A Chemically Defined Medium That Supports Mycotoxin Production by Stachybotrys chartarum Enabled Analysis of the Impact of Nitrogen and Carbon Sources on the Biosynthesis of Macrocyclic Trichothecenes and Stachybotrylactam.

Stachybotrys chartarum (Hypocreales, Ascomycota) is a toxigenic fungus that is frequently isolated from water-damaged buildings or improperly stored feed. The secondary metabolites formed by this mold have been associated with health problems in humans and animals. Several authors have studied the influence of environmental conditions on the production of mycotoxins, but these studies focused on undefined or complex substrates, such as building materials and media that impeded investigations of the influence of specific nutrients. In this study, a chemically defined cultivation medium was used to investigate the impact of several nitrogen and carbon sources on growth of S. chartarum and its production of macrocyclic trichothecenes (MTs) and stachybotrylactam (STLAC). Increasing concentrations of sodium nitrate were found to positively affect mycelial growth, the level of sporulation, and MT production, while ammonium nitrate and ammonium chloride had an inhibitory effect. Potato starch was the superior and most reliable carbon source tested. Additionally, we observed that the level of sporulation was correlated with the production of MTs but not with that of STLAC. In this study, we provide a chemically well-defined cultivation medium suitable for standardized in vitro testing of the capacity of S. chartarum isolates to produce macrocyclic trichothecenes. IMPORTANCE Macrocyclic trichothecenes (MTs) are highly toxic secondary metabolites that are produced by certain Stachybotrys chartarum strains, which consequently pose a risk for animals and humans. To identify hazardous, toxin-producing strains by analytical means, it is important to grow them under conditions that support MT production. Nutrients determine growth and development and thus the synthesis of secondary metabolites. Complex rich media are commonly used for diagnostics, but batch differences of supplements pose a risk for inconsistent data. We have established a chemically defined medium for S. chartarum and used it to analyze the impact of nitrogen and carbon sources. A key finding is that nitrate stimulates MT production, whereas ammonium suppresses it. Defining nutrients that support MT production will enable a more reliable identification of hazardous S. chartarum isolates. The new medium will also be instrumental in analyzing the biosynthetic pathways and regulatory mechanisms that control mycotoxin production in S. chartarum.

Microbial contamination and metabolite exposure assessment during waste and recyclable material collection.

Waste workers are exposed to bioaerosols when handling, lifting and dumping garbage. Bioaerosol exposure has been linked to health problems such as asthma, airway irritant symptoms, infectious, gastrointestinal and skin diseases, and cancer. Our objective was to characterize the exposure of urban collectors and drivers to inhalable bioaerosols and to measured the cytotoxic effect of air samples in order to evaluate their health risk. Personal and ambient air sampling were conducted during the summer of 2019. Workers from 12 waste trucks collecting recyclables, organic waste or compost were evaluated. Bacteria and fungi were cultured, molecular biology methods were used to detect microbial indicators, cytotoxic assays were performed and endotoxins and mycotoxins were quantified. Domestic waste collectors were exposed to concentrations of bacteria and endotoxins above the recommended limits, and Aspergillus section Fumigati was detected at critical concentrations in their breathing zones. Cytotoxic effects were observed in many samples, demonstrating the potential health risk for these workers. This study establishes evidence that waste workers are exposed to microbial health risks during collection. It also demonstrates the relevance of cytotoxic assays in documenting the general toxic risk found in air samples. Our results also suggest that exposures differ depending on the type of waste, job title and discharge/unloading locations.

Microbial contamination in waste collection: Unveiling this Portuguese occupational exposure scenario.

