Microorganisms, Volume 11, Issue 4 (April 2023) – 277 articles

Ecosystems are often resilient enough to fully recover following a natural disturbance, or to transform into a new equilibrium favourable to the surrounding flora and fauna. However, at a local level, whether this transformation will be beneficial or not depends strongly on the level of disturbance and the available mechanisms for recovery. The Arctic, however, provides a potentially extreme environment for microbial growth and this is reflected in the microbial biodiversity, the in-situ growth rates, the biogeochemical cycling and its sensitivity to environmental change. In this study, we evaluated the current microbial biodiversity and environmental conditions around the landfill site in Adventdalen, Svalbard to identify differences across bacterial communities that might promote or accelerate naturally occurring environmental recovery. Landfill sites can induce changes in the local environment through the input of exogenous chemicals (both organic and inorganic) and microorganisms. Leachate can flow with run-off from the primary location of the landfill site due to rain, snow or ice melt and spread material into soils surrounding the site. In this study we found a strong effect of the landfill site on the bacterial diversity in the local landscape. Intervention is highly desirable to enhance the environment and improve the restoration by subtly altering the conditions at the site (such as the pH or drainage courses) and by encouraging specific groups of naturally occurring indigenous microorganisms to bioremediate the site. Full article

Strains of the genus Delftia are poorly studied microorganisms. In this work, the complete genome of the naphthalene-degrading Delftia tsuruhatensis strain ULwDis3 isolated from seawater of the Gulf of Finland of the Baltic Sea was assembled. For the first time, genes encoding naphthalene cleavage pathways via salicylate and gentisate were identified in a strain of the genus Delftia. The genes are part of one operon (nag genes). Three open reading frames (ORFs) were found in the genome of D. tsuruhatensis strain ULwDis3 that encode gentisate 1.2-dioxygenase. One of the ORFs is part of the nag operon. The physiological and biochemical characteristics of the strain ULwDis3 when cultured in mineral medium with naphthalene as the sole source of carbon and energy were also studied. It was found that after 22 h of growth, the strain stopped consuming naphthalene, and at the same time, naphthalene 1.2-dioxygenase and salicylate 5-hydroxylase activities were not detected. Later, a decrease in the number of living cells and the death of the culture were observed. Gentisate 1.2-dioxygenase activity was detected from the time of gentisate formation until culture death. Full article

Modern food technology research has researched possible approaches to reducing the concentration of biogenic amines in food and thereby enhance and guarantee food safety. Applying adjunct cultures that can metabolise biogenic amines is a potential approach to reach the latter mentioned goal. Therefore, this study aims to study the crucial factors that could determine the decrease in biogenic amines concentration (histamine, tyramine, phenylethylamine, putrescine and cadaverine) in foodstuffs using Bacillus subtilis DEPE IB1 isolated from gouda-type cheese. The combined effects of cultivation temperature (8 °C, 23 °C and 30 °C) and the initial pH of the medium (5.0, 6.0, 7.0 and 8.0) under aerobic and also anaerobic conditions resulted in the decrease of the tested biogenic amines concentration during the cultivation time (another factor tested). Bacillus subtilis was cultivated (in vitro) in a medium supplemented with biogenic amines, and their degradation was detected using the high-performance liquid chromatography equipped with UV-detector. The course of biogenic amines degradation by Bacillus subtilis DEPE IB1 was significantly influenced by cultivation temperature and also the initial pH of the medium (p < 0.05). At the end of the cultivation, the concentration of all of the monitored biogenic amines was significantly reduced by 65–85% (p < 0.05). Therefore, this strain could be used for preventive purposes and contributes to food safety enhance. Full article

To study (16S rRNA-sequencing) the impact of gestational and corrected ages on the microbiota profile of human milk (HM) of mothers that delivered full-term and pre-term children, HM samples were obtained and classified according to the gestational age as group T (full-term births ≥37 weeks), and group P (pre-term births <37 weeks). Group P was longitudinally followed, and the samples were collected at the full-term corrected gestational age: when the chronological age plus the gestational age were ≥37 weeks (PT group). The HM microbiota composition differed depending on the gestational age (T vs. P). Group T had lower levels of Staphylococcus and higher levels of Rothia and Streptococcus, as compared to group P. The alpha Simpson diversity value was higher in group T than in P, whereas no differences were found between groups T and PT, suggesting a microbial evolution of the composition of group P towards group T over chronological age. Full-term delivery was associated with a greater diversity of microbes in HM. The microbial composition of pre-term HM, at the corrected age, did not show significant differences, as compared to the samples obtained from the full-term group, suggesting that it would be appropriate to consider the corrected age in terms of the composition and the diversity of the milk in future studies. Full article

