Editor's Choice Articles

Editor’s Choice articles are based on recommendations by the scientific editors of MDPI journals from around the world. Editors select a small number of articles recently published in the journal that they believe will be particularly interesting to authors, or important in this field. The aim is to provide a snapshot of some of the most exciting work published in the various research areas of the journal.

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Article
Betulinic Acid Ameliorates the T-2 Toxin-Triggered Intestinal Impairment in Mice by Inhibiting Inflammation and Mucosal Barrier Dysfunction through the NF-κB Signaling Pathway
Toxins 2020, 12(12), 794; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12120794 - 13 Dec 2020
Cited by 6
Abstract
T-2 toxin, a trichothecene mycotoxin produced by Fusarium, is widely distributed in crops and animal feed and frequently induces intestinal damage. Betulinic acid (BA), a plant-derived pentacyclic lupane-type triterpene, possesses potential immunomodulatory, antioxidant and anti-inflammatory biological properties. The current study aimed to [...] Read more.
T-2 toxin, a trichothecene mycotoxin produced by Fusarium, is widely distributed in crops and animal feed and frequently induces intestinal damage. Betulinic acid (BA), a plant-derived pentacyclic lupane-type triterpene, possesses potential immunomodulatory, antioxidant and anti-inflammatory biological properties. The current study aimed to explore the protective effect and molecular mechanisms of BA on intestinal mucosal impairment provoked by acute exposure to T-2 toxin. Mice were intragastrically administered BA (0.25, 0.5, or 1 mg/kg) daily for 2 weeks and then injected intraperitoneally with T-2 toxin (4 mg/kg) once to induce an intestinal impairment. BA pretreatment inhibited the loss of antioxidant capacity in the intestine of T-2 toxin-treated mice by elevating the levels of CAT, GSH-PX and GSH and reducing the accumulation of MDA. In addition, BA pretreatment alleviated the T-2 toxin-triggered intestinal immune barrier dysregulation by increasing the SIgA level in the intestine at dosages of 0.5 and 1 mg/kg, increasing IgG and IgM levels in serum at dosages of 0.5 and 1 mg/kg and restoring the intestinal C3 and C4 levels at a dosage of 1 mg/kg. BA administration at a dosage of 1 mg/kg also improved the intestinal chemical barrier by decreasing the serum level of DAO. Moreover, BA pretreatment improved the intestinal physical barrier via boosting the expression of ZO-1 and Occludin mRNAs and restoring the morphology of intestinal villi that was altered by T-2 toxin. Furthermore, treatment with 1 mg/kg BA downregulated the expression of p-NF-κB and p-IκB-α proteins in the intestine, while all doses of BA suppressed the pro-inflammatory cytokines expression of IL-1β, IL-6 and TNF-α mRNAs and increased the anti-inflammatory cytokine expression of IL-10 mRNA in the intestine of T-2 toxin-exposed mice. BA was proposed to exert a protective effect on intestinal mucosal disruption in T-2 toxin-stimulated mice by enhancing the intestinal antioxidant capacity, inhibiting the secretion of inflammatory cytokines and repairing intestinal mucosal barrier functions, which may be associated with BA-mediated inhibition of the NF-κB signaling pathway activation. Full article
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Article
Multi-Mycotoxin Occurrence in Dairy Cattle and Poultry Feeds and Feed Ingredients from Machakos Town, Kenya
Toxins 2020, 12(12), 762; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12120762 - 03 Dec 2020
Cited by 7
Abstract
Mycotoxins are common in grains in sub-Saharan Africa and negatively impact human and animal health and production. This study assessed occurrences of mycotoxins, some plant, and bacterial metabolites in 16 dairy and 27 poultry feeds, and 24 feed ingredients from Machakos town, Kenya, [...] Read more.
Mycotoxins are common in grains in sub-Saharan Africa and negatively impact human and animal health and production. This study assessed occurrences of mycotoxins, some plant, and bacterial metabolites in 16 dairy and 27 poultry feeds, and 24 feed ingredients from Machakos town, Kenya, in February and August 2019. We analyzed the samples using a validated multi-toxin liquid chromatography-tandem mass spectrometry method. A total of 153 mycotoxins, plant, and bacterial toxins, were detected in the samples. All the samples were co-contaminated with 21 to 116 different mycotoxins and/or metabolites. The commonly occurring and EU regulated mycotoxins reported were; aflatoxins (AFs) (70%; range 0.2–318.5 μg/kg), deoxynivalenol (82%; range 22.2–1037 μg/kg), ergot alkaloids (70%; range 0.4–285.7 μg/kg), fumonisins (90%; range 32.4–14,346 μg/kg), HT-2 toxin (3%; range 11.9–13.8 μg/kg), ochratoxin A (24%; range 1.1–24.3 μg/kg), T-2 toxin (4%; range 2.7–5.2 μg/kg) and zearalenone (94%; range 0.3–910.4 μg/kg). Other unregulated emerging mycotoxins and metabolites including Alternaria toxins, Aspergillus toxins, bacterial metabolites, cytochalasins, depsipeptides, Fusarium metabolites, metabolites from other fungi, Penicillium toxins, phytoestrogens, plant metabolites, and unspecific metabolites were also detected at varying levels. Except for total AFs, where the average contamination level was above the EU regulatory limit, all the other mycotoxins detected had average contamination levels below the limits. Ninety-six percent of all the samples were contaminated with more than one of the EU regulated mycotoxins. These co-occurrences may cause synergistic and additive health effects thereby hindering the growth of the Kenyan livestock sector. Full article
(This article belongs to the Section Mycotoxins)
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Article
Evidence for the Range Expansion of Ciguatera in French Polynesia: A Revisit of the 2009 Mass-Poisoning Outbreak in Rapa Island (Australes Archipelago)
Toxins 2020, 12(12), 759; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12120759 - 01 Dec 2020
Cited by 6
Abstract
Ciguatera poisoning (CP) results from the consumption of seafood contaminated with ciguatoxins (CTXs). This disease is highly prevalent in French Polynesia with several well-identified hotspots. Rapa Island, the southernmost inhabited island in the country, was reportedly free of CP until 2007. This study [...] Read more.
Ciguatera poisoning (CP) results from the consumption of seafood contaminated with ciguatoxins (CTXs). This disease is highly prevalent in French Polynesia with several well-identified hotspots. Rapa Island, the southernmost inhabited island in the country, was reportedly free of CP until 2007. This study describes the integrated approach used to investigate the etiology of a fatal mass-poisoning outbreak that occurred in Rapa in 2009. Symptoms reported in patients were evocative of ciguatera. Several Gambierdiscus field samples collected from benthic assemblages tested positive by the receptor binding assay (RBA). Additionally, the toxicity screening of ≈250 fish by RBA indicated ≈78% of fish could contain CTXs. The presence of CTXs in fish was confirmed by liquid chromatography tandem mass spectrometry (LC-MS/MS). The potential link between climate change and this range expansion of ciguatera to a subtropical locale of French Polynesia was also examined based on the analysis of temperature time-series data. Results are indicative of a global warming trend in Rapa area. A five-fold reduction in incidence rates was observed between 2009 and 2012, which was due in part to self-regulating behavior among individuals (avoidance of particular fish species and areas). Such observations underscore the prominent role played by community outreach in ciguatera risk management. Full article
(This article belongs to the Special Issue Ciguatoxins)
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Article
Insight into Unprecedented Diversity of Cyanopeptides in Eutrophic Ponds Using an MS/MS Networking Approach
Toxins 2020, 12(9), 561; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12090561 - 31 Aug 2020
Cited by 7
Abstract
Man-made shallow fishponds in the Czech Republic have been facing high eutrophication since the 1950s. Anthropogenic eutrophication and feeding of fish have strongly affected the physicochemical properties of water and its aquatic community composition, leading to harmful algal bloom formation. In our current [...] Read more.
Man-made shallow fishponds in the Czech Republic have been facing high eutrophication since the 1950s. Anthropogenic eutrophication and feeding of fish have strongly affected the physicochemical properties of water and its aquatic community composition, leading to harmful algal bloom formation. In our current study, we characterized the phytoplankton community across three eutrophic ponds to assess the phytoplankton dynamics during the vegetation season. We microscopically identified and quantified 29 cyanobacterial taxa comprising non-toxigenic and toxigenic species. Further, a detailed cyanopeptides (CNPs) profiling was performed using molecular networking analysis of liquid chromatography-tandem mass spectrometry (LC-MS/MS) data coupled with a dereplication strategy. This MS networking approach, coupled with dereplication, on the online global natural product social networking (GNPS) web platform led us to putatively identify forty CNPs: fourteen anabaenopeptins, ten microcystins, five cyanopeptolins, six microginins, two cyanobactins, a dipeptide radiosumin, a cyclooctapeptide planktocyclin, and epidolastatin 12. We applied the binary logistic regression to estimate the CNPs producers by correlating the GNPS data with the species abundance. The usage of the GNPS web platform proved a valuable approach for the rapid and simultaneous detection of a large number of peptides and rapid risk assessments for harmful blooms. Full article
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Article
Betulinic Acid Attenuates Oxidative Stress in the Thymus Induced by Acute Exposure to T-2 Toxin via Regulation of the MAPK/Nrf2 Signaling Pathway
Toxins 2020, 12(9), 540; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12090540 - 22 Aug 2020
Cited by 6
Abstract
T-2 toxin, the most toxic of the trichothecenes, is widely found in grains and feeds, and its intake poses serious risks to the health of humans and animals. An important cytotoxicity mechanism of T-2 toxin is the production of excess free radicals, which [...] Read more.
T-2 toxin, the most toxic of the trichothecenes, is widely found in grains and feeds, and its intake poses serious risks to the health of humans and animals. An important cytotoxicity mechanism of T-2 toxin is the production of excess free radicals, which in turn leads to oxidative stress. Betulinic acid (BA) has many biological activities, including antioxidant activity, which is a plant-derived pentacyclic triterpenoid. The protective effects and mechanisms of BA in blocking oxidative stress caused by acute exposure to T-2 toxin in the thymus of mice was studied. BA pretreatment reduced ROS production, decreased the MDA content, and increased the content of IgG in serum and the levels of SOD and GSH in the thymus. BA pretreatment also reduced the degree of congestion observed in histopathological tissue sections of the thymus induced by T-2 toxin. Besides, BA downregulated the phosphorylation of the p38, JNK, and ERK proteins, while it upregulated the expression of the Nrf2 and HO-1 proteins in thymus tissues. The results indicated that BA could protect the thymus against the oxidative damage challenged by T-2 toxin by activating Nrf2 and suppressing the MAPK signaling pathway. Full article
(This article belongs to the Special Issue Mycotoxins in Feed: Harm to Animals)
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Article
Investigation of the Efficacy of a Postbiotic Yeast Cell Wall-Based Blend on Newly-Weaned Pigs under a Dietary Challenge of Multiple Mycotoxins with Emphasis on Deoxynivalenol
Toxins 2020, 12(8), 504; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12080504 - 06 Aug 2020
Cited by 11
Abstract
Pigs are highly susceptible to mycotoxins. This study investigated the effects of a postbiotic yeast cell wall-based blend (PYCW; Nicholasville, KY, USA) on growth and health of newly-weaned pigs under dietary challenge of multiple mycotoxins. Forty-eight newly-weaned pigs (21 d old) were individually [...] Read more.
Pigs are highly susceptible to mycotoxins. This study investigated the effects of a postbiotic yeast cell wall-based blend (PYCW; Nicholasville, KY, USA) on growth and health of newly-weaned pigs under dietary challenge of multiple mycotoxins. Forty-eight newly-weaned pigs (21 d old) were individually allotted to four dietary treatments, based on a three phase-feeding, in a randomized complete block design (sex; initial BW) with two factors for 36 d. Two factors were dietary mycotoxins (deoxynivalenol: 2000 μg/kg supplemented in three phases; and aflatoxin: 200 μg/kg supplemented only in phase 3) and PYCW (0.2%). Growth performance (weekly), blood serum (d 34), and jejunal mucosa immune and oxidative stress markers (d 36) data were analyzed using MIXED procedure of SAS. Mycotoxins reduced (p < 0.05) average daily feed intake (ADFI) and average daily gain (ADG) during the entire period whereas PYCW did not affect growth performance. Mycotoxins reduced (p < 0.05) serum protein, albumin, creatinine, and alanine aminotransferase whereas PYCW decreased (p < 0.05) serum creatine phosphokinase. Neither mycotoxins nor PYCW affected pro-inflammatory cytokines and oxidative damage markers in the jejunal mucosa. No interaction was observed indicating that PYCW improved hepatic enzymes regardless of mycotoxin challenge. In conclusion, deoxynivalenol (2000 μg/kg, for 7 to 25 kg body weight) and aflatoxin B1 (200 μg/kg, for 16 to 25 kg body weight) impaired growth performance and nutrient digestibility of newly-weaned pigs, whereas PYCW could partially improve health of pigs regardless of mycotoxin challenge. Full article
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Article
Effect of Temperature, Water Activity and Carbon Dioxide on Fungal Growth and Mycotoxin Production of Acclimatised Isolates of Fusarium verticillioides and F. graminearum
Toxins 2020, 12(8), 478; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12080478 - 28 Jul 2020
Cited by 9
Abstract
Climate change is primarily manifested by elevated temperature and carbon dioxide (CO2) levels and is projected to provide suitable cultivation grounds for pests and pathogens in the otherwise unsuitable regions. The impacts of climate change have been predicted in many parts [...] Read more.
Climate change is primarily manifested by elevated temperature and carbon dioxide (CO2) levels and is projected to provide suitable cultivation grounds for pests and pathogens in the otherwise unsuitable regions. The impacts of climate change have been predicted in many parts of the world, which could threaten global food safety and food security. The aim of the present work was therefore to examine the interacting effects of water activity (aw) (0.92, 0.95, 0.98 aw), CO2 (400, 800, 1200 ppm) and temperature (30, 35 °C and 30, 33 °C for Fusarium verticillioides and F. graminearum, respectively) on fungal growth and mycotoxin production of acclimatised isolates of F. verticillioides and F. graminearum isolated from maize. To determine fungal growth, the colony diameters were measured on days 1, 3, 5, and 7. The mycotoxins produced were quantified using a quadrupole-time-of-flight mass spectrometer (QTOF-MS) combined with ultra-high-performance liquid chromatography (UHPLC) system. For F. verticillioides, the optimum conditions for growth of fumonisin B1 (FB1), and fumonisin B2 (FB2) were 30 °C + 0.98 aw + 400 ppm CO2. These conditions were also optimum for F. graminearum growth, and zearalenone (ZEA) and deoxynivalenol (DON) production. Since 30 °C and 400 ppm CO2 were the baseline treatments, it was hence concluded that the elevated temperature and CO2 levels tested did not seem to significantly impact fungal growth and mycotoxin production of acclimatised Fusarium isolates. To the best of our knowledge thus far, the present work described for the first time the effects of simulated climate change conditions on fungal growth and mycotoxin production of acclimatised isolates of F. verticillioides and F. graminearum. Full article
(This article belongs to the Special Issue Mycotoxins in Relation to Climate Change)
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Article
Development of an Improved Method of Sample Extraction and Quantitation of Multi-Mycotoxin in Feed by LC-MS/MS
Toxins 2020, 12(7), 462; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12070462 - 19 Jul 2020
Cited by 6
Abstract
A multi-mycotoxin chromatographic method was developed and validated for the simultaneous quantitation of aflatoxins (AFB1, AFB2, AFG1 and AFG2), ochratoxin A (OTA), zearalenone (ZON), deoxynivalenol (DON), nivalenol (NIV), diacetoxyscirpenol (DAS), fumonisins (FB1, FB2 and FB3), T-2 toxin (T-2) and HT-2 toxin (HT-2) in [...] Read more.
