Oxidative stress and viral Infections: rationale, experiences, and perspectives on N-acetylcysteine.

This article explores current evidence on the role of oxidative stress in viral infections, and on the use of antioxidant drugs as adjunctive treatment. MEDLINE/PubMed was searched for appropriate keywords, and preclinical and clinical studies with reviews were retrieved and examined by authors. Old and current evidence shows that GSH content reduction is the main mechanism of redox imbalance in viral-infected cells. Clinical studies found that GSH levels are depleted in patients with viral infections such as HIV and SARS-CoV. Viral infections activate inflammation through different pathways, and several of these mechanisms are related to oxidative stress. NAC is a precursor of GSH, and many of its intracellular effects are mediated by GSH replenishment, but it also activates some anti-inflammatory mechanisms. NAC has an excellent safety profile and better oral and topical bioavailability than GSH. These characteristics make NAC a suitable option as a repurposed drug. Adjunctive antioxidant treatment may improve the outcomes of antiviral therapies. Current evidence supports the rationale for this practice and some clinical experience showed encouraging results.

Diagnostic value of cutaneous manifestation of SARS-CoV-2 infection.

BACKGROUND: One of the challenging aspects of SARS-CoV-2 infection is its diverse multisystemic disease presentation. OBJECTIVES: To evaluate the diagnostic value of cutaneous manifestations of SARS-CoV-2 infection and investigate their duration and timing in relation to other COVID-19 symptoms. METHODS: We used data from 336 847 UK users of the COVID Symptom Study app to assess the diagnostic value of body rash or an acral rash in SARS-CoV-2 infection, and data from an independent online survey of 11 544 respondents to investigate skin-specific symptoms and collect their photographs. RESULTS: Using data from the app, we show significant association between skin rashes and a positive swab test result (odds ratio 1·67, 95% confidence interval 1·42-1·97). Strikingly, among the respondents of the independent online survey, we found that 17% of SARS-CoV-2-positive cases reported skin rashes as the first presentation, and 21% as the only clinical sign of COVID-19. Together with the British Association of Dermatologists, we have compiled a catalogue of images of the most common skin manifestations of COVID-19 from 400 individuals (https://covidskinsigns.com), which we have made publicly available to assist clinicians in recognition of this early clinical feature of COVID-19. CONCLUSIONS: Skin rashes cluster with other COVID-19 symptoms, are predictive of a positive swab test, and occur in a significant number of cases, either alone or before other classical symptoms. Recognizing rashes is important in identifying new and earlier cases of COVID-19.

A multicenter, observational, prospective study of self- and parent-reported quality of life in adolescent multiple sclerosis patients self-administering interferon-ß1a using RebiSmart™-the FUTURE study.

Besides the impact of disease per se, the use of immunomodulatory therapies in adolescents with relapsing-remitting multiple sclerosis (RRMS) may have an effect on quality of life (QL). The FUTURE (Quality of liFe in adolescent sUbjecTs affected by mUltiple sclerosis treated with immunomodulatoRy agEnt using self-injecting device) study was designed to evaluate the changes in QL of Italian adolescents with RRMS receiving treatment with IFN-ß1a (Rebif; 22 µg), administered subcutaneously three times weekly using the RebiSmart™ electronic autoinjection device over a 52-week period. Fifty adolescents with RRMS were enrolled and 40 completed the study. Changes from baseline to end of treatment (EoT) in adolescent self-reported and parent-reported QL were assessed using the Pediatric Quality of Life Inventory Multidimensional Fatigue Scale (PedsQL), which has been validated for use in pediatric MS and for which an Italian version is available. The adolescent self-reported total PedsQL4.0 score and all of its subscales tended to increase from baseline to EoT, the only exception being "Emotional functioning." In parent-reported measures, the total PedsQL4.0 score increased significantly from baseline to EoT (+ 5.27 points, p = 0.041). Significant increases were also evident for parent-reported "Psychosocial health summary score" (+ 5.90 points; p = 0.015) and "School functioning" (+ 7.84 points; p = 0.029). Our results indicate that adolescents with RRMS using the electronic injection device RebiSmart™ for self-administration of Rebif® can experience long-term improvements in QL.