Endophytic fungi are those that present part of their life cycle in healthy tissues of different plant hosts in symbiosis without causing harm. At the same time, fungus-plant symbiosis makes it possible for microorganisms to synthesize their own bioactive secondary metabolites while in the stationary stage. To accomplish this, the endophytic fungus Trichoderma asperellum was isolated from Bertholletia excelsa (Brazil nut) almonds. The fungus was cultivated and extracted with ethyl acetate, obtaining AM07Ac. Then, using HPTLC (High-performance thin-layer chromatography) and nuclear magnetic resonance (¹H NMR), β-amyrin, kaempferol, and brucine were identified as major compounds. Further in vivo assays in zebrafish demonstrated the activity of AM07Ac on melanogenesis by producing a concentration–response inhibitory effect, which, through an in silico study, proved to be related to the noted major compounds known to inhibit tyrosinase activity. The inhibition of tyrosinase prevents melanin accumulation in skin. Therefore, these results imply the importance of investigating microorganisms and their pharmacological activities, in particular the endophytic fungus Trichoderma asperellum as a generator of active metabolites for melanogenesis modulation. Full article

Rhizosheric bacteria with several abilities related to plant growth and health have been denominated Plant Growth-Promoting Rhizobacteria (PGPR). PGPR promote plant growth through several modes of action, be it directly or indirectly. The benefits provided by these bacteria can include increased nutrient availability, phytohormone production, shoot and root development, protection against several phytopathogens, and reduced diseases. Additionally, PGPR can help plants to withstand abiotic stresses such as salinity and drought and produce enzymes that detoxify plants from heavy metals. PGPR have become an important strategy in sustainable agriculture due to the possibility of reducing synthetic fertilizers and pesticides, promoting plant growth and health, and enhancing soil quality. There are many studies related to PGPR in the literature. However, this review highlights the studies that used PGPR for sustainable production in a practical way, making it possible to reduce the use of fertilizers such as phosphorus and nitrogen and fungicides, and to improve nutrient uptake. This review addresses topics such as unconventional fertilizers, seed microbiome for rhizospheric colonization, rhizospheric microorganisms, nitrogen fixation for reducing chemical fertilizers, phosphorus solubilizing and mineralizing, and siderophore and phytohormone production for reducing the use of fungicides and pesticides for sustainable agriculture. Full article

Lactic acid bacteria (LAB) share and provide several beneficial effects on human health, such as the release of bioactive metabolites, pathogen competition, and immune stimulation. The two major reservoirs of probiotic microorganisms are the human gastro-intestinal tract and fermented dairy products. However, other sources, such as plant-based foods, represent important alternatives thanks to their large distribution and nutritive value. Here, the probiotic potential of autochthonous Lactiplantibacillus plantarum PFA2018AU, isolated from carrots harvested in Fucino highland, Abruzzo (Italy), was investigated through in vitro and in vivo approaches. The strain was sent to the biobank of Istituto Zooprofilattico Sperimentale della Lombardia ed Emilia Romagna in Italy for the purpose of patent procedures under the Budapest Treaty. The isolate showed high survival capability under in vitro simulated gastro-intestinal conditions, antibiotic susceptibility, hydrophobicity, aggregation, and the ability to inhibit the in vitro growth of Salmonella enterica serovar Typhimurium, Listeria monocytogenes, Pseudomonas aeruginosa, and Staphylococcus aureus pathogens. Caenorhabditis elegans was used as the in vivo model in order to analyse prolongevity and anti-ageing effects. L. plantarum PFA2018AU significantly colonised the gut of the worms, extended their lifespan, and stimulated their innate immunity. Overall, these results showed that autochthonous LAB from vegetables, such as carrots, have functional features that can be considered novel probiotic candidates. Full article

A wide array of bacteria and fungi are known for their association with pests that impact the health of the olive tree. The latter presents the most economically important cultivation in Tunisia. The microbial diversity associated with olive orchards in Tunisia remains unknown and undetermined. This study investigated microbial diversity to elucidate the microbial interactions that lead to olive disease, and the bio-prospects for potential microbial biocontrol agents associated with insect pests of economic relevance for olive cultivation in the Mediterranean area. Bacterial and fungal isolation was made from soil and olive tree pests. A total of 215 bacterial and fungal strains were randomly isolated from eight different biotopes situated in Sfax (Tunisia), with different management practices. 16S rRNA and ITS gene sequencing were used to identify the microbial community. The majority of the isolated bacteria, Staphylococcus, Bacillus, Alcaligenes, and Providencia, are typical of the olive ecosystem and the most common fungi are Penicillium, Aspergillus, and Cladosporium. The different olive orchards depicted distinct communities, and exhibited dissimilar amounts of bacteria and fungi with distinct ecological functions that could be considered as promising resources in biological control. Full article

