Section Circumdati Research Articles

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.

Reduction in Ochratoxin A Occurrence in Coffee: From Good Practices to Biocontrol Agents.

Ochratoxin A (OTA) is a mycotoxin mainly produced by Aspergillus section Circumdati and section Nigri across the coffee chain. OTA is nephrotoxic and is a threat to human health. This review summarizes current knowledge on how to reduce OTA concentration in coffee from farm to cup. After a brief introduction to the OTA occurrence in coffee, current good management practices are introduced. The core of this review focuses on biocontrol and microbial decontamination by lactic acid bacteria, yeasts and fungi, and their associated enzymes currently reported in the literature. Special attention is given to publications closest to in vivo applications of biocontrol agents and microbial OTA adsorption or degradation agents. Finally, this review provides an opinion on which future techniques to promote within the coffee supply chain.

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.

Assessment of mycobiota in Thai pigmented rice: Insights into ochratoxin A and citrinin production by Aspergillus and Penicillium species

Pigmented rice has gained interest for its nutritional and health benefits attributed to its bioactive compounds. Despite these advantages, the vulnerability of pigmented rice to mycotoxin contamination poses a major concern. This study examined the mycobiota of seven Thai pigmented rice varieties, with a focus on fungi capable of producing ochratoxin A (OTA) and citrinin. The study analyzed 110 pigmented rice samples and found that the Aspergillus section Aspergillus was predominant (50–82.35%), followed by Penicillium species (12.50–70.59%). One isolate of A. carbonarius from Aspergillus section Nigri (1/112) and two isolates from Aspergillus section Circumdati (2/61)—A. steynii and A. flocculosus—produced OTA. However, none of the Penicillium species produced OTA, though citrinin production was notable in P. citrinum (128/280). The study found that the Black Jasmine rice and Red Jasmine rice varieties were more susceptible to mycotoxin production, while the Luem Pua glutinous rice and Hom Nil rice varieties displayed more resistance to the accumulation of both mycotoxins. These findings should guide targeted strategies to mitigate mycotoxin risks and ensure the safety of pigmented rice in the food chain.

Unveiling ochratoxin A and ochratoxigenic fungi in Brazilian artisanal Cheeses: Insights from production to consumption

Ochratoxin A (OTA) is a toxin produced by several Aspergillus species, mainly those belonging to section Circumdati and section Nigri. The presence of OTA in cheese has been reported recently in cave cheese in Italy. As artisanal cheese production in Brazil has increased, the aim of this study was to investigate the presence of ochratoxin A and related fungi in artisanal cheese consumed in Brazil.A total of 130 samples of artisanal cheeses with natural moldy rind at different periods of maturation were collected. Of this total, 79 samples were collected from 6 producers from Canastra region in the state of Minas Gerais, since this is the largest artisanal cheese producer region; 13 samples from one producer in the Amparo region in the state of São Paulo and 36 samples from markets located in these 2 states. Aspergillus section Circumdati occurred in samples of three producers and some samples from the markets. A. section Circumdati colony counts varied from 102 to 106 CFU/g. Molecular analysis revealed Aspergillus westerdijkiae (67 %) as the most frequent species, followed by Aspergillus ostianus (22 %), and Aspergillus steynii (11 %). All of these isolates of A. section Circumdati were able to produce OTA in Yeast Extract Sucrose Agar (YESA) at 25 °C/7 days. OTA was found in 22 % of the artisanal cheese samples, ranging from 1.0 to above 1000 µg/kg, but only five samples had OTA higher than 1000 µg/kg. These findings emphasize the significance of ongoing monitoring and quality control in the artisanal cheese production process to minimize potential health risks linked to OTA contamination.

Simultaneous detection of mycotoxigenic Aspergillus species of sections Circumdati and Flavi using multiplex digital PCR.

