Ochratoxin A Production Research Articles

Survey on the occurrence of Aspergillus section Nigri species in grapes cultivated in Umbria (central Italy) and influence of several factors on their distribution

AbstractSecondary bunch rots caused by Aspergillus species belonging to the section Nigri, commonly called black aspergilli, pose a serious threat to grapes yield and quality. Due to their ability to produce ochratoxin A (OTA), they are considered the primary source of contamination of this mycotoxin in grapes. In this study, a three-year survey was realized to assess the occurrence of black aspergilli in grapes harvested in the Umbria region, a very important grapevine cultivation area in central Italy, by fungal isolation and phylogenetic analysis. In addition, the influence of growing season, vineyard location, farming system and grapes growth stages on black aspergilli distribution was examined. Phylogenetic analysis of the 208 obtained isolates showed the presence of three black aspergilli species: Aspergillus tubingensis, Aspergillus welwitschiae and Aspergillus uvarum. Interestingly, A. carbonarius, one of the most important OTA producers, was not detected in the surveyed years. A. tubingensis, whose ability to produce OTA is controversial, was always the most isolated species followed by A. welwistichiae (OTA producer). The species A. uvarum (unable to produce OTA) was detected only in one surveyed year with a low incidence. A. tubingensis was always the species with the highest incidence, regardless of location, and farming system. Growth stage appeared to influence the incidence of the three species, that, in the case of A tubingensis, was lower during setting in comparison to berries pea-size and berries harvest-ripe. Finally, in the setting and berries pea-size stages, a stereomicroscope analysis, showed that flower debris was the substrate from which black aspergilli mainly developed prior to colonizing berries and bunches.

Essential oils from Cuminum cyminum and Laurus nobilis and their principal constituents: evaluation of antifungal and antimycotoxigenic potential in Aspergillus species.

The antifungal and antimycotoxigenic activities of the essential oils (EO) from Cuminum cyminum and Laurus nobilis, and their respective principal compounds, cuminaldehyde and 1,8-cineole, were evaluated against fungi of the genus Aspergillus: A.carbonarius, A.niger, A.ochraceus and A.westerdijkiae. The antifungal activity was determined by the contact method and the mycelial growth of the fungi was evaluated. Scanning Electron Microscopic (SEM) images were obtained to suggest modes of action of the compounds analyzed. The antimycotoxigenic activity was determined by HPLC. A.carbonarius was completely inhibited by cumin EO (500 µL L-1), by laurel EO and by cuminaldehyde (5000 µL L-1). The cumin EO (500 µL L-1) completely inhibited the growth of A.niger. All the samples inhibited the mycelial growth of A.ochraceus, especially cumin EO and cuminaldehyde (250 µL L-1). A.westerdijkiae was completely inhibited by cumin EO and cuminaldehyde (1000 µL L-1), by laurel EO and 1,8-cineole (10000 µL L-1). A decrease in the production of ochratoxin A (OTA) was observed post-treatment, except in A. ochraceus, only inhibited by laurel EO. SEM images showed morphological changes in fungal structures and spore inhibition post-treatment. The results confirmed the antifungal and antimycotoxigenic effect of EO and their principal constituents on fungi evaluated.

The Ecological Strategy Determines the Response of Fungi to Stress: A Study of the 2,4-diacetylphloroglucinol Activity Against Aspergillus and Fusarium Species.

Aspergillus and Fusarium are two economically important genera of fungi. They cause significant yield losses and contamination of crops with mycotoxins. In this study we aimed to evaluate the impact of 2,4-diacetylphloroglucinol (2,4-DAPG) on Aspergillus and Fusarium fungi. It is hypothesized that two fungal genera, which have different ecological strategies, react differently to stress caused by a secondary metabolite produced by rhizosphere Pseudomonas species. We found that 2,4-DAPG was able to reduce biofilm formation of Aspergillus and Fusarium, as reflected in biomass and its chemical composition. Furthermore, subinhibitory concentrations of 2,4-DAPG increased the levels of ergosterol and polysaccharides (α- and β-glucans, chitin) in the cell membrane and cell wall of Aspergillus, while decreasing them in Fusarium. 2,4-DAPG altered the production of secondary metabolites, especially mycotoxins and extracellular proteases. The production of ochratoxin A was decreased in A. ochraceus, and T-2 toxin and zearalenone, on the contrary, were increased in F. culmorum and F. sporotrichioides, respectively. Thus, using 2,4-DAPG we demonstrated that the ecological role of fungi determines their reaction to antibiotic substances produced by the plant microbiome. Our data contributes to understanding the molecular mechanisms behind symbiotic relationships in natural communities, which are mediated by the biosynthesis of antibiotics.