Multifarious plant growth-promoting Bacillus strains recovered from rhizospheric soils of the Indo Gangetic plains (IGPs) were identified as Bacillus licheniformis MNNITSR2 and Bacillus velezensis MNNITSR18 based on their biochemical characteristics and 16S rDNA gene analysis. Both strains exhibited the ability to produce IAA, siderophores, ammonia, lytic enzymes, HCN production, and phosphate solubilization capability and strongly inhibited the growth of phytopathogens such as Rhizoctonia solani and Fusariun oxysporum in vitro. In addition, these strains are also able to grow at a high temperature of 50 °C and tolerate up to 10–15% NaCl and 25% PEG 6000. The results of the pot experiment showed that individual seed inoculation and the coinoculation of multifarious plant growth promoting (PGP) Bacillus strains (SR2 and SR18) in rice fields significantly enhanced plant height, root length volume, tiller numbers, dry weight, and yield compared to the untreated control. This indicates that these strains are potential candidates for use as PGP inoculants/biofertilizers to increase rice productivity under field conditions for IGPs in Uttar Pradesh, India. Full article

As excellent biocontrol agents and plant growth promoters, Trichoderma species are agriculturally important. Trichoderma spp. cultures can be produced using solid-state or submerged cultivation, the latter being much less labor intensive and easier to control and automate. The aim of the study was to investigate the ability to increase the shelf-life of T. asperellum cells by optimizing cultivation media and upscaling the submerged cultivation process. Four different cultivation media were used with or without the addition of Tween 80 and stored with or without incorporation into peat, and viability, expressed as CFU/g, was assessed during one year of storage in an industrial warehouse. The addition of Tween 80 had a positive effect on the biomass yield. The culture medium played a major role in the ability of the mycelium to produce spores, which in turn influenced the amount of CFU. This effect was less pronounced when the biomass was mixed with peat prior to storage. A procedure that increases the number of CFU in a peat-based product formulation is recommended, namely, incubation of the mixture at 30 °C for 10 days prior to storage at 15 °C over an extended period of time. Full article

Neurodegenerative disorders (ND) are a group of conditions that affect the neurons in the brain and spinal cord, leading to their degeneration and eventually causing the loss of function in the affected areas. These disorders can be caused by a range of factors, including genetics, environmental factors, and lifestyle choices. Major pathological signs of these diseases are protein misfolding, proteosomal dysfunction, aggregation, inadequate degradation, oxidative stress, free radical formation, mitochondrial dysfunctions, impaired bioenergetics, DNA damage, fragmentation of Golgi apparatus neurons, disruption of axonal transport, dysfunction of neurotrophins (NTFs), neuroinflammatory or neuroimmune processes, and neurohumoral symptoms. According to recent studies, defects or imbalances in gut microbiota can directly lead to neurological disorders through the gut-brain axis. Probiotics in ND are recommended to prevent cognitive dysfunction, which is a major symptom of these diseases. Many in vivo and clinical trials have revealed that probiotics (Lactobacillus acidophilus, Bifidobacterium bifidum, and Lactobacillus casei, etc.) are effective candidates against the progression of ND. It has been proven that the inflammatory process and oxidative stress can be modulated by modifying the gut microbiota with the help of probiotics. As a result, this study provides an overview of the available data, bacterial variety, gut-brain axis defects, and probiotics’ mode of action in averting ND. A literature search on particular sites, including PubMed, Nature, and Springer Link, has identified articles that might be pertinent to this subject. The search contains the following few groups of terms: (1) Neurodegenerative disorders and Probiotics OR (2) Probiotics and Neurodegenerative disorders. The outcomes of this study aid in elucidating the relationship between the effects of probiotics on different neurodegenerative disorders. This systematic review will assist in discovering new treatments in the future, as probiotics are generally safe and cause mild side effects in some cases in the human body. Full article