Populations of ochratoxin-producing Aspergillus section Circumdati species and aflatoxin-producing Aspergillus section Flavi species frequently coexist in soil and are the main sources of mycotoxin contamination of tree nuts. Identification of mycotoxigenic Aspergillus species in these sections is difficult using traditional isolation and culture methods. We developed a multiplex digital PCR (dPCR) assay to detect and quantify Aspergillus ochraceus, Aspergillus westerdijkiae, and Aspergillus steynii (section Circumdati), as well as Aspergillus flavus and Aspergillus parasiticus (section Flavi), in environmental samples based on species-specific calmodulin gene sequences. Relative quantification of each species by dPCR of mixed-species templates correlated with corresponding DNA input ratios. Target species could be detected in soil inoculated with conidia from each species. Non-target species of sections Circumdati, Flavi, and Nigri were generally not detectable using this dPCR method. Detected non-target species (Aspergillus fresenii, Aspergillus melleus, Aspergillus sclerotiorum, and Aspergillus subramanianii) were discernible from A. ochraceus in dual-template dPCR reactions based on differential fluorescence intensity.

Effect of post-harvest management practices on the mycobiome and ochratoxin A contamination of differently processed Robusta coffees from Ivory Coast

Ochratoxin A (OTA) is a secondary metabolite produced primarily by the genus Aspergillus sp. sections Circumdati and Nigri. It can accumulate in coffee at post-harvest stage. In the present study five different dry processed coffees were sampled from Ivory Coast at harvesting and after drying. The OTA was detected in all the samples, from 3.62 μg/kg of coffee just after harvest to a higher-level of contamination in the dried coffee, between 11.04 and 760.24 μg/kg of coffee. Metabarcoding was used to further study the changes in the mycobiota. The alpha and beta diversity analysis revealed the presence of unique and more diverse fungal communities on the coffee after drying compared to fresh harvested coffee. Specially, coffee dried for a longer period showed a higher diversity. Fungal species such as Hypopichia sp. or Bernettozyma sp., were replaced in the top ten abundant species after drying by species such as Kurtzmaniella sp., Meyerozyma sp. and Fusarium sp. The increasing abundance of the ochratoxigenic fungi, A. carbonarius was correlated to the increased concentration of OTA on coffee. The construction of the negative correlation network shows that some yeasts could be good candidates for the biocontrol of A. carbonarius. The combination of DNA metabarcoding and OTA quantification was effective at deciphering the post-harvest origin of the OTA contamination. Overall, this study highlights the changes in the mycobiota under different drying conditions. Future perspectives include developing actions of prevention and control of the contamination with OTA during the post-harvest stages.

TOXIN PRODUCING MICROMYCETES OF THE GENUS PENICILLIUM AND ASPERGILLUS ON BERRIES, GRAPES, AND TOMATO FRUITS IN SLOVAK STORES

In this study, contamination by microscopic fungi (focused on Penicillium and Aspergillus species) of 75 fruit samples (strawberries, blackberries, blueberries, raspberries, grapes) and 29 tomato samples, which were obtained from Slovak stores was investigated. Totally 93.3% of fruit samples were contaminated with micromycetes (70.6% Botrytis spp., 52.0% Cladosporium spp., 42.7% Penicillium spp., 13.3% Rhizopus spp., 8.0% Alternaria spp., 6.7% Mucor spp., 5.3% Aspergillus spp., 4.0% Fusarium spp., and 1.3% Epicoccum spp.) and 6.7% of samples were contaminated with yeasts. The presence of 15 species of the genus Penicillium in fruits, namely P. atramentosum, P. aurantiogriseum, P. brevicompactum, P. bialowiezense, P. citrinum, P. crustosum, P. digitatum, P. expansum, P. fellutanum, P. glabrum, P. hordei, P. italicum, P. kiamense, P. olsonii, and P. purpurescens was found. With the highest frequency, P. brevicompactum (12.0%) and P. expansum (10.7%) were isolated from fruits. Four isolates of the genus Aspergillus were isolated from fruit samples (A. section Clavati, A. section Flavi, and two isolates of A. section Nigri). Thin layer chromatography (TLC) was used for testing of mycotoxin production by selected fungal isolates. Overall, 93.3% of tested Penicillium isolates and one tested Aspergillus of section Clavati proved the ability to produce at least one of the tested mycotoxins. As for tomato samples, 93.1% were contaminated with microscopic filamentous fungi (79.3% Penicillium spp., 24.1% Botrytis spp. and Alternaria spp., 10.3% Cladosporium spp., Mucor spp. and Rhizopus spp., 6.9% Aspergillus spp. and 3.4% Fusarium spp.) and 6.9% samples were contaminated with yeasts. In 65.5% of tomato samples, the occurrence of 8 species of the genus Penicillium (P. brevicompactum, P. crustosum, P. expansum, P. griseofulvum, P. italicum, P. olsonii, P. sublectaticum, P. thomii), and two species of the genus Aspergillus (from section Circumdati and Flavi) was recorded. P. olsonii (58.6%) and P. griseofulvum (10.3%) were isolated with the highest frequency. All 7 tested Penicillium isolates (100.0%) and 1 isolate of the genus Aspergillus (section Flavi), which were obtained from tomatoes, had the ability to produce at least one of the tested mycotoxins. Aspergillus section Flavi isolate (from tomatoes of Slovak origin) produced 5.5 µg.mL-1 of aflatoxin B1, 4.2 µg.mL-1 of aflatoxin B2, 154.4 µg.mL-1 of aflatoxin G1, and 5.6 µg.mL-1 of aflatoxin G2 on 14th day of cultivation on liquid YES medium at 25±1 °C in darkness, and detected by high pressure liquid chromatography (HPLC).

