Toxins

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Open AccessArticle

Organization and ELISA-Based Results of the First Proficiency Testing to Evaluate the Ability of European Union Laboratories to Detect Staphylococcal Enterotoxin Type B (SEB) in Buffer and Milk

by Yacine Nia, Mélanie Rodriguez, Reinhard Zeleny, Sabine Herbin, Frédéric Auvray, Uwe Fiebig, Marc-André Avondet, Amalia Munoz and Jacques-Antoine Hennekinne

Toxins 2016, 8(9), 268; https://doi.org/10.3390/toxins8090268 - 13 Sep 2016

Cited by 25 | Viewed by 6624

Abstract

The aim of this work was to organize the first proficiency test (PT) dedicated to staphylococcal enterotoxin B (SEB) detection in milk and buffer solutions. This paper describes the organization of the PT trial according to EN ISO 17043 requirements. Characterization of the [...] Read more.

The aim of this work was to organize the first proficiency test (PT) dedicated to staphylococcal enterotoxin B (SEB) detection in milk and buffer solutions. This paper describes the organization of the PT trial according to EN ISO 17043 requirements. Characterization of the SEB stock solution was performed using SDS-PAGE and SE-specific ELISA, and amino acid analysis was used to assign its protein concentration. The solution was then used to prepare six PT materials (four milk and two buffer batches) at a ng/g toxin level, which included one blank and one SEA-containing milk as specificity control. Suitable material homogeneity and stability were assessed using screening and quantitative ELISAs. Among the methods used by the participants, ELISA-based methods demonstrated their efficiency for the detection of SEB in both simple and complex matrices. The results serve as a basis for further improving the detection capabilities in expert laboratories and can therefore be considered as a contribution to biopreparedness. Full article

(This article belongs to the Special Issue Detection and identification of biological toxins in international proficiency tests)

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Open AccessArticle

Bioactivation and Regioselectivity of Pig Cytochrome P450 3A29 towards Aflatoxin B₁

by Jun Wu, Ruohong Chen, Caihui Zhang, Kangbai Li, Weiying Xu, Lijuan Wang, Qingmei Chen, Peiqiang Mu, Jun Jiang, Jikai Wen and Yiqun Deng

Toxins 2016, 8(9), 267; https://doi.org/10.3390/toxins8090267 - 12 Sep 2016

Cited by 23 | Viewed by 5731

Abstract

Due to unavoidable contaminations in feedstuff, pigs are easily exposed to aflatoxin B₁ (AFB₁) and suffer from poisoning, thus the poisoned products potentially affect human health. Heretofore, the metabolic process of AFB₁ in pigs remains to be clarified, especially [...] Read more.

Due to unavoidable contaminations in feedstuff, pigs are easily exposed to aflatoxin B₁ (AFB₁) and suffer from poisoning, thus the poisoned products potentially affect human health. Heretofore, the metabolic process of AFB₁ in pigs remains to be clarified, especially the principal cytochrome P450 oxidases responsible for its activation. In this study, we cloned CYP3A29 from pig liver and expressed it in Escherichia coli, and its activity has been confirmed with the typical P450 CO-reduced spectral characteristic and nifedipine-oxidizing activity. The reconstituted membrane incubation proved that the recombinant CYP3A29 was able to oxidize AFB₁ to form AFB₁-exo-8,9-epoxide in vitro. The structural basis for the regioselective epoxidation of AFB₁ by CYP3A29 was further addressed. The T309A mutation significantly decreased the production of AFBO, whereas F304A exhibited an enhanced activation towards AFB₁. In agreement with the mutagenesis study, the molecular docking simulation suggested that Thr309 played a significant role in stabilization of AFB₁ binding in the active center through a hydrogen bond. In addition, the bulk phenyl group of Phe304 potentially imposed steric hindrance on the binding of AFB₁. Our study demonstrates the bioactivation of pig CYP3A29 towards AFB₁ in vitro, and provides the insight for understanding regioselectivity of CYP3A29 to AFB₁. Full article

(This article belongs to the Collection Aflatoxins)

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Open AccessArticle

Neutralization of Botulinum Neurotoxin Type E by a Humanized Antibody

by Yağmur Derman, Katja Selby, Sebastian Miethe, André Frenzel, Yvonne Liu, Christine Rasetti-Escargueil, Arnaud Avril, Thibaut Pelat, Remi Urbain, Alexandre Fontayne, Philippe Thullier, Dorothea Sesardic, Miia Lindström, Michael Hust and Hannu Korkeala

Toxins 2016, 8(9), 257; https://doi.org/10.3390/toxins8090257 - 12 Sep 2016

Cited by 13 | Viewed by 6291

Abstract

Botulinum neurotoxins (BoNTs) cause botulism and are the deadliest naturally-occurring substances known to humans. BoNTs have been classified as one of the category A agents by the Centers for Disease Control and Prevention, indicating their potential use as bioweapons. To counter bio-threat and [...] Read more.