Fusarium wilt of lettuce is found throughout the world, causing significant yield losses. Lettuce is the most-cultivated leafy vegetable in Greece, affected by a large number of foliar and soil-borne pathogens. In this study, 84 isolates of Fusarium oxysporum, obtained from soil-grown lettuce plants exhibiting wilt symptoms, were characterized as belonging to race 1 of F. oxysporum f. sp. lactucae based on sequence analysis of the translation elongation factor 1-alpha (TEF1-α) gene and the rDNA intergenic spacer (rDNA-IGS) region. The isolates were also assigned to one single race through PCR assays with specific primers targeting race 1 and race 4 of the pathogen. In addition, four representative isolates were confirmed to be associated with race 1 based on the pathogenicity tests with a set of differential lettuce cultivars. Artificial inoculations on the most commonly cultivated lettuce cultivars in Greece revealed that the tested cultivars varied regarding their susceptibility to F. oxysporum f. sp. lactucae race 1. Cultivars (cvs.) “Cencibel” and “Lugano” were found to be highly susceptible, while cvs. “Sandalina” and “Starfighter” were the most resistant ones. Expression analysis of 10 defense-related genes (PRB1, HPL1, LTC1, SOD, ERF1, PAL1, LOX, MPK, BG, and GST) was carried out on artificially inoculated lettuce plants of the four above cultivars at different time points after inoculation. In resistant cultivars, a higher induction rate was observed for all the tested genes in comparison with the susceptible ones. Moreover, in resistant cultivars, all genes except LTC1, MPK, and GST showed their highest induction levels in their earliest stages of infection. The results of this study are expected to contribute to the implementation of an integrated management program to control Fusarium wilt of lettuce, based mainly on the use of resistant cultivars. Full article

Lyme borreliosis (LB) is not notifiable in many European countries, and accurate data on the incidence are often lacking. This study aimed to determine the seroprevalence of Borrelia burgdorferi sensu lato (s.l.)-specific antibodies in the general population of The Netherlands, and to determine risk factors associated with seropositivity. Sera and questionnaires were obtained from participants (n = 5592, aged 0–88 years) enrolled in a nationwide serosurveillance study. The sera were tested for B. burgdorferi s.l.-specific IgM and IgG antibodies using ELISA and immunoblot. Seroprevalence was estimated controlling for the survey design. Risk factors for seropositivity were analyzed using a generalized linear mixed-effect model. In 2016/2017, the seroprevalence in The Netherlands was 4.4% (95% CI 3.5–5.2). Estimates were higher in men (5.7% [95% CI 4.4–7.2]) than in women (3.1% [95% CI 2.0–4.0]), and increased with age from 2.6% (95% CI 1.4–4.4) in children to 7.7% (95% CI 5.9–7.9) in 60- to 88-year-olds. The seroprevalence for B. burgdorferi s.l. in the general population in The Netherlands was comparable to rates reported in European countries. The main risk factors for seropositivity were increasing age, being male and the tick bite frequency. The dynamics of LB infection are complex and involve variables from various disciplines. This could be further elucidated using infectious disease modelling. Full article

Microbial resources from the human gut may find use in various applications, such as empirical research on the microbiome, the development of probiotic products, and bacteriotherapy. Due to the development of “culturomics”, the number of pure bacterial cultures obtained from the human gut has significantly increased since 2012. However, there is still a considerable number of human gut microbes to be isolated and cultured. Thus, to improve the efficiency of obtaining microbial resources from the human gut, some constraints of the current methods, such as labor burden, culture condition, and microbial targetability, still need to be optimized. Here, we overview the general knowledge and recent development of culturomics for human gut microorganisms. Furthermore, we discuss the optimization of several parts of culturomics including sample collection, sample processing, isolation, and cultivation, which may improve the current strategies. Full article

The use of venoarterial (VA) extracorporeal membrane oxygenation therapy (ECMO) in patients admitted to cardiac intensive care units (CICU) has increased. Data regarding infections in this population are scarce. In this retrospective study, we analyzed the risk factors, outcome, and predictors of in-hospital mortality due to nosocomial infections in patients with ECMO admitted to a single coronary intensive care unit between July 2013 and March 2019 treated with VA-ECMO for >48 h. From 69 patients treated with VA-ECMO >48 h, (median age 58 years), 29 (42.0%) patients developed 34 episodes of infections with an infection rate of 0.92/1000 ECMO days. The most frequent were ventilator-associated pneumonia (57.6%), tracheobronchitis (9.1%), bloodstream infections (9.1%), skin and soft tissue infections (9.1%), and cytomegalovirus reactivation (9.1%). In-hospital mortality was 47.8%, but no association with nosocomial infections was found (p = 0.75). The number of days on ECMO (OR 1.14, 95% CI 1.01–1.30, p = 0.029) and noninfectious complications were higher in the infected patients (OR: 3.8 95% CI = 1.05–14.1). A higher baseline creatinine value (OR: 8.2 95% CI = 1.12–60.2) and higher blood lactate level at 4 h after ECMO initiation (OR: 2.0 95% CI = 1.23–3.29) were significant and independent risk factors for mortality. Conclusions: Nosocomial infections in medical patients treated with VA-ECMO are very frequent, mostly Gram-negative respiratory infections. Preventive measures could play an important role for these patients. Full article