Investigating the Ochratoxin Production Potential of Aspergillus spp in Indian Rice: A Comprehensive Study

Rice (Oryza sativa), world’s second most important cereal crop and most commonly consumed grains in the world. Though more than 100,000 species of fungi are known to exist, majority of mycotoxigenic fungi belong to the species of Aspergillus, Penicillium, and Fusarium. Among Aspergillus spp, Aspergillus niger and Aspergillus ochraceus are known to produce Ochratoxin A (OTA), a notable mycotoxin having adverse effects due to ubiquitous presence, renal toxicity and lengthy persistence. Totally eighty-one samples including unpolished (n = 36) and polished rice samples (n = 45) were collected in various districts of Tamil Nadu to identify ochratoxigenic fungi particularly Aspergillus spp. Out of 81 samples, 62 % (50/81) of samples including unpolished (n = 24) and polished rice (n = 26) documented the occurrence of ochratoxigenic fungi. Extraction of OTA was carried out by agar plug method and the ability of fungus to produce OTA was quantified by RP HPLC-FLD analysis. Among the fungal genera, Aspergillus spp (80 %) recorded the predominant fungus in rice. Molecular confirmation of ochratoxigenic fungi A. niger and A. ochraceus isolates was performed by 18s rDNA analysis. OTA producing ability by RP HPLC-FLD analysis revealed A. ochraceus of section Circumdati recorded the highest OTA production than A. niger isolates of section Nigri. The concentration range of OTA by A. ochraceus vary between 12.33 - 196.84 ng/g and 0.18 - 2.82 ng/g respectively in A. niger isolates. Among the A. ochraceus isolates, potent isolate AO 9 documented the highest OTA production (196.84 ng/g) followed by AO 6 (104.74 ng/g). Similarly, in A. niger AN 1 showed highest production of 2.82 ng/g followed by AN 5 isolate (1.25 ng/g) respectively. The objective of the study is to determine and quantify the ochratoxin production potential of Aspergillus spp in Indian rice by RP HPLC-FLD analysis. The occurrence of Aspergillus spp in rice and the subsequent ochratoxin production raise concern about possible health risk to animal and human environment upon consumption. Hence there is need for improved storage practices and regular monitoring to prevent the prevalence of Aspergillus fungus and OTA contamination in rice supply chain.

Inhibition of Aspergillus spp. growth and ochratoxin A production in Conilon and Arabica coffees based-medium by Saccharomyces cerevisiae

Saccharomyces cerevisiae CCMA 0159 is reported as a promising biocontrol agent against ochratoxin A (OTA)-producing fungi in coffee. Coffea arabica and Coffea canephora (var. Conilon or Robusta) are the most widely consumed coffee species around the world, cultivated in tropical and subtropical regions, each exhibiting distinct physicochemical and sensory characteristics. The objective of this study was to compare the growth and OTA production by Aspergillus carbonarius, A. ochraceus, and A. westerdijkiae in C. arabica and C. canephora, along with assessing the efficiency of S. cerevisiae CCMA 0159 in biocontrolling ochratoxigenic fungi in both coffee varieties. A. carbonarius exhibited a higher growth rate and OTA production in both coffee varieties, with C. canephora showing particular susceptibility. Conversely, A. ochraceus and A. westerdijkiae demonstrated lower growth and OTA production. S. cerevisiae was effective in biocontrolling the fungal isolates, inhibiting over 80 % of A. carbonarius growth in both coffee varieties. Among the mechanisms of action of the biological control agent, the production of volatile organic compounds stands out. The results of this study confirm the significant potential of S. cerevisiae CCMA 0159 as a biocontrol agent against Aspergillus for application in coffee-producing areas.