To d , or not to d : recent developments and comparisons of regularization schemes.

We give an introduction to several regularization schemes that deal with ultraviolet and infrared singularities appearing in higher-order computations in quantum field theories. Comparing the computation of simple quantities in the various schemes, we point out similarities and differences between them.

Effect of alkaline cooking of maize on the content of fumonisins B1 and B2 and their hydrolysed forms.

The effect of nixtamalization on the content of fumonisins (FBs), hydrolysed (HFBs) and partially hydrolysed (PHFBs) fumonisins in maize was investigated at laboratory-scale. Maize naturally contaminated with FBs and PHFBs was cooked with lime. Starting raw maize, steeping and washing waters and final masa fractions were analysed for toxin content. Control-cooking experiments without lime were also carried out. The nixtamalization reduced the amount of FBs and PHFBs in masa and converted them to HFBs. However, the three forms of fumonisins collected in all fractions amounted to 183%, indicating that nixtamalization made available forms of matrix-associated fumonisins that were then converted to their hydrolysed forms. Control-cooking enhanced FBs and PHFBs reduction, due to the solubility of fumonisins in water during the steeping process, but did not form HFBs. These findings indicate that benefits associated with enhancing the nutritional value of nixtamalized maize are also associated with a safer product in terms of fumonisin contamination.

Dose-dependent lipid peroxidation induction on ex vivo intestine tracts exposed to chyme samples from fumonisins contaminated corn samples.

Fumonisins (FBs), Fusarium mycotoxins common food contaminant, are a potent inducer of oxidative stress and lipid peroxidation in intestinal cells. In order to verify this toxic effect in intestine tract, the aim was to assess lipid peroxidation (as malondialdehyde MDA increased levels) on intestine rat samples exposed to chyme samples from in vitro digestion of FBs contaminated corn samples. Naturally (9.61±3.2 µg/gr), artificially (726±94 µg/gr) and spiked corn samples at EU permitted FBs levels were digested and added to luminal side of Ussing chamber for 120 min. Fumonisins-free corn sample was used as control. The MDA increase was observed just in 83% of intestine samples exposed at EU FBs levels and the digestion process seems to reduce this incidence (50% of samples). Malondialdehyde levels were FBs dose- and subject-related and ranged from 0.07±0.01 to 3.59±0.6 nmol/mg. Highest incidence and MDA % increment (I) were found when intestine tracts were exposed to chymes from artificially corn sample. The induction of lipid peroxidation induced by FBs could be due to interactions between FBs and intestinal membranes, with consequent modifications in membrane permeability and oxygen diffusion-concentration, as suggested by other authors.

Development of loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Penicillium nordicum in dry-cured meat products.

The need of powerful diagnostic tools for rapid, simple, and cost-effective detection of food-borne fungi has become very important in the area of food safety. Currently, several isothermal nucleic acid amplification methods have been developed as an alternative to PCR-based analyses. Loop-mediated isothermal amplification (LAMP) is one of these innovative methods; it requires neither gel electrophoresis to separate and visualize the products nor expensive laboratory equipment and it has been applied already for detection of pathogenic organisms. In the current study, we developed a LAMP assay for the specific detection of Penicillium nordicum, the major causative agent of ochratoxin A contamination in protein-rich food, especially dry-cured meat products. The assay was based on targeting otapksPN gene, a key gene in the biosynthesis of ochratoxin A (OTA) in P. nordicum. Amplification of DNA during the reaction was detected directly in-tube by color transition of hydroxynaphthol blue from violet to sky blue, visible to the naked eye, avoiding further post amplification analyses. Only DNAs isolated from several P. nordicum strains led to positive results and no amplification was observed from non-target OTA and non OTA-producing strains. The assay was able to detect down to 100 fg of purified targeted genomic DNA or 10(2) conidia/reaction within 60 min. The LAMP assay for detection and identification of P. nordicum was combined with a rapid DNA extraction method set up on serially diluted conidia, providing an alternative rapid, specific and sensitive DNA-based method suitable for application directly "on-site", notably in key steps of dry-cured meat production.

