessEditor’s ChoiceArticle

The Unusual Metalloprotease-Rich Venom Proteome of the Australian Elapid Snake Hoplocephalus stephensii

by Theo Tasoulis, C. Ruth Wang, Joanna Sumner, Nathan Dunstan, Tara L. Pukala and Geoffrey K. Isbister

Toxins 2022, 14(5), 314; https://doi.org/10.3390/toxins14050314 - 28 Apr 2022

Cited by 2 | Viewed by 3318

Abstract

The Australasian region is home to the most diverse elapid snake radiation on the planet (Hydrophiinae). Many of these snakes have evolved into unique ecomorphs compared to elapids on other continents; however, their venom compositions are poorly known. The Australian elapid Hoplocephalus stephensii [...] Read more.

The Australasian region is home to the most diverse elapid snake radiation on the planet (Hydrophiinae). Many of these snakes have evolved into unique ecomorphs compared to elapids on other continents; however, their venom compositions are poorly known. The Australian elapid Hoplocephalus stephensii (Stephen’s banded snake) is an arboreal snake with a unique morphology. Human envenoming results in venom-induced consumption coagulopathy, without neurotoxicity. Using transcriptomics and a multi-step fractionation method involving reverse-phase high-performance liquid chromatography, sodium dodecyl sulfate polyacrylamide gel electrophoresis and bottom-up proteomics, we characterized the venom proteome of H. stephensii. 92% of the total protein component of the venom by weight was characterized, and included all dominant protein families and 4 secondary protein families. Eighteen toxins made up 76% of the venom, four previously characterized and 14 new toxins. The four dominant protein families made up 77% of the venom, including snake venom metalloprotease (SVMP; 36.7%; three identified toxins), phospholipase A₂ (PLA₂; 24.0%; five identified toxins), three-finger toxin (3FTx; 10.2%; two toxins) and snake venom serine protease (SVSP; 5.9%; one toxin; Hopsarin). Secondary protein families included L-amino acid oxidase (LAAO; 10.8%; one toxin), natriuretic peptide (NP; 0.8%; two toxins), cysteine-rich secretory protein (CRiSP; 1.7%; two toxins), c-type lectin (CTL; 1.1%; one toxin), and one minor protein family, nerve growth factor (NGF; 0.8%; one toxin). The venom composition of H. stephensii differs to other elapids, with a large proportion of SVMP and LAAO, and a relatively small amount of 3FTx. H. stephensii venom appeared to have less toxin diversity than other elapids, with only 18 toxins making up three-quarters of the venom. Full article

(This article belongs to the Section Animal Venoms)

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22 pages, 2430 KiB

Open AccessEditor’s ChoiceArticle

Different Resistance to DON versus HT2 + T2 Producers in Nordic Oat Varieties

by Ingerd Skow Hofgaard, Guro Brodal, Marit Almvik, Morten Lillemo, Aina Lundon Russenes, Simon Graham Edwards and Heidi Udnes Aamot

Toxins 2022, 14(5), 313; https://doi.org/10.3390/toxins14050313 - 28 Apr 2022

Cited by 8 | Viewed by 3028

Abstract

Over recent decades, the Norwegian cereal industry has had major practical and financial challenges associated with the occurrence of Fusarium head blight (FHB) pathogens and their associated mycotoxins in cereal grains. Deoxynivalenol (DON) is one of the most common Fusarium-mycotoxins in Norwegian [...] Read more.

Over recent decades, the Norwegian cereal industry has had major practical and financial challenges associated with the occurrence of Fusarium head blight (FHB) pathogens and their associated mycotoxins in cereal grains. Deoxynivalenol (DON) is one of the most common Fusarium-mycotoxins in Norwegian oats, however T-2 toxin (T2) and HT-2 toxin (HT2) are also commonly detected. The aim of our study was to rank Nordic spring oat varieties and breeding lines by content of the most commonly occurring Fusarium mycotoxins (DON and HT2 + T2) as well as by the DNA content of their respective producers. We analyzed the content of mycotoxins and DNA of seven fungal species belonging to the FHB disease complex in grains of Nordic oat varieties and breeding lines harvested from oat field trials located in the main cereal cultivating district in South-East Norway in the years 2011–2020. Oat grains harvested from varieties with a high FHB resistance contained on average half the levels of mycotoxins compared with the most susceptible varieties, which implies that choice of variety may indeed impact on mycotoxin risk. The ranking of oat varieties according to HT2 + T2 levels corresponded with the ranking according to the DNA levels of Fusarium langsethiae, but differed from the ranking according to DON and Fusarium graminearum DNA. Separate tests are therefore necessary to determine the resistance towards HT2 + T2 and DON producers in oats. This creates practical challenges for the screening of FHB resistance in oats as today’s screening focuses on resistance to F. graminearum and DON. We identified oat varieties with generally low levels of both mycotoxins and FHB pathogens which should be preferred to mitigate mycotoxin risk in Norwegian oats. Full article

(This article belongs to the Special Issue Selected Papers from the 15th European Fusarium Seminar)

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14 pages, 2116 KiB

Open AccessEditor’s ChoiceArticle

Biocontrol of Fusarium graminearum, a Causal Agent of Fusarium Head Blight of Wheat, and Deoxynivalenol Accumulation: From In Vitro to In Planta

by Asmaa Abbas and Tapani Yli-Mattila

Toxins 2022, 14(5), 299; https://doi.org/10.3390/toxins14050299 - 22 Apr 2022

Cited by 17 | Viewed by 4547

Abstract

Crop diseases caused by Fusarium graminearum threaten crop production in both commercial and smallholder farming. F. graminearum produces deoxynivalenol mycotoxin, which is stable during food and feed processing. Therefore, the best way to prevent the sporulation of pathogens is to develop new [...] Read more.

Crop diseases caused by Fusarium graminearum threaten crop production in both commercial and smallholder farming. F. graminearum produces deoxynivalenol mycotoxin, which is stable during food and feed processing. Therefore, the best way to prevent the sporulation of pathogens is to develop new prevention strategies. Plant-based pesticides, i.e., natural fungicides, have recently gained interest in crop protection as alternatives to synthetic fungicides. Herein we show that treatment with the methanolic extract of medicinal plant Zanthoxylum bungeanum (M20 extract), decreased F. graminearum growth and abrogated DON production. The F. graminearum DNA levels were monitored by a quantitative TaqMan real-time PCR, while DON accumulation was assessed by HPLC quantification. This M20 extract was mainly composed of four flavonoids: quercetin, epicatechin, kaempferol-3-O-rhamnoside, and hyperoside. The in vitro bioassay, which measured the percent inhibition of fungal growth, showed that co-inoculation of four F. graminearum strains with the M20 extract inhibited the fungal growth up to 48.5%. After biocontrol treatments, F. graminearum DNA level was reduced up to 85.5% compared to that of wheat heads, which received F. graminearum mixture only. Moreover, DON production was decreased in wheat heads by 73% after biocontrol treatment; meanwhile in wheat heads inoculated with F. graminearum conidia, an average of 2.263 ± 0.8 mg/kg DON was detected. Overall, this study is a successful case from in vitro research to in planta, giving useful information for wheat protection against F. graminearum responsible for Fusarium Head Blight and DON accumulation in grains. Further studies are needed to study the mechanism by which M20 extract inhibited the DON production and what changes happened to the DON biosynthetic pathway genes. Full article

(This article belongs to the Special Issue New Insight into Fusarium Toxins and Aflatoxins)

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20 pages, 6271 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Clinical and Evolutionary Implications of Dynamic Coagulotoxicity Divergences in Bothrops (Lancehead Pit Viper) Venoms

by Lachlan Allan Bourke, Christina N. Zdenek, Anita Mitico Tanaka-Azevedo, Giovanni Perez Machado Silveira, Sávio Stefanini Sant’Anna, Kathleen Fernandes Grego, Caroline Fabri Bittencourt Rodrigues and Bryan Grieg Fry

Toxins 2022, 14(5), 297; https://doi.org/10.3390/toxins14050297 - 22 Apr 2022

Cited by 9 | Viewed by 2757

Abstract

Despite coagulotoxicity being a primary weapon for prey capture by Bothrops species (lancehead pit vipers) and coagulopathy being a major lethal clinical effect, a genus-wide comparison has not been undertaken. To fill this knowledge gap, we used thromboelastography to compare 37 venoms, from [...] Read more.