Previous studies anticipated that microorganisms and their metabolites in waste will increase as a consequence of a decreased collection frequency and due to differences in what kind of waste is bagged before collection leading to an increased exposure of workers handling the waste. This study aim was to investigate the microbial contamination present in the waste collection trucks (WCT) and in the support facilities (waste collection station - WCS). It was applied a multi-approach protocol using active (air sampling by impingement and impaction) and passive (surface swabs, electrostatic dust cloths and settled dust) sampling methods. The screening of azole-resistance, the investigation of mycotoxins and the assessment of the elicited biological responses in vitro were also carried out aiming recognizing the possible health effects of waste collection drivers. SARS-CoV-2 detection was also performed. In WCS only air samples had contamination in all the four sampling sites (canteen, operational removal core, operational removal center, and administrative service). Among all the analyzed matrices from the WCT a higher percentage of total bacterial counts and Gram-was detected in swabs (66.93%; 99.36%). In WCS the most common species were Penicillium sp. (43.98%) and Cladosporium sp. (24.68%), while on WCT Aspergillus sp. (4.18%) was also one of the most found. In the azole resistance screening Aspergillus genera was not observed in the azole-supplemented media. SARS-CoV-2 was not detected in any of the environmental samples collected, but Aspergillus section Fumigati was detected in 5 samples. Mycotoxins were not detected in EDC from WCS, while in WCT they were detected in filters (N = 1) and in settled dust samples (N = 16). In conclusion, our study reveals that a comprehensive sampling approach using active and passive sampling (e.g. settled dust sampling for a representative mycotoxin evaluation) and combined analytic methods (i.e., culture-based and molecular) is an important asset in microbial exposure assessments. Concerning the waste collection exposure scenario, the results of this study unveiled a complex exposure, particularly to fungi and their metabolites. Aspergillus section Fumigati highlight the significance of targeting this section in the waste management industry as an indicator of occupational health risk.

Bioburden contamination and Staphylococcus aureus colonization associated with firefighter's ambulances.

Ambulance vehicles are an essential part of emergency clinical services. Bioburden control in ambulances, through cleaning and disinfection, is crucial to minimize hospital-acquired infections, cross contamination and exposure of patients and ambulances' crew. In Portugal, firefighter crews are responsible, besides fire extinction, for first aid and urgent pre-hospital treatment. This study assessed the bioburden in Portuguese firefighters' ambulances with a multi-approach protocol using active and passive sampling methods. Fungal resistance profile and mycotoxins detection in ambulances' ambient, and S. aureus (SA) prevalence and resistance profile in ambulances' ambient and colonization in workers were also investigated. Toxigenic fungi with clinical relevance, namely Aspergillus section Fumigati, were found on ambulance's air in the hazardous dimension range. Interestingly, surface contamination was higher after cleaning in several sampling sites. Prevalence of S. aureus was 3% in environmental samples, of which 2% were methicillin-sensitive (MSSA) and 1% methicillin-resistant (MRSA). About 2.07 fungal species were able to grow in at least one azole, ranging from one (44% samples) to five (6% samples) species in each azole. Mycotoxins were detected in mops and electrostatic dust cloths. Colonization by S. aureus in the firefighter crew was observed with a high associated prevalence, namely 48%, with a 24% prevalence of MSSA (8/33) and 21% of MRSA (7/33). Additional studies are needed to determine the potential risk of infection transmission between different vehicle fleets and under varying conditions of use. This will strengthen the paramedic sector's mission to save lives without putting their own health and safety at risk.

Settled dust assessment in clinical environment: useful for the evaluation of a wider bioburden spectrum.

The collection and analysis of settled dust samples from indoor environments has become one of several environmental sampling methods used to assess bioburden indoors. The aim of the study was to characterize the bioburden in vacuumed settled dust from 10 Primary Health Care Centers by culture based and molecular methods. Results for bacterial load ranged from 1 to 12 CFU.g-1 of dust and Gram-negative bacteria ranged between 1 to 344 CFU.g-1 of dust. Fungal load ranged from 0 CFU.g-1 of dust to uncountable. Aspergillus section Fumigati was detected in 4 sampling sites where culture base-methods could not identify this section. Mucorales (Rhizopus sp.) was observed on 1 mg/L voriconazole. Three out of 10 settled dust samples were contaminated by mycotoxins. Settled dust sampling coupled with air sampling in a routine way might provide useful information about bioburden exposure.