A multi-mycotoxin chromatographic method was developed and validated for the simultaneous quantitation of aflatoxins (AFB1, AFB2, AFG1 and AFG2), ochratoxin A (OTA), zearalenone (ZON), deoxynivalenol (DON), nivalenol (NIV), diacetoxyscirpenol (DAS), fumonisins (FB1, FB2 and FB3), T-2 toxin (T-2) and HT-2 toxin (HT-2) in feed. The three most popular sample preparation techniques for determination of mycotoxins have been evaluated, and the method with highest recoveries was selected and optimized. This modified QuEChERS (quick, easy, cheap, effective, rugged and safe) approach was based on the extraction with acetonitrile, salting-out and cleanup with lipid removal. A reconstitution process in methanol/water was used to improve the MS responses and then the extracts were analyzed by LC-MS/MS. In this method, the recovery range is 70–100% for DON, DAS, FB1, FB2, FB3, HT-2, T-2, OTA, ZON, AFG1, AFG2, AFB1 and AFB2 and 55% for NIV in the spike range of 2–80 µg/kg. Method robustness was determined with acceptable z-scores in proficiency tests and validation experiments. Full article
(This article belongs to the Special Issue Rapid Detection of Mycotoxin Contamination)
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Article
AFLA-PISTACHIO: Development of a Mechanistic Model to Predict the Aflatoxin Contamination of Pistachio Nuts
Toxins 2020, 12(7), 445; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12070445 - 10 Jul 2020
Cited by 8
Abstract
In recent years, very many incidences of contamination with aflatoxin B1 (AFB1) in pistachio nuts have been reported as a major global problem for the crop. In Europe, legislation is in force and 12 μg/kg of AFB1 is the [...] Read more.
In recent years, very many incidences of contamination with aflatoxin B1 (AFB1) in pistachio nuts have been reported as a major global problem for the crop. In Europe, legislation is in force and 12 μg/kg of AFB1 is the maximum limit set for pistachios to be subjected to physical treatment before human consumption. The goal of the current study was to develop a mechanistic, weather-driven model to predict Aspergillus flavus growth and the AFB1 contamination of pistachios on a daily basis from nut setting until harvest. The planned steps were to: (i) build a phenology model to predict the pistachio growth stages, (ii) develop a prototype model named AFLA-pistachio (model transfer from AFLA-maize), (iii) collect the meteorological and AFB1 contamination data from pistachio orchards, (iv) run the model and elaborate a probability function to estimate the likelihood of overcoming the legal limit, and (v) manage a preliminary validation. The internal validation of AFLA-pistachio indicated that 75% of the predictions were correct. In the external validation with an independent three-year dataset, 95.6% of the samples were correctly predicted. According to the results, AFLA-pistachio seems to be a reliable tool to follow the dynamic of AFB1 contamination risk throughout the pistachio growing season. Full article
(This article belongs to the Collection Understanding Mycotoxin Occurrence in Food and Feed Chains)
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Article
Potential of Bacteriocins from Lactobacillus taiwanensis for Producing Bacterial Ghosts as a Next Generation Vaccine
Toxins 2020, 12(7), 432; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12070432 - 01 Jul 2020
Cited by 8
Abstract
Bacteriocins are functionally diverse toxins produced by most microbes and are potent antimicrobial peptides (AMPs) for bacterial ghosts as next generation vaccines. Here, we first report that the AMPs secreted from Lactobacillus taiwanensis effectively form ghosts of pathogenic bacteria and are identified as [...] Read more.
Bacteriocins are functionally diverse toxins produced by most microbes and are potent antimicrobial peptides (AMPs) for bacterial ghosts as next generation vaccines. Here, we first report that the AMPs secreted from Lactobacillus taiwanensis effectively form ghosts of pathogenic bacteria and are identified as diverse bacteriocins, including novel ones. In detail, a cell-free supernatant from L. taiwanensis exhibited antimicrobial activities against pathogenic bacteria and was observed to effectively cause cellular lysis through pore formation in the bacterial membrane using scanning electron microscopy (SEM). The treatment of the cell-free supernatant with proteinase K or EDTA proved that the antimicrobial activity is mediated by AMPs, and the purification of AMPs using Sep-Pak columns indicated that the cell-free supernatant includes various amphipathic peptides responsible for the antimicrobial activity. Furthermore, the whole-genome sequencing of L. taiwanensis revealed that the strain has diverse bacteriocins, confirmed experimentally to function as AMPs, and among them are three novel bacteriocins, designated as Tan 1, Tan 2, and Tan 3. We also confirmed, using SEM, that Tan 2 effectively produces bacterial ghosts. Therefore, our data suggest that the bacteriocins from L. taiwanensis are potentially useful as a critical component for the preparation of bacterial ghosts. Full article
(This article belongs to the Special Issue Drug Development Using Natural Toxins)
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Article
Low Doses of Mycotoxin Mixtures below EU Regulatory Limits Can Negatively Affect the Performance of Broiler Chickens: A Longitudinal Study
Toxins 2020, 12(7), 433; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12070433 - 01 Jul 2020
Cited by 7
Abstract
Several studies have reported a wide range of severe health effects as well as clinical signs, when livestock animals are exposed to high concentration of mycotoxins. However, little is known regarding health effects of mycotoxins at low levels. Thus, a long-term feeding trial [...] Read more.
Several studies have reported a wide range of severe health effects as well as clinical signs, when livestock animals are exposed to high concentration of mycotoxins. However, little is known regarding health effects of mycotoxins at low levels. Thus, a long-term feeding trial (between May 2017 and December 2019) was used to evaluate the effect of low doses of mycotoxin mixtures on performance of broiler chickens fed a naturally contaminated diet. In total, 18 successive broiler performance trials were carried out during the study period, with approximately 2200 one-day-old Ross-308 chicks used for each trial. Feed samples given to birds were collected at the beginning of each trial and analysed for multi-mycotoxins using a validated LC-MS/MS method. Furthermore, parameters including feed intake, body weight and feed efficiency were recorded on a weekly basis. In total, 24 mycotoxins were detected in samples analysed with deoxynivalenol (DON), zearalenone (ZEN), fumonisins (FBs), apicidin, enniatins (ENNs), emodin and beauvericin (BEV), the most prevalent mycotoxins. Furthermore, significantly higher levels (however below EU guidance values) of DON, ZEN, FBs, BEV, ENNs and diacetoxyscirpenol (DAS) were detected in 6 of the 18 performance trials. A strong positive relationship was observed between broilers feed efficiency and DON (R2 = 0.85), FBs (R2 = 0.53), DAS (R2 = 0.86), ZEN (R2 = 0.92), ENNs (R2 = 0.60) and BEV (R2 = 0.73). Moreover, a three-way interaction regression model revealed that mixtures of ZEN, DON and FBs (p = 0.01, R2 = 0.84) and ZEN, DON and DAS (p = 0.001, R2 = 0.91) had a statistically significant interaction effect on the birds’ feed efficiency. As farm animals are often exposed to low doses of mycotoxin mixtures (especially fusarium mycotoxins), a cumulative risk assessment in terms of measuring and mitigating against the economic, welfare and health impacts is needed for this group of compounds. Full article
(This article belongs to the Section Mycotoxins)
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Article
The Efficacy of Hydrogen Peroxide in Mitigating Cyanobacterial Blooms and Altering Microbial Communities across Four Lakes in NY, USA
Toxins 2020, 12(7), 428; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12070428 - 29 Jun 2020
Cited by 7
Abstract
Hydrogen peroxide (H2O2) has been proposed as an agent to mitigate toxic cyanobacterial blooms due to the heightened sensitivity of cyanobacteria to reactive oxygen species relative to eukaryotic organisms. Here, experiments were conducted using water from four diverse, eutrophic [...] Read more.
Hydrogen peroxide (H2O2) has been proposed as an agent to mitigate toxic cyanobacterial blooms due to the heightened sensitivity of cyanobacteria to reactive oxygen species relative to eukaryotic organisms. Here, experiments were conducted using water from four diverse, eutrophic lake ecosystems to study the effects of H2O2 on cyanobacteria and non-target members of the microbial community. H2O2 was administered at 4 µg L−1 and a combination of fluorometry, microscopy, flow cytometry, and high throughput DNA sequencing were used to quantify the effects on eukaryotic and prokaryotic plankton communities. The addition of H2O2 resulted in a significant reduction in cyanobacteria levels in nearly all experiments (10 of 11), reducing their relative abundance from, on average, 85% to 29% of the total phytoplankton community with Planktothrix being highly sensitive, Microcystis being moderately sensitive, and Cylindrospermopsis being most resistant. Concurrently, eukaryotic algal levels increased in 75% of experiments. The bacterial phyla Actinobacteria, cyanobacteria, Planctomycetes, and Verrucomicrobia were most negatively impacted by H2O2, with Actinobacteria being the most sensitive. The ability of H2O2 to reduce, but not fully eliminate, cyanobacteria from the eutrophic water bodies studied here suggests it may not be an ideal mitigation approach in high biomass ecosystems. Full article
(This article belongs to the Special Issue Removal of Cyanobacteria and Cyanotoxins in Waters)
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Article
Reduced Membrane-Bound Alkaline Phosphatase Does Not Affect Binding of Vip3Aa in a Heliothis virescens Resistant Colony
Toxins 2020, 12(6), 409; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12060409 - 19 Jun 2020
Cited by 6
Abstract
The Vip3Aa insecticidal protein from Bacillus thuringiensis (Bt) is produced by specific transgenic corn and cotton varieties for efficient control of target lepidopteran pests. The main threat to this technology is the evolution of resistance in targeted insect pests and understanding the mechanistic [...] Read more.
The Vip3Aa insecticidal protein from Bacillus thuringiensis (Bt) is produced by specific transgenic corn and cotton varieties for efficient control of target lepidopteran pests. The main threat to this technology is the evolution of resistance in targeted insect pests and understanding the mechanistic basis of resistance is crucial to deploy the most appropriate strategies for resistance management. In this work, we tested whether alteration of membrane receptors in the insect midgut might explain the >2000-fold Vip3Aa resistance phenotype in a laboratory-selected colony of Heliothis virescens (Vip-Sel). Binding of 125I-labeled Vip3Aa to brush border membrane vesicles (BBMV) from 3rd instar larvae from Vip-Sel was not significantly different from binding in the reference susceptible colony. Interestingly, BBMV from Vip-Sel larvae showed dramatically reduced levels of membrane-bound alkaline phosphatase (mALP) activity, which was further confirmed by a strong downregulation of the membrane-bound alkaline phosphatase 1 (HvmALP1) gene. However, the involvement of HvmALP1 as a receptor for the Vip3Aa protein was not supported by results from ligand blotting and viability assays with insect cells expressing HvmALP1. Full article
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Article
The Comparative Toxicity of 10 Microcystin Congeners Administered Orally to Mice: Clinical Effects and Organ Toxicity
Toxins 2020, 12(6), 403; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12060403 - 18 Jun 2020
Cited by 8
Abstract
Microcystins (MCs) are common cyanobacterial toxins that occur in freshwaters worldwide. Only two of the >200 MC variants have been tested for potential toxicity after oral exposure. This paper reports on the toxicity of 10 different MC congeners identified in algal blooms, microcystin-LR [...] Read more.
Microcystins (MCs) are common cyanobacterial toxins that occur in freshwaters worldwide. Only two of the >200 MC variants have been tested for potential toxicity after oral exposure. This paper reports on the toxicity of 10 different MC congeners identified in algal blooms, microcystin-LR (MCLR), MCLA, MCLF, MCLW, MCLY, MCRR, [Asp3]MCRR, [Asp3,Dhb7]MCRR, MCWR, and MCYR after single administrations to BALB/c mice. In a preliminary MCLR dose–response study of 3 to 9 mg/kg doses, ≥5 mg/kg induced clinical changes, increased serum levels of ALT, AST, and GLDH, liver congestion, increased liver/body weight ratios, and reduced serum glucose and total protein. Based on the extent of these effects, the 10 congeners were administered as single 7 mg/kg oral doses and toxicity evaluated. The greatest toxicity was observed with MCLA and MCLR including a high percentage of moribundity. In addition to eliciting effects similar to those listed above for MCLR, MCLA also induced serum alterations indicative of jaundice. MCLY, and MCYR induced changes like those noted with MCLR, but to lesser extents. MCLW and MCLF exhibited some serum and morphological changes associated with hepatic toxicity, while there were few indications of toxicity after exposures to MCRR, [Asp3]MCRR, [Asp3,Dhb7]MCRR, or MCWR. These data illustrate a wide spectrum of hepatic effects and different potencies of these MC congeners. Full article
(This article belongs to the Section Marine and Freshwater Toxins)
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Article
Genes Encoding the Virulence and the Antimicrobial Resistance in Enterotoxigenic and Shiga-toxigenic E. coli Isolated from Diarrheic Calves
Toxins 2020, 12(6), 383; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12060383 - 10 Jun 2020
Cited by 17
Abstract
Calf diarrhea is one of the considerable infectious diseases in calves, which results in tremendous economic losses globally. To determine the prevalence of Shiga-toxigenic E. coli (STEC) and Enterotoxigenic E. coli (ETEC) incriminated in calf diarrhea, with special reference to Shiga- toxins genes [...] Read more.
Calf diarrhea is one of the considerable infectious diseases in calves, which results in tremendous economic losses globally. To determine the prevalence of Shiga-toxigenic E. coli (STEC) and Enterotoxigenic E. coli (ETEC) incriminated in calf diarrhea, with special reference to Shiga- toxins genes (stx1 and stx2) and enterotoxins genes (lt and sta) that govern their pathogenesis, as well as the virulence genes; eaeA (intimin) and f41(fimbrial adhesion), and the screening of their antibiogram and antimicrobial resistance genes; aadB, sul1, and bla-TEM, a total of 274 fecal samples were collected (April 2018–Feb 2019) from diarrheic calves at different farms in El-Sharqia Governorate, Egypt. The bacteriological examination revealed that the prevalence of E. coli in diarrheic calves was 28.8%. The serotyping of the isolated E. coli revealed 7 serogroups; O26, O128, O111, O125, O45, O119 and O91. Furthermore, the Congo red binding test was carried out, where 89.8% of the examined strains (n = 71) were positive. The antibiogram of the isolated strains was investigated; the majority of E. coli serotypes exhibit multidrug resistance (MDR) to four antimicrobial agents; neomycin, gentamycin, streptomycin, and amikacin. Polymerase chain reaction (PCR) was used to detect the prevalence of the virulence genes; stx1, stx2 lt, sta, f41 and eaeA, as well as the antimicrobial resistance genes; aadB, sul1, and bla-TEM. The prevalence of STEC was 20.2% (n = 16), while the prevalence of ETEC was 30.4% (n = 24). Briefly, the Shiga toxins genes; stx1 and stx2, are the most prevalent virulence genes associated with STEC, which are responsible for the pathogenesis of the disease and helped by the intimin gene (eaeA). In addition, the lt gene is the most prevalent enterotoxin gene accompanied by the ETEC strains, either alone or in combination with sta and/or f41 genes. The majority of pathogenic E. coli incriminated in calf diarrhea possesses the aadB resistance gene, followed by the sul1 gene. Enrofloxacin, florfenicol, amoxicillin-clavulanic acid, and ampicillin-sulbactam, are the most effective antimicrobial agents against the isolated STEC and ETEC strains. Full article
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Article
Cylindrospermopsin-Microcystin-LR Combinations May Induce Genotoxic and Histopathological Damage in Rats
Toxins 2020, 12(6), 348; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12060348 - 26 May 2020
Cited by 8
Abstract
Cylindrospermopsin (CYN) and microcystins (MC) are cyanotoxins that can occur simultaneously in contaminated water and food. CYN/MC-LR mixtures previously investigated in vitro showed an induction of micronucleus (MN) formation only in the presence of the metabolic fraction S9. When this is the case, [...] Read more.
Cylindrospermopsin (CYN) and microcystins (MC) are cyanotoxins that can occur simultaneously in contaminated water and food. CYN/MC-LR mixtures previously investigated in vitro showed an induction of micronucleus (MN) formation only in the presence of the metabolic fraction S9. When this is the case, the European Food Safety Authority recommends a follow up to in vivo testing. Thus, rats were orally exposed to 7.5 + 75, 23.7 + 237, and 75 + 750 μg CYN/MC-LR/kg body weight (b.w.). The MN test in bone marrow was performed, and the standard and modified comet assays were carried out to measure DNA strand breaks or oxidative DNA damage in stomach, liver, and blood cells. The results revealed an increase in MN formation in bone marrow, at all the assayed doses. However, no DNA strand breaks nor oxidative DNA damage were induced, as shown in the comet assays. The histopathological study indicated alterations only in the highest dose group. Liver was the target organ showing fatty degeneration and necrotic hepatocytes in centrilobular areas, as well as a light mononuclear inflammatory periportal infiltrate. Additionally, the stomach had flaking epithelium and mild necrosis of epithelial cells. Therefore, the combined exposure to cyanotoxins may induce genotoxic and histopathological damage in vivo. Full article
(This article belongs to the Special Issue Freshwater Algal Toxins: Monitoring and Toxicity Profile)
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Article
The First Proteomic Study of Nostoc sp. PCC 7120 Exposed to Cyanotoxin BMAA under Nitrogen Starvation
Toxins 2020, 12(5), 310; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12050310 - 09 May 2020
Cited by 7
Abstract
The oldest prokaryotic photoautotrophic organisms, cyanobacteria, produce many different metabolites. Among them is the water-soluble neurotoxic non-protein amino acid beta-N-methylamino-L-alanine (BMAA), whose biological functions in cyanobacterial metabolism are of fundamental scientific and practical interest. An early BMAA inhibitory effect on nitrogen fixation and [...] Read more.