Despite coagulotoxicity being a primary weapon for prey capture by Bothrops species (lancehead pit vipers) and coagulopathy being a major lethal clinical effect, a genus-wide comparison has not been undertaken. To fill this knowledge gap, we used thromboelastography to compare 37 venoms, from across the full range of geography, taxonomy, and ecology, for their action upon whole plasma and isolated fibrinogen. Potent procoagulant toxicity was shown to be the main venom effect of most of the species tested. However, the most basal species (B. pictus) was strongly anticoagulant; this is consistent with procoagulant toxicity being a novel trait that evolved within Bothrops subsequent to their split from anticoagulant American pit vipers. Intriguingly, two of the arboreal species studied (B. bilineatus and B. taeniatus) lacked procoagulant venom, suggesting differential evolutionary selection pressures. Notably, some terrestrial species have secondarily lost the procoagulant venom trait: the Mogi Mirim, Brazil locality of B. alternatus; San Andres, Mexico locality of B. asper; B. diporus; and the São Roque of B. jararaca. Direct action on fibrinogen was extremely variable; this is consistent with previous hypotheses regarding it being evolutionary decoupled due to procoagulant toxicity being the primary prey-capture weapon. However, human patients live long enough for fibrinogen depletion to be clinically significant. The extreme variability may be reflective of antivenom variability, with these results thereby providing a foundation for such future work of clinical relevance. Similarly, the venom diversification trends relative to ecological niche will also be useful for integration with natural history data, to reconstruct the evolutionary pressures shaping the venoms of these fascinating snakes. Full article

(This article belongs to the Special Issue Insights into the Action and Application of Animal Toxins)

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10 pages, 1961 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

The Effect of Australian and Asian Commercial Antivenoms in Reversing the Post-Synaptic Neurotoxicity of O. hannah, N. naja and N. kaouthia Venoms In Vitro

by Tam M. Huynh, Wayne C. Hodgson, Geoffrey K. Isbister and Anjana Silva

Toxins 2022, 14(4), 277; https://doi.org/10.3390/toxins14040277 - 13 Apr 2022

Cited by 3 | Viewed by 3005

Abstract

Despite antivenoms being the only established specific treatment for neuromuscular paralysis arising from snake envenoming, their ability to reverse the post-synaptic neurotoxicity in snake envenoming is poorly understood. We investigated the ability of five commercial antivenoms i.e., King cobra monovalent, Thai cobra monovalent, [...] Read more.

Despite antivenoms being the only established specific treatment for neuromuscular paralysis arising from snake envenoming, their ability to reverse the post-synaptic neurotoxicity in snake envenoming is poorly understood. We investigated the ability of five commercial antivenoms i.e., King cobra monovalent, Thai cobra monovalent, Thai neuro polyvalent, Indian polyvalent and Australian polyvalent antivenoms to reverse neurotoxicity induced by the venoms of King cobra (Ophiophagus hannah, 3 µg/mL), Indian cobra (Naja naja, 5 µg/mL) and Thai cobra (Naja kaouthia, 3 µg/mL) using the in vitro chick-biventer cervicis nerve–muscle preparation. All three venoms displayed post-synaptic neurotoxicity, which was prevented by all tested antivenoms (40 µL/mL) added to the bath prior to venom. All antivenoms partially reversed the established post-synaptic neuromuscular block after the addition of the three venoms during a 180 min observation period, but to varying degrees and at different rates. The neurotoxic effects of O. hannah venom recovered to a greater magnitude (based on twitch height restoration) and faster than the neurotoxicity of N. kaouthia venom, which recovered to a lower magnitude more slowly. The recovery of post-synaptic neurotoxicity by N. naja venom was hindered due to the likely presence of cytotoxins in the venom, which cause direct muscle damage. The observations made in this study provide further evidence that the commercial antivenoms are likely to actively reverse established α-neurotoxin-mediated neuromuscular paralysis in snake envenoming, and there is cross-neutralisation with different antivenoms. Full article

(This article belongs to the Special Issue Toxinology and Pharmacology of Snake Venoms)

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21 pages, 1574 KiB

Open AccessEditor’s ChoiceArticle

Providing Biological Plausibility for Exposure–Health Relationships for the Mycotoxins Deoxynivalenol (DON) and Fumonisin B1 (FB1) in Humans Using the AOP Framework

by Annick D. van den Brand, Lola Bajard, Inger-Lise Steffensen, Anne Lise Brantsæter, Hubert A. A. M. Dirven, Jochem Louisse, Ad Peijnenburg, Sophie Ndaw, Alberto Mantovani, Barbara De Santis and Marcel J. B. Mengelers

Toxins 2022, 14(4), 279; https://doi.org/10.3390/toxins14040279 - 13 Apr 2022

Cited by 8 | Viewed by 4402

Abstract

Humans are chronically exposed to the mycotoxins deoxynivalenol (DON) and fumonisin B1 (FB1), as indicated by their widespread presence in foods and occasional exposure in the workplace. This exposure is confirmed by human biomonitoring (HBM) studies on (metabolites of) these mycotoxins in human [...] Read more.

Humans are chronically exposed to the mycotoxins deoxynivalenol (DON) and fumonisin B1 (FB1), as indicated by their widespread presence in foods and occasional exposure in the workplace. This exposure is confirmed by human biomonitoring (HBM) studies on (metabolites of) these mycotoxins in human matrices. We evaluated the exposure–health relationship of the mycotoxins in humans by reviewing the available literature. Since human studies did not allow the identification of unequivocal chronic health effects upon exposure to DON and FB1, the adverse outcome pathway (AOP) framework was used to structure additional mechanistic evidence from in vitro and animal studies on the identified adverse effects. In addition to a preliminary AOP for DON resulting in the adverse outcome (AO) ‘reduced body weight gain’, we developed a more elaborated AOP for FB1, from the molecular initiating event (MIE) ‘inhibition of ceramide synthases’ leading to the AO ‘neural tube defects’. The mechanistic evidence from AOPs can be used to support the limited evidence from human studies, to focus FB1- and DON-related research in humans to identify related early biomarkers of effect. In order to establish additional human exposure–health relationships in the future, recommendations are given to maximize the information that can be obtained from HBM. Full article

(This article belongs to the Special Issue Human Biomonitoring and Risk Assessment of Mycotoxins)

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15 pages, 357 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

Resistance Allele Frequency to Cry1Ab and Vip3Aa20 in Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in Louisiana and Three Other Southeastern U.S. States

by Shucong Lin, Isaac Oyediran, Ying Niu, Sebe Brown, Don Cook, Xinzhi Ni, Yan Zhang, Francis P. F. Reay-Jones, Jeng Shong Chen, Zhimou Wen, Marcelo Dimase and Fangneng Huang

Toxins 2022, 14(4), 270; https://doi.org/10.3390/toxins14040270 - 11 Apr 2022

Cited by 12 | Viewed by 2411

Abstract

The corn earworm/bollworm, Helicoverpa zea (Boddie), is a pest species that is targeted by both Bacillus thuringiensis (Bt) maize and cotton in the United States. Cry1Ab and Vip3Aa20 are two common Bt toxins that are expressed in transgenic maize. The objective of this [...] Read more.