Exploring a Cheese Ripening Process That Hinders Ochratoxin A Production by Penicillium nordicum and Penicillium verrucosum.

A lack of control of the technological abiotic parameters apparent during cheese manufacture, including temperature and relative humidity, results in this dairy product being prone to mold contamination. Sometimes, inoculant molds are used to obtain the characteristic sensory properties of this type of product. However, during the maturation process, some unwanted molds can colonize the ripening cheese and produce mycotoxins. Mycotoxigenic molds such as Penicillium nordicum and Penicillium verrucosum can colonize ripened cheeses, contaminating them with ochratoxin A (OTA), a nephrotoxic 2B toxin. Thus, the presence of OTA in cheeses could represent a hazard to consumers' health. This study has evaluated the growth and OTA production of P. nordicum and P. verrucosum on a cheese analogue under simulated ripening conditions of 10 and 15 °C and 0.96 water activity (aw). Ecophysiological, molecular, and analytical tools assessed the mold growth, gene expression, and OTA production under these environmental conditions. Both species were able to effectively colonize the cheese under these ripening conditions. However, neither species expressed the otapks and otanps biosynthetic genes or produced phenotypic OTA. Therefore, these results suggest a relatively low risk of exposure to OTA for consumers of this type of cheese product. The conditions used were thus appropriate for cheese ripening to minimize the potential for contamination with such mycotoxins. An appropriate adjustment of the technological ripening parameters during such cheese manufacture could contribute to OTA-free cheeses.

Saccharomyces cerevisiae CCMA 0159 showed a high capacity to inhibit OTA-producing Aspergillus species.

The use of yeast to biologically control the production of ochratoxin A, which is caused by filamentous fungal growth, in coffee farms is a promising alternative to conventional methods. However, yeast strains, environmental field conditions, and the chemical composition of grains / fruits can influence the effectiveness of biological control agents. We tested the efficiency of different yeast strains in controlling three species of ochratoxigenic fungi in a coffee-based medium. The reasons and mechanisms behind the varying sensitivities of different Aspergillus species to biological control remains unclear. Aspergillus carbonarius rapidly assimilated carbon sources, giving it an advantage over yeasts in substrate colonisation. Saccharomyces cerevisiae CCMA 0159 inhibited all three fungal species, achieving efficiencies of 74.4%, 100%, and 80.9% against A. carbonarius, A. westerdijkiae, and A. ochraceus, respectively. The strategies employed by S. cerevisiae CCMA 0159 to inhibit the growth of ochratoxigenic fungi included competition, production of fungicidal volatile compounds, and alteration of the substrate's physicochemical properties. Our results indicate that among the yeast isolates tested, S. cerevisiae CCMA 0159 is the most effective in inhibiting ochratoxigenic fungi in coffee, including the more resistant A. carbonarius.

Quantification using a HPLC-FLD-based Detector and Molecular Identifi-cation of Ochratoxin-A Producing Aspergillus Species Isolated from Stored Grain Samples.

Quantification using an HPLC-FLD based detector and Molecular identification of Ochratoxin-A producing Aspergillus Species isolated from stored grain samples. Fungi are cosmopolitan in origin and are known to grow in any suitable substra-tum. In the present investigation, Aspergillus species isolated from stored grain samples were analyzed for ochratoxin-A production. The objective of this study is the quantification of Ochratoxin and identification of ochratoxigenic fungi. A total of n=34 black Aspergilli and n=1 Ochre Aspergilli were isolated from grain samples of Bihar, India and it was tested for OTA production. The limit of detection (LOD) is found to be 0.33μg/Kg and the limit of quantification (LOQ) is found to be 1μg/Kg for OTA in HPLC-FLD. In the present study, out of all the fungal isolates, only TiB fungal isolate was able to produce the ochratoxin-A above the level of LOQ. The positive isolate TiB obtained from stored sesame seed samples was able to produce 25.54 μg/Kg of OTA. ITS sequence analysis of TiB isolate was able to matche 100% with Aspergillus welwitschiae and Aspergillus foetidus. This is the initial report of ochratoxigenic Aspergillus fungal isolate isolated from stored sesame seed samples of flood-prone areas of Bihar, India.