The oldest prokaryotic photoautotrophic organisms, cyanobacteria, produce many different metabolites. Among them is the water-soluble neurotoxic non-protein amino acid beta-N-methylamino-L-alanine (BMAA), whose biological functions in cyanobacterial metabolism are of fundamental scientific and practical interest. An early BMAA inhibitory effect on nitrogen fixation and heterocyst differentiation was shown in strains of diazotrophic cyanobacteria Nostoc sp. PCC 7120, Nostoc punctiforme PCC 73102 (ATCC 29133), and Nostoc sp. strain 8963 under conditions of nitrogen starvation. Herein, we present a comprehensive proteomic study of Nostoc (also called Anabaena) sp. PCC 7120 in the heterocyst formation stage affecting by BMAA treatment under nitrogen starvation conditions. BMAA disturbs proteins involved in nitrogen and carbon metabolic pathways, which are tightly co-regulated in cyanobacteria cells. The presented evidence shows that exogenous BMAA affects a key nitrogen regulatory protein, PII (GlnB), and some of its protein partners, as well as glutamyl-tRNA synthetase gltX and other proteins that are involved in protein synthesis, heterocyst differentiation, and nitrogen metabolism. By taking into account the important regulatory role of PII, it becomes clear that BMAA has a severe negative impact on the carbon and nitrogen metabolism of starving Nostoc sp. PCC 7120 cells. BMAA disturbs carbon fixation and the carbon dioxide concentrating mechanism, photosynthesis, and amino acid metabolism. Stress response proteins and DNA repair enzymes are upregulated in the presence of BMAA, clearly indicating severe intracellular stress. This is the first proteomic study of the effects of BMAA on diazotrophic starving cyanobacteria cells, allowing a deeper insight into the regulation of the intracellular metabolism of cyanobacteria by this non-protein amino acid. Full article
(This article belongs to the Special Issue Biological Role of Cyanotoxins: Experimental and In-Field Evidence)
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Article
Efficacy of Mycotoxin Detoxifiers on Health and Growth of Newly-Weaned Pigs under Chronic Dietary Challenge of Deoxynivalenol
Toxins 2020, 12(5), 311; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12050311 - 09 May 2020
Cited by 14
Abstract
The efficacy of yeast-based mycotoxin detoxifiers on health and growth performance of newly-weaned pigs (27-d-old) fed diets naturally contaminated with deoxynivalenol was investigated. Sixty pigs were individually assigned to five treatments for 34 d: NC (negative control, 1.2 mg/kg of deoxynivalenol); PC (positive [...] Read more.
The efficacy of yeast-based mycotoxin detoxifiers on health and growth performance of newly-weaned pigs (27-d-old) fed diets naturally contaminated with deoxynivalenol was investigated. Sixty pigs were individually assigned to five treatments for 34 d: NC (negative control, 1.2 mg/kg of deoxynivalenol); PC (positive control, 3.2 mg/kg of deoxynivalenol); CYC (PC + clay/yeast culture-based product, 0.2%); CYE (PC + clay/yeast cell wall/plant extracts/antioxidants-based product, 0.2%); and CYB (PC + clay/inactivated yeast/botanicals/antioxidants-based product, 0.2%). Blood and jejunal mucosa were sampled, and data were analyzed using Proc Mixed of SAS with pre-planned contrasts. Deoxynivalenol reduced the average daily gain (ADG) in phase 3. Pigs fed CYC had greater overall ADG, average daily feed intake during phase 3, and gain to feed ratio during phase 2 than PC. At d 14, deoxynivalenol reduced blood urea nitrogen/creatinine and tended to reduce blood urea nitrogen. Pigs fed CYB tended to have greater aspartate aminotransferase than PC. At d 34, pigs fed CYC and CYB tended to have lower serum creatine phosphokinase than PC. Pigs fed CYE had lower blood urea nitrogen/creatinine than PC. In jejunal mucosa, deoxynivalenol tended to increase malondialdehydes and decrease glutathione. Pigs fed CYE and CYB had lower malondialdehydes, pigs fed CYB had greater glutathione and tended to have lower immunoglobulin A than PC. Pigs fed CYC and CYE tended to have lower interleukin 8 than PC. In summary, deoxynivalenol challenge (1.2 vs. 3.2 mg/kg) mildly compromised growth performance and increased the oxidative stress of pigs. Mycotoxin detoxifiers could partially overcome deoxynivalenol toxicity enhancing liver health, whereas CYE and CYB reduced oxidative stress, and CYC and CYB reduced immune activation. In conclusion, yeast-based detoxifiers with functional components as clay/inactivated yeast/botanicals/antioxidants had increased detoxifying properties in newly-weaned pigs challenged with deoxynivalenol, potentially by enhancing adsorbability, immune function, gut health, and reducing oxidative stress. Full article
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Article
Repurposing Cancer Drugs Batimastat and Marimastat to Inhibit the Activity of a Group I Metalloprotease from the Venom of the Western Diamondback Rattlesnake, Crotalus atrox
Toxins 2020, 12(5), 309; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12050309 - 09 May 2020
Cited by 10
Abstract
Snakebite envenomation causes over 140,000 deaths every year, predominantly in developing countries. As a result, it is one of the most lethal neglected tropical diseases. It is associated with incredibly complex pathophysiology due to the vast number of unique toxins/proteins present in the [...] Read more.
Snakebite envenomation causes over 140,000 deaths every year, predominantly in developing countries. As a result, it is one of the most lethal neglected tropical diseases. It is associated with incredibly complex pathophysiology due to the vast number of unique toxins/proteins present in the venoms of diverse snake species found worldwide. Here, we report the purification and functional characteristics of a Group I (PI) metalloprotease (CAMP-2) from the venom of the western diamondback rattlesnake, Crotalus atrox. Its sensitivity to matrix metalloprotease inhibitors (batimastat and marimastat) was established using specific in vitro experiments and in silico molecular docking analysis. CAMP-2 shows high sequence homology to atroxase from the venom of Crotalus atrox and exhibits collagenolytic, fibrinogenolytic and mild haemolytic activities. It exerts a mild inhibitory effect on agonist-induced platelet aggregation in the absence of plasma proteins. Its collagenolytic activity is completely inhibited by batimastat and marimastat. Zinc chloride also inhibits the collagenolytic activity of CAMP-2 by around 75% at 50 μM, while it is partially potentiated by calcium chloride. Molecular docking studies have demonstrated that batimastat and marimastat are able to bind strongly to the active site residues of CAMP-2. This study demonstrates the impact of matrix metalloprotease inhibitors in the modulation of a purified, Group I metalloprotease activities in comparison to the whole venom. By improving our understanding of snake venom metalloproteases and their sensitivity to small molecule inhibitors, we can begin to develop novel and improved treatment strategies for snakebites. Full article
(This article belongs to the Special Issue Novel Strategies for the Diagnosis and Treatment of Snakebites)
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Article
Use of Mass Spectrometry to Determine the Diversity of Toxins Produced by Gambierdiscus and Fukuyoa Species from Balearic Islands and Crete (Mediterranean Sea) and the Canary Islands (Northeast Atlantic)
Toxins 2020, 12(5), 305; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12050305 - 07 May 2020
Cited by 8
Abstract
Over the last decade, knowledge has significantly increased on the taxonomic identity and distribution of dinoflagellates of the genera Gambierdiscus and Fukuyoa. Additionally, a number of hitherto unknown bioactive metabolites have been described, while the role of these compounds in ciguatera poisoning [...] Read more.
Over the last decade, knowledge has significantly increased on the taxonomic identity and distribution of dinoflagellates of the genera Gambierdiscus and Fukuyoa. Additionally, a number of hitherto unknown bioactive metabolites have been described, while the role of these compounds in ciguatera poisoning (CP) remains to be clarified. Ciguatoxins and maitotoxins are very toxic compounds produced by these dinoflagellates and have been described since the 1980s. Ciguatoxins are generally described as the main contributors to this food intoxication. Recent reports of CP in temperate waters of the Canary Islands (Spain) and the Madeira archipelago (Portugal) triggered the need for isolation and cultivation of dinoflagellates from these areas, and their taxonomic and toxicological characterization. Maitotoxins, and specifically maitotoxin-4, has been described as one of the most toxic compounds produced by these dinoflagellates (e.g., G. excentricus) in the Canary Islands. Thus, characterization of toxin profiles of Gambierdiscus species from adjacent regions appears critical. The combination of liquid chromatography coupled to either low- or high-resolution mass spectrometry allowed for characterization of several strains of Gambierdiscus and Fukuyoa from the Mediterranean Sea and the Canary Islands. Maitotoxin-3, two analogues tentatively identified as gambieric acid C and D, a putative gambierone analogue and a putative gambieroxide were detected in all G. australes strains from Menorca and Mallorca (Balearic Islands, Spain) while only maitotoxin-3 was present in an F. paulensis strain of the same region. An unidentified Gambierdiscus species (Gambierdiscus sp.2) from Crete (Greece) showed a different toxin profile, detecting both maitotoxin-3 and gambierone, while the availability of a G. excentricus strain from the Canary Islands (Spain) confirmed the presence of maitotoxin-4 in this species. Overall, this study shows that toxin profiles not only appear to be species-specific but probably also specific to larger geographic regions. Full article
(This article belongs to the Special Issue Ciguatoxins)
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Article
Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active on Voltage Gated Sodium Channels (VGSCs)
Toxins 2020, 12(5), 281; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12050281 - 27 Apr 2020
Cited by 7
Abstract
The neuroblastoma cell-based assay (CBA-N2a) is widely used for the detection of marine biotoxins in seafood products, yet a consensus protocol is still lacking. In this study, six key parameters of CBA-N2a were revisited: cell seeding densities, cell layer viability after 26 h [...] Read more.
The neuroblastoma cell-based assay (CBA-N2a) is widely used for the detection of marine biotoxins in seafood products, yet a consensus protocol is still lacking. In this study, six key parameters of CBA-N2a were revisited: cell seeding densities, cell layer viability after 26 h growth, MTT incubation time, Ouabain and Veratridine treatment and solvent and matrix effects. A step-by-step protocol was defined identifying five viability controls for the validation of CBA-N2a results. Specific detection of two voltage gated sodium channel activators, pacific ciguatoxin (P-CTX3C) and brevetoxin (PbTx3) and two inhibitors, saxitoxin (STX) and decarbamoylsaxitoxin (dc-STX) was achieved, with EC50 values of 1.7 ± 0.35 pg/mL, 5.8 ± 0.9 ng/mL, 3 ± 0.5 ng/mL and 15.8 ± 3 ng/mL, respectively. When applied to the detection of ciguatoxin (CTX)-like toxicity in fish samples, limit of detection (LOD) and limit of quantification (LOQ) values were 0.031 ± 0.008 and 0.064 ± 0.016 ng P-CTX3C eq/g of flesh, respectively. Intra and inter-assays comparisons of viability controls, LOD, LOQ and toxicity in fish samples gave coefficients of variation (CVs) ranging from 3% to 29%. This improved test adaptable to either high throughput screening or composite toxicity estimation is a useful starting point for a standardization of the CBA-N2a in the field of marine toxin detection. Full article
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Article
Molecular Identification and Mycotoxin Production by Alternaria Species Occurring on Durum Wheat, Showing Black Point Symptoms
Toxins 2020, 12(4), 275; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040275 - 23 Apr 2020
Cited by 7
Abstract
Black point is a fungal disease of wheat, mainly associated with mycotoxigenic Alternaria species. Affected wheat kernels are characterized by dark brown discolouration of the embryo region and reduction of grain quality. Potential risk is the possible accumulation of Alternaria mycotoxins, alternariol (AOH), [...] Read more.
Black point is a fungal disease of wheat, mainly associated with mycotoxigenic Alternaria species. Affected wheat kernels are characterized by dark brown discolouration of the embryo region and reduction of grain quality. Potential risk is the possible accumulation of Alternaria mycotoxins, alternariol (AOH), alternariol-monomethyl ether (AME), tenuazonic acid (TA), and altenuene (ALT), provided by haemato-toxic, genotoxic, and mutagenic activities. One hundred and twenty durum wheat samples belonging to 30 different genotypes grown in Bologna and Modena areas, in Italy, showing black point symptoms, were analyzed for Alternaria species and their mycotoxin contamination. Alternariol was selected as an indicator of the capability of the Alternaria species to produce mycotoxin in vivo in field conditions. The data showed that Alternaria species occurred in 118 out of 120 wheat kernels samples, with the incidence of infected kernels ranging between 1% and 26%. Moreover, AOH was detected by using a HPLC with a diode array detector (LC-DAD) in 98 out of 120 samples with values ranging between 24 and 262 µg Kg−1. Ninety-two Alternaria representative strains, previously identified morphologically, were identified at species/section level using gene sequencing, and therefore were analyzed for their mycotoxin profiles. Eighty-four strains, phylogenetically grouped in the Alternaria section, produced AOH, AME, and TA with values up to 8064, 14,341, and 3683 µg g−1, respectively, analyzed by using a LC-DAD. On the other hand, eight Alternaria strains, included in Infectoriae Section, showed a very low or no capability to produce mycotoxins. Full article
(This article belongs to the Special Issue Mycotoxins in Food: Origin and Management of Risk)
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Article
Oligomer Formation and Insecticidal Activity of Bacillus thuringiensis Vip3Aa Toxin
Toxins 2020, 12(4), 274; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040274 - 23 Apr 2020
Cited by 6
Abstract
Bacillus thuringiensis (Bt) Vip3A proteins are important insecticidal proteins used for control of lepidopteran insects. However, the mode of action of Vip3A toxin is still unclear. In this study, the amino acid residue S164 in Vip3Aa was identified to be critical for the [...] Read more.
Bacillus thuringiensis (Bt) Vip3A proteins are important insecticidal proteins used for control of lepidopteran insects. However, the mode of action of Vip3A toxin is still unclear. In this study, the amino acid residue S164 in Vip3Aa was identified to be critical for the toxicity in Spodoptera litura. Results from substitution mutations of the S164 indicate that the insecticidal activity of Vip3Aa correlated with the formation of a >240 kDa complex of the toxin upon proteolytic activation. The >240 kDa complex was found to be composed of the 19 kDa and the 65 kDa fragments of Vip3Aa. Substitution of the S164 in Vip3Aa protein with Ala or Pro resulted in loss of the >240 kDa complex and loss of toxicity in Spodoptera litura. In contrast, substitution of S164 with Thr did not affect the >240 kDa complex formation, and the toxicity of the mutant was only reduced by 35%. Therefore, the results from this study indicated that formation of the >240 kDa complex correlates with the toxicity of Vip3Aa in insects and the residue S164 is important for the formation of the complex. Full article
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Article
Critical Comparison of Analytical Performances of Two Immunoassay Methods for Rapid Detection of Aflatoxin M1 in Milk
Toxins 2020, 12(4), 270; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040270 - 22 Apr 2020
Cited by 6
Abstract
Aflatoxin B1 (AFB1) is a secondary metabolite produced by some Aspergillus spp. fungi affecting many crops and feed materials. Aflatoxin M1 (AFM1), the 4-hydroxylated metabolite of AFB1, is the main AFB1-related compound present [...] Read more.