The corn earworm/bollworm, Helicoverpa zea (Boddie), is a pest species that is targeted by both Bacillus thuringiensis (Bt) maize and cotton in the United States. Cry1Ab and Vip3Aa20 are two common Bt toxins that are expressed in transgenic maize. The objective of this study was to determine the resistance allele frequency (RAF) to Cry1Ab and Vip3Aa20 in H. zea populations that were collected during 2018 and 2019 from four southeastern U.S. states: Louisiana, Mississippi, Georgia, and South Carolina. By using a group-mating approach, 104 F₂ iso-lines of H. zea were established from field collections with most iso-lines (85) from Louisiana. These F₂ iso-lines were screened for resistance alleles to Cry1Ab and Vip3Aa20, respectively. There was no correlation in larval survivorship between Cry1Ab and Vip3Aa20 when the iso-lines were exposed to these two toxins. RAF to Cry1Ab maize was high (0.256) and the RAFs were similar between Louisiana and the other three states and between the two sampling years. In contrast, no functional major resistance allele (RA) that allowed resistant insects to survive on Vip3Aa20 maize was detected and the expected RAF of major RAs with 95% probability was estimated to 0 to 0.0073. However, functional minor RAs to Vip3Aa20 maize were not uncommon; the estimated RAF for minor alleles was 0.028. The results provide further evidence that field resistance to Cry1Ab maize in H. zea has widely occurred, while major RAs to Vip3Aa20 maize are uncommon in the southeastern U.S. region. Information that was generated from this study should be useful in resistance monitoring and refinement of resistance management strategies to preserve Vip3A susceptibility in H. zea. Full article

(This article belongs to the Special Issue Insecticidal Toxins from Bacillus thuringiensis 2021–2022)

16 pages, 1576 KiB

Open AccessEditor’s ChoiceArticle

Cytotoxicity and Antiviral Properties of Alkaloids Isolated from Pancratium maritimum

by Marco Masi, Roberta Di Lecce, Natacha Mérindol, Marie-Pierre Girard, Lionel Berthoux, Isabel Desgagné-Penix, Viola Calabrò and Antonio Evidente

Toxins 2022, 14(4), 262; https://doi.org/10.3390/toxins14040262 - 7 Apr 2022

Cited by 10 | Viewed by 4164

Abstract

Ten Amaryllidaceae alkaloids (AAs) were isolated for the first time from Pancratium maritimum collected in Calabria region, Italy. They belong to different subgroups of this family and were identified as lycorine, which is the main alkaloid, 9-O-demethyllycorine, haemanthidine, haemanthamine, 11-hydroxyvittatine, homolycorine, [...] Read more.

Ten Amaryllidaceae alkaloids (AAs) were isolated for the first time from Pancratium maritimum collected in Calabria region, Italy. They belong to different subgroups of this family and were identified as lycorine, which is the main alkaloid, 9-O-demethyllycorine, haemanthidine, haemanthamine, 11-hydroxyvittatine, homolycorine, pancracine, obliquine, tazettine and vittatine. Haemanthidine was isolated as a scalar mixture of two 6-epimers, as already known also for other 6-hydroxycrinine alkaloids, but for the first time they were separated as 6,11-O,O′-di-p-bromobenzoyl esters. The evaluation of the cytotoxic and antiviral potentials of all isolated compounds was undertaken. Lycorine and haemanthidine showed cytotoxic activity on Hacat cells and A431 and AGS cancer cells while, pancracine exhibited selective cytotoxicity against A431 cells. We uncovered that in addition to lycorine and haemanthidine, haemanthamine and pancracine also possess antiretroviral abilities, inhibiting pseudotyped human immunodeficiency virus (HIV)−1 with EC50 of 25.3 µM and 18.5 µM respectively. Strikingly, all the AAs isolated from P. maritimum were able to impede dengue virus (DENV) replication (EC₅₀ ranged from 0.34–73.59 µM) at low to non-cytotoxic concentrations (CC₅₀ ranged from 6.25 µM to >100 µM). Haemanthamine (EC50 = 337 nM), pancracine (EC₅₀ = 357 nM) and haemanthidine (EC₅₀ = 476 nM) were the most potent anti-DENV inhibitors. Thus, this study uncovered new antiviral properties of P. maritimum isolated alkaloids, a significant finding that could lead to the development of new therapeutic strategies to fight viral infectious diseases. Full article

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18 pages, 1084 KiB

Open AccessEditor’s ChoiceArticle

Multiplex Detection of 24 Staphylococcal Enterotoxins in Culture Supernatant Using Liquid Chromatography Coupled to High-Resolution Mass Spectrometry

by Donatien Lefebvre, Kevin Blanco-Valle, Jacques-Antoine Hennekinne, Stéphanie Simon, François Fenaille, François Becher and Yacine Nia

Toxins 2022, 14(4), 249; https://doi.org/10.3390/toxins14040249 - 31 Mar 2022

Cited by 16 | Viewed by 5499

Abstract

Staphylococcal food poisoning outbreaks are caused by the ingestion of food contaminated with staphylococcal enterotoxins (SEs). Among the 27 SEs described in the literature to date, only a few can be detected using immuno-enzymatic-based methods that are strongly dependent on the availability of [...] Read more.

Staphylococcal food poisoning outbreaks are caused by the ingestion of food contaminated with staphylococcal enterotoxins (SEs). Among the 27 SEs described in the literature to date, only a few can be detected using immuno-enzymatic-based methods that are strongly dependent on the availability of antibodies. Liquid chromatography, coupled to high-resolution mass spectrometry (LC-HRMS), has, therefore, been put forward as a relevant complementary method, but only for the detection of a limited number of enterotoxins. In this work, LC-HRMS was developed for the detection and quantification of 24 SEs. A database of 93 specific signature peptides and LC-HRMS parameters was optimized using sequences from 24 SEs, including their 162 variants. A label-free quantification protocol was established to overcome the absence of calibration standards. The LC-HRMS method showed high performance in terms of specificity, sensitivity, and accuracy when applied to 49 enterotoxin-producing strains. SE concentrations measured depended on both SE type and the coagulase-positive staphylococci (CPS) strain. This study indicates that LC-MS is a relevant alternative and complementary tool to ELISA methods. The advantages of LC-MS clearly lie in both the multiplex analysis of a large number of SEs, and the automated analysis of a high number of samples. Full article

(This article belongs to the Special Issue Foodborne Intoxications and Toxicoinfections—Major Pathogens and Challenges)

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25 pages, 6391 KiB

Open AccessEditor’s ChoiceArticle

Differences in PLA₂ Constitution Distinguish the Venom of Two Endemic Brazilian Mountain Lanceheads, Bothrops cotiara and Bothrops fonsecai

by Pedro G. Nachtigall, Luciana A. Freitas-de-Sousa, Andrew J. Mason, Ana M. Moura-da-Silva, Felipe G. Grazziotin and Inácio L. M. Junqueira-de-Azevedo

Toxins 2022, 14(4), 237; https://doi.org/10.3390/toxins14040237 - 25 Mar 2022

Cited by 7 | Viewed by 3183

Abstract

Interspecific differences in snake venom compositions can result from distinct regulatory mechanisms acting in each species. However, comparative analyses focusing on identifying regulatory elements and patterns that led to distinct venom composition are still scarce. Among venomous snakes, Bothrops cotiara and Bothrops fonsecai [...] Read more.

Interspecific differences in snake venom compositions can result from distinct regulatory mechanisms acting in each species. However, comparative analyses focusing on identifying regulatory elements and patterns that led to distinct venom composition are still scarce. Among venomous snakes, Bothrops cotiara and Bothrops fonsecai represent ideal models to complement our understanding of the regulatory mechanisms of venom production. These recently diverged species share a similar specialized diet, habitat, and natural history, but each presents a distinct venom phenotype. Here, we integrated data from the venom gland transcriptome and miRNome and the venom proteome of B. fonsecai and B. cotiara to better understand the regulatory mechanisms that may be acting to produce differing venom compositions. We detected not only the presence of similar toxin isoforms in both species but also distinct expression profiles of phospholipases A

_{2}

(PLA2) and some snake venom metalloproteinases (SVMPs) and snake venom serine proteinases (SVSPs) isoforms. We found evidence of modular expression regulation of several toxin isoforms implicated in venom divergence and observed correlated expression of several transcription factors. We did not find strong evidence for miRNAs shaping interspecific divergence of the venom phenotypes, but we identified a subset of toxin isoforms whose final expression may be fine-tuned by specific miRNAs. Sequence analysis on orthologous toxins showed a high rate of substitutions between PLA2s, which indicates that these toxins may be under strong positive selection or represent paralogous toxins in these species. Our results support other recent studies in suggesting that gene regulation is a principal mode of venom evolution across recent timescales, especially among species with conserved ecotypes. Full article

(This article belongs to the Section Animal Venoms)

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18 pages, 3965 KiB

Open AccessEditor’s ChoiceArticle

Bitiscetin-3, a Novel C-Type Lectin-like Protein Cloned from the Venom Gland of the Viper Bitis arietans, Induces Platelet Agglutination and Inhibits Binding of Von Willebrand Factor to Collagen

by Youhei Nashimoto, Fumio Matsushita, Johannes M. Dijkstra, Yuta Nakamura, Hidehiko Akiyama, Jiharu Hamako, Takashi Morita, Satohiko Araki and Taei Matsui

Toxins 2022, 14(4), 236; https://doi.org/10.3390/toxins14040236 - 25 Mar 2022

Cited by 2 | Viewed by 2488

Abstract

Bitiscetin-1 (aka bitiscetin) and bitiscetin-2 are C-type lectin-like proteins purified from the venom of Bitis arietans (puff adder). They bind to von Willebrand factor (VWF) and—at least bitiscetin-1—induce platelet agglutination via enhancement of VWF binding to platelet glycoprotein Ib (GPIb). Bitiscetin-1 and -2 [...] Read more.