Influence on functional parameters of intestinal tract induced by short-term exposure to fumonisins contaminated corn chyme samples.

The gut is a possible target toward mycotoxin fumonisins (FBs) exposure. The study aims to investigate the effects induced by FBs contaminated-corn chyme samples on functional parameters of human and rat intestine by using Ussing chamber. Fumonisins-contaminated corn and processed corn samples were undergone to in vitro digestion process and then added to luminal side. A reduction (about 90%) of short circuit current (Isc µA/cm(2)) during exposure of human colon tissues to fumonisins-free corn chyme samples was observed, probably related to increased chyme osmolality. This hyperosmotic stress could drain water towards the luminal compartment, modifying Na(+) and Cl(-) transports. The presence of FBs in corn chyme samples, independently to their concentration, did not affect significantly the Isc, probably related to their interference towards epithelial Na(+) transport, as assessed by using a specific inhibitor (Amiloride). The rat colon tract represents a more accessible model to study FBs toxicity showing a similar functional response to human. In the rat small intestine a significant reduction (about 15%) of Isc parameter during exposure to uncontaminated or FBs contaminated corn chyme samples was observed; therefore such model was not suitable to assess the FBs toxicity, probably because the prevalent glucose and amino acids electrogenic absorption overwhelmed the FBs influence on ionic transport.

Effect of variability in lighting and temperature environments for mature gilts housed in gestation crates on measures of reproduction and animal well-being.

The effects of room temperature and light intensity before breeding and into early gestation were evaluated on the reproductive performance and well-being of gilts housed individually in crates. In eight replicates, estrus was synchronized in mature gilts (n = 198) and after last feeding of Matrix were randomly assigned to a room temperature of 15°C (COLD), 21°C (NEUTRAL), or 30°C (HOT) and a light intensity of 11 (DIM) or 433 (BRIGHT) lx. Estrous detection was performed daily and gilts inseminated twice. Blood samples were collected before and after breeding for determination of immune measures and cortisol concentrations. Gilt ADFI, BW, and body temperature were measured. On d 30 postbreeding, gilts were slaughtered to recover reproductive tracts to evaluate pregnancy and litter characteristics. There were no temperature × light intensity interactions for any response variable. Reproductive measures of follicle development, expression of estrus, ovulation rate, pregnancy rate (83.2%), litter size (14.3 ± 0.5), and fetal measures were not affected by temperature or lighting (P > 0.10). Gilts in COLD (37.6°C) had a lower (P < 0.05) rectal temperature than those in NEUTRAL (38.2°C) and HOT (38.6 ± 0.04°C). Both BW gain and final BW were greater (P < 0.0001) for gilts kept in HOT than those in NEUTRAL or COLD environments. Cortisol was greater (P < 0.01) for gilts kept in COLD compared with those kept in the HOT room. Gilts housed in the HOT environment made more postural changes (P < 0.05) than did those kept in either COLD or NEUTRAL temperatures. Gilts kept in the HOT temperature spent more total time lying and more time lying ventrally compared with those gilts housed in the NEUTRAL or COLD rooms. Total white blood cells and the percentage of neutrophils as well as neutrophil-to-lymphocyte ratio were all influenced (P < 0.05) by temperature but there was no effect (P > 0.10) of light or interaction with temperature on other immune cells or measures. These results indicate that temperatures in the range of 15 to 30°C or light intensity at 11 to 433 lx do not impact reproduction during the follicular phase and into early gestation for mature gilts housed in gestation crates. However, room temperature does impact physiological, behavioral, and immune responses of mature gilts and should be considered as a potential factor that may influence gilt well-being during the first 30 d postbreeding.