87 Portuguese Groceries: A Potential Indoor Environment to Be Used as Public Health Sentinel for Fungal and Mycotoxins Contamination

Abstract This study aimed to characterize microbial contamination in Portuguese grocery stores. The research was carried out in 15 grocery stores in Cascais, Portugal. Electrostatic dust cloths (EDC) and surface swabs were used on 3 sampling sites: checkout, fruits/vegetable, and warehouse/dispenser. Fungal contamination was characterized by culture-based methods, through the inoculation onto two different culture media: malt extract agar (MEA) supplemented with chloramphenicol (0.05 %) and dichloran-glycerol agar (DG18). Screening of azole resistance was performed by inoculation of EDC extracts´ in azole-supplemented Sabouraud dextrose agar (SDA) media, according to EUCAST guideline. A total of 39 samples were screened for mycotoxins contamination. The highest fungal contamination was obtained from fruits/vegetables in swabs (76 % MEA; 71 % DG18). The most prevalent genera in swabs was Cladosporium sp. (54 % MEA). Regarding EDC, besides Penicillium sp. (63 %) the most prevalent were Aspergillus sections Circumdati (25 %) and Aspergilli (35 %). Penicillium sp. was highly prevalent in the checkout (61 % SDA; 87 % voriconazole), and warehouse/dispenser sectors (73% voriconazole; 52% posaconazole). Circumdati and Fumigati were the most prevalent Aspergillus sections in fruits/vegetables (73 % and 64 % in voriconazole, respectively). Four mycotoxins (fumonisin B1, B2, B3) were detected simultaneously in most of the 36 samples. Identifying the most critical workplaces in groceries concerning fungal and mycotoxins contamination is of upmost importance to human health (workers and consumers) when using a One Health approach.

Identification of Green Coffee Contaminated Microfungi of the Genus <i>Aspergillus</i> on the Basis of Polyphasic Approach