Importance: Although highly effective in treating recurrent Clostridioides difficile infection (RCDI), the mechanisms of action of fecal microbial transplantation (FMT) are not fully understood. Aim: The aim of this study was to explore microbially derived products or pathways that could contribute to the therapeutic efficacy of FMT. Methods: Stool shotgun metagenomic sequencing data from 18 FMT-treated RCDI patients at 4 points in time were used for the taxonomic and functional profiling of their gut microbiome. The abundance of the KEGG orthology (KO) groups was subjected to univariate linear mixed models to assess the significance of the observed differences between 0 (pre-FMT), 1, 4, and 12 weeks after FMT. Results: Of the 59,987 KO groups identified by shotgun metagenomic sequencing, 27 demonstrated a statistically significant change after FMT. These KO groups are involved in many cellular processes, including iron homeostasis, glycerol metabolism, and arginine regulation, all of which have been implicated to play important roles in bacterial growth and virulence in addition to modulating the intestinal microbial composition. Conclusion: Our findings suggest potential changes in key KO groups post-FMT, which may contribute to FMT efficacy beyond the restored microbial composition/diversity and metabolism of bile acids and short-chain fatty acids. Future larger studies that include a fecal metabolomics analysis combined with animal model validation work are required to further elucidate the molecular mechanisms. Full article

Bacteria use an array of sigma factors to regulate gene expression during different stages of their life cycles. Full-length, atomic-level structures of sigma factors have been challenging to obtain experimentally as a result of their many regions of intrinsic disorder. AlphaFold has now supplied plausible full-length models for most sigma factors. Here we discuss the current understanding of the structures and functions of sigma factors in the model organism, Bacillus subtilis, and present an X-ray crystal structure of a region of B. subtilis SigE, a sigma factor that plays a critical role in the developmental process of spore formation. Full article

Fungemia is a fatal systemic infection that can occur in immunocompromised patients. Despite that, antifungal stewardship is spreading widely, but the mortality rate is extremely high, showing 40–60%. Loderomyces elongiporus is a newly morphologically detected pathogen, first described in 1994, followed by isolation in humans in 2008. It has been misrecognized as Candida parapsilosis. Recently, fever attributable to L. elongisporus fungemia cases has been reported, and the etiology and clinical features are still unknown. Here, we present three successfully treated L. elongisporus fungemia cases by echinocandin. In total, 11 cases were reviewed, including ours. Six of the eleven cases (55%) had external devices. All cases had some immunocompromised conditions or underlying diseases, such as diabetes mellitus, lung cancer, etc. Six patients survived, and the remaining five died. Seven patients who had received echinocandin initially survived. Risk factors for L. elongiporus fungemia overlap with those of candidemia. Even though there is no breakpoint for L. elongiporus, echinocandin can be a helpful treatment regimen for L. elongiporus fungemia. Full article

Global warming and acidification of the global ocean are two important manifestations of the ongoing climate change. To characterize their joint impact on Vibrio adaptation and fitness, we analyzed the temperature-dependent adaptation of Vibrio harveyi at different pHs (7.0, 7.5, 8.0, 8.3 and 8.5) that mimic the pH of the world ocean in the past, present and future. Comparison of V. harveyi growth at 20, 25 and 30 °C show that higher temperature per se facilitates the logarithmic growth of V. harveyi in nutrient-rich environments in a pH-dependent manner. Further survival tests carried out in artificial seawater for 35 days revealed that cell culturability declined significantly upon incubation at 25 °C and 30 °C but not at 20 °C. Moreover, although acidification displayed a negative impact on cell culturability at 25 °C, it appeared to play a minor role at 30 °C, suggesting that elevated temperature, rather than pH, was the key player in the observed reduction of cell culturability. In addition, analyses of the stressed cell morphology and size distribution by epifluorescent microscopy indicates that V. harveyi likely exploits different adaptation strategies (e.g., acquisition of coccoid-like morphology) whose roles might differ depending on the temperature–pH combination. Full article

High counts of bacteria are present in beach sand, and human health threats attributable to contact with sand have been reported. In this study, we investigated fecal indicator bacteria in the top surface sand of coastal beaches. Monitoring investigations were performed during a monsoon when rainfall occurs randomly, and the composition of the coliforms was analyzed. The coliform count in the top surface sand (depth < 1 cm) increased by approximately 100 fold (26–2.23 × 10³ CFU/100 g) with increasing water content because of precipitation. The composition of the coliforms in the top surface sand changed within 24 h of rainfall, with Enterobacter comprising more than 40% of the coliforms. Estimation of factors that changed the bacterial counts and composition revealed that coliform counts tended to increase with increasing water content in the top surface sand. However, the abundance of Enterobacter was independent of the sand surface temperature and water content. Coliform counts in the top surface sand rapidly increased and the composition showed remarkable variations because of the supply of water to the beach following rainfall. Among them, some bacteria with suspected pathogenicity were present. Controlling bacteria in coastal beaches is important for improving public health for beachgoers. Full article