Bitiscetin-1 (aka bitiscetin) and bitiscetin-2 are C-type lectin-like proteins purified from the venom of Bitis arietans (puff adder). They bind to von Willebrand factor (VWF) and—at least bitiscetin-1—induce platelet agglutination via enhancement of VWF binding to platelet glycoprotein Ib (GPIb). Bitiscetin-1 and -2 bind the VWF A1 and A3 domains, respectively. The A3 domain includes the major site of VWF for binding collagen, explaining why bitiscetin-2 blocks VWF-to-collagen binding. In the present study, sequences for a novel bitiscetin protein—bitiscetin-3—were identified in cDNA constructed from the B. arietans venom gland. The deduced amino acid sequences of bitiscetin-3 subunits α and β share 79 and 80% identity with those of bitiscetin-1, respectively. Expression vectors for bitiscetin-3α and -3β were co-transfected to 293T cells, producing the heterodimer protein recombinant bitiscetin-3 (rBit-3). Functionally, purified rBit-3 (1) induced platelet agglutination involving VWF and GPIb, (2) did not compete with bitiscetin-1 for binding to VWF, (3) blocked VWF-to-collagen binding, and (4) lost its platelet agglutination inducing ability in the presence of an anti-VWF monoclonal antibody that blocked VWF-to-collagen binding. These combined results suggest that bitiscetin-3 binds to the A3 domain, as does bitiscetin-2. Except for a small N-terminal fragment of a single subunit—which differs from that of both bitiscetin-3 subunits—the sequences of bitiscetin-2 have never been determined. Therefore, by identifying and analyzing bitiscetin-3, the present study is the first to present the full-length α- and β-subunit sequences and recombinant expression of a bitiscetin-family toxin that blocks the binding of VWF to collagen. Full article

(This article belongs to the Section Animal Venoms)

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19 pages, 2651 KiB

Open AccessEditor’s ChoiceArticle

Venomics Reveals a Non-Compartmentalised Venom Gland in the Early Diverged Vermivorous Conus distans

by Jutty Rajan Prashanth, Sebastien Dutertre, Subash Kumar Rai and Richard J. Lewis

Toxins 2022, 14(3), 226; https://doi.org/10.3390/toxins14030226 - 19 Mar 2022

Cited by 2 | Viewed by 2675

Abstract

The defensive use of cone snail venom is hypothesised to have first arisen in ancestral worm-hunting snails and later repurposed in a compartmentalised venom duct to facilitate the dietary shift to molluscivory and piscivory. Consistent with its placement in a basal lineage, we [...] Read more.

The defensive use of cone snail venom is hypothesised to have first arisen in ancestral worm-hunting snails and later repurposed in a compartmentalised venom duct to facilitate the dietary shift to molluscivory and piscivory. Consistent with its placement in a basal lineage, we demonstrate that the C. distans venom gland lacked distinct compartmentalisation. Transcriptomics revealed C. distans expressed a wide range of structural classes, with inhibitory cysteine knot (ICK)-containing peptides dominating. To better understand the evolution of the venom gland compartmentalisation, we compared C. distans to C. planorbis, the earliest diverging species from which a defence-evoked venom has been obtained, and fish-hunting C. geographus from the Gastridium subgenus that injects distinct defensive and predatory venoms. These comparisons support the hypothesis that venom gland compartmentalisation arose in worm-hunting species and enabled repurposing of venom peptides to facilitate the dietary shift from vermivory to molluscivory and piscivory in more recently diverged cone snail lineages. Full article

(This article belongs to the Special Issue Predatory and Defensive Venom Peptides)

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12 pages, 1471 KiB

Open AccessEditor’s ChoiceArticle

A Potent Inhibitor of the Cystic Fibrosis Transmembrane Conductance Regulator Blocks Disease and Morbidity Due to Toxigenic Vibrio cholerae

by Fabian Rivera-Chávez, Bradley T. Meader, Sinan Akosman, Vuk Koprivica and John J. Mekalanos

Toxins 2022, 14(3), 225; https://doi.org/10.3390/toxins14030225 - 18 Mar 2022

Cited by 8 | Viewed by 5021

Abstract

Vibrio cholerae uses cholera toxin (CT) to cause cholera, a severe diarrheal disease in humans that can lead to death within hours of the onset of symptoms. The catalytic activity of CT in target epithelial cells increases cellular levels of 3′,5′-cyclic AMP (cAMP), [...] Read more.

Vibrio cholerae uses cholera toxin (CT) to cause cholera, a severe diarrheal disease in humans that can lead to death within hours of the onset of symptoms. The catalytic activity of CT in target epithelial cells increases cellular levels of 3′,5′-cyclic AMP (cAMP), leading to the activation of the cystic fibrosis transmembrane conductance regulator (CFTR), an apical ion channel that transports chloride out of epithelial cells, resulting in an electrolyte imbalance in the intestinal lumen and massive water loss. Here we report that when administered perorally, benzopyrimido-pyrrolo-oxazinedione, (R)-BPO-27), a potent small molecule inhibitor of CFTR, blocked disease symptoms in a mouse model for acute diarrhea caused by toxigenic V. cholerae. We show that both (R)-BPO-27 and its racemic mixture, (R/S)-BPO-27, are able to protect mice from CT-dependent diarrheal disease and death. Furthermore, we show that, consistent with the ability of the compound to block the secretory diarrhea induced by CT, BPO-27 has a measurable effect on suppressing the gut replication and survival of V. cholerae, including a 2010 isolate from Haiti that is representative of the most predominant ‘variant strains’ that are causing epidemic and pandemic cholera worldwide. Our results suggest that BPO-27 should advance to human Phase I studies that could further address its safety and efficacy as therapeutic or preventative drug intervention for diarrheal syndromes, including cholera, that are mediated by CFTR channel activation. Full article

(This article belongs to the Special Issue Protein Toxins of Pathogenic Vibrio Species)

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16 pages, 2170 KiB

Open AccessEditor’s ChoiceArticle

Warming and Salt Intrusion Affect Microcystin Production in Tropical Bloom-Forming Microcystis

by Bui Trung, Marlies E. Vollebregt and Miquel Lürling

Toxins 2022, 14(3), 214; https://doi.org/10.3390/toxins14030214 - 16 Mar 2022

Cited by 5 | Viewed by 2627

Abstract

The Vietnamese Mekong Delta is predicted to be one of the regions most impacted by climate change, causing increased temperature and salinity in inland waters. We hypothesized that the increase in temperature and salinity may impact the microcystin (MC) production of two Microcystis [...] Read more.