Development of a rapid method for determination of OchratoxinA in grape mash and wine.

The occurrence of Ochratoxin A (OTA) in wine is commonly known, but there is only limited information about its occurrence in grape mash and wines of German origin. Climate change has led to higher temperatures in the southern regions of Germany, which may increase the growth of fungi associated with the production of OTA and increase the content of this mycotoxin in grapes. A safe and rapid UHPLC-FLD method was developed and validated to assess the contamination of grape mash and wine with OTA. A total of 71 samples of grape mash and 30 wines from various wine producers in Baden-Württemberg, Germany, were analysed for OTA content. The results showed that no samples contained OTA in concentrations above the limit of detection. Further monitoring of samples from different vintages is needed.

Impact of OTAbZIP on Ochratoxin A production, mycelium growth and pathogenicity of Aspergillus westerdijkiae under water activity stress

ABSTRACT Aspergillus westerdijkiae is a major producer of ochratoxin A (OTA), a highly toxic and carcinogenic mycotoxin found in various food and feed products. A. westerdijkiae produces excessive amount of OTA under various water activity (aw) conditions that occur during food and feed storage. The biosynthetic gene clusters associated with OTA production include OTAbZIP, which plays a key role in controlling mycotoxin production in response to environmental conditions. This study explored the regulation of OTA biosynthesis in A. westerdijkiae fc-1, focusing on the OTAbZIP gene’s influence under aw stress. The mycelium growth of A. westerdijkiae fc-1 wild type and OTAbZIP mutant strains increased by 40.7% and 50.5% under high water activity (0.96 aw) respectively, at 6 days post-inoculation (dpi), indicating a stress on A. westerdijkiae fc-1. While OTAbZIP mutant did not produce OTA under both high and moderate aW conditions. The wild type produced OTA and OTA biosynthetic gene expression levels were downregulated under high (0.96 aw) and moderate (0.91 aw) water activity. The expression level of hog1 gene in OTAbZIP mutant was significantly lower than in the wild type. Pathogenicity tests revealed that deletion of OTAbZIP did not significantly affect disease infection. This study shows that deleting OTAbZIP gene greatly reduces OTA production, affecting the strain’s adaptability to water activity stress.

Ochratoxin A Contamination of California Pistachios and Identification of Causal Agents.

Ochratoxin A (OTA) is a potent mycotoxin produced by Aspergillus and Penicillium spp., which contaminates many crops, including pistachios. Pistachios contaminated with OTA may be subjected to border rejections resulting in significant economic losses to the United States agricultural revenues. The current study examined prevalence of OTA in California-grown pistachios and identified its causal agents. OTA was detected in 20% of samples from 2018 to 2021 (n = 809), with 18% of samples exceeding the European Union regulatory limit of 5 μg/kg. Fungi potentially responsible for OTA contamination were isolated from leaves, nuts, and soil collected from 14 pistachio orchards across California. A total of 1,882 isolates of Aspergillus section Nigri and 85 isolates of section Circumdati were recovered. Within section Nigri, 216 (11.5%) isolates were identified as potential OTA producers using a boscalid-resistance assay. Phylogenetic analyses of partial gene sequences for β-tubulin and calmodulin genes resolved section Circumdati into four species: A. ochraceus (33%), A. melleus (28%), A. bridgeri (21%), and A. westerdijkiae (19%). A. westerdijkiae produced the highest levels of OTA in inoculated pistachios (47 μg/g), followed by A. ochraceus (9.6 μg/g) and A. melleus (3.3 μg/g). A. bridgeri did not produce OTA. OTA production by section Circumdati was optimal from 20 to 30°C. All 216 boscalid-resistant isolates from section Nigri were identified as A. tubingensis, and representative isolates (n = 130) produced 3.8 μg/kg OTA in inoculated pistachios. This is the first detailed report on OTA contamination and causal fungi in California pistachios and will be helpful in devising effective management strategies.