Botulinum neurotoxins (BoNTs) cause botulism and are the deadliest naturally-occurring substances known to humans. BoNTs have been classified as one of the category A agents by the Centers for Disease Control and Prevention, indicating their potential use as bioweapons. To counter bio-threat and naturally-occurring botulism cases, well-tolerated antibodies by humans that neutralize BoNTs are relevant. In our previous work, we showed the neutralizing potential of macaque (Macaca fascicularis)-derived scFv-Fc (scFv-Fc ELC18) by in vitro endopeptidase immunoassay and ex vivo mouse phrenic nerve-hemidiaphragm assay by targeting the light chain of the botulinum neurotoxin type E (BoNT/E). In the present study, we germline-humanized scFv-Fc ELC18 into a full IgG hu8ELC18 to increase its immunotolerance by humans. We demonstrated the protection and prophylaxis capacity of hu8ELC18 against BoNT/E in a mouse model. A concentration of 2.5 ng/mouse of hu8ELC18 protected against 5 mouse lethal dose (MLD) in a mouse protection assay and complete neutralization of 1 LD50 of pure BoNT/E toxin was achieved with 8 ng of hu8ELC18 in mouse paralysis assay. Furthermore, hu8ELC18 protected mice from 5 MLD if injected up to 14 days prior to intraperitoneal BoNT/E administration. This newly-developed humanized IgG is expected to have high tolerance in humans. Full article

(This article belongs to the Special Issue Botulinum Neurotoxins Antibody and Vaccine)

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Open AccessArticle

Tetanus Neurotoxin Neutralizing Antibodies Screened from a Human Immune scFv Antibody Phage Display Library

by Han Wang, Rui Yu, Ting Fang, Ting Yu, Xiangyang Chi, Xiaopeng Zhang, Shuling Liu, Ling Fu, Changming Yu and Wei Chen

Toxins 2016, 8(9), 266; https://doi.org/10.3390/toxins8090266 - 11 Sep 2016

Cited by 25 | Viewed by 6666

Abstract

Tetanus neurotoxin (TeNT) produced by Clostridium tetani is one of the most poisonous protein substances. Neutralizing antibodies against TeNT can effectively prevent and cure toxicosis. Using purified Hc fragments of TeNT (TeNT-Hc) as an antigen, three specific neutralizing antibody clones recognizing different epitopes [...] Read more.

Tetanus neurotoxin (TeNT) produced by Clostridium tetani is one of the most poisonous protein substances. Neutralizing antibodies against TeNT can effectively prevent and cure toxicosis. Using purified Hc fragments of TeNT (TeNT-Hc) as an antigen, three specific neutralizing antibody clones recognizing different epitopes were selected from a human immune scFv antibody phage display library. The three antibodies (2-7G, 2-2D, and S-4-7H) can effectively inhibit the binding between TeNT-Hc and differentiated PC-12 cells in vitro. Moreover, 2-7G inhibited TeNT-Hc binding to the receptor via carbohydrate-binding sites of the W pocket while 2-2D and S-4-7H inhibited binding of the R pocket. Although no single mAb completely protected mice from the toxin, they could both prolong survival when challenged with 20 LD₅₀s (50% of the lethal dose) of TeNT. When used together, the mAbs completely neutralized 1000 LD₅₀s/mg Ab, indicating their high neutralizing potency in vivo. Antibodies recognizing different carbohydrate-binding pockets could have higher synergistic toxin neutralization activities than those that recognize the same pockets. These results could lead to further production of neutralizing antibody drugs against TeNT and indicate that using TeNT-Hc as an antigen for screening human antibodies for TeNT intoxication therapy from human immune antibody library was convenient and effective. Full article

(This article belongs to the Section Bacterial Toxins)

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Open AccessArticle

Effective Botulinum Toxin Injection Guide for Treatment of Temporal Headache

by You-Jin Choi, Won-Jae Lee, Hyung-Jin Lee, Kang-Woo Lee, Hee-Jin Kim and Kyung-Seok Hu

Toxins 2016, 8(9), 265; https://doi.org/10.3390/toxins8090265 - 8 Sep 2016

Cited by 21 | Viewed by 12608

Abstract

This study involved an extensive analysis of published research on the morphology of the temporalis muscle in order to provide an anatomical guideline on how to distinguish the temporalis muscle and temporalis tendon by observing the surface of the patient’s face. Twenty-one hemifaces [...] Read more.

This study involved an extensive analysis of published research on the morphology of the temporalis muscle in order to provide an anatomical guideline on how to distinguish the temporalis muscle and temporalis tendon by observing the surface of the patient’s face. Twenty-one hemifaces of cadavers were used in this study. The temporalis muscles were dissected clearly for morphological analysis between the temporalis muscle and tendon. The posterior border of the temporalis tendon was classified into three types: in Type I the posterior border of the temporalis tendon is located in front of reference line L2 (4.8%, 1/21), in Type II it is located between reference lines L2 and L3 (85.7%, 18/21), and in Type III it is located between reference lines L3 and L4 (9.5%, 2/21). The vertical distances between the horizontal line passing through the jugale (LH) and the temporalis tendon along each of reference lines L0, L1, L2, L3, and L4 were 29.7 ± 6.8 mm, 45.0 ± 8.8 mm, 37.7 ± 11.1 mm, 42.5 ± 7.5 mm, and 32.1 ± 0.4 mm, respectively. BoNT-A should be injected into the temporalis muscle at least 45 mm vertically above the zygomatic arch. This will ensure that the muscle region is targeted and so produce the greatest clinical effect with the minimum concentration of BoNT-A. Full article

(This article belongs to the Collection Botulinum Toxins on Human Pain)

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Open AccessArticle

Early Activation of MAPK p44/42 Is Partially Involved in DON-Induced Disruption of the Intestinal Barrier Function and Tight Junction Network

by Alexandra Springler, Sabine Hessenberger, Gerd Schatzmayr and Elisabeth Mayer

Toxins 2016, 8(9), 264; https://doi.org/10.3390/toxins8090264 - 8 Sep 2016

Cited by 46 | Viewed by 6435

Abstract

Deoxynivalenol (DON), produced by the plant pathogens Fusarium graminearum and Fusarium culmorum, is one of the most common mycotoxins, contaminating cereal and cereal-derived products. Although worldwide contamination of food and feed poses health threats to humans and animals, pigs are particularly susceptible [...] Read more.