The present study describes an epidemiological investigation into a carbapenem-resistant Acinetobacter baumannii (CRAB) outbreak, which had occurred in a neonatal intensive care unit (NICU), and the subsequent strengthening of infection control interventions. Upon the onset of the outbreak, existing infection control interventions were reviewed, and a set of containment measures were instituted. All CRAB isolates were characterized in terms of antimicrobial susceptibility testing and their genetic relatedness. The investigation process identified gaps within the NICU’s existing infection control measures, which had likely resulted in the outbreak. CRAB was isolated from nine preterm infants: five colonized and four infected. All five colonized patients were discharged well. However, three out of four of the infected infants died. Outbreak investigation and genomic subtyping of environmental swabs revealed that mini syringe drivers shared between patients and a sink in the milk preparation room had served as CRAB reservoirs with possible transmission via the hands of healthcare workers. Implementation of immediate actions such as reinforcement of hand hygiene practices, intensified environmental cleaning, geographical cohorting, reviewing of milk handling practices and sink management protocol had resulted in no further CRAB isolation. The CRAB outbreak in the NICU underlines the importance of consistent compliance with infection-control interventions. Integration of epidemiological and microbiological data, together with comprehensive preventive measures, successfully brought the outbreak to a halt. Full article

Water monitor lizards (WMLs) reside in unhygienic and challenging ecological surroundings and are routinely exposed to various pathogenic microorganisms. It is possible that their gut microbiota produces substances to counter microbial infections. Here we determine whether selected gut bacteria of water monitor lizards (WMLs) possess anti-amoebic properties using Acanthamoeba castellanii of the T4 genotype. Conditioned media (CM) were prepared from bacteria isolated from WML. The CM were tested using amoebicidal, adhesion, encystation, excystation, cell cytotoxicity and amoeba-mediated host cell cytotoxicity assays in vitro. Amoebicidal assays revealed that CM exhibited anti-amoebic effects. CM inhibited both excystation and encystation in A. castellanii. CM inhibited amoebae binding to and cytotoxicity of host cells. In contrast, CM alone showed limited toxic effects against human cells in vitro. Mass spectrometry revealed several antimicrobials, anticancer, neurotransmitters, anti-depressant and other metabolites with biological functions. Overall, these findings imply that bacteria from unusual places, such as WML gut, produce molecules with anti-acanthamoebic capabilities. Full article

Identifying fungal clones propagated during outbreaks in hospital settings is a problem that increasingly confronts biologists. Current tools based on DNA sequencing or microsatellite analysis require specific manipulations that are difficult to implement in the context of routine diagnosis. Using deep learning to classify the mass spectra obtained during the routine identification of fungi by MALDI-TOF mass spectrometry could be of interest to differentiate isolates belonging to epidemic clones from others. As part of the management of a nosocomial outbreak due to Candida parapsilosis in two Parisian hospitals, we studied the impact of the preparation of the spectra on the performance of a deep neural network. Our purpose was to differentiate 39 otherwise fluconazole-resistant isolates belonging to a clonal subset from 56 other isolates, most of which were fluconazole-susceptible, collected during the same period and not belonging to the clonal subset. Our study carried out on spectra obtained on four different machines from isolates cultured for 24 or 48 h on three different culture media showed that each of these parameters had a significant impact on the performance of the classifier. In particular, using different culture times between learning and testing steps could lead to a collapse in the accuracy of the predictions. On the other hand, including spectra obtained after 24 and 48 h of growth during the learning step restored the good results. Finally, we showed that the deleterious effect of the device variability used for learning and testing could be largely improved by including a spectra alignment step during preprocessing before submitting them to the neural network. Taken together, these experiments show the great potential of deep learning models to identify spectra of specific clones, providing that crucial parameters are controlled during both culture and preparation steps before submitting spectra to a classifier. Full article

Bacillus subtilis is one of the commonly used industrial strains for riboflavin production. High-throughput screening is useful in biotechnology, but there are still an insufficient number of articles focusing on improving the riboflavin production of B. subtilis by this powerful tool. With droplet-based microfluidics technology, single cells can be encapsulated in droplets. The screening can be carried out by detecting the fluorescence intensity of secreted riboflavin. Thus, an efficient and high-throughput screening method suitable for riboflavin production strain improvement could be established. In this study, droplet-based microfluidics screening was applied, and a more competitive riboflavin producer U3 was selected from the random mutation library of strain S1. The riboflavin production and biomass of U3 were higher than that of S1 in flask fermentation. In addition, the results of fed-batch fermentation showed that the riboflavin production of U3 was 24.3 g/L, an 18% increase compared with the parent strain S1 (20.6 g/L), and the yield (g riboflavin/100 g glucose) increased by 19%, from 7.3 (S1) to 8.7 (U3). Two mutations of U3 (sinR^G89R and icd^D28E) were identified through whole genome sequencing and comparison. Then they were introduced into BS168DR (parent of S1) for further analysis, which also caused riboflavin production to increase. This paper provides protocols for screening riboflavin-producing B. subtilis with droplet-based microfluidics technology and reveals mutations in riboflavin overproduction strains. Full article