Diversity of spore-forming bacteria and identification of Bacillus amyloliquefaciens as a species frequently associated with the ropy spoilage of bread.

This study examines the diversity of spore-forming bacteria isolated from raw materials/bread using molecular methods along with a rapid and innovative technology, the FT-NIR spectroscopy. Microbiological analysis showed that 23% of semolina and 42% of other raw materials (including grain, brewer yeast, improvers) contained more than 100 spores/g and more than 50% of each kind of sample was contaminated at a level ranging from 1 to 100 spores/g. A high bacterial diversity characterized raw materials. In total 176 isolates were collected and characterized: 13 bacterial species belonging to Bacillus (10) and Paenibacillus (3) genera were identified by sequencing of 16S rRNA, gyrA or gyrB genes. The two closely related species Bacillus amyloliquefaciens (strain N45.1) and Bacillus subtilis (strain S63) were also analyzed by the spectroscopic technique FT-NIR. This analysis gave clear discrimination between the strains in the score plot obtained by the PCA and allowed to identify the spectral region 5600-4000 cm(-1) as the information-rich region for discrimination. B. amyloliquefaciens, possibly misidentified as B. subtilis in previous studies, was recognized as the most frequent species, found also in ropy bread. Moreover, the screening test for rope production indicated that mainly B. amyloliquefaciens, together with B. subtilis and Bacillus pumilus, could cause spoilage in bread, even if the last two species were represented by a low number of isolates. The Bacillus cereus group and Bacillus megaterium showed a lower percentage (30-70%) of isolates potentially able to cause the rope, but considering the high number of B. cereus group isolates detected in this study, this bacterial group should also be considered important in rope spoilage. In conclusion, results demonstrate that raw materials used to produce bread represent a rich source of spore-forming bacteria, therefore their microbiological quality should be monitored before use. Moreover, this study highlights for the first time the importance of the species B. amyloliquefaciens in rope spoilage and indicates that other species may also cause this alteration although strains of the same species may behave differently.

Towards the quantification of residual milk allergens in caseinate-fined white wines using HPLC coupled with single-stage Orbitrap mass spectrometry.

A method based on LC-ESI-high-resolution (HR)-MS analysis, using a single-stage Orbitrap mass spectrometer, was developed for the quantification of casein allergens potentially present in white wines as a result of fining by caseinate. The method consists of (1) extraction from the matrix by ultrafiltration, (2) digestion with trypsin and (3) detection/quantification of residual caseins, obtained by monitoring the LC-MS response of representative tryptic peptides (peak areas in extracted-ion chromatograms). Method linearity was assessed first on caseinate solutions prepared either in water or in wine matrix (the ultrafiltration residue of a protein-free white wine). Limits of detection (LOD) ranged from 0.1 to 0.3 µg ml(-1) (S/N = 3) in water, and between 0.15 and 0.7 µg ml(-1) in wine matrix, depending on the selected peptide. Method repeatability and reproducibility, measured as response variability (standard deviation) due to LC-MS analysis alone and to both enzymatic digestion and LC-MS analysis, were assessed on caseinate standard solutions in water and ranged from 5 to 12% and from 8 to 20%, respectively. A higher variability was usually observed for the peptide marker response in the case of matrix-matched samples, the only exception being peptide GPFPIIV from ß-casein, the marker also providing the highest sensitivity. The method was finally applied to a casein-free white wine ('Greco di Tufo') fined with caseinate at different concentrations, after discarding the precipitate due to casein-wine components aggregation. Minimum detectable added caseinate concentrations (i.e. those corresponding to responses with S/N = 3) were estimated between 39 and 51 µg ml(-1), according to the peptide marker chosen. These limits are compatible with caseinate concentrations typically adopted for wine-fining purposes. Moreover, a cross-check with the calibration performed in wine matrix led to an estimation of the concentration of dissolved caseinate to be in the low ng ml(-1) range.