Deoxynivalenol (DON), produced by the plant pathogens Fusarium graminearum and Fusarium culmorum, is one of the most common mycotoxins, contaminating cereal and cereal-derived products. Although worldwide contamination of food and feed poses health threats to humans and animals, pigs are particularly susceptible to this mycotoxin. DON derivatives, such as deepoxy-deoxynivalenol (DOM-1), are produced by bacterial transformation of certain intestinal bacteria, which are naturally occurring or applied as feed additives. Intestinal epithelial cells are the initial barrier against these food- and feed-borne toxins. The present study confirms DON-induced activation of MAPK p44/42 and inhibition of p44/42 by MAPK-inhibitor U0126 monoethanolate. Influence of DON and DOM-1 on transepithelial electrical resistance (TEER), viability and expression of seven tight junction proteins (TJ), as well as the potential of U0126 to counteract DON-induced effects, was assessed. While DOM-1 showed no effect, DON significantly reduced TEER of differentiated IPEC-J2 and decreased expression of claudin-1 and -3, while leaving claudin-4; ZO-1, -2, and -3 and occludin unaffected. Inhibition of p44/42 counteracted DON-induced TEER decrease and restored claudin-3, but not claudin-1 expression. Therefore, effects of DON on TEER and claudin-3 are at least partially p44/42 mediated, while effects on viability and claudin-1 are likely mediated via alternative pathways. Full article

(This article belongs to the Section Mycotoxins)

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Open AccessArticle

Beyond Ribosomal Binding: The Increased Polarity and Aberrant Molecular Interactions of 3-epi-deoxynivalenol

by Yousef I. Hassan, Honghui Zhu, Yan Zhu and Ting Zhou

Toxins 2016, 8(9), 261; https://doi.org/10.3390/toxins8090261 - 8 Sep 2016

Cited by 14 | Viewed by 5267

Abstract

Deoxynivalenol (DON) is a secondary fungal metabolite and contaminant mycotoxin that is widely detected in wheat and corn products cultivated around the world. Bio-remediation methods have been extensively studied in the past two decades and promising ways to reduce DON-associated toxicities have been [...] Read more.

Deoxynivalenol (DON) is a secondary fungal metabolite and contaminant mycotoxin that is widely detected in wheat and corn products cultivated around the world. Bio-remediation methods have been extensively studied in the past two decades and promising ways to reduce DON-associated toxicities have been reported. Bacterial epimerization of DON at the C3 carbon was recently reported to induce a significant loss in the bio-toxicity of the resulting stereoisomer (3-epi-DON) in comparison to the parental compound, DON. In an earlier study, we confirmed the diminished bio-potency of 3-epi-DON using different mammalian cell lines and mouse models and mechanistically attributed it to the reduced binding of 3-epi-DON within the ribosomal peptidyl transferase center (PTC). In the current study and by inspecting the chromatographic behavior of 3-epi-DON and its molecular interactions with a well-characterized enzyme, Fusarium graminearum Tri101 acetyltransferase, we provide the evidence that the C3 carbon epimerization of DON influences its molecular interactions beyond the abrogated PTC binding. Full article

(This article belongs to the Special Issue Promising Detoxification Strategies to Mitigate Mycotoxins in Food and Feed)

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Open AccessArticle

A Tricky Trait: Applying the Fruits of the “Function Debate” in the Philosophy of Biology to the “Venom Debate” in the Science of Toxinology

by Timothy N. W. Jackson and Bryan G. Fry

Toxins 2016, 8(9), 263; https://doi.org/10.3390/toxins8090263 - 7 Sep 2016

Cited by 30 | Viewed by 7711

Abstract

The “function debate” in the philosophy of biology and the “venom debate” in the science of toxinology are conceptually related. Venom systems are complex multifunctional traits that have evolved independently numerous times throughout the animal kingdom. No single concept of function, amongst those [...] Read more.

The “function debate” in the philosophy of biology and the “venom debate” in the science of toxinology are conceptually related. Venom systems are complex multifunctional traits that have evolved independently numerous times throughout the animal kingdom. No single concept of function, amongst those popularly defended, appears adequate to describe these systems in all their evolutionary contexts and extant variations. As such, a pluralistic view of function, previously defended by some philosophers of biology, is most appropriate. Venom systems, like many other functional traits, exist in nature as points on a continuum and the boundaries between “venomous” and “non-venomous” species may not always be clearly defined. This paper includes a brief overview of the concept of function, followed by in-depth discussion of its application to venom systems. A sound understanding of function may aid in moving the venom debate forward. Similarly, consideration of a complex functional trait such as venom may be of interest to philosophers of biology. Full article

(This article belongs to the Collection Evolution of Venom Systems)

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Open AccessCommunication

The Contents of Ustiloxins A and B along with Their Distribution in Rice False Smut Balls

by Xiaohan Wang, Xiaoxiang Fu, Fengke Lin, Weibo Sun, Jiajia Meng, Ali Wang, Daowan Lai, Ligang Zhou and Yang Liu

Toxins 2016, 8(9), 262; https://doi.org/10.3390/toxins8090262 - 6 Sep 2016

Cited by 36 | Viewed by 7636

Abstract

Ustiloxins are cyclopeptide mycotoxins isolated from rice false smut balls (FSBs), the ball-like colonies transformed from the individual grains through the filament infection by the fungal pathogen Villosiclava virens. There were no obvious relations between ustiloxin content and any of the collection [...] Read more.