First report of Aspergillus species in green pistachio of Bronte

Aspergillus contamination of pistachios causes significant product losses and potential presence of mycotoxins, particularly aflatoxin B1 (AFB1), and ochratoxin A (OTA). These toxins, which threaten human health, are strictly monitored by most nations. Italian pistachios produced in Bronte, Sicily, have high nutritional value and unique organoleptic properties, but the extent to which they contain these contaminants is unknown. Aspergillus spp. isolated from Bronte pistachios (cultivar Napoletana) were assessed for their ability to synthesize OTA or AFB1. Aspergillus occurrence in pistachio samples was measured at 1137 cfu g-1 for in shell pistachios and 770 cfu g-1 for kernels. The predominant isolated Aspergillus species was A. niger representing 74% of section Nigri (black isolates) and 47% of all Aspergillus isolates. Within section Flavi, A. flavus comprised 83% of green isolates. Only one black isolate (identified as A. carbonarius) had high OTA production, but all the A. flavus isolates had potential to produce AFG1 and AFB1, with AFB1 produced amount ranging from 0.1 to 8498 ng mL-1 of culture filtrate.

Production of Ochratoxin A and Citrinin and the Expression of Their Biosynthetic Genes from Penicillium verrucosum in Liquid Culture.

Penicillium verrucosum is a fungal pathogen capable of producing two mycotoxins of concern, ochratoxin A (OTA) and citrinin (CIT). The production profile of these two mycotoxins is not well understood but could help mitigate co-contamination in the food supply. As such, the production of OTA and CIT from P. verrucosum DAOMC 242724 was investigated under different growing conditions in liquid culture. We found that among the different liquid media chosen, liquid YES (yeast extract sucrose) medium induced the highest production of both OTA and CIT, when P. verrucosum DAOMC 242724 was cultured in stationary mode. Shake culture significantly reduced the amounts of OTA and CIT produced. Among all culture conditions tested, far greater amounts of CIT were produced compared to OTA. Consequently, upon transcriptomic data analysis, a statistically significant increase in the expression of CIT biosynthetic genes was easier to detect than the expression of OTA biosynthetic genes. Our study also revealed that the putative biosynthetic gene clusters of OTA and CIT in P. verrusocum DAOMC 242724 are likely distinct from each other. It appears that despite sharing a highly similar structure, the isocoumarin rings of OTA and CIT are each assembled by a specialized polyketide synthase enzyme. Our data identified a putative nonreducing polyketide synthase responsible for assembling the carbo-skeleton of CIT. In contrast, a highly reducing polyketide synthase appears to be involved in the biosynthesis of OTA.

Occurrence of ochratoxin A in Sichuan bacon from different geographical regions and characterization and biocontrol of ochratoxigenic Aspergillus westerdijkiae strain 21G2-1A

Sichuan bacon represents the most prevalent dry-cured meat product across Southwest China, but it is vulnerable to fungal spoilage. In the present study, a total of 47 Sichuan bacons were obtained from different regions of the Sichuan Province and analyzed for the presence of ochratoxin A (OTA), yielding a positive rate of 23.4 % (11/47). All the observed OTA concentrations exceeded the maximum admissible dose in meat products (1 μg/kg) established by some EU countries, with the highest OTA concentration being 250.75 μg/kg, which raises a food safety concern and reveals the need for a standardized scientific processing protocol. Then, an OTA-producing fungus named 21G2-1A was isolated from positive samples and found to be Aspergillus westerdijkiae. Further characterization suggested a positive correlation between fungal growth and OTA production. The optimal temperature for the former was 25 °C, while it was 20 °C for the latter. Although the A. westerdijkiae strain 21G2-1A demonstrated greater mycelium growth in the presence of NaCl, OTA production was significantly dismissed when the salinity was greater than 5 %. Four lactic acid bacteria (LAB) were screened out as antagonists against the ochratoxigenic fungus. In vitro evaluation of the antagonists revealed that live cells inhibited fungal growth, and adsorption also contributed to OTA removal at different levels. This study sheds some light on OTA control in Sichuan bacon through a biological approach.