Green nanotechnology has made the synthesis of nanoparticles a possible approach. Nanotechnology has a significant impact on several scientific domains and has diverse applications in different commercial areas. The current study aimed to develop a novel and green approach for the biosynthesis of silver oxide nanoparticles (Ag₂ONPs) utilizing Parieteria alsinaefolia leaves extract as a reducing, stabilizing and capping agent. The change in color of the reaction mixture from light brown to reddish black determines the synthesis of Ag₂ONPs. Further, different techniques were used to confirm the synthesis of Ag₂ONPs, including UV-Visible spectroscopy, Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), scanning electron microscope (SEM), Energy-dispersive X-ray spectroscopy (EDX), zeta potential and dynamic light scattering (DLS) analyses. The Scherrer equation determined a mean crystallite size of ~22.23 nm for Ag₂ONPs. Additionally, different in vitro biological activities have been investigated and determined significant therapeutic potentials. Radical scavenging DPPH assay (79.4%), reducing power assay (62.68 ± 1.77%) and total antioxidant capacity (87.5 ± 4.8%) were evaluated to assess the antioxidative potential of Ag₂ONPs. The disc diffusion method was adopted to evaluate the antibacterial and antifungal potentials of Ag₂ONPs using different concentrations (125–1000 μg/mL). Moreover, the brine shrimp cytotoxicity assay was investigated and the LC₅₀ value was calculated as 2.21 μg/mL. The biocompatibility assay using red blood cells (<200 μg/mL) confirmed the biosafe and biocompatible nature of Ag₂ONPs. Alpha-amylase inhibition assay was performed and reported 66% inhibition. In conclusion, currently synthesized Ag₂ONPs have exhibited strong biological potential and proved as an attractive eco-friendly candidate. In the future, this preliminary research work will be a helpful source and will open new avenues in diverse fields, including the pharmaceutical, biomedical and pharmacological sectors. Full article

Recent bacteriological investigations of freshwater mussel mortality events in the southeastern United States have identified a variety of bacteria and differences in bacterial communities between sick and healthy mussels. In particular, Yokenella regensburgei and Aeromonas spp. have been shown to be associated with moribund mussels, although it remains unclear whether these bacteria are causes or consequences of disease. To further understand the role of bacteria in mussel epizootics, we investigated mortality events that occurred in the upper Midwest in the Embarrass River (Wisconsin) and the Huron River (Michigan). For comparison, we also studied mussels from an unaffected population in the St. Croix River (Wisconsin). Diverse bacterial genera were identified from these sites, including Y. regensburgei from moribund mussels in the Embarrass River (Wisconsin). This bacterium has also been consistently isolated during ongoing mortality events in the Clinch River (Virginia). Subsequently, we developed and validated molecular assays for the detection of Yokenella to use in future investigations of mussel mortality events and to identify environmental reservoirs of this bacterium. Full article

The fall armyworm, Spodoptera frugiperda (Noctuidae; Lepidoptera), is a serious threat to food security as it has the potential to feed on over 353 plant species. To control this insect pest, endophytic colonization of entomopathogenic fungi (EPF) in plants is being considered as a safer and more effective alternative. This study evaluated the efficacy of two EPFs, Beauveria bassiana and Metarhizium anisopliae, for endophytic colonization using foliar spray and seed treatment methods on maize plants, and their impact on the survival, development, and fecundity of S. frugiperda. Both EPF effectively colonized the maize plants with foliar spray and seed treatment methods, resulting in 72–80% and 50–60% colonization rates, respectively, 14 days after inoculation. The EPF negatively impacted the development and fecundity of S. frugiperda. Larvae feeding on EPF-inoculated leaves had slower development (21.21 d for M. anisopliae and 20.64 d for B. bassiana) than the control treatment (20.27 d). The fecundity rate was also significantly reduced to 260.0–290.1 eggs/female with both EPF applications compared with the control treatment (435.6 eggs/female). Age-stage-specific parameters showed lower fecundity, life expectancy, and survival of S. frugiperda when they fed on both EPF-inoculated leaves compared with untreated leaves. Furthermore, both EPFs had a significant effect on population parameters such as intrinsic (r = 0.127 d⁻¹ for B. bassiana, and r = 0.125 d⁻¹ for M. anisopliae) and finite rate (λ = 1.135 d⁻¹ for B. bassiana, and λ = 1.1333 d⁻¹ for M. anisopliae) of S. frugiperda compared with the control (r = 0.133 d⁻¹ and λ = 1.146 d⁻¹). These findings suggest that EPF can be effectively used for the endophytic colonization of maize plants to control S. frugiperda. Therefore, these EPFs should be integrated into pest management programs for this pest. Full article