Ustiloxins are cyclopeptide mycotoxins isolated from rice false smut balls (FSBs), the ball-like colonies transformed from the individual grains through the filament infection by the fungal pathogen Villosiclava virens. There were no obvious relations between ustiloxin content and any of the collection areas, collection times, or average weight of each FSB. The rice false smut balls at early, middle, and late maturity stages were respectively divided into different parts (glume, chlamydospores, mycelia, and pseudoparenchyma). The highest content of ustiloxins A and B of rice FSBs was found at the early maturity stage. Both ustiloxins A and B were mainly distributed in the middle layer containing mycelia and immature chlamydospores of the FSBs. When the rice FSBs were at the early maturity stage, the total yield of ustiloxins A and B in the middle layer of each ball was 48.3 µg, which was 3.20-fold of the yield (15.1 µg) of the inner part of the ball. The rice FSBs at the early maturity stage are the appropriate materials for the production of ustiloxins A and B. Full article

(This article belongs to the Special Issue Selected papers from 1^st China Mycotoxin Conference: Current trends in mycotoxin prevention and control)

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Open AccessArticle

miR-541 Contributes to Microcystin-LR-Induced Reproductive Toxicity through Regulating the Expression of p15 in Mice

by Xiannan Meng, Ling Zhang, Xiang Chen, Zou Xiang, Dongmei Li and Xiaodong Han

Toxins 2016, 8(9), 260; https://doi.org/10.3390/toxins8090260 - 6 Sep 2016

Cited by 15 | Viewed by 5697

Abstract

Microcystin-leucine arginine (MC-LR) is a harmful cyanotoxin produced by cyanobacteria. MC-LR can exert endocrine-disrupting activities in many organisms. We have previously demonstrated that MC-LR exerts both acute and chronic reproductive toxicity in male mice, resulting in a decline in sperm quality and damage [...] Read more.

Microcystin-leucine arginine (MC-LR) is a harmful cyanotoxin produced by cyanobacteria. MC-LR can exert endocrine-disrupting activities in many organisms. We have previously demonstrated that MC-LR exerts both acute and chronic reproductive toxicity in male mice, resulting in a decline in sperm quality and damage to testicular structure. Moreover, we also observed extensive alterations in a panel of microRNAs in spermatogonial cells after exposure to MC-LR. In this study, we have confirmed that miR-541 was significantly increased both in GC-1 cells (in vitro) and in mouse testes (in vivo) after exposure to MC-LR. Our data support that p15 was the target gene of miR-541. Increase in miR-541 led to a reduction of p15 and murine double minute2 (MDM2), promoting the activation of p53 signaling and MC-LR-mediated cell apoptosis. Moreover, cells responded to MC-LR with reduced viability and increased apoptosis. Consistently, inhibiting miR-541 could upregulate the expression of p15 and MDM2, resulting in the downregulation of phospho-p53. Downregulation of miR-541 promoted cell viability by reducing MC-LR-induced cell apoptosis. In conclusion, we demonstrate here a crucial role for miR-541 in MC-LR-induced toxic effects on the reproductive system, in an attempt to provide a rational strategy for the diagnosis and treatment of MC-LR-induced impairment in the reproductive system. Full article

(This article belongs to the Collection Freshwater HABs and Health in a Changing World)

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Open AccessCommunication

A Novel Zak Knockout Mouse with a Defective Ribotoxic Stress Response

by Dakshina M. Jandhyala, John Wong, Nicholas J. Mantis, Bruce E. Magun, John M. Leong and Cheleste M. Thorpe

Toxins 2016, 8(9), 259; https://doi.org/10.3390/toxins8090259 - 2 Sep 2016

Cited by 11 | Viewed by 4704

Abstract

Ricin activates the proinflammatory ribotoxic stress response through the mitogen activated protein 3 kinase (MAP3K) ZAK, resulting in activation of mitogen activated protein kinases (MAPKs) p38 and JNK1/2. We had a novel zak−/− mouse generated to study the role of ZAK signaling in [...] Read more.

Ricin activates the proinflammatory ribotoxic stress response through the mitogen activated protein 3 kinase (MAP3K) ZAK, resulting in activation of mitogen activated protein kinases (MAPKs) p38 and JNK1/2. We had a novel zak−/− mouse generated to study the role of ZAK signaling in vivo during ricin intoxication. To characterize this murine strain, we intoxicated zak−/− and zak+/+ bone marrow–derived murine macrophages with ricin, measured p38 and JNK1/2 activation by Western blot, and measured zak, c-jun, and cxcl-1 expression by qRT-PCR. To determine whether zak−/− mice differed from wild-type mice in their in vivo response to ricin, we performed oral ricin intoxication experiments with zak+/+ and zak−/− mice, using blinded histopathology scoring of duodenal tissue sections to determine differences in tissue damage. Unlike macrophages derived from zak+/+ mice, those derived from the novel zak−/− strain fail to activate p38 and JNK1/2 and have decreased c-jun and cxcl-1 expression following ricin intoxication. Furthermore, compared with zak+/+ mice, zak−/− mice have decreased duodenal damage following in vivo ricin challenge. zak−/− mice demonstrate a distinct ribotoxic stress–associated phenotype in response to ricin and therefore provide a new animal model for in vivo studies of ZAK signaling. Full article

(This article belongs to the Section Plant Toxins)

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Open AccessArticle

Confirmation and Fine Mapping of a Major QTL for Aflatoxin Resistance in Maize Using a Combination of Linkage and Association Mapping

by Yu Zhang, Min Cui, Jimin Zhang, Lei Zhang, Chenliu Li, Xin Kan, Qian Sun, Dexiang Deng and Zhitong Yin

Toxins 2016, 8(9), 258; https://doi.org/10.3390/toxins8090258 - 2 Sep 2016

Cited by 21 | Viewed by 6712

Abstract

Maize grain contamination with aflatoxin from Aspergillus flavus (A. flavus) is a serious health hazard to animals and humans. To map the quantitative trait loci (QTLs) associated with resistance to A. flavus, we employed a powerful approach that differs from [...] Read more.