Assessment of the microbial contamination of mechanical protection gloves used on waste sorting industry: A contribution for the risk characterization.

In Portugal, mechanical protection gloves (MPG) are of mandatory use and during their use sweat is released and, consequently, the humidity of the material increases leading to conditions favorable to the growth of microorganisms. However, no studies have been conducted in MPG to assess the bioburden. This study intended to determine the bioburden present in MPG and their biological effects, and to discuss the possibility to use MPG as a passive method to assess occupational exposure to microbial contamination. Fungal burden was characterized through molecular tools for fungal toxigenic species, and antifungal resistance and mycotoxins profiles were determined. Cell viability was determined in swine kidney (SK) monolayer and hepatocellular carcinoma (Hep G2) cell lines. All MPG samples presented Gram-negative bacteria. The fungal contamination ranged from 0 CFU.m-2 in both MEA and DG18, to 5.09 × 106 and 2.75 × 106 and the most commonly fungi found was Aspergillus spp. (50.46%). Azole resistant Aspergillus sections were found in azole supplemented media. Aspergillus sections (Circumdati, Flavi, Fumigati and Versicolores) were detected by molecular tools in 66 out of 67 samples. The most reported mycotoxin was mycophenolic acid (89.6%). HepG2 cells appear to be more sensitive to MPG contamination, with high cytotoxicity (IC50 < 0.05 mm2/ml) observed for 18 out of 57 gloves. MPG can be used in passive sampling to assess occupational exposure to bioburden in waste sorting industries and contribute for risk characterization. Some contaminants of MPG had cytotoxic potential and affected the biology of hepatic cells more than renal cells.

Exposure assessment in one central hospital: A multi-approach protocol to achieve an accurate risk characterization.

The bioburden in a Hospital building originates not only from patients, visitors and staff, but is also disseminated by several indoor hospital characteristics and outdoor environmental sources. This study intends to assess the exposure to bioburden in one central Hospital with a multi-approach protocol using active and passive sampling methods. The microbial contamination was also characterized through molecular tools for toxigenic species, antifungal resistance and mycotoxins and endotoxins profile. Two cytotoxicity assays (MTT and resazurin) were conducted with two cell lines (Calu-3 and THP-1), and in vitro pro-inflammatory potential was assessed in THP-1 cell line. Out of the 15 sampling locations 33.3% did not comply with Portuguese legislation regarding bacterial contamination, whereas concerning fungal contamination 60% presented I/O > 1. Toxigenic fungal species were observed in 27% of the sampled rooms (4 out of 15) and qPCR analysis successfully amplified DNA from the Aspergillus sections Flavi and Fumigati, although mycotoxins were not detected. Growth of distinct fungal species was observed on Sabouraud dextrose agar with triazole drugs, such as Aspergillus section Versicolores on 1 mg/L VORI. The highest concentrations of endotoxins were found in settled dust samples and ranged from 5.72 to 23.0 EU.mg-1. While a considerable cytotoxic effect (cell viability < 30%) was observed in one HVAC filter sample with Calu-3 cell line, it was not observed with THP-1 cell line. In air samples a medium cytotoxic effect (61-68% cell viability) was observed in 3 out of 15 samples. The cytokine responses produced a more potent average cell response (46.8 ± 12.3 ρg/mL IL-1ß; 90.8 ± 58.5 ρg/mL TNF-α) on passive samples than air samples (25.5 ± 5.2 ρg/mL IL-1ß and of 19.4 ± 5.2 ρg/mL TNF-α). A multi-approach regarding parameters to assess, sampling and analysis methods should be followed to characterize the biorburden in the Hospital indoor environment. This study supports the importance of considering exposure to complex mixtures in indoor environments.

Mycotoxins feed contamination in a dairy farm - potential implications for milk contamination and workers' exposure in a One Health approach.