The Aspergillus species are widespread in the environment, able to grow at high temperatures and minimal humidity, including in regions with a hot tropical climate. Some species have the potential to produce toxins. This causes the risk of contamination by fungi of the genus Aspergillus and the mycotoxins (MT) produced by them of plant materials and food products, which is possible at any stage of production, transportation and storage. In the volume of coffee imported to the Russian Federation, 85% is accounted for by raw materials (green coffee), for which the risks of mold damage remain at all stages preceding the roasting stage. It is relevant to study the species composition and toxinogenic properties of Aspergillus spp., which contaminate food raw materials for the production of mass consumption foodstuffs. Contaminated products include coffee, which is one of the basic products of the consumer basket. Reliable data on species identification and toxigenic potential of Aspergillus spp. can be obtained only with an integrated approach based on polyphasic taxonomy. The purpose of this work is to study the species composition of fungi of the genus Aspergillus isolated from green coffee using an integrated approach based on polyphasic taxonomy. The species composition of fungi of the genus Aspergillus from the internal mycoflora of 16 samples of green coffee beans of Arabica and Robusta. The species belonging of the isolated 34 singlspore isolates of Aspergillus spp. was determined by cultural and morphological methods and confirmed by molecular genetic analysis, i.e., RT-PCR with DNA markers of conservative sequences (ITS, CaM, β-tub), studied in vitro profile of produced secondary toxic metabolites. The dominance of species of the Niger section was established (A. niger, 90%, and A. tubingensis, A. carbonarius); then, in decreasing order, the species of section Flavi followed (A. flavus, 100%), sections Circumdati (A. ochraceus, 40% and A. westerdijkiae, 60%). In section Fumigati there was one strain of A. fumigatus. Analysis of the profile of toxic metabolites by HPLC-MS/MS in the multi-detection mode showed the production of mycotoxins by the following species: A. niger – fumonisin B2 and ochratoxin A, A. flavus – aflatoxins B1 and B2 together with sterigmatocystin, A. westerdijkiae – ochratoxin A and penicillic acid, A. ochraceus – penicillic acid. Amounts of produced MT show a high toxinogenic potential of Aspergillus spp. Thus, 20 out of 34 strains produced significant amounts of dangerous, regulated mycotoxins: AFL B1, OTA, FB2. Non-toxinogenic isolates were represented by the species A. niger, A. carbonarius, A. tubingensis, A. flavus, and A. fumigatus. A study of the species composition and toxinogenic properties of green coffee contaminants of the genus Aspergillus using a polyphasic approach was carried out in Russia for the first time.

Deep Genotypic Species Delimitation of Aspergillus Section Flavi Isolated from Brazilian Foodstuffs and the Description of Aspergillus annui sp. nov. and Aspergillus saccharicola sp. nov.

Aspergillus section Flavi is a fungal group that is important in food because it contains spoilage and potentially aflatoxigenic species. Aflatoxins are metabolites that are harmful to human and animal health and have been recognized as the primary natural contaminant in food. Therefore, recognizing the biodiversity of this group in food is necessary to reduce risks to public health. Our study aimed to investigate the diversity of Aspergillus section Flavi isolated from Brazilian foodstuffs such as cassava, sugarcane, black pepper, paprika, Brazil nuts, yerba-mate, peanuts, rice, and corn. A polyphasic approach integrating phenotypic data and multilocus genotypic analyses (CaM, BenA, and RPB2) was performed for 396 strains. Two new species in the Aspergillus subgenus Circumdati section Flavi are proposed using maximum-likelihood analysis, Bayesian inference, and coalescence-based methods: Aspergillus saccharicola sp. nov. and Aspergillus annui sp. nov. A. saccharicola sp. nov. belongs to the series Flavi, is a potentially aflatoxigenic species (B1, B2, G1, and G2), closely related to Aspergillus arachidicola, and was found mostly in sugarcane. A. annui sp. nov. was isolated from samples of sweet paprika. To accommodate A. annui sp. nov., a new series Annuorum was proposed.

Small NRPS-like enzymes in Aspergillus sections Flavi and Circumdati selectively form substituted pyrazinone metabolites.

Aspergillus fungi produce mycotoxins that are detrimental to human and animal health. Two sections of aspergilli are of particular importance to cereal food crops such as corn and barley. Aspergillus section Flavi species like A. flavus and A. parasiticus produce aflatoxins, while section Circumdati species like A. ochraceus and A. sclerotiorum produce ochratoxin A. Mitigating these toxins in food and feed is a critical and ongoing worldwide effort. We have previously investigated biosynthetic gene clusters in Aspergillus flavus that are linked to fungal virulence in corn. We found that one such cluster, asa, is responsible for the production of aspergillic acid, an iron-binding, hydroxamic acid-containing pyrazinone metabolite. Furthermore, we found that the asa gene cluster is present in many other aflatoxin- and ochratoxin-producing aspergilli. The core gene in the asa cluster encodes the small nonribosomal peptide synthetase-like (NRPS-like) protein AsaC. We have swapped the asaC ortholog from A. sclerotiorum into A. flavus, replacing its native copy, and have also cloned both asaC orthologs into Saccharomyces cerevisiae. We show that AsaC orthologs in section Flavi and section Circumdati, while only containing adenylation-thiolation-reductase (ATR) domains, can selectively biosynthesize distinct pyrazinone natural products: deoxyaspergillic acid and flavacol, respectively. Because pyrazinone natural products and the gene clusters responsible for their production are implicated in a variety of important microbe-host interactions, uncovering the function and selectivity of the enzymes involved could lead to strategies that ultimately benefit human health.