Quantitative analysis of mycotoxins in cereal foods by collision cell fragmentation-high-resolution mass spectrometry: performance and comparison with triple-stage quadrupole detection.

A liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method for the simultaneous determination of aflatoxins (B(1), B(2), G(1), G(2)), ochratoxin A, deoxynivalenol, zearalenone, T-2 and HT-2 toxins in wheat flour, barley flour and crisp bread was developed. Mycotoxin fragmentation patterns obtained by high-energy collision dissociation (HCD) were investigated to obtain quantitative and confirmatory information (two characteristic masses per mycotoxin) using Orbitrap™-based high-resolution mass spectrometry. LC-HRMS (full-scan) detection carried out by HCD allows the monitoring of the pseudo-molecular ion and an additional characteristic fragment (for each mycotoxin) with mass accuracy in the range 0.1-3.9 ppm, meeting current European regulatory requirements for LC-MS confirmatory analysis. A sample preparation procedure based on polymeric solid-phase extraction cartridges was applied, allowing recoveries higher than 74% for nine mycotoxins, with a relative standard deviation lower than 13%. Detection limits in the range 0.5-3.4 µg kg(-1) were obtained for three cereal matrices. A critical comparison between the proposed method and a validated method based on triple quadrupole mass spectrometry showed similar performance in terms of detection limits, recoveries and repeatability, and matrix effects. Based on an efficient sample extraction and clean-up, the LC-HCD-HRMS method reported here represents a reliable and robust alternative tool for mycotoxin analysis in food matrices as compared with well-established triple quadrupole-based approaches.

Comparison of methods and optimisation of the analysis of fumonisins B1 and B2 in masa flour, an alkaline cooked corn product.

A comparison study of different extraction and clean-up procedures for the liquid chromatographic analysis of fumonisins B(1) (FB(1)) and B(2) (FB(2)) in corn masa flour was performed. The procedures included extraction (heat or room temperature) with acidic conditions or EDTA-containing solvents, and clean-up by immunoaffinity or C18 solid-phase extraction columns. Thereafter an analytical method was optimised using extraction with an acidic mixture of methanol-acetonitrile-citrate/phosphate buffer, clean-up through the immunoaffinity column and determination of fumonisins by liquid chromatography with automated pre-column derivatisation with o-phthaldialdehyde reagent. Recovery experiments performed on yellow, white and blue masa flours at spiking levels of 400, 800 and 1200 µg kg(-1) FB(1) and of 100, 200 and 300 µg kg(-1) FB(2) gave overall mean recoveries of 99% (±6%) for FB(1) and 88% (±6%) for FB(2). Good recoveries (higher than 90% for both FB(1) and FB(2)) were also obtained with corn tortilla chips. The limits of quantification of the method (signal-to-noise ratio of 10) were 25 µg kg(-1) for FB(1) and 17 µg kg(-1) for FB(2). The method was tested on different commercial corn masa flours as well as on white and yellow corn tortilla chips, showing fumonisin contamination levels (FB(1) + FB(2)) up to 1800 µg kg(-1) (FB(1) + FB(2)) in masa flour and 960 µg kg(-1) in tortilla chips. Over 30% of masa flours originating from Mexico exceeded the European Union maximum permitted level.

Serum elements and oxidative status in clinically isolated syndromes: imbalance and predictivity.