Aflatoxin B1 (AFB1) is a secondary metabolite produced by some Aspergillus spp. fungi affecting many crops and feed materials. Aflatoxin M1 (AFM1), the 4-hydroxylated metabolite of AFB1, is the main AFB1-related compound present in milk, and it is categorized by the International Agency for Research on Cancer (IARC) as a “group 1 human carcinogen”. The aim of this work was to evaluate and compare the analytical performances of two commercial immunoassays widely applied for the detection of AFM1 in milk, namely strip test immunoassay and enzyme linked immunosorbent assay (ELISA). Assay validation included samples at AFM1 levels of 25, 50, 75 ng/kg and blank samples (AFM1 < 0.5 ng/kg). With respect to a screening target concentration (STC) of 50 ng/kg the two assays showed cut-off values of 37.7 ng/kg and 47.5 ng/kg for strip test and ELISA, respectively, a false suspect rate for blanks <0.1% (for both assays) and a false negative rate for samples containing AFM1 at levels higher than STC, of 0.4% (for both assays). The intermediate precision (RSDip) was <32% for the strip test and <15% for the ELISA. Method verification through long-term intra-laboratory quality control (QC) measurements confirmed the results from the validation study. Furthermore, a satisfactory correlation of the results obtained with both immunoassays and the AOAC Official Method 2000.08 was obtained for the analysis of cow milk samples naturally contaminated with AFM1 at levels within “not detected” (< 0.5 ng/kg) and 50 ng/kg. Finally, the extension of the scope of the strip test method to goat and sheep milk was evaluated by applying the experimental design foreseen in the EU regulation. Full article
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Communication
Liquid Chromatography Coupled to High-Resolution Mass Spectrometry for the Confirmation of Caribbean Ciguatoxin-1 as the Main Toxin Responsible for Ciguatera Poisoning Caused by Fish from European Atlantic Coasts
Toxins 2020, 12(4), 267; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040267 - 21 Apr 2020
Cited by 9
Abstract
Ciguatera poisoning (CP) is a common seafood intoxication mainly caused by the consumption of fish contaminated by ciguatoxins. Recent studies showed that Caribbean ciguatoxin-1 (C-CTX1) is the main toxin causing CP through fish caught in the Northeast Atlantic, e.g., Canary Islands (Spain) and [...] Read more.
Ciguatera poisoning (CP) is a common seafood intoxication mainly caused by the consumption of fish contaminated by ciguatoxins. Recent studies showed that Caribbean ciguatoxin-1 (C-CTX1) is the main toxin causing CP through fish caught in the Northeast Atlantic, e.g., Canary Islands (Spain) and Madeira (Portugal). The use of liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) combined with neuroblastoma cell assay (N2a) allowed the initial confirmation of the presence of C-CTX1 in contaminated fish samples from the abovementioned areas, nevertheless the lack of commercially available reference materials for these particular ciguatoxin (CTX) analogues has been a major limitation to progress research. The EuroCigua project allowed the preparation of C-CTX1 laboratory reference material (LRM) from fish species (Seriola fasciata) from the Madeira archipelago (Portugal). This reference material was used to implement a liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS) for the detection of C-CTX1, acquisition of full-scan as well as collision-induced mass spectra of this particular analogue. Fragmentation pathways were proposed based on fragments obtained. The optimized LC-HRMS method was then applied to confirm C-CTX1 in fish (Bodianus scrofa) caught in the Selvagens Islands (Portugal). Full article
(This article belongs to the Special Issue Ciguatoxins)
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Article
The Importance of Allelopathic Picocyanobacterium Synechococcus sp. on the Abundance, Biomass Formation, and Structure of Phytoplankton Assemblages in Three Freshwater Lakes
Toxins 2020, 12(4), 259; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040259 - 16 Apr 2020
Cited by 6
Abstract
The contribution of picocyanobacteria to summer phytoplankton blooms, accompanied by an ecological crisis, is a new phenomenon in Europe. This issue requires careful investigation. We studied allelopathic activity of freshwater picocyanobacterium Synechococcus sp. on phytoplankton assemblages from three freshwater lakes. In this study, [...] Read more.
The contribution of picocyanobacteria to summer phytoplankton blooms, accompanied by an ecological crisis, is a new phenomenon in Europe. This issue requires careful investigation. We studied allelopathic activity of freshwater picocyanobacterium Synechococcus sp. on phytoplankton assemblages from three freshwater lakes. In this study, the allelopathic activity of the Synechococcus sp. on the total abundance, biomass, as well as structure of the phytoplankton assemblages were investigated. Our results indicated that addition of exudates obtained from Synechococcus sp. affected the number of cells and biomass of the phytoplankton communities; the degree of inhibition or stimulation was different for each species, causing a change in the phytoplankton abundance and dominance during the experiment. We observed that some group of organisms (especially cyanobacteria from the genus Aphanothece, Limnothrix, Microcystis, and Synechococcus) showed tolerance for allelopathic compounds produced and released by Synechococcus sp. It is also worth noting that in some samples, Bacillariophyceae (e.g., Amphora pediculus, Navicula pygmaea, and Nitzschia paleacea) were completely eliminated in the experimental treatments, while present in the controls. This work demonstrated that the allelopathic activity exhibited by the Synechococcus sp. is probably one of the major competitive strategies affecting some of the coexisting phytoplankton species in freshwater ecosystems. To our best knowledge this is the first report of the allelopathic activity of Synechococcus sp. in the freshwater reservoirs, and one of the few published works showing allelopathic properties of freshwater picocyanobacteria on coexisting phytoplankton species. Full article
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Article
CRISPR-Mediated Knockout of the ABCC2 Gene in Ostrinia furnacalis Confers High-Level Resistance to the Bacillus thuringiensis Cry1Fa Toxin
Toxins 2020, 12(4), 246; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040246 - 11 Apr 2020
Cited by 10
Abstract
The adoption of transgenic crops expressing Bacillus thuringiensis (Bt) insecticidal crystalline (Cry) proteins has reduced insecticide application, increased yields, and contributed to food safety worldwide. However, the efficacy of transgenic Bt crops is put at risk by the adaptive resistance evolution of target [...] Read more.
The adoption of transgenic crops expressing Bacillus thuringiensis (Bt) insecticidal crystalline (Cry) proteins has reduced insecticide application, increased yields, and contributed to food safety worldwide. However, the efficacy of transgenic Bt crops is put at risk by the adaptive resistance evolution of target pests. Previous studies indicate that resistance to Bacillus thuringiensis Cry1A and Cry1F toxins was genetically linked with mutations of ATP-binding cassette (ABC) transporter subfamily C gene ABCC2 in at least seven lepidopteran insects. Several strains selected in the laboratory of the Asian corn borer, Ostrinia furnacalis, a destructive pest of corn in Asian Western Pacific countries, developed high levels of resistance to Cry1A and Cry1F toxins. The causality between the O. furnacalis ABCC2 (OfABCC2) gene and resistance to Cry1A and Cry1F toxins remains unknown. Here, we successfully generated a homozygous strain (OfC2-KO) of O. furnacalis with an 8-bp deletion mutation of ABCC2 by the CRISPR/Cas9 approach. The 8-bp deletion mutation results in a frame shift in the open reading frame of transcripts, which produced a predicted protein truncated in the TM4-TM5 loop region. The knockout strain OfC2-KO showed much more than a 300-fold resistance to Cry1Fa, and low levels of resistance to Cry1Ab and Cry1Ac (<10-fold), but no significant effects on the toxicities of Cry1Aa and two chemical insecticides (abamectin and chlorantraniliprole), compared to the background NJ-S strain. Furthermore, we found that the Cry1Fa resistance was autosomal, recessive, and significantly linked with the 8-bp deletion mutation of OfABCC2 in the OfC2-KO strain. In conclusion, in vivo functional investigation demonstrates the causality of the OfABCC2 truncating mutation with high-level resistance to the Cry1Fa toxin in O. furnacalis. Our results suggest that the OfABCC2 protein might be a functional receptor for Cry1Fa and reinforces the association of this gene to the mode of action of the Cry1Fa toxin. Full article
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Article
Phylogenomic Analysis of Secondary Metabolism in the Toxic Cyanobacterial Genera Anabaena, Dolichospermum and Aphanizomenon
Toxins 2020, 12(4), 248; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040248 - 11 Apr 2020
Cited by 8
Abstract
Cyanobacteria produce an array of toxins that pose serious health risks to humans and animals. The closely related diazotrophic genera, Anabaena, Dolichospermum and Aphanizomenon, frequently form poisonous blooms in lakes and brackish waters around the world. These genera form a complex [...] Read more.
Cyanobacteria produce an array of toxins that pose serious health risks to humans and animals. The closely related diazotrophic genera, Anabaena, Dolichospermum and Aphanizomenon, frequently form poisonous blooms in lakes and brackish waters around the world. These genera form a complex now termed the Anabaena, Dolichospermum and Aphanizomenon (ADA) clade and produce a greater array of toxins than any other cyanobacteria group. However, taxonomic confusion masks the distribution of toxin biosynthetic pathways in cyanobacteria. Here we obtained 11 new draft genomes to improve the understanding of toxin production in these genera. Comparison of secondary metabolite pathways in all available 31 genomes for these three genera suggests that the ability to produce microcystin, anatoxin-a, and saxitoxin is associated with specific subgroups. Each toxin gene cluster was concentrated or even limited to a certain subgroup within the ADA clade. Our results indicate that members of the ADA clade encode a variety of secondary metabolites following the phylogenetic clustering of constituent species. The newly sequenced members of the ADA clade show that phylogenetic separation of planktonic Dolichospermum and benthic Anabaena is not complete. This underscores the importance of taxonomic revision of Anabaena, Dolichospermum and Aphanizomenon genera to reflect current phylogenomic understanding. Full article
(This article belongs to the Section Marine and Freshwater Toxins)
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Article
Protein-Bound Uremic Toxins in Hemodialysis Patients Relate to Residual Kidney Function, Are Not Influenced by Convective Transport, and Do Not Relate to Outcome
Toxins 2020, 12(4), 234; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040234 - 07 Apr 2020
Cited by 10
Abstract
Protein-bound uremic toxins (PBUTs) are predominantly excreted by renal tubular secretion and hardly removed by traditional hemodialysis (HD). Accumulation of PBUTs is proposed to contribute to the increased morbidity and mortality of patients with end-stage kidney disease (ESKD). Preserved PBUT excretion in patients [...] Read more.
Protein-bound uremic toxins (PBUTs) are predominantly excreted by renal tubular secretion and hardly removed by traditional hemodialysis (HD). Accumulation of PBUTs is proposed to contribute to the increased morbidity and mortality of patients with end-stage kidney disease (ESKD). Preserved PBUT excretion in patients with residual kidney function (RKF) and/or increased PBUT clearance with improved dialysis techniques might improve the prognosis of patients with ESKD. The aims of this study are to explore determinants of PBUTs in HD patients, and investigate whether hemodiafiltration (HDF) lowers PBUT plasma concentrations, and whether PBUTs are related to the outcome. Predialysis total plasma concentrations of kynurenine, kynurenic acid, indoxyl sulfate, indole-3-acetic acid, p-cresyl sulfate, p-cresyl glucuronide, and hippuric acid were measured by UHPLC-MS at baseline and after 6 months of follow-up in the first 80 patients participating in the CONvective TRAnsport Study (CONTRAST), a randomized controlled trial that compared the effects of online HDF versus low-flux HD on all-cause mortality and new cardiovascular events. RKF was inversely related to kynurenic acid (p < 0.001), indoxyl sulfate (p = 0.001), indole-3-acetic acid (p = 0.024), p-cresyl glucuronide (p = 0.004) and hippuric acid (p < 0.001) plasma concentrations. Only indoxyl sulfate decreased by 8.0% (−15.3 to 34.6) in patients treated with HDF and increased by 11.9% (−15.4 to 31.9) in HD patients after 6 months of follow-up (HDF vs. HD: p = 0.045). No independent associations were found between PBUT plasma concentrations and either risk of all-cause mortality or new cardiovascular events. In summary, in the current population, RKF is an important determinant of PBUT plasma concentrations in HD patients. The addition of convective transport did not consistently decrease PBUT plasma concentrations and no relation was found between PBUTs and cardiovascular endpoints. Full article
(This article belongs to the Section Uremic Toxins)
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Article
Cylindrospermopsin- and Deoxycylindrospermopsin-Producing Raphidiopsis raciborskii and Microcystin-Producing Microcystis spp. in Meiktila Lake, Myanmar
Toxins 2020, 12(4), 232; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040232 - 07 Apr 2020
Cited by 7
Abstract
Meiktila Lake is a shallow reservoir located close to Meiktila city in central Myanmar. Its water is used for irrigation, domestic purposes and drinking water. No detailed study of the presence of cyanobacteria and their potential toxin production has been conducted so far. [...] Read more.
Meiktila Lake is a shallow reservoir located close to Meiktila city in central Myanmar. Its water is used for irrigation, domestic purposes and drinking water. No detailed study of the presence of cyanobacteria and their potential toxin production has been conducted so far. To ascertain the cyanobacterial composition and presence of cyanobacterial toxins in Meiktila Lake, water samples were collected in March and November 2017 and investigated for physico-chemical and biological parameters. Phytoplankton composition and biomass determination revealed that most of the samples were dominated by the cyanobacterium Raphidiopsis raciborskii. In a polyphasic approach, seven isolated cyanobacterial strains were classified morphologically and phylogenetically as R. raciborskii, and Microcystis spp. and tested for microcystins (MCs), cylindrospermopsins (CYNs), saxitoxins and anatoxins by enzyme-linked immunosorbent assay (ELISA) and liquid chromatography–mass spectrometry (LC–MS). ELISA and LC–MS analyses confirmed CYNs in three of the five Raphidiopsis strains between 1.8 and 9.8 μg mg−1 fresh weight. Both Microcystis strains produced MCs, one strain 52 congeners and the other strain 20 congeners, including 22 previously unreported variants. Due to the presence of CYN- and MC-producing cyanobacteria, harmful effects on humans, domestic and wild animals cannot be excluded in Meiktila Lake. Full article
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Article
Effects of Nutrient Limitation on the Synthesis of N-Rich Phytoplankton Toxins: A Meta-Analysis
Toxins 2020, 12(4), 221; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040221 - 01 Apr 2020
Cited by 10
Abstract
Eutrophication has played a major role in the worldwide increase of harmful algal blooms (HABs). Higher input of key nutrients, such as nitrogen (N) and phosphorus (P), can stimulate the growth of harmful algal species in freshwater, estuarine, and coastal marine ecosystems. Some [...] Read more.
Eutrophication has played a major role in the worldwide increase of harmful algal blooms (HABs). Higher input of key nutrients, such as nitrogen (N) and phosphorus (P), can stimulate the growth of harmful algal species in freshwater, estuarine, and coastal marine ecosystems. Some HAB-forming taxa, particularly several cyanobacteria and dinoflagellate species, are harmful through the production of N-rich toxins that have detrimental effects on the environment and human health. Here, we test how changes in nutrient availability affect N-rich toxin synthesis in cyanobacteria and dinoflagellates using a meta-analysis approach. Overall, N-rich toxin content showed an increase with P limitation, while it tended to decrease with N limitation, but we also observed substantial variation in responses both within and across genera and toxin groups. For instance, in response to N limitation, microcystin content varied from a 297% decrease up to a 273% increase, and paralytic shellfish poisoning (PSP) toxin content varied from a 204% decrease to an 82% increase. Cylindrospermopsin, produced by N2-fixing cyanobacteria, showed no clear direction in response to nutrient limitation, and cellular contents of this compound may thus vary independently of nutrient fluctuations. Our results confirm earlier reported stoichiometric regulation of N-rich phytoplankton toxins, showing increased toxin content with an increase in cellular N:P ratios, and vice versa. Thus, changes in N-rich toxin content largely follow the changes in relative cellular N content. Consequently, although nutrient limitation may limit bloom biomass and thereby bloom toxicity, our results warn that P limitation can cause accumulation of cellular toxins and thus lead to unexpected increases in bloom toxicity. Full article
(This article belongs to the Special Issue Environmental Drivers of Algal and Cyanobacterial Toxin Dynamics)
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Article
Individual and Combined Effect of Zearalenone Derivates and Beauvericin Mycotoxins on SH-SY5Y Cells
Toxins 2020, 12(4), 212; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12040212 - 27 Mar 2020
Cited by 15
Abstract
Beauvericin (BEA) and zearalenone derivatives, α-zearalenol (α-ZEL), and β-zearalenol (β-ZEL), are produced by several Fusarium species. Considering the impact of various mycotoxins on human’s health, this study determined and evaluated the cytotoxic effect of individual, binary, and tertiary mycotoxin treatments consisting of α-ZEL, [...] Read more.
Beauvericin (BEA) and zearalenone derivatives, α-zearalenol (α-ZEL), and β-zearalenol (β-ZEL), are produced by several Fusarium species. Considering the impact of various mycotoxins on human’s health, this study determined and evaluated the cytotoxic effect of individual, binary, and tertiary mycotoxin treatments consisting of α-ZEL, β-ZEL, and BEA at different concentrations over 24, 48, and 72 h on SH-SY5Y neuronal cells, by using the MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetrazoliumbromide). Subsequently, the isobologram method was applied to elucidate if the mixtures produced synergism, antagonism, or additive effects. Ultimately, we determined the amount of mycotoxin recovered from the media after treatment using liquid chromatography coupled with electrospray ionization–quadrupole time-of-flight mass spectrometry (LC–ESI–qTOF-MS). The IC50 values detected at all assayed times ranged from 95 to 0.2 μM for the individual treatments. The result indicated that β-ZEL was the most cytotoxic mycotoxin when tested individually. The major effect detected for all combinations assayed was synergism. Among the combinations assayed, α-ZEL + β-ZEL + BEA and α-ZEL + BEA presented the highest cytotoxic potential with respect to the IC value. In individual treatment, α-ZEL was the most recovered mycotoxin; while, this was observed for BEA in binary combination α-ZEL + BEA. Full article
(This article belongs to the Special Issue Mycotoxins Study: Toxicology, Identification and Control)
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Article
No More Tears: Mining Sequencing Data for Novel Bt Cry Toxins with CryProcessor
Toxins 2020, 12(3), 204; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030204 - 23 Mar 2020
Cited by 7
Abstract
Bacillus thuringiensis (Bt) is a natural pathogen of insects and some other groups of invertebrates that produces three-domain Cry (3d-Cry) toxins, which are highly host-specific pesticidal proteins. These proteins represent the most commonly used bioinsecticides in the world and are used [...] Read more.