BACKGROUND: Dairy farming feed can be contaminated with mycotoxins, affecting animals' health and milk quality. Dairy farming is also prone to occupational exposure to mycotoxins, and feed is recognized as a source of contamination in the workplace. An exploratory study was developed in a dairy farm located in Portugal intending to assess the mycotoxins present in the feed. RESULTS: All the samples analyzed presented contamination by at least two mycotoxins and up to a maximum of 13 mycotoxins in the same sample. Zearalenone (ZEA) was detected in all the samples (n = 10) followed by deoxynivalenol (DON), which was reported in eight samples, and ochratoxin A (OTA), reported in five samples. CONCLUSION: The results point to the possible contamination of milk by several mycotoxins and raise the possibility of occupational exposure to mycotoxins due to feed contamination. An adequate One Health approach for dairy production should address these issues through effective preventive actions such as avoiding the use of feed contaminated with mycotoxins. This represents an important challenge due to climate change. It requires proper attention and accurate management measures. © 2019 Society of Chemical Industry.

Analysis of Mycotoxins and Cytotoxicity of Airborne Molds Isolated from the Zoological Garden-Screening Research.

OBJECTIVE: The objective of this paper was to assess the airborne mold contamination, secondary metabolite profiles, and cytotoxicity of the dominant fungal species isolated from the air in selected rooms at a Zoological Garden. MATERIALS AND METHODS: Fungal concentrations were measured with MAS-100 air samplers. The collected airborne fungi were identified using a combination of morphological and molecular methods. The cytotoxicity of 84 strains belonging to two Penicillium and Aspergillus genera was determined using the quantitative colorimetric MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium salt) assay. The mycotoxins were detected using high-performance liquid chromatography (HPLC) with a mass spectrometry detector. RESULTS: The ITS gene was amplified and sequenced to identify the 132 species. For mycotoxicological and cytotoxicity analyses, 52 Penicillium isolates and 32 Aspergillus representatives were selected. Cytotoxicity was confirmed in 97.6% of cases analyzed. Using the LC-MS/MS method, 42 out of 84 strains produced at least one of the following toxins: ochratoxin A, ochratoxin B, patulin, gliotoxin, roquefortine C, griseofulvin, sterigmatocystin, fumonisin B2, moniliformin, and mycophenolic acid. CONCLUSIONS: Analytical methods for assessing the presence of mycotoxins in fungal isolates collected directly from the air have proven to be an effective tool. Our research provides new information on the occurrence of potentially toxin-producing molds within a zoo.

Microbial Assessment in A Rare Norwegian Book Collection: A One Health Approach to Cultural Heritage.

Microbial contamination poses a threat to both the preservation of library and archival collections and the health of staff and users. This study investigated the microbial communities and potential health risks associated with the UNESCO-classified Norwegian Sea Trade Archive (NST Archive) collection exhibiting visible microbial colonization and staff health concerns. Dust samples from book surfaces and the storage environment were analysed using culturing methods, qPCR, Next Generation Sequencing, and mycotoxin, cytotoxicity, and azole resistance assays. Penicillium sp., Aspergillus sp., and Cladosporium sp. were the most common fungi identified, with some potentially toxic species like Stachybotrys sp., Toxicladosporium sp., and Aspergillus section Fumigati. Fungal resistance to azoles was not detected. Only one mycotoxin, sterigmatocystin, was found in a heavily contaminated book. Dust extracts from books exhibited moderate to high cytotoxicity on human lung cells, suggesting a potential respiratory risk. The collection had higher contamination levels compared to the storage environment, likely due to improved storage conditions. Even though overall low contamination levels were obtained, these might be underestimated due to the presence of salt (from cod preservation) that could have interfered with the analyses. This study underlines the importance of monitoring microbial communities and implementing proper storage measures to safeguard cultural heritage and staff well-being.

Assessment of waste workers occupational risk to microbial agents and cytotoxic effects of mixed contaminants present in the air of waste truck cabin and ventilation filters.