Microbial Contamination in the Coffee Industry: An Occupational Menace besides a Food Safety Concern?

Respiratory abnormalities among workers at coffee roasting and packaging facilities have already been reported; however, little is known about microbiological contamination inside coffee production facilities. This study intends to assess the microbial contamination (fungi and bacteria) in two coffee industries from Brazil with a multi-approach protocol for sampling and for subsequent analyses using four main sources of samples: filtering respiratory protection devices (FRPD) used by workers, settled dust, electrostatic dust cloths (EDC) and coffee beans. The fungal contamination in the assessed industries was also characterized through the molecular detection of toxigenic species and antifungal resistance. Total bacteria contamination presented the highest values in FRPD collected from both industries (7.45 × 104 CFU·m−2; 1.09 × 104 CFU·m−2). Aspergillus genera was widespread in all the environmental samples collected and sections with clinical relevance (Fumigati) and with toxigenic potential (Nigri and Circumdati) were recovered from FRPD. Circumdati section was observed in 4 mg/mL itraconazole. Sections Circumdati (EDC, coffee beans and settled dust) and Nidulantes (EDC, coffee beans and FRPD) were detected by qPCR. Some of the targeted Aspergillus sections that have been identified microscopically were not detected by qPCR and vice-versa. Overall, this study revealed that microbial contamination is a potential occupational risk in the milling stage and should be tackled when assessing exposure and performing risk assessment. In addition, a multi-sampling campaign should be the approach to follow when assessing microbial contamination and FRPD should be included in this campaign. Occupational exposure to mycotoxins should be considered due to high fungal diversity and contamination. A One Health approach should address these issues in order to prevent consumption of coffee crops and beans infected by fungi and, more specifically, to avoid widespread azole resistance.

The Secondary Metabolites and Biosynthetic Diversity From Aspergillus ochraceus.

Aspergillus ochraceus, generally known as a food spoilage fungus, is the representative species in Aspergillus section Circumdati. A. ochraceus strains are widely distributed in nature, and usually isolated from cereal, coffee, fruit, and beverage. Increasing cases suggest A. ochraceus acts as human and animal pathogens due to producing the mycotoxins. However, in terms of benefits to mankind, A. ochraceus is the potential source of industrial enzymes, and has excellent capability to produce diverse structural products, including polyketides, nonribosomal peptides, diketopiperazine alkaloids, benzodiazepine alkaloids, pyrazines, bis-indolyl benzenoids, nitrobenzoyl sesquiterpenoids, and steroids. This review outlines recent discovery, chemical structure, biosynthetic pathway, and bio-activity of the natural compounds from A. ochraceus.

Respiratory Failure With Aspergillus submaranianii and Lymphoplasmacystic Vasculitis.

Aspergillus subramanianii is a very rare form of Aspergillus in section Circumdati. Aspergillus subramanianii has been rarely reported or ever published. IgG4 disease is an emerging disease entity that can be a diagnostic challenge. We report a critically ill patient who had invasive A subramanianii pulmonary aspergillosis and posthumous diagnosis of IgG subclass 4 interstitial lung disease.

Genome and Metabolome MS-Based Mining of a Marine Strain of Aspergillus affinis.