Bacillus thuringiensis (Bt) is a natural pathogen of insects and some other groups of invertebrates that produces three-domain Cry (3d-Cry) toxins, which are highly host-specific pesticidal proteins. These proteins represent the most commonly used bioinsecticides in the world and are used for commercial purposes on the market of insecticides, being convergent with the paradigm of sustainable growth and ecological development. Emerging resistance to known toxins in pests stresses the need to expand the list of known toxins to broaden the horizons of insecticidal approaches. For this purpose, we have elaborated a fast and user-friendly tool called CryProcessor, which allows productive and precise mining of 3d-Cry toxins. The only existing tool for mining Cry toxins, called a BtToxin_scanner, has significant limitations such as limited query size, lack of accuracy and an outdated database. In order to find a proper solution to these problems, we have developed a robust pipeline, capable of precise 3d-Cry toxin mining. The unique feature of the pipeline is the ability to search for Cry toxins sequences directly on assembly graphs, providing an opportunity to analyze raw sequencing data and overcoming the problem of fragmented assemblies. Moreover, CryProcessor is able to predict precisely the domain layout in arbitrary sequences, allowing the retrieval of sequences of definite domains beyond the bounds of a limited number of toxins presented in CryGetter. Our algorithm has shown efficiency in all its work modes and outperformed its analogues on large amounts of data. Here, we describe its main features and provide information on its benchmarking against existing analogues. CryProcessor is a novel, fast, convenient, open source (https://github.com/lab7arriam/cry_processor), platform-independent, and precise instrument with a console version and elaborated web interface (https://lab7.arriam.ru/tools/cry_processor). Its major merits could make it possible to carry out massive screening for novel 3d-Cry toxins and obtain sequences of specific domains for further comprehensive in silico experiments in constructing artificial toxins. Full article
(This article belongs to the Section Bacterial Toxins)
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Article
An Appetite for Destruction: Detecting Prey-Selective Binding of α-Neurotoxins in the Venom of Afro-Asian Elapids
Toxins 2020, 12(3), 205; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030205 - 23 Mar 2020
Cited by 14
Abstract
Prey-selective venoms and toxins have been documented across only a few species of snakes. The lack of research in this area has been due to the absence of suitably flexible testing platforms. In order to test more species for prey specificity of their [...] Read more.
Prey-selective venoms and toxins have been documented across only a few species of snakes. The lack of research in this area has been due to the absence of suitably flexible testing platforms. In order to test more species for prey specificity of their venom, we used an innovative taxonomically flexible, high-throughput biolayer interferometry approach to ascertain the relative binding of 29 α-neurotoxic venoms from African and Asian elapid representatives (26 Naja spp., Aspidelaps scutatus, Elapsoidea boulengeri, and four locales of Ophiophagus hannah) to the alpha-1 nicotinic acetylcholine receptor orthosteric (active) site for amphibian, lizard, snake, bird, and rodent targets. Our results detected prey-selective, intraspecific, and geographical differences of α-neurotoxic binding. The results also suggest that crude venom that shows prey selectivity is likely driven by the proportions of prey-specific α-neurotoxins with differential selectivity within the crude venom. Our results also suggest that since the α-neurotoxic prey targeting does not always account for the full dietary breadth of a species, other toxin classes with a different pathophysiological function likely play an equally important role in prey immobilisation of the crude venom depending on the prey type envenomated. The use of this innovative and taxonomically flexible diverse assay in functional venom testing can be key in attempting to understanding the evolution and ecology of α-neurotoxic snake venoms, as well as opening up biochemical and pharmacological avenues to explore other venom effects. Full article
(This article belongs to the Section Animal Venoms)
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Article
Fangs for the Memories? A Survey of Pain in Snakebite Patients Does Not Support a Strong Role for Defense in the Evolution of Snake Venom Composition
Toxins 2020, 12(3), 201; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030201 - 22 Mar 2020
Cited by 7
Abstract
Animals use venoms for multiple purposes, most prominently for prey acquisition and self-defense. In snakes, venom composition often evolves as a result of selection for optimization for local diet. However, whether selection for a defensive function has also played a role in driving [...] Read more.
Animals use venoms for multiple purposes, most prominently for prey acquisition and self-defense. In snakes, venom composition often evolves as a result of selection for optimization for local diet. However, whether selection for a defensive function has also played a role in driving the evolution of venom composition has remained largely unstudied. Here, we use an online survey of snakebite victims to test a key prediction of a defensive function, that envenoming should result in the rapid onset of severe pain. From the analysis of 584 snakebite reports, involving 192 species of venomous snake, we find that the vast majority of bites do not result in severe early pain. Phylogenetic comparative analysis shows that where early pain after a bite evolves, it is often lost rapidly. Our results, therefore, do not support the hypothesis that natural selection for antipredator defense played an important role in the origin of venom or front-fanged delivery systems in general, although there may be intriguing exceptions to this rule. Full article
(This article belongs to the Special Issue Evolutionary Ecology of Venom)
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Article
Natural Occurrence of Deoxynivalenol and Its Acetylated Derivatives in Chinese Maize and Wheat Collected in 2017
Toxins 2020, 12(3), 200; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030200 - 22 Mar 2020
Cited by 9
Abstract
Deoxynivalenol (DON), along with 3-acetyl-deoxynivalenol (3-ADON) and 15-acetyl-deoxynivalenol (15-ADON), occur in grains and cereal products and is often hazardous to humans and livestock. In this study, 579 wheat samples and 606 maize samples intended for consumption were collected from China in 2017 and [...] Read more.
Deoxynivalenol (DON), along with 3-acetyl-deoxynivalenol (3-ADON) and 15-acetyl-deoxynivalenol (15-ADON), occur in grains and cereal products and is often hazardous to humans and livestock. In this study, 579 wheat samples and 606 maize samples intended for consumption were collected from China in 2017 and analyzed to determine the co-occurrence of type-B trichothecenes (DON, 3-ADON, and 15-ADON). All the wheat samples tested positive for DON, while 99.83% of the maize samples were DON-positive with mean DON concentrations of 165.87 and 175.30 μg/kg, respectively. Per the Chinese standard limits for DON, 3.63% of wheat and 2.97% of the maize samples were above the maximum limit of 1000 μg/kg. The DON derivatives (3-ADON and 15-ADON) were less frequently found and were present at lower levels than DON in wheat. 3-ADON and 15-ADON had incidences of 13.53% and 76.40%, respectively, in maize. By analyzing the distribution ratio of DON and its derivatives in wheat and maize, DON (95.51%) was the predominant toxin detected in wheat samples, followed by 3.97% for the combination of DON + 3-ADON, while DON + 3-ADON + 15-ADON and DON + 15-ADON were only found in 0.17% and 0.35% of wheat samples, respectively. Additionally, a large amount of the maize samples were contaminated with DON + 15-ADON (64.19%) and DON (22.11%). The samples with a combination of DON + 3-ADON and DON + 3-ADON + 15-ADON accounted for 1.32% and 12.21%, respectively. Only one maize sample did not contain all three mycotoxins. Our study shows the necessity of raising awareness of the co-occurrence of mycotoxin contamination in grains from China to protect consumers from the risk of exposure to DON and its derivatives. Full article
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Article
Multi-Mycotoxin Occurrence and Exposure Assessment Approach in Foodstuffs from Algeria
Toxins 2020, 12(3), 194; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030194 - 19 Mar 2020
Cited by 10
Abstract
A survey on 120 cereal samples (barley, maize, rice and wheat) from Algerian markets has been carried out to evaluate the presence of 15 mycotoxins (ochratoxin A, deoxynivalenol, fumonisin B1 and B2, T-2 and HT-2 toxins, zearalenone, fusarenon X, citrinin, sterigmatocystin, enniatins A, [...] Read more.
A survey on 120 cereal samples (barley, maize, rice and wheat) from Algerian markets has been carried out to evaluate the presence of 15 mycotoxins (ochratoxin A, deoxynivalenol, fumonisin B1 and B2, T-2 and HT-2 toxins, zearalenone, fusarenon X, citrinin, sterigmatocystin, enniatins A, A1, B and B1, and beauvericin). With this purpose, a QuEChERS-based extraction and ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) were used. Analytical results showed that 78 cereal samples (65%) were contaminated with at least one toxin, while 50% were contaminated with three to nine mycotoxins. T-2 toxin, citrinin, beauvericin and deoxynivalenol were the most commonly found mycotoxins (frequency of 50%, 41.6%, 40.8% and 33.3%, respectively). Fumonisins (B1 + B2), enniatins B and B1, deoxynivalenol and zearalenone registered high concentrations (289–48878 µg/kg, 1.2–5288 µg/kg, 15–4569 µg/kg, 48–2055 µg/kg and 10.4–579 µg/kg, respectively). Furthermore, concentrations higher than those allowed by the European Union (EU) were observed in 21, 8 and 1 samples for fumonisins, zearalenone and deoxinivalenol, respectively. As a conclusion, the high levels of fumonisins (B1 + B2) in maize and deoxynivalenol, zearalenone and HT-2 + T-2 toxins in wheat, represent a health risk for the average adult consumer in Algeria. These results pointed out the necessity of a consistent control and the definition of maximum allowed levels for mycotoxins in Algerian foodstuffs. Full article
(This article belongs to the Special Issue Application of LC-MS/MS in the Mycotoxins Studies)
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Article
Deoxynivalenol Induces Inflammation in IPEC-J2 Cells by Activating P38 Mapk And Erk1/2
Toxins 2020, 12(3), 180; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030180 - 13 Mar 2020
Cited by 6
Abstract
Fusarium-derived mycotoxin deoxynivalenol (DON) usually induces diarrhea, vomiting and gastrointestinal inflammation. We studied the cytotoxic effect of DON on porcine small intestinal epithelium using the intestinal porcine epithelial cell line IPEC-J2. We screened out differentially expressed genes (DEGs) using RNA-seq and identified 320 [...] Read more.
Fusarium-derived mycotoxin deoxynivalenol (DON) usually induces diarrhea, vomiting and gastrointestinal inflammation. We studied the cytotoxic effect of DON on porcine small intestinal epithelium using the intestinal porcine epithelial cell line IPEC-J2. We screened out differentially expressed genes (DEGs) using RNA-seq and identified 320 upregulated genes and 160 downregulated genes. The enrichment pathways of these DEGs focused on immune-related pathways. DON induced proinflammatory gene expression, including cytokines, chemokines and other inflammation-related genes. DON increased IL1A, IL6 and TNF-α release and DON activated the phosphorylation of extracellular signal-regulated kinase-1 and-2 (ERK1/2), JUN N-terminal kinase (JNK) and p38 MAPK. A p38 inhibitor attenuated DON-induced IL6, TNF-α, CXCL2, CXCL8, IL12A, IL1A, CCL20, CCL4 and IL15 production, while an ERK1/2 inhibitor had only a small inhibitory effect on IL15 and IL6. An inhibitor of p38 MAPK decreased the release of IL1A, IL6 and TNF-α and an inhibitor of ERK1/2 partly attenuated protein levels of IL6. These data demonstrate that DON induces proinflammatory factor production in IPEC-J2 cells by activating p38 and ERK1/2. Full article
(This article belongs to the Special Issue Toxicological Effects of Mycotoxins on Target Cells)
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Article
Efficacy of Divinylbenzenic Resin in Removing Indoxyl Sulfate and P-cresol Sulfate in Hemodialysis Patients: Results from an In Vitro Study and an In Vivo Pilot Trial (xuanro4-Nature 3.2)
Toxins 2020, 12(3), 170; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030170 - 10 Mar 2020
Cited by 9
Abstract
High serum levels of microbiota-derived uremic toxins, indoxyl sulfate (IS) and p-cresyl sulfate (PCS), are associated with chronic kidney disease (CKD) progression and cardiovascular complications. IS and PCS cannot be efficiently removed by conventional hemodialysis (HD), due to their high binding affinity for [...] Read more.
High serum levels of microbiota-derived uremic toxins, indoxyl sulfate (IS) and p-cresyl sulfate (PCS), are associated with chronic kidney disease (CKD) progression and cardiovascular complications. IS and PCS cannot be efficiently removed by conventional hemodialysis (HD), due to their high binding affinity for albumin. This study evaluates the efficacy of a divinylbenzene-polyvinylpyrrolidone (DVB-PVP) cartridge and a synbiotic to reduce uremic toxins in HD patients. First, the in vitro efficacy of DVB-PVP in adsorbing IS and PCS was evaluated. Second, a randomized, placebo-controlled pilot study in HD patients was carried out to establish whether the administration of a synbiotic, either individually and in association with DVB-PVP-HD, could reduce the production of uremic toxins. In vitro data showed that DVB-PVP resin removed a mean of 56% PCS and around 54% IS, after 6 h of perfusion. While, in the in vivo study, the DVB-PVP cartridge showed its adsorbing efficacy only for IS plasma levels. The combination of synbiotic treatment with DVB-PVP HD decreased IS and PCS both at pre- and post-dialysis levels. In conclusion, this study provides the first line of evidence on the synergistic action of gut microbiota modulation and an innovative absorption-based approach in HD patients, aimed at reducing plasma levels of IS and PCS. Full article
(This article belongs to the Section Uremic Toxins)
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Article
Biological Control of Aflatoxin in Maize Grown in Serbia
Toxins 2020, 12(3), 162; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030162 - 05 Mar 2020
Cited by 16
Abstract
Aspergillus flavus is the main producer of aflatoxin B1, one of the most toxic contaminants of food and feed. With global warming, climate conditions have become favourable for aflatoxin contamination of agricultural products in several European countries, including Serbia. The infection of maize [...] Read more.
Aspergillus flavus is the main producer of aflatoxin B1, one of the most toxic contaminants of food and feed. With global warming, climate conditions have become favourable for aflatoxin contamination of agricultural products in several European countries, including Serbia. The infection of maize with A. flavus, and aflatoxin synthesis can be controlled and reduced by application of a biocontrol product based on non-toxigenic strains of A. flavus. Biological control relies on competition between atoxigenic and toxigenic strains. This is the most commonly used biological control mechanism of aflatoxin contamination in maize in countries where aflatoxins pose a significant threat. Mytoolbox Af01, a native atoxigenic A. flavus strain, was obtained from maize grown in Serbia and used to produce a biocontrol product that was applied in irrigated and non-irrigated Serbian fields during 2016 and 2017. The application of this biocontrol product reduced aflatoxin levels in maize kernels (51–83%). The biocontrol treatment had a highly significant effect of reducing total aflatoxin contamination by 73%. This study showed that aflatoxin contamination control in Serbian maize can be achieved through biological control methods using atoxigenic A. flavus strains. Full article
(This article belongs to the Special Issue Biocontrol Agents and Natural Compounds against Mycotoxinogenic Fungi)
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Article
Occurrence and Risk Assessment of Pyrrolizidine Alkaloids in Spices and Culinary Herbs from Various Geographical Origins
Toxins 2020, 12(3), 155; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030155 - 01 Mar 2020
Cited by 12
Abstract
Pyrrolizidine alkaloids (PA) and their N-oxides (PANO) are a group of toxic secondary plant metabolites occurring predominantly as contaminants in (herbal) teas, honeys and food supplements, as well as in spices and culinary herbs. Depending on the botanical origin of the contaminating [...] Read more.