The Vietnamese Mekong Delta is predicted to be one of the regions most impacted by climate change, causing increased temperature and salinity in inland waters. We hypothesized that the increase in temperature and salinity may impact the microcystin (MC) production of two Microcystis strains isolated in this region from a freshwater pond (strain MBC) and a brackish water pond (strain MTV). The Microcystis strains were grown at low (27 °C), medium (31 °C), high (35 °C) and extremely high (37 °C) temperature in flat photobioreactors (Algaemist). At each temperature, when cultures reached a stable state, sea salt was added to increase salinity to 4‰, 8‰, 12‰ and 16‰. MC concentrations and cell quota were reduced at high and extremely high temperatures. Salinity, in general, had comparable effects on MC concentrations and quota. At a salinity of 4‰ and 8‰, concentrations of MC per mL of culture and MC cell quota (based on chlorophyll, dry-weight and particle counts) were higher than at 0.5‰, while at the highest salinities (12‰ and 16‰) these were strongly reduced. Strain MBC produced five MC variants of which MC-RR and MC-LR were most abundant, followed by MC-YR and relatively low amounts of demethylated variants dmMC-RR and dmMC-LR. In strain MTV, MC-RR was most abundant, with traces of MC-YR and dmMC-RR only in cultures grown at 16‰ salinity. Overall, higher temperature led to lower MC concentrations and cell quota, low salinity seemed to promote MC production and high salinity reduced MC production. Hence, increased temperature and higher salinity could lead to less toxic Microcystis, but since these conditions might favour Microcystis over other competitors, the overall biomass gain could offset a lower toxicity. Full article

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

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16 pages, 2557 KiB

Open AccessEditor’s ChoiceArticle

Assessing the Validity of Normalizing Aflatoxin B₁-Lysine Albumin Adduct Biomarker Measurements to Total Serum Albumin Concentration across Multiple Human Population Studies

by Joshua W. Smith, Derek K. Ng, Christian S. Alvarez, Patricia A. Egner, Sean M. Burke, Jian-Guo Chen, Thomas W. Kensler, Jill Koshiol, Alvaro Rivera-Andrade, María F. Kroker-Lobos, Manuel Ramírez-Zea, Katherine A. McGlynn and John D. Groopman

Toxins 2022, 14(3), 162; https://doi.org/10.3390/toxins14030162 - 23 Feb 2022

Cited by 4 | Viewed by 2357

Abstract

The assessment of aflatoxin B₁ (AFB₁) exposure using isotope-dilution liquid chromatography-mass spectrometry (LCMS) of AFB₁-lysine adducts in human serum albumin (HSA) has proven to be a highly productive strategy for the biomonitoring of AFB₁ exposure. To compare [...] Read more.

The assessment of aflatoxin B₁ (AFB₁) exposure using isotope-dilution liquid chromatography-mass spectrometry (LCMS) of AFB₁-lysine adducts in human serum albumin (HSA) has proven to be a highly productive strategy for the biomonitoring of AFB₁ exposure. To compare samples across different individuals and settings, the conventional practice has involved the normalization of raw AFB₁-lysine adduct concentrations (e.g., pg/mL serum or plasma) to the total circulating HSA concentration (e.g., pg/mg HSA). It is hypothesized that this practice corrects for technical error, between-person variance in HSA synthesis or AFB₁ metabolism, and other factors. However, the validity of this hypothesis has been largely unexamined by empirical analysis. The objective of this work was to test the concept that HSA normalization of AFB₁-lysine adduct concentrations effectively adjusts for biological and technical variance and improves AFB₁ internal dose estimates. Using data from AFB₁-lysine and HSA measurements in 763 subjects, in combination with regression and Monte Carlo simulation techniques, we found that HSA accounts for essentially none of the between-person variance in HSA-normalized (R² = 0.04) or raw AFB₁-lysine measurements (R² = 0.0001), and that HSA normalization of AFB₁-lysine levels with empirical HSA values does not reduce measurement error any better than does the use of simulated data (n = 20,000). These findings were robust across diverse populations (Guatemala, China, Chile), AFB₁ exposures (10⁵ range), HSA assays (dye-binding and immunoassay), and disease states (healthy, gallstones, and gallbladder cancer). HSA normalization results in arithmetic transformation with the addition of technical error from the measurement of HSA. Combined with the added analysis time, cost, and sample consumption, these results suggest that it may be prudent to abandon the practice of normalizing adducts to HSA concentration when measuring any HSA adducts—not only AFB₁-lys adducts—when using LCMS in serum/plasma. Full article

(This article belongs to the Special Issue Mycotoxin Biomarkers: Innovation and Utility)

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19 pages, 2250 KiB

Open AccessEditor’s ChoiceArticle

ExoU Induces Lung Endothelial Cell Damage and Activates Pro-Inflammatory Caspase-1 during Pseudomonas aeruginosa Infection

by Kierra S. Hardy, Amanda N. Tuckey, Phoibe Renema, Mita Patel, Abu-Bakr Al-Mehdi, Domenico Spadafora, Cody A. Schlumpf, Robert A. Barrington, Mikhail F. Alexeyev, Troy Stevens, Jean-Francois Pittet, Brant M. Wagener, Jon D. Simmons, Diego F. Alvarez and Jonathon P. Audia

Toxins 2022, 14(2), 152; https://doi.org/10.3390/toxins14020152 - 18 Feb 2022

Cited by 9 | Viewed by 3007

Abstract

The Gram-negative, opportunistic pathogen Pseudomonas aeruginosa utilizes a type III secretion system to inject exoenzyme effectors into a target host cell. Of the four best-studied exoenzymes, ExoU causes rapid cell damage and death. ExoU is a phospholipase A₂ (PLA₂) that [...] Read more.

The Gram-negative, opportunistic pathogen Pseudomonas aeruginosa utilizes a type III secretion system to inject exoenzyme effectors into a target host cell. Of the four best-studied exoenzymes, ExoU causes rapid cell damage and death. ExoU is a phospholipase A₂ (PLA₂) that hydrolyses host cell membranes, and P. aeruginosa strains expressing ExoU are associated with poor outcomes in critically ill patients with pneumonia. While the effects of ExoU on lung epithelial and immune cells are well studied, a role for ExoU in disrupting lung endothelial cell function has only recently emerged. Lung endothelial cells maintain a barrier to fluid and protein flux into tissue and airspaces and regulate inflammation. Herein, we describe a pulmonary microvascular endothelial cell (PMVEC) culture infection model to examine the effects of ExoU. Using characterized P. aeruginosa strains and primary clinical isolates, we show that strains expressing ExoU disrupt PMVEC barrier function by causing substantial PMVEC damage and lysis, in a PLA₂-dependent manner. In addition, we show that strains expressing ExoU activate the pro-inflammatory caspase-1, in a PLA₂-dependent manner. Considering the important roles for mitochondria and oxidative stress in regulating inflammatory responses, we next examined the effects of ExoU on reactive oxygen species production. Infection of PMVECs with P. aeruginosa strains expressing ExoU triggered a robust oxidative stress compared to strains expressing other exoenzyme effectors. We also provide evidence that, intriguingly, ExoU PLA₂ activity was detectable in mitochondria and mitochondria-associated membrane fractions isolated from P. aeruginosa-infected PMVECs. Interestingly, ExoU-mediated activation of caspase-1 was partially inhibited by reactive oxygen species scavengers. Together, these data suggest ExoU exerts pleiotropic effects on PMVEC function during P. aeruginosa infection that may inhibit endothelial barrier and inflammatory functions. Full article

(This article belongs to the Section Bacterial Toxins)

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17 pages, 1691 KiB

Open AccessEditor’s ChoiceArticle

Efficacy of Fumonisin Esterase in Piglets as Animal Model for Fumonisin Detoxification in Humans: Pilot Study Comparing Intraoral to Intragastric Administration

by Kaat Neckermann, Gunther Antonissen, Barbara Doupovec, Dian Schatzmayr, James Gathumbi, Véronique Delcenserie, Silvio Uhlig and Siska Croubels

Toxins 2022, 14(2), 136; https://doi.org/10.3390/toxins14020136 - 11 Feb 2022

Cited by 2 | Viewed by 2590

Abstract

Fumonisins, a group of highly prevalent and toxic mycotoxins, are suspected to be causal agents of several diseases in animals and humans. In the animal feed industry, fumonisin esterase is used as feed additive to prevent mycotoxicosis caused by fumonisins. In humans, a [...] Read more.