Workers in the waste-processing industry are potentially exposed to high concentrations of biological contaminants, leading to respiratory and digestive problems and skin irritations. However, few data on the exposure of waste collection truck (WCT) drivers are available. The goal was to document the microbial risk of the waste collection truck (WCT) workers while in the vehicle cab. Long-period sampling using the truck air filters (CAF) and short time ambient air sampling in the cab were used. The potential release of microbial particles from CAFs was also investigated since it could contribute to the microbial load of the cabin air. A combination of analytical methods also helped assess the complex mixture of the biological agents. Aspergillus sections Fumigati and Flavi, E. coli, Enterobacter spp. and Legionella spp. were detected in the CAF of trucks collecting three types of waste. The highest levels of bacteria and fungi were found in the CAF from organic WCT. The highest endotoxin concentrations in CAF were 300 EU/cm2. Most of the CAF showed cytotoxic effects on both lung cells and hepatocytes. Only one mycotoxin was detected in a CAF. The maximal concentrations in the ambient WCT air varied according to the type of waste collected. The highest proportion (84%) of the air samples without cytotoxic effects on the lungs cells was for the recyclable material WCTs. The results revealed the potential microbial risk to workers from a complex mixture of bio-contaminants in the cabs of vehicles collecting all types of waste. The sustained cytotoxic effect indicates the potential adverse health-related impact of mixed contaminants (biological and non-biological) for the workers. Overall, this study highlights the benefits of using complementary sampling strategy and combined analytical methods for a the assessment of the microbial risk in work environments and the need to implement protective measures for the workers.Implications: Exposure to microbial agents is a well-known occupational hazard in the waste management sector. No previous study had evaluated the cytotoxicity of ambient air and ventilation filters to document worker exposure to a combination of contaminants during waste collection. This research confirms the usefulness of ventilation filters for long-term characterization of exposure to infectious agents, azole-resistant fungi, coliform bacteria and mycotoxin. Overall, this study highlights the importance of using several sampling and analysis methods for a comprehensive assessment of microbial risk in work environments, as well as the need to implement appropriate protective measures for collection workers.

Complementary sampling strategy and combined analytical methods are helpful in risk assessment.Air filter analysis (long-term sampling) assesses the presence of airborne biological contaminants over a long period.The type of waste collected influences the microbiological hazard of the workers.Waste collection workers are potentially exposed to infectious and mycotoxin-producing fungi.Cytotoxic assays revealed that waste collection workers are potentially.

Tea contamination by mycotoxins and azole-resistant mycobiota - The need of a One Health approach to tackle exposures.

Despite tea beneficial health effects, there is a substantial risk of tea contamination by harmful pathogens and mycotoxins. A total of 40 tea samples (17 green (raw) tea; 13 black (fermented) tea; 10 herbal infusions or white tea) were purchased from different markets located in Lisbon district during 2020. All products were directly available to consumers either in bulk (13) and or in individual packages (27). Bacterial analysis was performed by inoculating 150 µL of samples extracts in tryptic soy agar (TSA) supplemented with 0.2 % nystatin medium for mesophilic bacteria, and in Violet Red bile agar (VRBA) medium for coliforms (Gram-negative bacteria). Fungal research was performed by spreading 150 µL of samples in malt extract agar (MEA) supplemented with 0.05 % chloramphenicol and in dichloran-glycerol agar (DG18) media. The molecular detection of the Aspergillus sections Fumigati, Nidulantes, Circumdati and Flavi was carried out by Real Time PCR (qPCR). Detection of mycotoxins was performed using high performance liquid chromatograph (HPLC) with a mass spectrometry detector. Azole resistance screening was achieved following the EUCAST guidelines. The highest counts of total bacteria (TSA) were obtained in green raw tea (81.6 %), while for coliform counts (VRBA) were found in samples from black raw tea (96.2 %). The highest fungal counts were obtained in green raw tea (87.7 % MEA; 69.6 % DG18). Aspergillus sp. was the most prevalent genus in all samples on MEA (54.3 %) and on DG18 (56.2 %). In the raw tea 23 of the samples (57.5 %) presented contamination by one to five mycotoxins in the same sample. One Aspergillus section Fumigati isolate from green tea beverage recovered form itraconazole-Sabouraud dextrose agar (SDA) medium, presented itraconazole and posaconazole E-test MICs above MIC90 values. Our findings open further discussion regarding the One-Health approach and the necessary investment in researching biological hazards and azole-resistance associated with the production and consumption of tea (in particular green tea).