Maize grain contamination with aflatoxin from Aspergillus flavus (A. flavus) is a serious health hazard to animals and humans. To map the quantitative trait loci (QTLs) associated with resistance to A. flavus, we employed a powerful approach that differs from previous methods in one important way: it combines the advantages of the genome-wide association analysis (GWAS) and traditional linkage mapping analysis. Linkage mapping was performed using 228 recombinant inbred lines (RILs), and a highly significant QTL that affected aflatoxin accumulation, qAA8, was mapped. This QTL spanned approximately 7 centi-Morgan (cM) on chromosome 8. The confidence interval was too large for positional cloning of the causal gene. To refine this QTL, GWAS was performed with 558,629 single nucleotide polymorphisms (SNPs) in an association population comprising 437 maize inbred lines. Twenty-five significantly associated SNPs were identified, most of which co-localised with qAA8 and explained 6.7% to 26.8% of the phenotypic variation observed. Based on the rapid linkage disequilibrium (LD) and the high density of SNPs in the association population, qAA8 was further localised to a smaller genomic region of approximately 1500 bp. A high-resolution map of the qAA8 region will be useful towards a marker-assisted selection (MAS) of A. flavus resistance and a characterisation of the causal gene. Full article

(This article belongs to the Collection Aflatoxins)

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Open AccessCommunication

Presence of the Cyanotoxin Microcystin in Arctic Lakes of Southwestern Greenland

by Jessica V. Trout-Haney, Zachary T. Wood and Kathryn L. Cottingham

Toxins 2016, 8(9), 256; https://doi.org/10.3390/toxins8090256 - 31 Aug 2016

Cited by 21 | Viewed by 5708

Abstract

Cyanobacteria and their toxins have received significant attention in eutrophic temperate and tropical systems where conspicuous blooms of certain planktonic taxa release toxins into fresh water, threatening its potability and safe use for recreation. Although toxigenic cyanobacteria are not confined to high nutrient [...] Read more.

Cyanobacteria and their toxins have received significant attention in eutrophic temperate and tropical systems where conspicuous blooms of certain planktonic taxa release toxins into fresh water, threatening its potability and safe use for recreation. Although toxigenic cyanobacteria are not confined to high nutrient environments, bloom-forming species, or planktonic taxa, these other situations are studied les often studied. For example, toxin production in picoplankton and benthic cyanobacteria—the predominant photoautotrophs found in polar lakes—is poorly understood. We quantified the occurrence of microcystin (MC, a hepatotoxic cyanotoxin) across 18 Arctic lakes in southwestern Greenland. All of the focal lakes contained detectable levels of MC, with concentrations ranging from 5 ng·L⁻¹ to >400 ng·L⁻¹ during summer, 2013–2015. These concentrations are orders of magnitude lower than many eutrophic systems, yet the median lake MC concentration in Greenland (57 ng·L⁻¹) was still 6.5 times higher than the median summer MC toxicity observed across 50 New Hampshire lakes between 1998 and 2008 (8.7 ng·L⁻¹). The presence of cyanotoxins in these Greenlandic lakes demonstrates that high latitude lakes can support toxigenic cyanobacteria, and suggests that we may be underestimating the potential for these systems to develop high levels of cyanotoxins in the future. Full article

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

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Open AccessArticle

Discovery of Novel Bacterial Cell-Penetrating Phylloseptins in Defensive Skin Secretions of the South American Hylid Frogs, Phyllomedusa duellmani and Phyllomedusa coelestis

by Nan Yang, Lei Li, Di Wu, Yitian Gao, Xinping Xi, Mei Zhou, Lei Wang, Tianbao Chen and Chris Shaw

Toxins 2016, 8(9), 255; https://doi.org/10.3390/toxins8090255 - 31 Aug 2016

Cited by 9 | Viewed by 5674

Abstract

Phylloseptin (PS) peptides, derived from South American hylid frogs (subfamily Phyllomedusinae), have been found to have broad-spectrum antimicrobial activities and relatively low haemolytic activities. Although PS peptides have been identified from several well-known and widely-distributed species of the Phyllomedusinae, there remains merit in [...] Read more.

Phylloseptin (PS) peptides, derived from South American hylid frogs (subfamily Phyllomedusinae), have been found to have broad-spectrum antimicrobial activities and relatively low haemolytic activities. Although PS peptides have been identified from several well-known and widely-distributed species of the Phyllomedusinae, there remains merit in their study in additional, more obscure and specialised members of this taxon. Here, we report the discovery of two novel PS peptides, named PS-Du and PS-Co, which were respectively identified for the first time and isolated from the skin secretions of Phyllomedusa duellmani and Phyllomedusa coelestis. Their encoding cDNAs were cloned, from which it was possible to deduce the entire primary structures of their biosynthetic precursors. Reversed-phase high-performance liquid chromatography (RP-HPLC) and tandem mass spectrometry (MS/MS) analyses were employed to isolate and structurally-characterise respective encoded PS peptides from skin secretions. The peptides had molecular masses of 2049.7 Da (PS-Du) and 1972.8 Da (PS-Co). They shared typical N-terminal sequences and C-terminal amidation with other known phylloseptins. The two peptides exhibited growth inhibitory activity against E. coli (NCTC 10418), as a standard Gram-negative bacterium, S. aureus (NCTC 10788), as a standard Gram-positive bacterium and C. albicans (NCPF 1467), as a standard pathogenic yeast, all as planktonic cultures. Moreover, both peptides demonstrated the capability of eliminating S. aureus biofilm. Full article