Fumonisins, a group of highly prevalent and toxic mycotoxins, are suspected to be causal agents of several diseases in animals and humans. In the animal feed industry, fumonisin esterase is used as feed additive to prevent mycotoxicosis caused by fumonisins. In humans, a popular dosage form for dietary supplements, with high patient acceptance for oral intake, is capsule ingestion. Thus, fumonisin esterase provided in a capsule could be an effective strategy against fumonisin intoxication in humans. To determine the efficacy of fumonisin esterase through capsule ingestion, two modes of application were compared using piglets in a small-scale preliminary study. The enzyme was administered intraorally (in-feed analogue) or intragastrically (capsule analogue), in combination with fumonisin B1 (FB1). Biomarkers for FB1 exposure; namely FB1, hydrolysed FB1 (HFB1) and partially hydrolysed forms (pHFB1a and pHFB1b), were measured both in serum and faeces using a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, and toxicokinetic parameters were calculated. Additionally, the serum sphinganine/sphingosine (Sa/So) ratio, a biomarker of effect, was determined using LC-MS/MS. A significantly higher Sa/So ratio was shown in the placebo group compared to both esterase treatments, demonstrating the efficacy of the esterase. Moreover, a significant decrease in serum FB1 area under the concentration-time curve (AUC) and an increase of faecal HFB1 AUC were observed after intraoral esterase administration. However, these effects were not observed with statistical significance after intragastric esterase administration with the current sample size. Full article

(This article belongs to the Special Issue Mycotoxins and Their Chromatographic-Based Detection Technology)

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12 pages, 348 KiB

Open AccessEditor’s ChoiceArticle

Analysis of Mycotoxin and Secondary Metabolites in Commercial and Traditional Slovak Cheese Samples

by Luana Izzo, Petra Mikušová, Sonia Lombardi, Michael Sulyok and Alberto Ritieni

Toxins 2022, 14(2), 134; https://doi.org/10.3390/toxins14020134 - 10 Feb 2022

Cited by 11 | Viewed by 3156

Abstract

Cheese represents a dairy product extremely inclined to fungal growth and mycotoxin production. The growth of fungi belonging to Aspergillus, Penicillium, Fusarium, Claviceps, Alternaria, and Trichoderma genera in or on cheese leads to undesirable changes able to affect [...] Read more.

Cheese represents a dairy product extremely inclined to fungal growth and mycotoxin production. The growth of fungi belonging to Aspergillus, Penicillium, Fusarium, Claviceps, Alternaria, and Trichoderma genera in or on cheese leads to undesirable changes able to affect the quality of the final products. In the present investigation, a total of 68 types of commercial and traditional Slovak cheeses were analyzed to investigate the occurrence of fungal metabolites. Altogether, 13 fungal metabolites were identified and quantified. Aflatoxin M1, the only mycotoxin regulated in milk and dairy products, was not detected in any case. However, the presence of metabolites that have never been reported in cheeses, such as tryptophol at a maximum concentration level from 13.4 to 7930 µg/kg (average: 490 µg/kg), was recorded. Out of all detected metabolites, enniatin B represents the most frequently detected mycotoxin (0.06–0.71 µg/kg) in the analyzed samples. Attention is drawn to the lack of data on mycotoxins’ origin from Slovak cheeses; in fact, this is the first reported investigation. Our results indicate the presence of fungal mycotoxin contamination for which maximum permissible levels are not established, highlighting the importance of monitoring the source and producers of contamination in order to protect consumers’ health. Full article

(This article belongs to the Special Issue Mycotoxins in Food and Feed: Detection and Identification)

25 pages, 3961 KiB

Open AccessEditor’s ChoiceArticle

Host Genotype and Weather Effects on Fusarium Head Blight Severity and Mycotoxin Load in Spring Barley

by Felix Hoheneder, Eva Maria Biehl, Katharina Hofer, Johannes Petermeier, Jennifer Groth, Markus Herz, Michael Rychlik, Michael Heß and Ralph Hückelhoven

Toxins 2022, 14(2), 125; https://doi.org/10.3390/toxins14020125 - 8 Feb 2022

Cited by 7 | Viewed by 2848

Abstract

Epidemiology of Fusarium Head Blight (FHB) of spring barley is relatively little understood. In a five-year study, we assessed quantitative resistance to FHB in an assortment of 17 spring barley genotypes in the field in southern Germany. To this end, we used soil [...] Read more.

Epidemiology of Fusarium Head Blight (FHB) of spring barley is relatively little understood. In a five-year study, we assessed quantitative resistance to FHB in an assortment of 17 spring barley genotypes in the field in southern Germany. To this end, we used soil and spray inoculation of plants with F. culmorum and F. avenaceum. This increased disease pressure and provoked genotypic differentiation. To normalize effects of variable weather conditions across consecutive seasons, we used a disease ranking of the genotypes based on quantification of fungal DNA contents and multiple Fusarium toxins in harvested grain. Together, this allowed for assessment of stable quantitative FHB resistance of barley in several genotypes. Fungal DNA contents were positively associated with species-specific Fusarium toxins in single years and over several years in plots with soil inoculation. In those plots, plant height limited FHB; however, this was not observed after spray inoculation. A multiple linear regression model of recorded weather parameter and fungal DNA contents over five years identified time periods during the reproductive phase of barley, in which weather strongly influenced fungal colonization measured in mature barley grain. Environmental conditions before heading and late after anthesis showed strongest associations with F. culmorum DNA in all genotypes, whereas for F. avenaceum, this was less consistent where we observed weather-dependent associations, depending on the genotype. Based on this study, we discuss aspects of practical resistance breeding in barley relevant to improve quantitative resistance to FHB and associated mycotoxin contaminations. Full article

(This article belongs to the Special Issue Selected Papers from the 15th European Fusarium Seminar)

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12 pages, 1313 KiB

Open AccessEditor’s ChoiceArticle

Different Algicidal Modes of the Two Bacteria Aeromonas bestiarum HYD0802-MK36 and Pseudomonas syringae KACC10292^T against Harmful Cyanobacteria Microcystis aeruginosa

by Bum Soo Park, Chong-Sung Park, Yuna Shin, Sungae Yoon, Myung-Soo Han and Yoon-Ho Kang

Toxins 2022, 14(2), 128; https://doi.org/10.3390/toxins14020128 - 8 Feb 2022

Cited by 7 | Viewed by 2713

Abstract

Blooms of harmful cyanobacteria Microcystis aeruginosa lead to an adverse effect on freshwater ecosystems, and thus extensive studies on the control of this cyanobacteria’s blooms have been conducted. Throughout this study, we have found that the two bacteria Aeromonas bestiarum HYD0802-MK36 and Pseudomonas [...] Read more.

Blooms of harmful cyanobacteria Microcystis aeruginosa lead to an adverse effect on freshwater ecosystems, and thus extensive studies on the control of this cyanobacteria’s blooms have been conducted. Throughout this study, we have found that the two bacteria Aeromonas bestiarum HYD0802-MK36 and Pseudomonas syringae KACC10292^T are capable of killing M. aeruginosa. Interestingly, these two bacteria showed different algicidal modes. Based on an algicidal range test using 15 algal species (target and non-target species), HYD0802-MK36 specifically attacked only target cyanobacteria M. aeruginosa, whereas the algicidal activity of KACC10292^T appeared in a relatively broad algicidal range. HYD0802-MK36, as a direct attacker, killed M. aeruginosa cells when direct cell (bacterium)-to-cell (cyanobacteria) contact happens. KACC10292^T, as an indirect attacker, released algicidal substance which is located in cytoplasm. Interestingly, algicidal activity of KACC10292^T was enhanced according to co-cultivation with the host cyanobacteria, suggesting that quantity of algicidal substance released from this bacterium might be increased via interaction with the host cyanobacteria. Full article

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

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11 pages, 1516 KiB

Open AccessEditor’s ChoiceArticle

Ultra-Long-Term Therapy of Benign Essential Blepharospasm with Botulinumtoxin A—30 Years of Experience in a Tertiary Care Center

by Bettina Wabbels, Rolf Fimmers and Peter Roggenkämper

Toxins 2022, 14(2), 120; https://doi.org/10.3390/toxins14020120 - 7 Feb 2022

Cited by 3 | Viewed by 2171

Abstract

Aim of this study was to investigate the long-term results of botulinum toxin A (BoNT-A) injections for the treatment of benign essential blepharospasm (BEB) and to report our experience with (ultra-)long-term treatment with onabotulinumtoxin-A. We conducted a retrospective cross-sectional analysis at a university [...] Read more.