Microbial contamination in grocery stores from Portugal and Spain - The neglected indoor environment to be tackled in the scope of the One Health approach.

Microbial contamination in grocery shops (GS) should be evaluated since food commodities are commonly handled by workers and customers increasing the risk of food contamination and disease transmission. The aim of this study was to evaluate the microbial contamination in Portuguese and Spanish GS with a multi-approach protocol using passive (electrostatic dust cloths and surface swabs) sampling methods. The molecular detection of Aspergillus sections, mycotoxin analysis, screening of azole resistance as well as cytotoxicity measurement were conducted to better estimate the potential health risks of exposure and to identify possible relations between the risk factors studied. Fruits/vegetables sampling location was the one identified has being the most contaminated (bacteria and fungi) area in GS from both countries. Aspergillus section Fumigati and Fusarium species were observed in samples from Portuguese groceries with reduced susceptibilities to azoles commonly used in the clinical treatment of fungal infections. Fumonisin B2 was detected in Portuguese GS possible unveiling this emergent threat concerning occupational exposure and food safety. Overall, the results obtained raise concerns regarding human health and food safety and must be surveilled applying a One Health approach.

Six Feet under Microbiota: Microbiologic Contamination and Toxicity Profile in Three Urban Cemeteries from Lisbon, Portugal.

Cemeteries are potential environmental reservoirs of pathogenic microorganisms from organic matter decomposition. This study aimed to characterize the microbial contamination in three cemeteries, and more specifically in grave diggers' facilities. One active sampling method (impingement method) and several passive sampling methods (swabs, settled dust, settled dust filters and electrostatic dust cloths-EDC) were employed. The molecular detection of Aspergillus sections and SARS-CoV-2, as well as mycotoxin analysis, screening of azole resistance, and cytotoxicity measurement were also conducted. Total bacteria contamination was 80 CFU·m-2 in settled dust samples, reached 849 CFU·m-2 in EDC and 20,000 CFU·m-2 in swabs, and ranged from 5000 to 10,000 CFU·m-2 in filters. Gram-negative bacteria (VRBA) were only observed in in settled dust samples (2.00 × 105 CFU·m-2). Regarding Aspergillus sp., the highest counts were obtained in DG18 (18.38%) and it was not observed in azole-supplemented SDA media. SARS-CoV-2 and the targeted Aspergillus sections were not detected. Mycophenolic acid was detected in one settled dust sample. Cytotoxic effects were observed for 94.4% filters and 5.6% EDC in A549 lung epithelial cells, and for 50.0% filters and 5.6% EDC in HepG2 cells. Future studies are needed in this occupational setting to implement more focused risk management measures.

The Evolution of the Satratoxin and Atranone Gene Clusters of Stachybotrys chartarum.

Stachybotrys chartarum is frequently isolated from damp building materials or improperly stored animal forage. Human and animal exposure to the secondary metabolites of this mold is linked to severe health effects. The mutually exclusive production of either satratoxins or atranones defines the chemotypes A and S. Based upon the genes (satratoxin cluster, SC1-3, sat or atranone cluster, AC1, atr) that are supposed to be essential for satratoxin and atranone production, S. chartarum can furthermore be divided into three genotypes: the S-type possessing all sat- but no atr-genes, the A-type lacking the sat- but harboring all atr-genes, and the H-type having only certain sat- and all atr-genes. We analyzed the above-mentioned gene clusters and their flanking regions to shed light on the evolutionary relationship. Furthermore, we performed a deep re-sequencing and LC-MS/MS (Liquid chromatography-mass spectrometry) analysis. We propose a first model for the evolution of the S. chartarum genotypes. We assume that genotype H represents the most ancient form. A loss of the AC1 and the concomitant acquisition of the SC2 led to the emergence of the genotype S. According to our model, the genotype H also developed towards genotype A, a process that was accompanied by a loss of SC1 and SC3.