Pyrrolizidine alkaloids (PA) and their N-oxides (PANO) are a group of toxic secondary plant metabolites occurring predominantly as contaminants in (herbal) teas, honeys and food supplements, as well as in spices and culinary herbs. Depending on the botanical origin of the contaminating plant, the pattern of PA/PANO can strongly vary within a sample. The current study aimed to broaden the existing data on the occurrence of PA/PANO in spices and culinary herbs. For this, 305 authentic samples covering 15 different matrices mainly harvested in 2016 or 2017 and originating from 36 countries were investigated for the presence of 44 PA/PANO. Fifty-eight percent of the samples contained at least one PA/PANO. The average sum content over all samples was 323 µg/kg (median of 0.9 µg/kg, 95% percentile of 665 µg/kg). The highest amount of 24.6 mg/kg was detected in an oregano sample. Additionally, conspicuous analyte patterns were discovered in samples from similar cultivation regions, indicating related botanical sources of PA/PANO contaminations. Particularly, oregano and cumin from Turkey often contained high amounts of PA/PANO. The results were used to assess the acute and chronic health risks related to PA/PANO intake via spices and culinary herbs, indicating a potential health risk in particular for adults and children with high consumption or when considering worst-case contamination scenarios of a sum content of 5500 µg/kg. Full article
(This article belongs to the Section Plant Toxins)
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Article
Cyanotoxins Occurrence in Portugal: A New Report on Their Recent Multiplication
Toxins 2020, 12(3), 154; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030154 - 29 Feb 2020
Cited by 6
Abstract
Historical reports show that in Portugal, cyanotoxins reports were mainly in the Center (cylindrospermopsins) and South (cylindrospermopsins, saxitoxins) regions of the country apart from the well distributed microcystins. Therefore, in our study, seven freshwater ecosystems located in the North and Center Regions of [...] Read more.
Historical reports show that in Portugal, cyanotoxins reports were mainly in the Center (cylindrospermopsins) and South (cylindrospermopsins, saxitoxins) regions of the country apart from the well distributed microcystins. Therefore, in our study, seven freshwater ecosystems located in the North and Center Regions of Portugal were screened between April and September of 2017 for the main cyanotoxins (microcystins, cylindrospermopsins, anatoxin-a, and saxitoxins) by a two methods approach that combined the application of molecular (PCR) and immunological (ELISA) assays. Results from our survey reveal that both methods revealed the presence of all main cyanotoxins. ELISA results showed that 48% of the samples were above (1.6–18.8 μg/L) the guideline value established for microcystins (1 μg/L), while in the remaining cyanotoxins, 33% of the samples were above (1.1–6.8 μg/L) the guideline value established for anatoxin–a (1 μg/L). Further, for saxitoxins, only one sample gave a value above (4.3 μg/L) the guideline (3 μg/L) and this corresponded to a North Region ecosystem. In the cytotoxin cylindrospermopsins, none of the samples were above the guideline established value (1 μg/L). This study will improve the risk assessment strategy in Portugal, as well as advance water quality and water management. Full article
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Article
In Silico and In Vitro Studies of Mycotoxins and Their Cocktails; Their Toxicity and Its Mitigation by Silibinin Pre-Treatment
Toxins 2020, 12(3), 148; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030148 - 28 Feb 2020
Cited by 11
Abstract
Mycotoxins found in randomly selected commercial milk thistle dietary supplement were evaluated for their toxicity in silico and in vitro. Using in silico methods, the basic physicochemical, pharmacological, and toxicological properties of the mycotoxins were predicted using ACD/Percepta. The in vitro cytotoxicity of [...] Read more.
Mycotoxins found in randomly selected commercial milk thistle dietary supplement were evaluated for their toxicity in silico and in vitro. Using in silico methods, the basic physicochemical, pharmacological, and toxicological properties of the mycotoxins were predicted using ACD/Percepta. The in vitro cytotoxicity of individual mycotoxins was determined in mouse macrophage (RAW 264.7), human hepatoblastoma (HepG2), and human embryonic kidney (HEK 293T) cells. In addition, we studied the bioavailability potential of mycotoxins and silibinin utilizing an in vitro transwell system with differentiated human colon adenocarcinoma cells (Caco-2) simulating mycotoxin transfer through the intestinal epithelial barrier. The IC50 values for individual mycotoxins in studied cells were in the biologically relevant ranges as follows: 3.57–13.37 nM (T-2 toxin), 5.07–47.44 nM (HT-2 toxin), 3.66–17.74 nM (diacetoxyscirpenol). Furthermore, no acute toxicity was obtained for deoxynivalenol, beauvericin, zearalenone, enniatinENN-A, enniatin-A1, enniatin-B, enniatin-B1, alternariol, alternariol-9-methyl ether, tentoxin, and mycophenolic acid up to the 50 nM concentration. The acute toxicity of these mycotoxins in binary combinations exhibited antagonistic effects in the combinations of T-2 with DON, ENN-A1, or ENN-B, while the rest showed synergistic or additive effects. Silibinin had a significant protective effect against both the cytotoxicity of three mycotoxins (T-2 toxin, HT-2 toxin, DAS) and genotoxicity of AME, AOH, DON, and ENNs on HEK 293T. The bioavailability results confirmed that AME, DAS, ENN-B, TEN, T-2, and silibinin are transported through the epithelial cell layer and further metabolized. The bioavailability of silibinin is very similar to mycotoxins poor penetration. Full article
(This article belongs to the Special Issue Toxicological Effects of Mycotoxins on Target Cells)
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Article
Variation of Fusarium Free, Masked, and Emerging Mycotoxin Metabolites in Maize from Agriculture Regions of South Africa
Toxins 2020, 12(3), 149; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030149 - 28 Feb 2020
Cited by 12
Abstract
The presence of mycotoxins in cereal grain is a very important food safety issue with the occurrence of masked mycotoxins extensively investigated in recent years. This study investigated the variation of different Fusarium metabolites (including the related regulated, masked, and emerging mycotoxin) in [...] Read more.
The presence of mycotoxins in cereal grain is a very important food safety issue with the occurrence of masked mycotoxins extensively investigated in recent years. This study investigated the variation of different Fusarium metabolites (including the related regulated, masked, and emerging mycotoxin) in maize from various agriculture regions of South Africa. The relationship between the maize producing regions, the maize type, as well as the mycotoxins was established. A total of 123 maize samples was analyzed by a LC-MS/MS multi-mycotoxin method. The results revealed that all maize types exhibited a mixture of free, masked, and emerging mycotoxins contamination across the regions with an average of 5 and up to 24 out of 42 investigated Fusarium mycotoxins, including 1 to 3 masked forms at the same time. Data obtained show that fumonisin B1, B2, B3, B4, and A1 were the most prevalent mycotoxins and had maximum contamination levels of 8908, 3383, 990, 1014, and 51.5 µg/kg, respectively. Deoxynivalenol occurred in 50% of the samples with a mean concentration of 152 µg/kg (max 1380 µg/kg). Thirty-three percent of the samples were contaminated with zearalenone at a mean concentration of 13.6 µg/kg (max 146 µg/kg). Of the masked mycotoxins, DON-3-glucoside occurred at a high incidence level of 53%. Among emerging toxins, moniliformin, fusarinolic acid, and beauvericin showed high occurrences at 98%, 98%, and 83%, and had maximum contamination levels of 1130, 3422, and 142 µg/kg, respectively. Significant differences in the contamination pattern were observed between the agricultural regions and maize types. Full article
(This article belongs to the Special Issue Application of LC-MS/MS in the Mycotoxins Studies)
Article
Assessment of the Effect of Satureja montana and Origanum virens Essential Oils on Aspergillus flavus Growth and Aflatoxin Production at Different Water Activities
Toxins 2020, 12(3), 142; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12030142 - 25 Feb 2020
Cited by 8
Abstract
Aflatoxin contamination of foodstuffs poses a serious risk to food security, and it is essential to search for new control methods to prevent these toxins entering the food chain. Several essential oils are able to reduce the growth and mycotoxin biosynthesis of toxigenic [...] Read more.
Aflatoxin contamination of foodstuffs poses a serious risk to food security, and it is essential to search for new control methods to prevent these toxins entering the food chain. Several essential oils are able to reduce the growth and mycotoxin biosynthesis of toxigenic species, although their efficiency is strongly influenced by the environmental conditions. In this work, the effectiveness of Satureja montana and Origanum virens essential oils to control Aspergillus flavus growth was evaluated under three water activity levels (0.94, 0.96 and 0.98 aw) using a Bioscreen C, a rapid in vitro spectrophotometric technique. The aflatoxin concentrations at all conditions tested were determined by HPLC-FLD. Aspergillus flavus growth was delayed by both essential oil treatments. However, only S. montana essential oil was able to significantly affect aflatoxin production, although the inhibition percentages widely differed among water activities. The most significant reduction was observed at 0.96 aw, which is coincident with the conditions in which A. flavus reached the highest levels of aflatoxin production. On the contrary, the treatment with S. montana essential oil was not effective in significantly reducing aflatoxin production at 0.94 aw. Therefore, it is important to study the interaction of the new control compounds with environmental factors before their application in food matrices, and in vitro ecophysiological studies are a good option since they provide accurate and rapid results. Full article
(This article belongs to the Special Issue Aflatoxins: Food Sources, Occurrence and Toxicological Effects)
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Article
Effect of Ozone and Electron Beam Irradiation on Degradation of Zearalenone and Ochratoxin A
Toxins 2020, 12(2), 138; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020138 - 24 Feb 2020
Cited by 6
Abstract
Zearalenone (ZEN) and ochratoxin A (OTA) are key concerns of the food industry because of their toxicity and pollution scope. This study investigated the effects of ozone and electron beam irradiation (EBI) on the degradation of ZEN and OTA. Results demonstrated that 2 [...] Read more.
Zearalenone (ZEN) and ochratoxin A (OTA) are key concerns of the food industry because of their toxicity and pollution scope. This study investigated the effects of ozone and electron beam irradiation (EBI) on the degradation of ZEN and OTA. Results demonstrated that 2 mL of 50 μg/mL ZEN was completely degraded after 10 s of treatment by 2.0 mg/L ozone. The degradation rate of 1 μg/mL ZEN by 16 kGy EBI was 92.76%. Methanol was superior to acetonitrile in terms of degrading ZEN when the irradiation dose was higher than 6 kGy. The degradation rate of 2 mL of 5 μg/mL OTA by 50 mg/L ozone at 180 s was 34%, and that of 1 μg/mL OTA by 16 kGy EBI exceeded 90%. Moreover, OTA degraded more rapidly in acetonitrile. Ozone performed better in the degradation of ZEN, whereas EBI was better for OTA. The conclusions provide theoretical and practical bases for the degradation of different fungal toxins. Full article
(This article belongs to the Special Issue Mycotoxins Study: Toxicology, Identification and Control)
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Article
A Lateral Flow Strip Based on a Truncated Aptamer-Complementary Strand for Detection of Type-B Aflatoxins in Nuts and Dried Figs
Toxins 2020, 12(2), 136; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020136 - 22 Feb 2020
Cited by 8
Abstract
Type-B aflatoxins (AFB1 and AFB2) frequently contaminate food, especially nuts and fried figs, and seriously threaten human health; hence, it is necessary for the newly rapid and sensitive detection methods to prevent the consumption of potentially contaminated food. Here, a [...] Read more.
Type-B aflatoxins (AFB1 and AFB2) frequently contaminate food, especially nuts and fried figs, and seriously threaten human health; hence, it is necessary for the newly rapid and sensitive detection methods to prevent the consumption of potentially contaminated food. Here, a lateral flow aptasensor for the detection of type-B aflatoxins was developed. It is based on the use of fluorescent dye Cy5 as a label for the aptamer, and on the competition between type-B aflatoxins and the complementary DNA of the aptamer. This is the first time that the complementary strand of the aptamer has been used as the test line (T-line) to detect type-B aflatoxins. In addition, the truncated aptamer was used to improve the affinity with type-B aflatoxins in our study. Therefore, the lengths of aptamer and cDNA probe were optimized as key parameters for higher sensitivity. In addition, binding buffer and organic solvent were investigated. The results showed that the best pair for achieving improved sensitivity and accuracy in detecting AFB1 was formed by a shorter aptamer (32 bases) coupled with the probe complementary to the AFB1 binding region of the aptamer. Under the optimal experimental conditions, the test strip showed an excellent linear relationship in the range from 0.2 to 20 ng/mL with a limit of detection of 0.16 ng/mL. This aptamer-based strip was successfully applied to the determination of type-B aflatoxins in spiked and commercial peanuts, almonds, and dried figs, and the recoveries of the spiked samples were from 93.3%−112.0%. The aptamer-complementary strand-based lateral flow test strip is a potential alternative tool for the rapid and sensitive detection of type-B aflatoxins in nuts and dried figs. It is of help for monitoring aflatoxins to avoid the consumption of unsafe food. Full article
(This article belongs to the Section Mycotoxins)
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Article
Toxicity Characterisation of Gambierdiscus Species from the Canary Islands
Toxins 2020, 12(2), 134; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020134 - 21 Feb 2020
Cited by 10
Abstract
In the last decade, several outbreaks of ciguatera fish poisoning (CFP) have been reported in the Canary Islands (central northeast Atlantic Ocean), confirming ciguatera as an emerging alimentary risk in this region. Five Gambierdiscus species, G. australes, G. excentricus, G. silvae [...] Read more.
In the last decade, several outbreaks of ciguatera fish poisoning (CFP) have been reported in the Canary Islands (central northeast Atlantic Ocean), confirming ciguatera as an emerging alimentary risk in this region. Five Gambierdiscus species, G. australes, G. excentricus, G. silvae, G. carolinianus and G. caribaeus, have been detected in macrophytes from this area and are known to produce the ciguatoxins (CTXs) that cause CFP. A characterization of the toxicity of these species is the first step in identifying locations in the Canary Islands at risk of CFP. Therefore, in this study the toxicity of 63 strains of these five Gambierdiscus species were analysed using the erythrocyte lysis assay to evaluate their maitotoxin (MTX) content. In addition, 20 of the strains were also analysed in a neuroblastoma Neuro-2a (N2a) cytotoxicity assay to determine their CTX-like toxicity. The results allowed the different species to be grouped according to their ratios of CTX-like and MTX-like toxicity. MTX-like toxicity was especially high in G. excentricus and G. australes but much lower in the other species and lowest in G. silvae. CTX-like toxicity was highest in G. excentricus, which produced the toxin in amounts ranging between 128.2 ± 25.68 and 510.6 ± 134.2 fg CTX1B equivalents (eq) cell−1 (mean ± SD). In the other species, CTX concentrations were as follows: G. carolinianus (100.84 ± 18.05 fg CTX1B eq cell−1), G. australes (31.1 ± 0.56 to 107.16 ± 21.88 fg CTX1B eq cell−1), G. silvae (12.19 ± 0.62 to 76.79 ± 4.97 fg CTX1B eq cell−1) and G. caribaeus (<LOD to 90.37 ± 15.89 fg CTX1B eq cell−1). Unlike the similar CTX-like toxicity of G. australes and G. silvae strains from different locations, G. excentricus and G. caribaeus differed considerably according to the origin of the strain. These differences emphasise the importance of species identification to assess the regional risk of CFP. Full article
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Article
Varespladib (LY315920) and Methyl Varespladib (LY333013) Abrogate or Delay Lethality Induced by Presynaptically Acting Neurotoxic Snake Venoms
Toxins 2020, 12(2), 131; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020131 - 20 Feb 2020
Cited by 22
Abstract
The phospholipase A2 (PLA2) inhibitor Varespladib (LY315920) and its orally bioavailable prodrug, methyl-Varespladib (LY333013) inhibit PLA2 activity of a wide variety of snake venoms. In this study, the ability of these two forms of Varespladib to halt or delay [...] Read more.
The phospholipase A2 (PLA2) inhibitor Varespladib (LY315920) and its orally bioavailable prodrug, methyl-Varespladib (LY333013) inhibit PLA2 activity of a wide variety of snake venoms. In this study, the ability of these two forms of Varespladib to halt or delay lethality of potent neurotoxic snake venoms was tested in a mouse model. The venoms of Notechis scutatus, Crotalus durissus terrificus, Bungarus multicinctus, and Oxyuranus scutellatus, all of which have potent presynaptically acting neurotoxic PLA2s of variable quaternary structure, were used to evaluate simple dosing regimens. A supralethal dose of each venom was injected subcutaneously in mice, followed by the bolus intravenous (LY315920) or oral (LY333013) administration of the inhibitors, immediately and at various time intervals after envenoming. Control mice receiving venom alone died within 3 h of envenoming. Mice injected with O. scutellatus venom and treated with LY315920 or LY333013 survived the 24 h observation period, whereas those receiving C. d. terrificus and B. multicinctus venoms survived at 3 h or 6 h with a single dose of either form of Varespladib, but not at 24 h. In contrast, mice receiving N. scutatus venom and then the inhibitors died within 3 h, similarly to the control animals injected with venom alone. LY315920 was able to reverse the severe paralytic manifestations in mice injected with venoms of O. scutellatus, B. multicinctus, and C. d. terrificus. Overall, results suggest that the two forms of Varespladib are effective in abrogating, or delaying, neurotoxic manifestations induced by some venoms whose neurotoxicity is mainly dependent on presynaptically acting PLA2s. LY315920 is able to reverse paralytic manifestations in severely envenomed mice, but further work is needed to understand the significance of species-specific differences in animal models as they compare to clinical syndromes in human and for potential use in veterinary medicine. Full article
(This article belongs to the Section Animal Venoms)
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Article
Phytotoxic Metabolites Isolated from Neufusicoccum batangarum, the Causal Agent of the Scabby Canker of Cactus Pear (Opuntia ficus-indica L.)