Accurate and appropriate extrapulmonary tuberculosis (EPTB) diagnosis remains challenging due to its paucibacillary nature, requirement of invasive collection procedures, and lack of sensitive tests. This study investigated the diagnostic performance of different methods for the diagnosis of EPTB. A total of 1340 EPTB specimens were collected from presumptive EPTB patients from four different hospitals between November 2015 and March 2017. The collected specimens were tested with AFB microscopy, culture, Xpert MTB/RIF assay (Xpert), and MTBDRplus assay. Among the 1340 EPTB specimens, 49 (3.66%), 141 (10.52%), 166 (12.39%), and 154 (11.49%) were positive in AFB microscopy, culture, Xpert MTB/RIF, and MTBDRplus assay, respectively. A total of 194 (14.9%) cases were found positive in at least one of these methods. Using culture as a reference standard, the sensitivity and specificity of AFB microscopy, Xpert MTB/RIF, and MTBDRplus assay were: 27.0%/99.1%, 83.7%/96.0%, and 79.4%/96.5%, respectively. Compared to the composite reference standard, the sensitivity of culture, AFB microscopy, Xpert MTB/RIF, and MTBDRplus assay was 72.7%, 25.3%, 85.6%, and 79.4%, respectively, with a specificity of 100% for all the methods. The Xpert MTB/RIF assay showed the highest sensitivity compared to other methods. Considering the short turnaround time and promising findings, Xpert MTB/RIF assay should be integrated into national TB guidelines as a routine diagnostic test. Full article

Milk, due to its diversity in terms of its nutritional content, is an important element of the human diet, as well as a good medium for the development of bacteria. The genus Bacillus contains ubiquitous aerobic, rod-shaped, endospore-producing gram-positive bacteria. Representatives of the Bacillus cereus group and the Bacillus subtilis group contribute to shortening the shelf life of milk and dairy products by degrading milk components and its additives. They also produce a number of heat-stable toxins and can cause a number of ailments, mainly in the digestive system. The aim of this research was to identify Bacillus sp. strains isolated from raw milk and to determine their antibiotic resistance. Strains isolated from raw milk samples (n = 45) were identified by MALDI-TOF MS. Ninety strains of Bacillus sp. were identified, for which the antibiotic resistance phenotype was determined. A total of 90 strains of Bacillus were classified in five groups (the Bacillus cereus group (n = 35), B. licheniformis (n = 7), the B. subtilis group (n = 29), B. pumilus (n = 16), and Bacillus sp. (n = 3). All isolates were susceptible to chloramphenicol and meropenem. The antibiotic resistance profiles of the tested groups of Bacillus spp. differed from each other, which is of particular concern in relation to multidrug-resistant representatives of the B. cereus group resistant to cefotaxime (94.29%), ampicillin (88.57%), rifampicin (80%), and norfloxacin (65.71%). Our study provides data on the prevalence and antibiotic sensitivity of Bacillus sp. In raw milk, suggesting a potential risk to health and the dairy industry. Full article

The ability of a Penicillium bilaiae strain to support acid production and simultaneously solubilize inorganic sources of phosphate in conditions of submerged, solid-state fermentation (SSF) and immobilized cell system was examined in this study. Abiotic stress factors such as NaCl and different values of pH were introduced into the different fermentation process schemes to measure the fungal response. The results showed a higher tolerance of P. bilaiae when the fermentation process was carried out in solid-state and immobilized-cell conditions, which mimics the natural state of the soil microorganisms. The acidic culture conditions were not found to be suitable for fungal growth, which increased at a higher pH, with values of 4.0 and 6.0 being optimal for all types of fermentation. The presence of increasing amounts of NaCl provoked low biomass growth, titratable acidity, and simultaneous phosphate (P) solubilization. These results were, however, less pronounced at pH 4.0 and 6.0, particularly in conditions of SSF. Studying stress-tolerant microbial characteristics, particularly in different conditions and combinations of stress factors, is of great importance for further managing the overall microbial inoculants’ production and formulation process as well as their applications in specific soil–plant systems. Full article