Unveiling the Occupational Exposure to Microbial Contamination in Conservation-Restoration Settings.

Assuring a proper environment for the fulfillment of professional activities is one of the Sustainable Development Goals and is contemplated in the One Health approach assumed by the World Health Organization. This particular study is applied to an often neglected sector of our society-the conservators/restorers-despite the many health issues reported by these professionals. Three different specialties (textiles, paintings and wood sculpture) and locations were selected for evaluation by placement of electrostatic dust cloths. After treatment of the samples, bacterial and fungal contamination were assessed, as well as mycotoxin determination, the presence of azole-resistant strains and cytotoxicity of the microorganisms encountered. Bacteria were only present in one of medias used and showed relatively low numbers. The highest level of contamination by fungi was identified in one of the textiles settings. The textile area also showed the highest variability for fungi. Aspergillus sp. are one indicator of possible environmental issues, and A. sections Fumigati and Circumdati were particularly relevant in two of the settings and identified in all of them. No mycotoxins were detected and the large majority of the fungi identified were non-cytotoxic. Overall, these can be considered low-contaminated environments but attention should be given to the Aspergillus sp. contamination. Additional studies are needed not only to make these results more robust, but also to test if the environmental sampling alone is the best approach in a setting where there is very little movement and dust displacement and where professionals are in very close proximity to the artefacts being treated, which may suggest the existence of a micro-atmosphere worth evaluating and comparing to the obtained results.

The impact of the Fusarium mycotoxin deoxynivalenol on the health and performance of broiler chickens.

The aim of the present experiment was to investigate the effects of feeding grains naturally contaminated with Fusarium mycotoxins on morphometric indices of jejunum and to follow the passage of deoxynivalenol (DON) through subsequent segments of the digestive tract of broilers. A total of 45 1-d-old broiler chickens (Ross 308 males) were randomly allotted to three dietary treatments (15 birds/treatment): (1) control diet; (2) diet contaminated with 1 mg DON/kg feed; (3) diet contaminated with 5 mg DON/kg feed for five weeks. None of the zootechnical traits (body weight, body weight gain, feed intake, and feed conversion) responded to increased DON levels in the diet. However, DON at both dietary levels (1 mg and 5 mg DON/kg feed) significantly altered the small intestinal morphology. In the jejunum, the villi were significantly (P < 0.01) shorter in both DON treated groups compared with the controls. Furthermore, the dietary inclusion of DON decreased (P < 0.05) the villus surface area in both DON treated groups. The absolute or relative organ weights (liver, heart, proventriculus, gizzard, small intestine, spleen, pancreas, colon, cecum, bursa of Fabricius and thymus) were not altered (P > 0.05) in broilers fed the diet containing DON compared with controls. DON and de-epoxy-DON (DOM-1) were analyzed in serum, bile, liver, feces and digesta from consecutive segments of the digestive tract (gizzard, cecum, and rectum). Concentrations of DON and its metabolite DOM-1 in serum, bile, and liver were lower than the detection limits of the applied liquid chromatography coupled with mass spectrometry (LC-MS/MS) method. Only about 10 to 12% and 6% of the ingested DON was recovered in gizzard and feces, irrespective of the dietary DON-concentration. However, the DON recovery in the cecum as percentage of DON-intake varied between 18 to 22% and was not influenced by dietary DON-concentration. Interestingly, in the present trial, DOM-1 did not appear in the large intestine and in feces. The results indicate that deepoxydation in the present study hardly occurred in the distal segments of the digestive tract, assuming that the complete de-epoxydation occurs in the proximal small intestine where the majority of the parent toxin is absorbed. In conclusion, diets with DON contamination below levels that induce a negative impact on performance could alter small intestinal morphology in broilers. Additionally, the results confirm that the majority of the ingested DON quickly disappears through the gastrointestinal tract.