BACKGROUND: Metals are suspected of being involved in the pathogenesis of various neurologic diseases. We previously found a complex imbalance in serum chemical elements and oxidative status in patients with clinically definite multiple sclerosis (CDMS). OBJECTIVE: To understand whether this imbalance affects people with clinically isolated syndrome (CIS) and, if so, whether it predicts conversion to CDMS. METHODS: We studied 22 chemical elements and the oxidative status in 49 patients with CIS, 49 patients with CDMS, and 49 healthy donors (HD). Univariate and multivariate approaches were used to identify profiles for each group. A logistic regression analysis was used to identify the predictive potential of baseline data (elements, oxidative status, and MRI findings) for conversion to CDMS over 36 months. RESULTS: Several elements and oxidative status values differed significantly among the 3 groups. Discriminant analysis revealed a major contribution of Ca, Fe, Sn, Zn, serum antioxidant capacity, and serum oxidative status, which resulted in distinct profiles (the prediction of group membership was 96% [cross-validated 92%] for HD, 92% [cross-validated 92%] for CDMS, and 90% [cross-validated 86%] for CIS). A weighted combination of element concentrations and oxidative status values, adjusting for all other predictors, would predict a reduction in the risk of conversion to CDMS within 3 years (odds ratio 0.37; 95% confidence interval 0.18-0.76; p = 0.007), thereby proving more effective than MRI at baseline. CONCLUSIONS: The peculiar imbalance in serum elements and oxidative status that characterizes patients with CIS and may predict conversion to CDMS warrants studies on larger sample sizes.

Identification of allergenic milk proteins markers in fined white wines by capillary liquid chromatography-electrospray ionization-tandem mass spectrometry.

A method based on capillary liquid chromatography combined with electrospray ionization-tandem mass spectrometry (CapLC-ESI-MS-MS) for the detection and identification of casein deriving peptides in fined white wine is described. This is the first step towards the development of a liquid chromatography mass spectrometric method for the detection/identification of markers of potentially allergenic milk proteins used as wine fining agents. The method demonstrated to be capable of detecting some peptides arising from alpha and beta casein (with the relative aminoacidic sequences elucidated) in extracts of white wine fined with casein at 100 and 1000 microg/mL. This MS based approach appears to be a useful tool for screening purposes as well as a confirmatory tool for the unequivocal identification of caseins in ELISA positive samples.

Natural occurrence of fumonisin B2 in red wine from Italy.

The potential risk of exposure to fumonisin B(2) (FB(2)) in the grape-wine chain has recently been revealed after a report of Aspergillus niger in grapes and its ability to produce FB(2) and FB(4). The occurrence of these two fumonisins in wine was investigated by LC/MS/MS in 51 market samples (45 red, five white and one rose wine) produced in various Italian regions. Nine samples of red wine were found to be contaminated by fumonisin B(2) at levels ranging from 0.4 to 2.4 ng/ml, while FB(4) was not detected in any of the tested samples. This is the first report on the natural occurrence of FB(2) in wine, indicating that, although at low levels, there is a potential risk of FB(2) exposure for the wine-consumer.

Riluzole in cerebellar ataxia: a randomized, double-blind, placebo-controlled pilot trial.