(This article belongs to the Section Animal Venoms)

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Open AccessArticle

A Novel Population of Fusarium fujikuroi Isolated from Southeastern U.S. Winegrapes Reveals the Need to Re-Evaluate the Species’ Fumonisin Production

by Stephanie L. Bolton, Phillip M. Brannen and Anthony E. Glenn

Toxins 2016, 8(9), 254; https://doi.org/10.3390/toxins8090254 - 31 Aug 2016

Cited by 18 | Viewed by 5376

Abstract

Mycotoxins pose a challenge to a safe food supply worldwide, and their threat is expected to worsen with our changing climate. The need for diligence is exemplified by the discovery of fumonisin B2 in wine, which joins ochratoxin A as a mycotoxin of [...] Read more.

Mycotoxins pose a challenge to a safe food supply worldwide, and their threat is expected to worsen with our changing climate. The need for diligence is exemplified by the discovery of fumonisin B2 in wine, which joins ochratoxin A as a mycotoxin of concern in the grape-wine chain. To elucidate the mycotoxin risk in southeastern American wine, grape samples were collected from vineyards during harvest in 2013 and potentially mycotoxigenic fungi (Fusarium and Aspergillus) were isolated from the samples. Numerous Fusarium isolates were recovered and identified to the species level by comparison of translation elongation factor 1-α gene sequences to verified strains. Fusarium fujikuroi was the most abundant species recovered (239 isolates), followed by F. proliferatum (52), F. incarnatum-equiseti (14), F. oxysporum (7), F. concentricum (1), and F. solani (1). In vitro assays quantified fumonisin production for representative isolates via liquid chromatography-tandem mass spectrometry. Surprisingly, nearly all F. fujikuroi isolates produced fumonisins B1, B2, and B3 at levels comparable to both the F. proliferatum isolates and the positive control, Fusarium verticillioides. Such capacity for fumonisin production refutes the generally accepted notion that F. fujikuroi produces undetectable or low levels of fumonisins and provides evidence to reconsider this species as a mycotoxigenic threat to economically significant crops. Full article

(This article belongs to the Section Mycotoxins)

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Open AccessArticle

Development of Toxicological Risk Assessment Models for Acute and Chronic Exposure to Pollutants

by Elke S. Reichwaldt, Daniel Stone, Dani J. Barrington, Som C. Sinang and Anas Ghadouani

Toxins 2016, 8(9), 251; https://doi.org/10.3390/toxins8090251 - 31 Aug 2016

Cited by 12 | Viewed by 6928

Abstract

Alert level frameworks advise agencies on a sequence of monitoring and management actions, and are implemented so as to reduce the risk of the public coming into contact with hazardous substances. Their effectiveness relies on the detection of the hazard, but with many [...] Read more.

Alert level frameworks advise agencies on a sequence of monitoring and management actions, and are implemented so as to reduce the risk of the public coming into contact with hazardous substances. Their effectiveness relies on the detection of the hazard, but with many systems not receiving any regular monitoring, pollution events often go undetected. We developed toxicological risk assessment models for acute and chronic exposure to pollutants that incorporate the probabilities that the public will come into contact with undetected pollution events, to identify the level of risk a system poses in regards to the pollutant. As a proof of concept, we successfully demonstrated that the models could be applied to determine probabilities of acute and chronic illness types related to recreational activities in waterbodies containing cyanotoxins. Using the acute model, we identified lakes that present a ‘high’ risk to develop Day Away From Work illness, and lakes that present a ‘low’ or ‘medium’ risk to develop First Aid Cases when used for swimming. The developed risk models succeeded in categorising lakes according to their risk level to the public in an objective way. Modelling by how much the probability of public exposure has to decrease to lower the risks to acceptable levels will enable authorities to identify suitable control measures and monitoring strategies. We suggest broadening the application of these models to other contaminants. Full article

(This article belongs to the Collection Freshwater HABs and Health in a Changing World)

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Open AccessArticle

Exposure Assessment of Infants to Aflatoxin M₁ through Consumption of Breast Milk and Infant Powdered Milk in Brazil

by Angélica T. Ishikawa, Cássia R. Takabayashi-Yamashita, Elisabete Y. S. Ono, Artur K. Bagatin, Fabiana F. Rigobello, Osamu Kawamura, Elisa Y. Hirooka and Eiko N. Itano

Toxins 2016, 8(9), 246; https://doi.org/10.3390/toxins8090246 - 31 Aug 2016

Cited by 60 | Viewed by 7683

Abstract

Aflatoxin M₁ (AFM₁) is an important biomarker that can be used to evaluate aflatoxin exposure in both humans and animals. The aim of this study was to evaluate the exposure degree of infants to AFM₁ through consumption of breast [...] Read more.