Dietary Supplements Based on Red Yeast Rice-A Source of Citrinin?

Citrinin (CIT) is secondary metabolite of filamentous molds. This mycotoxin has nephrotoxic, hepatotoxic, embryocidal, and fetotoxic properties. It is also produced by several species of the three genera Penicillium spp., Aspergillus spp., and Monascus spp., which are used to make red yeast rice (RYR). The material for this study consisted of 15 dietary supplements containing an extract of fermented red rice, available on the Polish market. Samples were extracted using a MeOH-H2O mixture, cleaned-up with an immunoaffinity CitriTest HPLC column, and quantified by HPLC-FLD. None of the analyzed samples contained CIT above the established limit of detection (LOD). Studies on the presence of toxic metabolites in red yeast rice show the importance of regulating this product and of clear information on the label regarding the standardized amounts of monacolin.

Ochratoxin A levels in serum of Polish dialysis patients with chronic renal failure.

Ochratoxin A (OTA) is a mycotoxin produced by the fungi Aspergillus and Penicillium. It occurs naturally in many products of plant origin and in animals because of the carry-over from feed to meat or milk. Ochratoxin A has nephrotoxic, carcinogenic, hepatotoxic, neurotoxic, and genotoxic properties. Data on ochratoxin concentrations in blood or serum from patients with different kidney disorders are available for several European countries, as well as for Africa and Asia. In this study, we determined OTA concentrations in serum samples from chronic renal failure patients receiving dialysis and from healthy controls, collected in central Poland. Ochratoxin A was analyzed after extraction and purification using immunoaffinity columns by liquid chromatography with fluorescence detection (limit of quantification: 0.1 ng/mL) in 88 patients and 16 healthy volunteers. The dialysis group consisted of 40 women and 48 men aged between 23 and 85 years. The mean OTA concentrations were 0.75 ng/mL (maximum 2.78 ng/mL) in dialysis patients and 0.70 ng/mL (maximum 1.44 ng/mL) in healthy controls. The mean concentrations in patients treated by dialysis were 0.76 and 0.74 ng/ml for women and men, respectively (maximum 2.53 ng/ml for women and 2.78 ng/ml for men). Statistical analysis using Student's t-test showed no statistically significant differences between the control group (non-dialysis patients) and all dialysis patients.

Concentrations of zearalenone and its metabolites in female wild boars from woodlands and farmlands.

As a valuable food base and safe shelter, large agricultural fields create favourable living conditions for wild boars for most of the year. The occurrence of mould in these fields, causing hormonal disturbances, may lead to a lower age of sexual maturity and a prolonged breeding season, and therefore, the population growth in farmlands can be markedly higher than that in extensive woodlands. This study was initiated because of reproductive cycle disturbances in wild boar populations, which were presumably linked with habitat and food types, especially rotten maize. To determine if the major sources of zearalenone (ZEN) in female wild boars are maize cobs infected with mycotoxins, we compared the concentrations of ZEN and its metabolites in organs, tissues and body fluids of wild boars hunted in extensive maize fields and in extensive woodlands where no large maize fields were located within a distance of several km. Samples of blood, bile, liver, kidneys, muscles, urine, stomach and colon contents as well as ovaries from young female wild boars (40-60 kg) were collected in 2011-2014 and prepared for liquid chromatography tandem mass spectrometry (HPLC-MS/MS). The results showed that ZEN was present in most of the samples, from both farmlands and woodlands, but its concentration within individual types of the analysed body fluids or tissues differed depending mainly on habitat type. In nearly all the analysed cases, higher concentrations of ZEN and its derivatives were detected in the samples collected from wild boars in farmlands, suggesting that wild boars living and feeding in extensive maize fields take in markedly higher amounts of mycoestrogens.