BACKGROUND: The pleiotropic effects of riluzole may antagonize common mechanisms underlying chronic cerebellar ataxia, a debilitating and untreatable consequence of various diseases. METHODS: In a randomized, double-blind, placebo-controlled pilot trial, 40 patients presenting with cerebellar ataxias of different etiologies were randomly assigned to riluzole (100 mg/day) or placebo for 8 weeks. The following outcome measures were compared: proportion of patients with a decrease of at least 5 points in the International Cooperative Ataxia Rating Scale (ICARS) total score after 4 and 8 weeks compared with the baseline score; mean changes from the baseline to posttreatment ICARS (total score and subscores at 8 weeks); and occurrence of adverse events. RESULTS: Riluzole and placebo groups did not differ in baseline characteristics. The number of patients with a 5-point ICARS drop was significantly higher in the riluzole group than in the placebo group after 4 weeks (9/19 vs 1/19; odds ratio [OR] = 16.2; 95% confidence interval [CI ] 1.8-147.1) and 8 weeks (13/19 vs 1/19; OR = 39.0; 95% CI 4.2-364.2). The mean change in the riluzole group ICARS after treatment revealed a decrease (p < 0.001) in the total score (-7.05 [4.96] vs 0.16 [2.65]) and major subscores (-2.11 [2.75] vs 0.68 [1.94] for static function, -4.11 [2.96] vs 0.37 [2.0] for kinetic function, and -0.74 [0.81] vs 0.05 [0.40] for dysarthria). Sporadic, mild adverse events occurred. CONCLUSIONS: These findings indicate the potential effectiveness of riluzole as symptomatic therapy in diverse forms of cerebellar ataxia. CLASSIFICATION OF EVIDENCE: This study provides Class I evidence that riluzole reduces, by at least 5 points, the ICARS score in patients with a wide range of disorders that cause cerebellar ataxia (risk difference 63.2%, 95% CI 33.5%-79.9%).

HLA-class I markers and multiple sclerosis susceptibility in the Italian population.

Previous studies reported an association with multiple sclerosis (MS) of distinct HLA-class I markers, namely HLA-A*02, HLA-Cw*05 and MOG-142L. In this work, we tested the association with MS of A*02 and Cw*05 in 1273 Italian MS patients and 1075 matched controls, which were previously analyzed for MOG-142, and explored the relationship among these three markers in modulating MS risk. HLA-A*02 conferred a statistically robust MS protection (odds ratio, OR=0.61; 95% confidence intervals, CI=0.51-0.72, P<10(-9)), which was independent of DRB1*15 and of any other DRB1* allele and remained similar after accounting for the other two analyzed class I markers. Conversely, the protective effect we previously observed for MOG-142L was secondary to its linkage disequilibrium with A*02. Cw*05 was not associated considering the whole sample, but its presence significantly enhanced the protection in the HLA-A*02-positive group, independently of DRB1: the OR conferred by A*02 in Cw*05-positive individuals (0.22, 95% CI=0.13-0.38) was significantly lower than in Cw*05-negative individuals (0.69, 95% CI=0.58-0.83) with a significant (P=4.94 x 10(-5)) multiplicative interaction between the two markers. In the absence of A*02, Cw*05 behaved as a risk factor, particularly in combination with DRB1*03 (OR=3.89, P=0.0006), indicating that Cw*05 might be a marker of protective or risk haplotypes, respectively.

Effects of in vitro exposure to natural levels of zearalenone and its derivatives on chromatin structure stability in equine spermatozoa.

The purpose of this study was to assess the natural exposure of male horses (Equus caballus) to the mycotoxin zearalenone (ZEA) by using the ELISA test and to evaluate the effects of in vitro exposure of sperm cells to mycotoxin-containing urine extracts on sperm chromatin structure stability. Because of their occurrence in urine samples, ZEA and its derivatives were tested by sperm chromatin structure assay (SCSA) at natural levels detected by ELISA. Thirty-eight urine extracts of Italian (n = 11) and northeastern European (n = 27) horses were tested on frozen-thawed spermatozoa to evaluate the toxic effect of mycotoxin on their chromatin structure by flow cytometry. Different parameters of the DNA fragmentation index (DFI), such as the mean (X -DFI), the percentage (%-DFI), and the standard deviation (SD-DFI), were analyzed. Urine samples showed a mean level of 32.3 ng/mL ZEA with significantly higher concentrations in northeastern European samples than in Italian samples, probably in relation to climatic and feeding differences. The toxic effects of ZEA-containing urine samples on SCSA parameters were found at low ZEA concentrations and were mainly observed in Italian samples. By using mycotoxin standards, ZEA, alpha-zearalenol, and beta-zearalenol proved to be more toxic compounds for sperm chromatin stability than other tested derivatives. A nongenomic mechanism of action can be hypothesized.