Toxins 2020, 12(2), 126; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020126 - 18 Feb 2020
Cited by 9
Abstract
Six phytotoxins were obtained from the culture filtrates of the ascomycete Neofusicoccum batangarum, the causal agent of the scabby canker of cactus pear (Opuntia ficus-indica L.) in minor Sicily islands. The phytotoxins were identified as (−)-(R)-mellein (1); [...] Read more.
Six phytotoxins were obtained from the culture filtrates of the ascomycete Neofusicoccum batangarum, the causal agent of the scabby canker of cactus pear (Opuntia ficus-indica L.) in minor Sicily islands. The phytotoxins were identified as (−)-(R)-mellein (1); (±)-botryoisocoumarin A (2); (−)-(3R,4R)- and (−)-(3R,4S)-4-hydroxymellein (3 and 4); (−)-terpestacin (5); and (+)-3,4-dihydro-4,5,8-trihydroxy-3-methylisocoumarin, which we named (+)-neoisocoumarin (6). This identification was done by comparing their spectral and optical data with those already reported in literature. The absolute configuration (3R,4S) to (+)-neoisocoumarin (6) was determined using the advanced Mosher method. All six metabolites were shown to have phytotoxicity on the host (cactus pear) and non-host (tomato) plants, and the most active compounds were (±)-botryoisocoumarin A (2), (−)-terpestacin (5), and (+)-neoisocoumarin (6). Full article
(This article belongs to the Special Issue Microbial and Plant Phytotoxins)
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Article
Rapid, Sensitive, and Accurate Point-of-Care Detection of Lethal Amatoxins in Urine
Toxins 2020, 12(2), 123; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020123 - 15 Feb 2020
Cited by 8
Abstract
Globally, mushroom poisonings cause about 100 human deaths each year, with thousands of people requiring medical assistance. Dogs are also susceptible to mushroom poisonings and require medical assistance. Cyclopeptides, and more specifically amanitins (or amatoxins, here), are the mushroom poison that causes the [...] Read more.
Globally, mushroom poisonings cause about 100 human deaths each year, with thousands of people requiring medical assistance. Dogs are also susceptible to mushroom poisonings and require medical assistance. Cyclopeptides, and more specifically amanitins (or amatoxins, here), are the mushroom poison that causes the majority of these deaths. Current methods (predominantly chromatographic, as well as antibody-based) of detecting amatoxins are time-consuming and require expensive equipment. In this work, we demonstrate the utility of the lateral flow immunoassay (LFIA) for the rapid detection of amatoxins in urine samples. The LFIA detects as little as 10 ng/mL of α-amanitin (α-AMA) or γ-AMA, and 100 ng/mL of β-AMA in urine matrices. To demonstrate application of this LFIA for urine analysis, this study examined fortified human urine samples and urine collected from exposed dogs. Urine is sampled directly without the need for any pretreatment, detection from urine is completed in 10 min, and the results are read by eye, without the need for specialized equipment. Analysis of both fortified human urine samples and urine samples collected from intoxicated dogs using the LFIA correlated well with liquid chromatography–mass spectrometry (LC-MS) methods. Full article
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Article
Mitochondria and Lysosomes Participate in Vip3Aa-Induced Spodoptera frugiperda Sf9 Cell Apoptosis
Toxins 2020, 12(2), 116; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020116 - 13 Feb 2020
Cited by 7
Abstract
Vip3Aa, a soluble protein produced by certain Bacillus thuringiensis strains, is capable of inducing apoptosis in Sf9 cells. However, the apoptosis mechanism triggered by Vip3Aa is unclear. In this study, we found that Vip3Aa induces mitochondrial dysfunction, as evidenced by signs of collapse [...] Read more.
Vip3Aa, a soluble protein produced by certain Bacillus thuringiensis strains, is capable of inducing apoptosis in Sf9 cells. However, the apoptosis mechanism triggered by Vip3Aa is unclear. In this study, we found that Vip3Aa induces mitochondrial dysfunction, as evidenced by signs of collapse of mitochondrial membrane potential, accumulation of reactive oxygen species, release of cytochrome c, and caspase-9 and -3 activation. Meanwhile, our results indicated that Vip3Aa reduces the ability of lysosomes in Sf9 cells to retain acridine orange. Moreover, pretreatment with Z-Phe-Tyr-CHO (a cathepsin L inhibitor) or pepstatin (a cathepsin D inhibitor) increased Sf9 cell viability, reduced cytochrome c release, and decreased caspase-9 and -3 activity. In conclusion, our findings suggested that Vip3Aa promotes Sf9 cell apoptosis by mitochondrial dysfunction, and lysosomes also play a vital role in the action of Vip3Aa. Full article
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Article
Zearalenone Removal from Corn Oil by an Enzymatic Strategy
Toxins 2020, 12(2), 117; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020117 - 13 Feb 2020
Cited by 7
Abstract
The estrogen-like mycotoxin zearalenone (ZEN) is one of the most widely distributed contaminants especially in maize and its commodities, such as corn oil. ZEN degrading enzymes possess the potential for counteracting the negative effect of ZEN and its associated high safety risk in [...] Read more.
The estrogen-like mycotoxin zearalenone (ZEN) is one of the most widely distributed contaminants especially in maize and its commodities, such as corn oil. ZEN degrading enzymes possess the potential for counteracting the negative effect of ZEN and its associated high safety risk in corn oil. Herein, we targeted enhancing the secretion of ZEN degrading enzyme by Pichia pastoris through constructing an expression plasmid containing three optimized expression cassettes of zlhy-6 codon and signal peptides. Further, we explored various parameters of enzymatic detoxification in neutralized oil and analyzed tocopherols and sterols losses in the corn oil. In addition, the distribution of degraded products was demonstrated as well by Agilent 6510 Quadrupole Time-of-Flight mass spectrometry. P. pastoris GSZ with the glucoamylase signal was observed with the highest ZLHY-6 secretion yield of 0.39 mg/mL. During the refining of corn oil, ZEN in the crude oil was reduced from 1257.3 to 13 µg/kg (3.69% residual) after neutralization and enzymatic detoxification. Compared with the neutralized oil, no significant difference in the total tocopherols and sterols contents was detected after enzymatic detoxification. Finally, the degraded products were found to be entirely eliminated by washing. This study presents an enzymatic strategy for efficient and safe ZEN removal with relatively low nutrient loss, which provides an important basis for further application of enzymatic ZEN elimination in the industrial process of corn oil production. Full article
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Article
Ultra-High-Performance Liquid Chromatography Coupled with Quadrupole Orbitrap High-Resolution Mass Spectrometry for Multi-Residue Analysis of Mycotoxins and Pesticides in Botanical Nutraceuticals
Toxins 2020, 12(2), 114; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020114 - 12 Feb 2020
Cited by 14
Abstract
Cannabidiol (CBD) food supplements made of Cannabis sativa L. extracts have quickly become popular products due to their health-promoting effects. However, potential contaminants, such as mycotoxins and pesticides, can be coextracted during the manufacturing process and placed into the final product. Accordingly, a [...] Read more.
Cannabidiol (CBD) food supplements made of Cannabis sativa L. extracts have quickly become popular products due to their health-promoting effects. However, potential contaminants, such as mycotoxins and pesticides, can be coextracted during the manufacturing process and placed into the final product. Accordingly, a novel methodology using ultra-high-performance liquid chromatography coupled with quadrupole Orbitrap high-resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS) was developed to quantify 16 mycotoxins produced by major C. sativa fungi, followed by a post-target screening of 283 pesticides based on a comprehensive spectral library. The validated procedure was applied to ten CBD-based products. Up to six different Fusarium mycotoxins were found in seven samples, the most prevalent being zearalenone (60%) and enniatin B1 (30%), both found at a maximum level of 11.6 ng/g. Co-occurrence was observed in four samples, including one with enniatin B1, enniatin A and enniatin A1. On the other hand, 46 different pesticides were detected after retrospective analysis. Ethoxyquin (50%), piperonyl butoxide (40%), simazine (30%) and cyanazine (30%) were the major residues found. These results highlight the necessity of monitoring contaminants in food supplements in order to ensure a safe consumption, even more considering the increase trend in their use. Furthermore, the developed procedure is proposed as a powerful analytical tool to evaluate the potential mycotoxin profile of these particular products. Full article
(This article belongs to the Special Issue Application of LC-MS/MS in the Mycotoxins Studies)
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Article
The Effectiveness of Durian Peel as a Multi-Mycotoxin Adsorbent
Toxins 2020, 12(2), 108; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020108 - 08 Feb 2020
Cited by 7
Abstract
Durian peel (DP) is an agricultural waste that is widely used in dyes and for organic and inorganic pollutant adsorption. In this study, durian peel was acid-treated to enhance its mycotoxin adsorption efficacy. The acid-treated durian peel (ATDP) was assessed for simultaneous adsorption [...] Read more.
Durian peel (DP) is an agricultural waste that is widely used in dyes and for organic and inorganic pollutant adsorption. In this study, durian peel was acid-treated to enhance its mycotoxin adsorption efficacy. The acid-treated durian peel (ATDP) was assessed for simultaneous adsorption of aflatoxin B1 (AFB1), ochratoxin A (OTA), zearalenone (ZEA), deoxynivalenol (DON), and fumonisin B1 (FB1). The structure of the ATDP was also characterized by SEM–EDS, FT–IR, a zetasizer, and a surface-area analyzer. The results indicated that ATDP exhibited the highest mycotoxin adsorption towards AFB1 (98.4%), ZEA (98.4%), and OTA (97.3%), followed by FB1 (86.1%) and DON (2.0%). The pH significantly affected OTA and FB1 adsorption, whereas AFB1 and ZEA adsorption was not affected. Toxin adsorption by ATDP was dose-dependent and increased exponentially as the ATDP dosage increased. The maximum adsorption capacity (Qmax), determined at pH 3 and pH 7, was 40.7 and 41.6 mmol kg−1 for AFB1, 15.4 and 17.3 mmol kg−1 for ZEA, 46.6 and 0.6 mmol kg−1 for OTA, and 28.9 and 0.1 mmol kg−1 for FB1, respectively. Interestingly, ATDP reduced the bioaccessibility of these mycotoxins after gastrointestinal digestion using an in vitro, validated, static model. The ATDP showed a more porous structure, with a larger surface area and a surface charge modification. These structural changes following acid treatment may explain the higher efficacy of ATDP in adsorbing mycotoxins. Hence, ATDP can be considered as a promising waste material for mycotoxin biosorption. Full article
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Article
Chronic Microcystin-LR Exposure Induces Abnormal Lipid Metabolism via Endoplasmic Reticulum Stress in Male Zebrafish
Toxins 2020, 12(2), 107; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020107 - 07 Feb 2020
Cited by 6
Abstract
In order to explore effects of low levels of continuous microcystin-LR (MC-LR) (a cyanotoxin) exposure on hepatic lipid metabolism on the basis of the endoplasmic reticulum stress (ERS) pathway, we exposed adult male zebrafish to MC-LR (0, 1, 5, and 25 μg/L) for [...] Read more.
In order to explore effects of low levels of continuous microcystin-LR (MC-LR) (a cyanotoxin) exposure on hepatic lipid metabolism on the basis of the endoplasmic reticulum stress (ERS) pathway, we exposed adult male zebrafish to MC-LR (0, 1, 5, and 25 μg/L) for 60 days, and hepatic histopathology as well as lipid metabolic parameters were determined with mRNA levels of ERS signal molecules and downstream factors, along with genes associated with lipid metabolism in zebrafish liver. The results revealed that prolonged exposure to MC-LR remarkably altered the levels of hepatic total cholesterol and triglyceride and led to hepatic steatosis, which was also confirmed by hepatic cytoplasmic vacuolization in Hematoxylin/eosin (H&E) stain and lipid droplet accumulation in Oil Red O stain. The severity of hepatic damage and lipidation was increased in a dose-related manner. MC-LR exposure significantly upregulated transcriptional levels of ERS markers including hspa5, mapk8, and chop, indicating the occurrence of ERS in the liver of zebrafish. Concurrently, MC-LR significantly improved mRNA expression of unfolded protein response (UPR) pathway-related genes including atf6, eif2ak3, ern1, and xbp1s, suggesting that all of the three UPR branches were activated by MC-LR. MC-LR also induced significant upregulation of downstream lipid metabolism-related factors and genes including srebf1, srebf2, fatty acid synthase (fasn), acetyl-CoA carboxylase (acaca), stearoyl-CoA desaturase (scd), HMG CoA reductase (hmgcra), and HMG CoA synthase (hmgcs1), and downregulation of genes associated with lipolysis such as triglyceride hydrolase gene (atgl), hormone-sensitive enzyme gene (hsla), and carnitine palmitoyltransferase gene (cpt1aa). Our present results indicated that the cause of hepatic lipid accumulation by MC-LR was mainly by upregulating lipogenic and cholesterol genes but downregulating the expression of lipolytic genes through the induction of srebf1 and srebf2, which were involved in the activation of ERS signal pathways. Full article
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Article
In Vitro Rumen Simulations Show a Reduced Disappearance of Deoxynivalenol, Nivalenol and Enniatin B at Conditions of Rumen Acidosis and Lower Microbial Activity
Toxins 2020, 12(2), 101; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020101 - 05 Feb 2020
Cited by 9
Abstract
Ruminants are generally considered to be less susceptible to the effects of mycotoxins than monogastric animals as the rumen microbiota are capable of detoxifying some of these toxins. Despite this potential degradation, mycotoxin-associated subclinical health problems are seen in dairy cows. In this [...] Read more.
Ruminants are generally considered to be less susceptible to the effects of mycotoxins than monogastric animals as the rumen microbiota are capable of detoxifying some of these toxins. Despite this potential degradation, mycotoxin-associated subclinical health problems are seen in dairy cows. In this research, the disappearance of several mycotoxins was determined in an in vitro rumen model and the effect of realistic concentrations of those mycotoxins on fermentation was assessed by volatile fatty acid production. In addition, two hypotheses were tested: (1) a lower rumen pH leads to a decreased degradation of mycotoxins and (2) rumen fluid of lactating cows degrade mycotoxins better than rumen fluid of non-lactating cows. Maize silage was spiked with a mixture of deoxynivalenol (DON), nivalenol (NIV), enniatin B (ENN B), mycophenolic acid (MPA), roquefortine C (ROQ-C) and zearalenone (ZEN). Fresh rumen fluid of two lactating cows (L) and two non-lactating cows (N) was added to a buffer of normal pH (6.8) and low pH (5.8), leading to four combinations (L6.8, L5.8, N6.8, N5.8), which were added to the spiked maize substrate. In this study, mycotoxins had no effect on volatile fatty acid production. However, not all mycotoxins fully disappeared during incubation. ENN B and ROQ-C disappeared only partially, whereas MPA showed almost no disappearance. The disappearance of DON, NIV, and ENN B was hampered when pH was low, especially when the inoculum of non-lactating cows was used. For ZEN, a limited transformation of ZEN to α-ZEL and β-ZEL was observed, but only at pH 6.8. In conclusion, based on the type of mycotoxin and the ruminal conditions, mycotoxins can stay intact in the rumen. Full article
(This article belongs to the Special Issue Mycotoxins in Feed: Harm to Animals)
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Article
Aflatoxin M1 Determination in Infant Formulae Distributed in Monterrey, Mexico
Toxins 2020, 12(2), 100; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020100 - 04 Feb 2020
Cited by 8
Abstract
The occurrence of aflatoxin M1 (AFM1) in infant formulae commercialized in the metropolitan area of Monterrey (Nuevo León, Mexico) was determined by using immunoaffinity column clean-up followed by HPLC determination with fluorimetric detection. For this, 55 infant formula powders were [...] Read more.