Aim of this study was to investigate the long-term results of botulinum toxin A (BoNT-A) injections for the treatment of benign essential blepharospasm (BEB) and to report our experience with (ultra-)long-term treatment with onabotulinumtoxin-A. We conducted a retrospective cross-sectional analysis at a university hospital. Patients with BEB and BoNT-A treatment were assigned to the Total Blepharospasm Group, patients with ≥21 onabotulinumtoxin-A injections to the Ona Long-Term Group. The Total Blepharospasm Group (n = 1940) included 33,933 BoNT-A injections. The age of patients at symptom onset was (mean ± SD) 58.0 ± 13.1 years, and 70.4% were female. The Ona long-term group (n = 234) included 10,632 onabotulinumtoxin-A injections. In this group, patients received 45.4 ± 22.9 injections with a mean dose of 22.2 IU ± 0.5. The duration of treatment was 12.6 ± 5.4 years, ranging from 2.9 to 30.0 years. The effect–duration–dose quotient did not change during long-term treatment. The observed side effects were comparable in type and frequency to other studies, even with the (ultra-)long treatment with onabotulinumtoxin-A. Our results, based on one of the largest patient populations and a treatment duration of up to 30 years, impressively demonstrate that onabotulinumtoxin-A is a safe and effective therapy for essential blepharospasm, even in the ultra-long term. Full article

(This article belongs to the Special Issue Clinical Application of Botulinum Toxin)

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18 pages, 4066 KiB

Open AccessEditor’s ChoiceArticle

Susceptibility of Sea Bream (Sparus aurata) to AIP56, an AB-Type Toxin Secreted by Photobacterium damselae subsp. piscicida

by Inês Lua Freitas, Alexandra Teixeira, Inês Loureiro, Johnny Lisboa, Aurélia Saraiva, Nuno Miguel Simões dos Santos and Ana do Vale

Toxins 2022, 14(2), 119; https://doi.org/10.3390/toxins14020119 - 5 Feb 2022

Cited by 5 | Viewed by 2912

Abstract

Photobacterium damselae subsp. piscicida (Phdp) is a Gram-negative bacterium that infects a large number of marine fish species in Europe, Asia, and America, both in aquacultures and in the natural environment. Among the affected hosts are economically important cultured fish, such [...] Read more.

Photobacterium damselae subsp. piscicida (Phdp) is a Gram-negative bacterium that infects a large number of marine fish species in Europe, Asia, and America, both in aquacultures and in the natural environment. Among the affected hosts are economically important cultured fish, such as sea bream (Sparus aurata), sea bass (Dicentrarchus labrax), yellowtail (Seriola quinqueradiata), and cobia (Rachycentron canadum). The best characterized virulence factor of Phdp is the Apoptosis-Inducing Protein of 56 kDa (AIP56), a secreted AB-type toxin that has been shown to induce apoptosis of sea bass phagocytes during infection. AIP56 has an A subunit that displays metalloprotease activity against NF-kB p65 and a B subunit that mediates binding and internalization of the A subunit in susceptible cells. Despite the fact that the aip56 gene is highly prevalent in Phdp isolates from different fish species, the toxicity of AIP56 has only been studied in sea bass. In the present study, the toxicity of AIP56 for sea bream was evaluated. Ex vivo assays showed that sea bream phagocytes are resistant to AIP56 cytotoxicity and that resistance was associated with an inefficient internalization of the toxin by those cells. Accordingly, in vivo intoxication assays revealed that sea bream is much more resistant to AIP56-induced lethality than sea bass. These findings, showing that the effect of AIP56 is different in sea bass and sea bream, set the basis for future studies to characterize the effects of AIP56 and to fully elucidate its virulence role in different Phdp susceptible hosts. Full article

(This article belongs to the Special Issue Bacterial Exotoxins and Their Impact on Host–Pathogen Interaction in Animals and Humans)

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17 pages, 11262 KiB

Open AccessEditor’s ChoiceArticle

Regulated Mycotoxin Occurrence and Co-Occurrence in Croatian Cereals

by Marija Kovač, Mateja Bulaić, Ante Nevistić, Tomislav Rot, Jurislav Babić, Mario Panjičko, Tihomir Kovač and Bojan Šarkanj

Toxins 2022, 14(2), 112; https://doi.org/10.3390/toxins14020112 - 2 Feb 2022

Cited by 15 | Viewed by 2775

Abstract

A total of 209 samples of various cereal crops (maize, wheat, barley, rye and oats) grown in Croatian fields during 2016 and 2017 were collected to analyze and determine the occurrence and co-occurrence of EU regulated mycotoxins in cereals (AFB1, AFB2, AFG1, AFG2, [...] Read more.

A total of 209 samples of various cereal crops (maize, wheat, barley, rye and oats) grown in Croatian fields during 2016 and 2017 were collected to analyze and determine the occurrence and co-occurrence of EU regulated mycotoxins in cereals (AFB1, AFB2, AFG1, AFG2, DON, FB1, FB2, ZEA, T-2, HT-2 and OTA). The analysis, performed by a validated confirmatory LC-MS/MS method based on a dilute and shoot principle, highlighted Fusarium mycotoxins as the main contaminants, often co-occurring in samples from both years (50.0% in 2016 and 33.7% in 2017). DON was found to be the most frequent mycotoxin, present in 72.5% of the 2016 samples and 32.6% of the 2017 samples, while maize proved to be the most contaminated cereal type of both years with FUM as the most abundant mycotoxins, with an average concentration of 1180 µg/kg. Moderate temperatures with periods of high humidity favored the accumulation of DON in wheat samples instead of other Fusarium mycotoxins, while similar conditions favored maize contamination with FUM. A total of 8.3% of all the 2016 harvest samples and 7.9% of the 2017 harvest samples were assessed as non-compliant, containing mycotoxins in concentrations higher than the levels set by the EU legislation for food. Full article

(This article belongs to the Special Issue Environmental Stress on the Production of Mycotoxins)

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14 pages, 2401 KiB

Open AccessEditor’s ChoiceArticle

Tissular Genomic Responses to Oral FB1 Exposure in Pigs

by Léonie Dopavogui, Arnaud Polizzi, Anne Fougerat, Pascal Gourbeyre, Chloé Terciolo, Wendy Klement, Philippe Pinton, Joëlle Laffite, Anne-Marie Cossalter, Jean-Denis Bailly, Olivier Puel, Yannick Lippi, Claire Naylies, Hervé Guillou, Isabelle P. Oswald and Nicolas Loiseau

Toxins 2022, 14(2), 83; https://doi.org/10.3390/toxins14020083 - 22 Jan 2022

Cited by 3 | Viewed by 2629

Abstract

Fumonisin B1 (FB1) is a widespread mycotoxin produced by fungal Fusarium species—mainly in maize, one of the plants most commonly used for food and feed. Pigs and horses are the animal species most susceptible to this mycotoxin. FB1 exposure can cause highly diverse [...] Read more.

Fumonisin B1 (FB1) is a widespread mycotoxin produced by fungal Fusarium species—mainly in maize, one of the plants most commonly used for food and feed. Pigs and horses are the animal species most susceptible to this mycotoxin. FB1 exposure can cause highly diverse clinical symptoms, including hepatotoxicity, immunotoxicity, and intestinal barrier function disturbance. Inhibition of ceramide synthetase is a well-understood ubiquitous molecular mechanism of FB1 toxicity, but other more tissue-specific effects remain to be elucidated. To investigate the effects of FB1 in different exposed tissues, we cross-analyzed the transcriptomes of fours organs: liver, jejunum, jejunal Peyer’s patches, and spleen. During a four-week study period, pigs were fed a control diet or a FB1-contaminated diet (10 mg/kg feed). In response to oral FB1 exposure, we observed common biological processes in the four organs, including predominant and recurrent processes (extracellular matrix organization, integrin activation, granulocyte chemotaxis, neutrophil migration, and lipid and sterol homeostasis), as well as more tissue-specific processes that appeared to be related to lipid outcomes (cell cycle regulation in jejunum, and gluconeogenesis in liver). Full article

(This article belongs to the Special Issue Toxic Effect of Mycotoxins)

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15 pages, 782 KiB

Open AccessEditor’s ChoiceArticle

Use of Mustard Extracts Fermented by Lactic Acid Bacteria to Mitigate the Production of Fumonisin B₁ and B₂ by Fusarium verticillioides in Corn Ears

by Raquel Torrijos, Tiago de Melo Nazareth, Pilar Vila-Donat, Jordi Mañes and Giuseppe Meca

Toxins 2022, 14(2), 80; https://doi.org/10.3390/toxins14020080 - 21 Jan 2022

Cited by 4 | Viewed by 3885

Abstract

Corn (Zea mays) is a worldwide crop subjected to infection by toxigenic fungi such as Fusarium verticillioides during the pre-harvest stage. Fusarium contamination can lead to the synthesis of highly toxic mycotoxins, such as Fumonisin B₁ (FB₁) and [...] Read more.