Aflatoxin M₁ (AFM₁) is an important biomarker that can be used to evaluate aflatoxin exposure in both humans and animals. The aim of this study was to evaluate the exposure degree of infants to AFM₁ through consumption of breast milk and infant powdered milk in Brazil. For this purpose, the estimated daily intake (EDI) for infants was calculated based on the AFM₁ levels analyzed in 94 breast milk (BM) samples collected in Southern Brazil, and 16 infant powdered milk (IPM) samples commonly commercialized in Brazil. AFM₁ was detected in 5.3% (n = 5) and 43.8% (n = 7) of BM and IPM samples, with mean levels of 0.003 ng/g and 0.011 ng/g, respectively. All the IPM samples showed AFM₁ levels lower than those established by the Brazilian guidelines (5 ng/g), and in most of the samples (81.25%) levels were below the maximum limit tolerated by the European Commission (0.025 ng/g). The EDI of AFM₁ for infants aged zero to 12 months old showed values from 0.018 to 0.069 ng/kg body weight/day for BM, and 0.078 to 0.306 ng/kg body weight/day for IPM. Hazard index (HI) values for BM and IPM were less than one, except for IPM intended for infants up to one month. In conclusion, the exposure of infants to AFM₁ was low, but continuous monitoring of mycotoxin levels is essential to minimize infant health risk. Full article

(This article belongs to the Special Issue Exposure and Risk Assessment for Mycotoxins)

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172 KiB

Open AccessReply

Reply to the Letter “Create Guidelines for Characterization of Venom Peptides” from Dr. Volker Herzig

by Marcus Vinicius Gomez

Toxins 2016, 8(9), 253; https://doi.org/10.3390/toxins8090253 - 30 Aug 2016

Cited by 1 | Viewed by 3935

Abstract

Regarding our paper “PhTx3-4, a Spider Toxin Calcium Channel Blocker, Reduces NMDA-Induced Injury of the Retina”, published in Toxins 2016, 8, doi:10.3390/toxins8030070 [1].[...] Full article

(This article belongs to the Section Animal Venoms)

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Open AccessComment

Create Guidelines for Characterization of Venom Peptides

by Volker Herzig

Toxins 2016, 8(9), 252; https://doi.org/10.3390/toxins8090252 - 30 Aug 2016

Cited by 2 | Viewed by 4025

Abstract

In the course of my duties as a curator for the ArachnoServer database [1,2], I recently came across the article published by Binda et al. in Toxins [3].[...] Full article

(This article belongs to the Section Animal Venoms)

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Open AccessReview

Natural Inhibitors of Snake Venom Metalloendopeptidases: History and Current Challenges

by Viviane A. Bastos, Francisco Gomes-Neto, Jonas Perales, Ana Gisele C. Neves-Ferreira and Richard H. Valente

Toxins 2016, 8(9), 250; https://doi.org/10.3390/toxins8090250 - 26 Aug 2016

Cited by 27 | Viewed by 9267

Abstract

The research on natural snake venom metalloendopeptidase inhibitors (SVMPIs) began in the 18th century with the pioneering work of Fontana on the resistance that vipers exhibited to their own venom. During the past 40 years, SVMPIs have been isolated mainly from the sera [...] Read more.

The research on natural snake venom metalloendopeptidase inhibitors (SVMPIs) began in the 18th century with the pioneering work of Fontana on the resistance that vipers exhibited to their own venom. During the past 40 years, SVMPIs have been isolated mainly from the sera of resistant animals, and characterized to different extents. They are acidic oligomeric glycoproteins that remain biologically active over a wide range of pH and temperature values. Based on primary structure determination, mammalian plasmatic SVMPIs are classified as members of the immunoglobulin (Ig) supergene protein family, while the one isolated from muscle belongs to the ficolin/opsonin P35 family. On the other hand, SVMPIs from snake plasma have been placed in the cystatin superfamily. These natural antitoxins constitute the first line of defense against snake venoms, inhibiting the catalytic activities of snake venom metalloendopeptidases through the establishment of high-affinity, non-covalent interactions. This review presents a historical account of the field of natural resistance, summarizing its main discoveries and current challenges, which are mostly related to the limitations that preclude three-dimensional structural determinations of these inhibitors using “gold-standard” methods; perspectives on how to circumvent such limitations are presented. Potential applications of these SVMPIs in medicine are also highlighted. Full article

(This article belongs to the Special Issue Snake Venom Metalloproteinases)

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Open AccessCommunication

Varespladib (LY315920) Appears to Be a Potent, Broad-Spectrum, Inhibitor of Snake Venom Phospholipase A2 and a Possible Pre-Referral Treatment for Envenomation

by Matthew Lewin, Stephen Samuel, Janie Merkel and Philip Bickler

Toxins 2016, 8(9), 248; https://doi.org/10.3390/toxins8090248 - 25 Aug 2016

Cited by 158 | Viewed by 14148

Abstract

Snakebite remains a neglected medical problem of the developing world with up to 125,000 deaths each year despite more than a century of calls to improve snakebite prevention and care. An estimated 75% of fatalities from snakebite occur outside the hospital setting. Because [...] Read more.