The occurrence of aflatoxin M1 (AFM1) in infant formulae commercialized in the metropolitan area of Monterrey (Nuevo León, Mexico) was determined by using immunoaffinity column clean-up followed by HPLC determination with fluorimetric detection. For this, 55 infant formula powders were classified in two groups, starter (49 samples) and follow-on (6 samples) formulae. Eleven of the evaluated samples (20%) presented values above the permissible limit set by the European Union for infant formulae (25 ng/L), ranging from 40 to 450 ng/L. The estimated daily intake (EDI) for AFM1 was determined employing the average body weight (bw) of the groups of age in the ranges of 0–6 and 6–12 months, and 1–2 years. The results evidenced high intake values, ranging from 1.56 to 14 ng/kg bw/day, depending on the group. Finally, with the EDI value, the carcinogenic risk index was determined, presenting a high risk for all the evaluated groups. Based on these results, it is a necessary extra effort by the regulatory agencies to reduce the AFM1 presence in infant formulae consumed in Mexico. Full article
(This article belongs to the Special Issue Mycotoxins in Feed and Food Chain: Present Status and Future Concerns)
Article
Domain Shuffling between Vip3Aa and Vip3Ca: Chimera Stability and Insecticidal Activity against European, American, African, and Asian Pests
Toxins 2020, 12(2), 99; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020099 - 04 Feb 2020
Cited by 9
Abstract
The bacterium Bacillus thuringiensis produces insecticidal Vip3 proteins during the vegetative growth phase with activity against several lepidopteran pests. To date, three different Vip3 protein families have been identified based on sequence identity: Vip3A, Vip3B, and Vip3C. In this study, we report the [...] Read more.
The bacterium Bacillus thuringiensis produces insecticidal Vip3 proteins during the vegetative growth phase with activity against several lepidopteran pests. To date, three different Vip3 protein families have been identified based on sequence identity: Vip3A, Vip3B, and Vip3C. In this study, we report the construction of chimeras by exchanging domains between Vip3Aa and Vip3Ca, two proteins with marked specificity differences against lepidopteran pests. We found that some domain combinations made proteins insoluble or prone to degradation by trypsin as most abundant insect gut protease. The soluble and trypsin-stable chimeras, along with the parental proteins Vip3Aa and Vip3Ca, were tested against lepidopteran pests from different continents: Spodoptera exigua, Spodoptera littoralis, Spodoptera frugiperda, Helicoverpa armigera, Mamestra brassicae, Anticarsia gemmatalis, and Ostrinia furnacalis. The exchange of the Nt domain (188 N-terminal amino acids) had little effect on the stability and toxicity (equal or slightly lower) of the resulting chimeric protein against all insects except for S. frugiperda, for which the chimera with the Nt domain from Vip3Aa and the rest of the protein from Vip3Ca showed a significant increase in toxicity compared to the parental Vip3Ca. Chimeras with the C-terminal domain from Vip3Aa (from amino acid 510 of Vip3Aa to the Ct) with the central domain of Vip3Ca (amino acids 189–509 based on the Vip3Aa sequence) made proteins that could not be solubilized. Finally, the chimera including the Ct domain of Vip3Ca and the Nt and central domain from Vip3Aa was unstable. Importantly, an insect species tolerant to Vip3Aa but susceptible to Vip3Ca, such as Ostrinia furnacalis, was also susceptible to chimeras maintaining the Ct domain from Vip3Ca, in agreement with the hypothesis that the Ct region of the protein is the one conferring specificity to Vip3 proteins. Full article
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Article
Cultivation Area Affects the Presence of Fungal Communities and Secondary Metabolites in Italian Durum Wheat Grains
Toxins 2020, 12(2), 97; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020097 - 03 Feb 2020
Cited by 6
Abstract
In this study, durum wheat kernels harvested in three climatically different Italian cultivation areas (Emilia Romagna, Umbria and Sardinia) in 2015, were analyzed with a combination of different isolation methods to determine their fungal communities, with a focus on Fusarium head blight (FHB) [...] Read more.
In this study, durum wheat kernels harvested in three climatically different Italian cultivation areas (Emilia Romagna, Umbria and Sardinia) in 2015, were analyzed with a combination of different isolation methods to determine their fungal communities, with a focus on Fusarium head blight (FHB) complex composition, and to detect fungal secondary metabolites in the grains. The genus Alternaria was the main component of durum wheat mycobiota in all investigated regions, with the Central Italian cultivation area showing the highest incidence of this fungal genus and of its secondary metabolites. Fusarium was the second most prevalent genus of the fungal community in all cultivation environments, even if regional differences in species composition were detected. In particular, Northern areas showed the highest Fusarium incidence, followed by Central and then Southern cultivation areas. Focusing on the FHB complex, a predominance of Fusarium poae, in particular in Northern and Central cultivation areas, was found. Fusarium graminearum, in the analyzed year, was mainly detected in Emilia Romagna. Because of the highest Fusarium incidence, durum wheat harvested in the Northern cultivation area showed the highest presence of Fusarium secondary metabolites. These results show that durum wheat cultivated in Northern Italy may be subject to a higher FHB infection risk and to Fusarium mycotoxins accumulation. Full article
(This article belongs to the Section Mycotoxins)
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Article
Inflammatory Reaction Induced by Two Metalloproteinases Isolated from Bothrops atrox Venom and by Fragments Generated from the Hydrolysis of Basement Membrane Components
Toxins 2020, 12(2), 96; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020096 - 02 Feb 2020
Cited by 9
Abstract
Snake venom metalloproteinases (SVMPs) play an important role in local tissue damage of snakebite patients, mostly by hydrolysis of basement membrane (BM) components. We evaluated the proinflammatory activity of SVMPs Atroxlysin-Ia (ATXL) and Batroxrhagin (BATXH) from Bothrops atrox venom and their hydrolysis products [...] Read more.
Snake venom metalloproteinases (SVMPs) play an important role in local tissue damage of snakebite patients, mostly by hydrolysis of basement membrane (BM) components. We evaluated the proinflammatory activity of SVMPs Atroxlysin-Ia (ATXL) and Batroxrhagin (BATXH) from Bothrops atrox venom and their hydrolysis products of Matrigel. BALB/c mice were injected with SVMPs (2 μg), for assessment of paw edema and peritoneal leukocyte accumulation. Both SVMPs induced edema, representing an increase of ~70% of the paw size. Leukocyte infiltrates reached levels of 6 × 106 with ATXL and 5 × 106 with BATXH. TNF-α was identified in the supernatant of BATXH—or venom-stimulated MPAC cells. Incubation of Matrigel with the SVMPs generated fragments, including peptides from Laminin, identified by LC–MS/MS. The Matrigel hydrolysis peptides caused edema that increased 30% the paw size and promoted leukocyte accumulation (4–5 × 106) to the peritoneal cavity, significantly higher than Matrigel control peptides 1 and 4 h after injection. Our findings suggest that ATXL and BATXH are involved in the inflammatory reaction observed in B. atrox envenomings by direct action on inflammatory cells or by releasing proinflammatory peptides from BM proteins that may amplify the direct action of SVMPs through activation of endogenous signaling pathways. Full article
(This article belongs to the Section Animal Venoms)
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Article
A Liquid Chromatographic Method for Rapid and Sensitive Analysis of Aflatoxins in Laboratory Fungal Cultures
Toxins 2020, 12(2), 93; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020093 - 30 Jan 2020
Cited by 6
Abstract
Culture methods supplemented with high-performance liquid chromatography (HPLC) technique provide a rapid and simple tool for detecting levels of aflatoxins (AFs) produced by fungi. This study presents a robust method for simultaneous quantification of aflatoxin (AF) B1, B2, G1, and G2 levels in [...] Read more.
Culture methods supplemented with high-performance liquid chromatography (HPLC) technique provide a rapid and simple tool for detecting levels of aflatoxins (AFs) produced by fungi. This study presents a robust method for simultaneous quantification of aflatoxin (AF) B1, B2, G1, and G2 levels in several fungal cultivation states: submerged shake culture, liquid slant culture, and solid-state culture. The recovery of the method was evaluated by spiking a mixture of AFs at several concentrations to the test medium. The applicability of the method was evaluated by using aflatoxigenic and non-aflatoxigenic Aspergilli. A HPLC coupled with the diode array (DAD) and fluorescence (FLD) detectors was used to determine the presence and amounts of AFs. Both detectors showed high sensitivity in detecting spiked AFs or AFs produced in situ by toxigenic fungi. Our methods showed 76%–88% recovery from medium spiked with 2.5, 10, 50, 100, and 500 ng/mL AFs. The limit of quantification (LOQ) for AFs were 2.5 to 5.0 ng/mL with DAD and 0.025 to 2.5 ng/mL with FLD. In this work, we described in detail a protocol, which can be considered the foremost and only verified method, to extract, detect, and quantify AFs employing both aflatoxigenic and non-toxigenic Aspergilli. Full article
(This article belongs to the Special Issue Rapid Detection of Mycotoxin Contamination)
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Article
T-2 Toxin Induces Oxidative Stress, Apoptosis and Cytoprotective Autophagy in Chicken Hepatocytes
Toxins 2020, 12(2), 90; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020090 - 29 Jan 2020
Cited by 7
Abstract
T-2 toxin is type A trichothecenes mycotoxin, which produced by fusarium species in cereal grains. T-2 toxin has been shown to induce a series of toxic effects on the health of human and animal, such as immunosuppression and carcinogenesis. Previous study has proven [...] Read more.
T-2 toxin is type A trichothecenes mycotoxin, which produced by fusarium species in cereal grains. T-2 toxin has been shown to induce a series of toxic effects on the health of human and animal, such as immunosuppression and carcinogenesis. Previous study has proven that T-2 toxin caused hepatotoxicity in chicken, but the regulatory mechanism is unclear. In the present study, we assessed the toxicological effect of T-2 toxin on apoptosis and autophagy in hepatocytes. The total of 120 1-day-old healthy broilers were allocated randomly into four groups and reared for 21 day with complete feed containing 0 mg/kg, 0.5 mg/kg, 1 mg/kg or 2 mg/kg T-2 toxin, respectively. The results showed that the apoptosis rate and pathological changes degree hepatocytes were aggravated with the increase of T-2 toxin. At the molecular mechanism level, T-2 toxin induced mitochondria-mediated apoptosis by producing reactive oxygen species, promoting cytochrome c translocation between the mitochondria and cytoplasm, and thus promoting apoptosomes formation. Meanwhile, the expression of the autophagy-related protein, ATG5, ATG7 and Beclin-1, and the LC3-II/LC3-I ratio were increased, while p62 was downregulated, suggesting T-2 toxin caused autophagy in hepatocytes. Further experiments demonstrated that the PI3K/AKT/mTOR signal may be participated in autophagy induced by T-2 toxin in chicken hepatocytes. These data suggest a possible underlying molecular mechanism for T-2 toxin that induces apoptosis and autophagy in chicken hepatocytes Full article
(This article belongs to the Special Issue Toxicological Effects of Mycotoxins on Target Cells)
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Article
Acute Oral Toxicity of Pinnatoxin G in Mice
Toxins 2020, 12(2), 87; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020087 - 28 Jan 2020
Cited by 6
Abstract
Pinnatoxin G (PnTx-G) is a marine cyclic imine toxin produced by the dinoflagellate Vulcanodinium rugosum, frequently detected in edible shellfish from Ingril Lagoon (France). As other pinnatoxins, to date, no human poisonings ascribed to consumption of PnTx-G contaminated seafood have been reported, [...] Read more.
Pinnatoxin G (PnTx-G) is a marine cyclic imine toxin produced by the dinoflagellate Vulcanodinium rugosum, frequently detected in edible shellfish from Ingril Lagoon (France). As other pinnatoxins, to date, no human poisonings ascribed to consumption of PnTx-G contaminated seafood have been reported, despite its potent antagonism at nicotinic acetylcholine receptors and its high and fast-acting toxicity after intraperitoneal or oral administration in mice. The hazard characterization of PnTx-G by oral exposure is limited to a single acute toxicity study recording lethality and clinical signs in non-fasted mice treated by gavage or through voluntary food ingestion, which showed differences in PnTx-G toxic potency. Thus, an acute toxicity study was carried out using 3 h-fasted CD-1 female mice, administered by gavage with PnTx-G (8–450 µg kg−1). At the dose of 220 µg kg−1 and above, the toxin induced a rapid onset of clinical signs (piloerection, prostration, hypothermia, abdominal breathing, paralysis of the hind limbs, and cyanosis), leading to the death of mice within 30 min. Except for moderate mucosal degeneration in the small intestine recorded at doses of 300 µg kg−1, the toxin did not induce significant morphological changes in the other main organs and tissues, or alterations in blood chemistry parameters. This acute oral toxicity study allowed to calculate an oral LD50 for PnTx-G equal to 208 μg kg−1 (95% confidence limits: 155–281 µg kg−1) and to estimate a provisional NOEL of 120 µg kg−1. Full article
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Article
Molecular Epidemiology of Methicillin-Susceptible and Methicillin-Resistant Staphylococcus aureus in Wild, Captive and Laboratory Rats: Effect of Habitat on the Nasal S. aureus Population
Toxins 2020, 12(2), 80; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020080 - 24 Jan 2020
Cited by 6
Abstract
Rats are a reservoir of human- and livestock-associated methicillin-resistant Staphylococcus aureus (MRSA). However, the composition of the natural S. aureus population in wild and laboratory rats is largely unknown. Here, 144 nasal S. aureus isolates from free-living wild rats, captive wild rats and [...] Read more.
Rats are a reservoir of human- and livestock-associated methicillin-resistant Staphylococcus aureus (MRSA). However, the composition of the natural S. aureus population in wild and laboratory rats is largely unknown. Here, 144 nasal S. aureus isolates from free-living wild rats, captive wild rats and laboratory rats were genotyped and profiled for antibiotic resistances and human-specific virulence genes. The nasal S. aureus carriage rate was higher among wild rats (23.4%) than laboratory rats (12.3%). Free-living wild rats were primarily colonized with isolates of clonal complex (CC) 49 and CC130 and maintained these strains even in husbandry. Moreover, upon livestock contact, CC398 isolates were acquired. In contrast, laboratory rats were colonized with many different S. aureus lineages—many of which are commonly found in humans. Five captive wild rats were colonized with CC398-MRSA. Moreover, a single CC30-MRSA and two CC130-MRSA were detected in free-living or captive wild rats. Rat-derived S. aureus isolates rarely harbored the phage-carried immune evasion gene cluster or superantigen genes, suggesting long-term adaptation to their host. Taken together, our study revealed a natural S. aureus population in wild rats, as well as a colonization pressure on wild and laboratory rats by exposure to livestock- and human-associated S. aureus, respectively. Full article
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Article
Bioprospecting Sponge-Associated Marine Cyanobacteria to Produce Bioactive Compounds
Toxins 2020, 12(2), 73; https://0-doi-org.brum.beds.ac.uk/10.3390/toxins12020073 - 23 Jan 2020
Cited by 6
Abstract
Marine cyanobacteria are considered a prolific source of bioactive natural products with a range of biotechnological and pharmacological applications. However, data on the production of natural compounds from sponge-associated cyanobacteria are scarce. This study aimed to assess the potential of sponge-associated cyanobacteria strains [...] Read more.
Marine cyanobacteria are considered a prolific source of bioactive natural products with a range of biotechnological and pharmacological applications. However, data on the production of natural compounds from sponge-associated cyanobacteria are scarce. This study aimed to assess the potential of sponge-associated cyanobacteria strains representing different taxonomic groups for the production of bioactive compounds and the biological activity of their extracts. Phylogenetic analysis of sponge-associated cyanobacteria and screening for the presence of genes encoding non-ribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs) were performed. Methanol extracts of the sponge-associated strains were analyzed for cyanotoxin production and tested for antioxidant activity and cytotoxic activity against several human cancer cell lines and pathogenic bacteria. PKS were detected in all sponge-associated strains examined, indicating the metabolic potential of the isolates. PKS genes were more ubiquitous than NRPS genes. Cyanotoxins (i.e., cylindrospermopsin, anatoxin-a, nodularin, and microcystins) were not detected in any of the sponge-associated cyanobacterial strains. Strains belonging to Leptothoe, Pseudanabaena, and Synechococcus were found to have activity mainly against Staphylococcus aureus. In addition, sponge-associated Leptothoe strains (TAU-MAC 0915, 1015, 1115, and 1215) were found to be highly cytotoxic and in most cases more effective against human cancer cell lines than against normal cells. Extracts with the most promising bioactivity deserve further investigation in order to isolate and identify the bioactive molecule(s). Full article
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Article
Isolation and Characterization of [D-Leu1]microcystin-LY from Microcystis aeruginosa CPCC-464