Corn (Zea mays) is a worldwide crop subjected to infection by toxigenic fungi such as Fusarium verticillioides during the pre-harvest stage. Fusarium contamination can lead to the synthesis of highly toxic mycotoxins, such as Fumonisin B₁ (FB₁) and Fumonisin B₂ (FB₂), which compromises human and animal health. The work aimed to study the antifungal properties of fermented yellow and oriental mustard extracts using nine lactic acid bacteria (LAB) in vitro. Moreover, a chemical characterization of the main phenolic compounds and organic acids were carried out in the extracts. The results highlighted that the yellow mustard, fermented by Lactiplantibacillus plantarum strains, avoided the growth of Fusarium spp. in vitro, showing Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC) values, ranging from 7.8 to 15.6 g/L and 15.6 to 31.3 g/L, respectively. Then, the lyophilized yellow mustard fermented extract by L. plantarum TR71 was applied through spray-on corn ears contaminated with F. verticillioides to study the antimycotoxigenic activity. After 14 days of incubation, the control contained 14.71 mg/kg of FB₁, while the treatment reduced the content to 1.09 mg/kg (92.6% reduction). Moreover, no FB₂ was observed in the treated samples. The chemical characterization showed that lactic acid, 3-phenyllactic acid, and benzoic acid were the antifungal metabolites quantified in higher concentrations in the yellow mustard fermented extract with L. plantarum TR71. The results obtained confirmed the potential application of fermented mustard extracts as a solution to reduce the incidence of mycotoxins in corn ears. Full article

(This article belongs to the Special Issue Reduction and Control of Mycotoxins along Entire Food and Feed Chain)

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11 pages, 1888 KiB

Open AccessEditor’s ChoiceArticle

Discrimination of the Activity of Low-Affinity Wild-Type and High-Affinity Mutant Recombinant BoNT/B by a SIMA Cell-Based Reporter Release Assay

by Frank Neuschäfer-Rube, Andrea Pathe-Neuschäfer-Rube and Gerhard P. Püschel

Toxins 2022, 14(1), 65; https://doi.org/10.3390/toxins14010065 - 17 Jan 2022

Cited by 3 | Viewed by 2220

Abstract

Botulinum neurotoxin (BoNT) is used for the treatment of a number of ailments. The activity of the toxin that is isolated from bacterial cultures is frequently tested in the mouse lethality assay. Apart from the ethical concerns inherent to this assay, species-specific differences [...] Read more.

Botulinum neurotoxin (BoNT) is used for the treatment of a number of ailments. The activity of the toxin that is isolated from bacterial cultures is frequently tested in the mouse lethality assay. Apart from the ethical concerns inherent to this assay, species-specific differences in the affinity for different BoNT serotypes give rise to activity results that differ from the activity in humans. Thus, BoNT/B is more active in mice than in humans. The current study shows that the stimulus-dependent release of a luciferase from a differentiated human neuroblastoma–based reporter cell line (SIMA-hPOMC1-26-Gluc) was inhibited by clostridial and recombinant BoNT/A to the same extent, whereas both clostridial and recombinant BoNT/B inhibited the release to a lesser extent and only at much higher concentrations, reflecting the low activity of BoNT/B in humans. By contrast, the genetically modified BoNT/B-MY, which has increased affinity for human synaptotagmin, and the BoNT/B protein receptor inhibited luciferase release effectively and with an EC50 comparable to recombinant BoNT/A. This was due to an enhanced uptake into the reporter cells of BoNT/B-MY in comparison to the recombinant wild-type toxin. Thus, the SIMA-hPOMC1-26-Gluc cell assay is a versatile tool to determine the activity of different BoNT serotypes providing human-relevant dose-response data. Full article

(This article belongs to the Section Bacterial Toxins)

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6 pages, 241 KiB

Open AccessEditor’s ChoiceArticle

Safety of High-Dose Botulinum Toxin Injections for Parotid and Submandibular Gland Radioprotection

by Joerg Mueller, Thomas Langbein, Aditi Mishra and Richard P. Baum

Toxins 2022, 14(1), 64; https://doi.org/10.3390/toxins14010064 - 17 Jan 2022

Cited by 11 | Viewed by 4116

Abstract

Botulinum Toxin injections into salivary glands (SG) up to a total dose of 100 units IncobotulinumtoxinA (IncoA) represent the treatment of choice for sialorrhea. However, BTX might also protect SG against sialotoxic radioligand cancer therapies. The radioligand Actinium-225-PSMA effectively targets Prostate Cancer (PCa) [...] Read more.

Botulinum Toxin injections into salivary glands (SG) up to a total dose of 100 units IncobotulinumtoxinA (IncoA) represent the treatment of choice for sialorrhea. However, BTX might also protect SG against sialotoxic radioligand cancer therapies. The radioligand Actinium-225-PSMA effectively targets Prostate Cancer (PCa) metastases but inevitably destroys SG due to unintended gland uptake. A preliminary case series with regular-dose IncoA failed to reduce SG PSMA-radioligand uptake. We therefore increased IncoA dosage in combination with transdermal scopolamine until a clinically relevant SG PSMA-radioligand uptake reduction was achieved. Ten consecutive men with metastasized PCa refractory to all other cancer therapies received gradually increasing IncoA dosages as part of a compassionate use PSMA-radioligand-therapy trial. The parotid gland received six and the submandibular gland three injection points under ultrasound control, up to a maximum of 30 units IncoA per injection point. A maximum total dose of 250 units IncoA was applied with up to 170 units per parotid and 80 units per submandibular gland. Treatment was well tolerated and all side-effects were non-serious. The most frequent side-effect was dry mouth of mild severity. No dysphagia, facial weakness, chewing difficulties or systemic side-effects were observed. SG injections with IncoA up to a total dose of 250 units are safe when distributed among several injection-points under ultrasound control by an experienced physician. These preliminary findings lay the basis for future trials including BTX as major component for SG protection in established as well as newly emerging radioligand cancer therapies. Full article

(This article belongs to the Special Issue Botulinum Toxins: New Uses in the Treatment of Diseases)

16 pages, 3865 KiB

Open AccessEditor’s ChoiceArticle

Taqman qPCR Quantification and Fusarium Community Analysis to Evaluate Toxigenic Fungi in Cereals

by Elina Sohlberg, Vertti Virkajärvi, Päivi Parikka, Sari Rämö, Arja Laitila and Tuija Sarlin

Toxins 2022, 14(1), 45; https://doi.org/10.3390/toxins14010045 - 6 Jan 2022

Cited by 8 | Viewed by 3204

Abstract

Fusarium head blight (FHB) is an economically important plant disease. Some Fusarium species produce mycotoxins that cause food safety concerns for both humans and animals. One especially important mycotoxin-producing fungus causing FHB is Fusarium graminearum. However, Fusarium species form a disease complex [...] Read more.

Fusarium head blight (FHB) is an economically important plant disease. Some Fusarium species produce mycotoxins that cause food safety concerns for both humans and animals. One especially important mycotoxin-producing fungus causing FHB is Fusarium graminearum. However, Fusarium species form a disease complex where different Fusarium species co-occur in the infected cereals. Effective management strategies for FHB are needed. Development of the management tools requires information about the diversity and abundance of the whole Fusarium community. Molecular quantification assays for detecting individual Fusarium species and subgroups exist, but a method for the detection and quantification of the whole Fusarium group is still lacking. In this study, a new TaqMan-based qPCR method (FusE) targeting the Fusarium-specific elongation factor region (EF1α) was developed for the detection and quantification of Fusarium spp. The FusE method was proven as a sensitive method with a detection limit of 1 pg of Fusarium DNA. Fusarium abundance results from oat samples correlated significantly with deoxynivalenol (DON) toxin content. In addition, the whole Fusarium community in Finnish oat samples was characterized with a new metabarcoding method. A shift from F. culmorum to F. graminearum in FHB-infected oats has been detected in Europe, and the results of this study confirm that. These new molecular methods can be applied in the assessment of the Fusarium community and mycotoxin risk in cereals. Knowledge gained from the Fusarium community analyses can be applied in developing and selecting effective management strategies for FHB. Full article

(This article belongs to the Special Issue Selected Papers from the 15th European Fusarium Seminar)

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