Genomic analysis reveals genes that encode the synthesis of volatile compounds by a Bacillus velezensis-based biofungicide used in the treatment of grapes to control Aspergillus carbonarius

Fungal control strategies based on the use of Bacillus have emerged in agriculture as eco-friendly alternatives to replace/reduce the use of synthetic pesticides. Bacillus sp. P1 was reported as a new promising strain for control of Aspergillus carbonarius, a known producer of ochratoxin A, categorized as possible human carcinogen with high nephrotoxic potential. Grape quality can be influenced by vineyard management practices, including the use of fungal control agents. The aim of this study was to evaluate, for the first time, the quality parameters of Chardonnay grapes exposed to an antifungal Bacillus-based strategy for control of A. carbonarius, supporting findings by genomic investigations. Furthermore, genomic tools were used to confirm that the strain P1 belongs to the non-pathogenic species Bacillus velezensis and also to certify its biosafety. The genome of B. velezensis P1 harbors genes that are putatively involved in the production of volatiles and hydrolytic enzymes, which are responsible for releasing the free form of aroma compounds. In addition to promote biocontrol of phytopathogenic fungi and ochratoxins, the treatment with B. velezensis P1 did not change the texture (hardness and firmness), color and pH of the grapes. Heat map and hierarchical clustering analysis (HCA) of volatiles evaluated by GC/MS revealed that Bacillus-treated grapes showed higher levels of compounds with a pleasant odor descriptions such as 3-hydroxy-2-butanone, 2,3-butanediol, 3-methyl-1-butanol, 3,4-dihydro-β-ionone, β-ionone, dihydroactinidiolide, linalool oxide, and β-terpineol. The results of this study indicate that B. velezensis P1 presents desirable properties to be used as a biocontrol agent.

Biocontrol activities of yeasts or lactic acid bacteria isolated from Robusta coffee against Aspergillus carbonarius growth and ochratoxin A production in vitro

Biocontrol Agents (BCAs) can be an eco-friendly alternative to fungicides to reduce the contamination with mycotoxigenic fungi on coffee. In the present study, different strains of bacteria and yeasts were isolated from Ivorian Robusta coffee. Their ability to reduce fungal growth and Ochratoxin A (OTA) production during their confrontation against Aspergillus carbonarius was screened on solid media. Some strains were able to reduce growth and OTA production by 85 % and 90 % and were molecularly identified as two yeasts, Rhodosporidiobolus ruineniae and Meyerozyma caribbica. Subsequent tests on liquid media with A. carbonarius or solely with OTA revealed adhesion of R. ruineniae to the mycelium of A. carbonarius through Scanning Electron Microscopy, and an OTA adsorption efficiency of 50 %. For M. caribbica potential degradation of OTA after 24 h incubation was observed. Both yeasts could be potential BCAs good candidates for Ivorian Robusta coffee protection against A. carbonarius and OTA contamination.

Discovering potential biomarkers for Ochratoxin A production by Penicillium nordicum in dry-cured meat matrices through untargeted metabolomics

Ochratoxin A (OTA) is a mycotoxin produced by Penicillium nordicum, being this species a notable producer in dry-cured meat products. This toxin is one of the main fungal contaminants found in a variety of foods, including ripened meat products. Due to its harmful effects on health, it is essential to develop strategies from different approaches to identify whether products are contaminated and try to predict mycotoxin production during maturation to establish necessary corrective measures. In this study, targeted and untargeted metabolomic analyses were conducted to identify differential metabolites produced by three P. nordicum strains. These metabolites could be related to OTA production in agar-ham and dry-fermented sausage agar media under different water activity conditions. A Q Exactive™ Plus Hybrid Quadrupole-Orbitrap™ Mass Spectrometer was used to identify potential biomarkers and predictor-variables associated with OTA, employing: six comparatives that covered various aspects before and after OTA emergence, two analysis platforms (RStudio and MetaboAnalyst) and two approaches (imputation and non-imputation). The analysis was performed following two strategies, a Univariate and a Multivariate Statistical Analysis. As a result, several significant metabolites were identified as significantly linked to OTA production, including Asenjonamide C, (S,S)-Anacine, SPF-32629 A and Pseudouridine, which could potentially serve as biomarkers for OTA production. The study also highlighted the importance of using bioinformatic platforms, such as RStudio and MetaboAnalyst, for comprehensive analysis when integrating multiple outputs. This study paves the way for OTA prevention and taking corrective actions within the Hazard Analysis and Critical Control Points (HACCP) framework, after pointing out metabolites associated with OTA production, which deserve to be further studied in hams undergone to industrial ripening.