Snakebite remains a neglected medical problem of the developing world with up to 125,000 deaths each year despite more than a century of calls to improve snakebite prevention and care. An estimated 75% of fatalities from snakebite occur outside the hospital setting. Because phospholipase A2 (PLA2) activity is an important component of venom toxicity, we sought candidate PLA2 inhibitors by directly testing drugs. Surprisingly, varespladib and its orally bioavailable prodrug, methyl-varespladib showed high-level secretory PLA2 (sPLA2) inhibition at nanomolar and picomolar concentrations against 28 medically important snake venoms from six continents. In vivo proof-of-concept studies with varespladib had striking survival benefit against lethal doses of Micrurus fulvius and Vipera berus venom, and suppressed venom-induced sPLA2 activity in rats challenged with 100% lethal doses of M. fulvius venom. Rapid development and deployment of a broad-spectrum PLA2 inhibitor alone or in combination with other small molecule inhibitors of snake toxins (e.g., metalloproteases) could fill the critical therapeutic gap spanning pre-referral and hospital setting. Lower barriers for clinical testing of safety tested, repurposed small molecule therapeutics are a potentially economical and effective path forward to fill the pre-referral gap in the setting of snakebite. Full article

(This article belongs to the Section Animal Venoms)

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Open AccessArticle

The Dinoflagellate Toxin 20-Methyl Spirolide-G Potently Blocks Skeletal Muscle and Neuronal Nicotinic Acetylcholine Receptors

by Aurélie Couesnon, Rómulo Aráoz, Bogdan I. Iorga, Evelyne Benoit, Morgane Reynaud, Denis Servent and Jordi Molgó

Toxins 2016, 8(9), 249; https://doi.org/10.3390/toxins8090249 - 24 Aug 2016

Cited by 15 | Viewed by 6464

Abstract

The cyclic imine toxin 20-methyl spirolide G (20-meSPX-G), produced by the toxigenic dinoflagellate Alexandrium ostenfeldii/Alexandrium peruvianum, has been previously reported to contaminate shellfish in various European coastal locations, as revealed by mouse toxicity bioassay. The aim of the present study [...] Read more.

The cyclic imine toxin 20-methyl spirolide G (20-meSPX-G), produced by the toxigenic dinoflagellate Alexandrium ostenfeldii/Alexandrium peruvianum, has been previously reported to contaminate shellfish in various European coastal locations, as revealed by mouse toxicity bioassay. The aim of the present study was to determine its toxicological profile and its molecular target selectivity. 20-meSPX-G blocked nerve-evoked isometric contractions in isolated mouse neuromuscular preparations, while it had no action on contractions elicited by direct electrical stimulation, and reduced reversibly nerve-evoked compound muscle action potential amplitudes in anesthetized mice. Voltage-clamp recordings in Xenopus oocytes revealed that 20-meSPX-G potently inhibited currents evoked by ACh on Torpedo muscle-type and human α7 nicotinic acetylcholine receptors (nAChR), whereas lower potency was observed in human α4β2 nAChR. Competition-binding assays showed that 20-meSPX-G fully displaced [³H]epibatidine binding to HEK-293 cells expressing the human α3β2 (Ki = 0.040 nM), whereas a 90-fold lower affinity was detected in human α4β2 nAChR. The spirolide displaced [¹²⁵I]α-bungarotoxin binding to Torpedo membranes (Ki = 0.028 nM) and in HEK-293 cells expressing chick chimeric α7-5HT₃ nAChR (Ki = 0.11 nM). In conclusion, this is the first study to demonstrate that 20-meSPX-G is a potent antagonist of nAChRs, and its subtype selectivity is discussed on the basis of molecular docking models. Full article

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

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Open AccessArticle

Aflatoxin B₁ and M₁ Degradation by Lac2 from Pleurotus pulmonarius and Redox Mediators

by Martina Loi, Francesca Fanelli, Paolo Zucca, Vania C. Liuzzi, Laura Quintieri, Maria T. Cimmarusti, Linda Monaci, Miriam Haidukowski, Antonio F. Logrieco, Enrico Sanjust and Giuseppina Mulè

Toxins 2016, 8(9), 245; https://doi.org/10.3390/toxins8090245 - 23 Aug 2016

Cited by 105 | Viewed by 9320

Abstract

Laccases (LCs) are multicopper oxidases that find application as versatile biocatalysts for the green bioremediation of environmental pollutants and xenobiotics. In this study we elucidate the degrading activity of Lac2 pure enzyme form Pleurotus pulmonarius towards aflatoxin B₁ (AFB₁) and [...] Read more.

Laccases (LCs) are multicopper oxidases that find application as versatile biocatalysts for the green bioremediation of environmental pollutants and xenobiotics. In this study we elucidate the degrading activity of Lac2 pure enzyme form Pleurotus pulmonarius towards aflatoxin B₁ (AFB₁) and M₁ (AFM₁). LC enzyme was purified using three chromatographic steps and identified as Lac2 through zymogram and LC-MS/MS. The degradation assays were performed in vitro at 25 °C for 72 h in buffer solution. AFB₁ degradation by Lac2 direct oxidation was 23%. Toxin degradation was also investigated in the presence of three redox mediators, (2,2′-azino-bis-[3-ethylbenzothiazoline-6-sulfonic acid]) (ABTS) and two naturally-occurring phenols, acetosyringone (AS) and syringaldehyde (SA). The direct effect of the enzyme and the mediated action of Lac2 with redox mediators univocally proved the correlation between Lac2 activity and aflatoxins degradation. The degradation of AFB₁ was enhanced by the addition of all mediators at 10 mM, with AS being the most effective (90% of degradation). AFM₁ was completely degraded by Lac2 with all mediators at 10 mM. The novelty of this study relies on the identification of a pure enzyme as capable of degrading AFB₁ and, for the first time, AFM₁, and on the evidence that the mechanism of an effective degradation occurs via the mediation of natural phenolic compounds. These results opened new perspective for Lac2 application in the food and feed supply chains as a biotransforming agent of AFB₁ and AFM₁. Full article

(This article belongs to the Special Issue Selected Papers from the 5th International Symposium on Mycotoxins and Toxigenic Moulds: Challenges and Perspectives)

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Toxins, Volume 8, Issue 9 (September 2016) – 23 articles

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