Practical Strategies to Reduce Ochratoxin A in Foods.

Ochratoxin A (OTA), a potent nephrotoxin, is one of the most deleterious mycotoxins, with its prevalence in agricultural crops and their processed foods around the world. OTA is a major concern to food safety, as OTA exposure through dietary intake may lead to a significant level of accumulation in the body as a result of its long half-life (about 35 days). Its potent renal toxicity and high risk of exposure as well as the difficulty in controlling environmental factors OTA production has prompted the need for timely information on practical strategies for the food industry to effectively manage OTA contamination during food processing. The effects of various food processes, including both nonthermal and thermal methods, on the reduction in OTA were summarized in this review, with emphasis on the toxicity of residual OTA as well as its known and unknown degradation products. Since complete removal of OTA from foodstuffs is not feasible, additional strategies that may facilitate the reduction in OTA in food, such as adding baking soda and sugars, was also discussed, so that the industry may understand and apply practical measures to ensure the safety of its products destined for human consumption.

Genome analysis of Pseudomonas strain 4B with broad antagonistic activity against toxigenic fungi.

Pseudomonas sp. 4B isolated from the effluent pond of a bovine abattoir was investigated as antifungal against toxigenic fungi. The complete genome of Pseudomonas 4B was sequenced using the Illumina MiSeq platform. Phylogenetic analysis and genome comparisons indicated that the strain belongs to the Pseudomonas aeruginosa group. In silico investigation revealed gene clusters associated with the biosynthesis of several antifungals, including pyocyanin, rhizomide, thanamycin, and pyochelin. This bacterium was investigated through antifungal assays, showing an inhibitory effect against all toxigenic fungi tested. Bacterial cells reduced the diameter of fungal colonies, colony growth rate, and sporulation of each indicator fungi in 10-day simultaneous growing tests. The co-incubation of bacterial suspension and fungal spores in yeast extract-sucrose broth for 48h resulted in reduced spore germination. During simultaneous growth, decreased production of aflatoxin B1 and ochratoxin A by Aspergillus flavus and Aspergillus carbonarius, respectively, was observed. Genome analysis and in vitro studies showed the ability of P. aeruginosa 4B to reduce fungal growth parameters and mycotoxin levels, indicating the potential of this bacterium to control toxigenic fungi. The broad antifungal activity of this strain may represent a sustainable alternative for the exploration and subsequent use of its possible metabolites in order to control mycotoxin-producing fungi.

Effects of temperature on the production of ochratoxin A by Aspergillus niger

This study aimed to investigate the effects of temperature on OTA production by Aspergillus niger and characterized the underlying regulatory metabolism. The results indicated that 23 °C induced OTA production, when compared with a non-inducing temperature of 30 °C. Genes involved in the OTA biosynthesis cluster, including p450, nrps, and hal, were up-regulated; genes related to antioxidants, such as cat, pod, and gstA, were down-regulated; and genes participating in oxidative phosphorylation, including those encoding succinate dehydrogenase and ATP synthase, were up-regulated at 23 °C, when compared to 30 °C. Correspondingly, the content of H2O2 and O2−, the sensitivity to H2O2, the mitochondrial complex II activity and ATP content were increased at 23 °C. These results indicated that temperature altered the cellular redox homeostasis and oxidative phosphorylation, and up-regulated expressions of OTA biosynthesis genes, thus ultimately triggering OTA biosynthesis. Notably, the novel and most significantly up-regulated zinc finger transcriptional factor, An12g06480, which was essential for A. niger growth and OTA biosynthesis, was identified. In the An12g06480 deletion strain, OTA production was significantly inhibited, and decreased by 89.47%. In addition, deletion of An12g06480 decreased mitochondrial complex II activity and ATP content by 81.41% and 16.17%, indicating that An12g06480 positively affected OTA biosynthesis by regulating cellular oxidative phosphorylation.