Aspergillus section Circumdati encompasses several species that express both beneficial (e.g., biochemical transformation of steroids and alkaloids, enzymes and metabolites) and harmful compounds (e.g., production of ochratoxin A (OTA)). Given their relevance, it is important to analyze the genetic and metabolic diversity of the species of this section. We sequenced the genome of Aspergillus affinis CMG 70, isolated from sea water, and compared it with the genomes of species from section Circumdati, including A. affinis’s strain type. The A. affinis genome was characterized considering secondary metabolites biosynthetic gene clusters (BGCs), carbohydrate-active enzymes (CAZymes), and transporters. To uncover the biosynthetic potential of A. affinis CMG 70, an untargeted metabolomics (LC-MS/MS) approach was used. Cultivating the fungus in the presence and absence of sea salt showed that A. affinis CMG 70 metabolite profiles are salt dependent. Analyses of the methanolic crude extract revealed the presence of both unknown and well-known Aspergillus compounds, such as ochratoxin A, anti-viral (e.g., 3,5-Di-tert-butyl-4-hydroxybenzoic acid and epigallocatechin), anti-bacterial (e.g., 3-Hydroxybenzyl alcohol, l-pyroglutamic acid, lecanoric acid), antifungal (e.g., lpyroglutamic acid, 9,12,13-Trihydroxyoctadec-10-enoic acid, hydroxyferulic acid), and chemotherapeutic (e.g., daunomycinone, mitoxantrone) related metabolites. Comparative analysis of 17 genomes from 16 Aspergillus species revealed abundant CAZymes (568 per species), secondary metabolite BGCs (73 per species), and transporters (1359 per species). Some BGCs are highly conserved in this section (e.g., pyranonigrin E and UNII-YC2Q1O94PT (ACR toxin I)), while others are incomplete or completely lost among species (e.g., bikaverin and chaetoglobosins were found exclusively in series Sclerotiorum, while asperlactone seemed completely lost). The results of this study, including genome analysis and metabolome characterization, emphasize the molecular diversity of A. affinis CMG 70, as well as of other species in the section Circumdati.

Storage fungi and ochratoxin A associated with arabica coffee bean in postharvest processes in Northern Thailand

Arabica coffee is one of the most popular varieties globally; however, during postharvest and storage, it can be contaminated by fungi which produce mycotoxins such as ochratoxin A (OTA). The goal of this study was to determine fungal contamination and OTA levels of arabica coffee produced from Chiang Rai and Chiang Mai provinces in Northern Thailand. Aspergillus and Penicillium were mainly isolated and evaluated from coffee samples in the postharvest process. Coffee cherries, parchment coffee, and green coffee beans were commonly found contaminated with fungi. Coffee beans were contaminated with Penicillium, Aspergillus section Flavi, Aspergillus section Nigri, and Aspergillus section Circumdati at 1.2–10%. Fungal contamination in parchment coffee was studied using three drying methods. The results showed that coffee dried in a greenhouse had higher fungal contamination than samples dried on concrete ground or on a bamboo table. Whether basic storage packaging of green coffee beans affects fungal contamination was also investigated. Green coffee beans in three packaging types showed similar levels of Penicillium, while of the three packaging types, coffee in a polypropylene woven bag showed significantly less Aspergillus than those in a ramie sack or a plastic mesh bag. Fourteen strains of Aspergillus section Circumdati were isolated from coffee samples and 5 isolates showed green-blue fluorescence under ultraviolet light suggesting a potential of OTA production. The morphology of Aspergillus section Circumdati isolates were studied and five isolates with similar morphological characters were further identified to the species level based on ITS and the β-tubulin gene regions. Sequencing and phylogenetic analysis suggested that four isolates were grouped with A. westerdijkiae and one isolate with A. melleus. Analysis of OTA in parchment coffee, green coffee beans, and roasted coffee was performed using HPLC. The results showed that OTA was detected in only one sample of roasted coffee at 0.39 μg kg−1.

Diversity of Mycobiota in Spanish Grape Berries and Selection of Hanseniaspora uvarum U1 to Prevent Mycotoxin Contamination.

The occurrence of mycotoxins on grapes poses a high risk for food safety; thus, it is necessary to implement effective prevention methods. In this work, a metagenomic approach revealed the presence of important mycotoxigenic fungi in grape berries, including Aspergillus flavus, Aspergillus niger aggregate species, or Aspergillus section Circumdati. However, A. carbonarius was not detected in any sample. One of the samples was not contaminated by any mycotoxigenic species, and, therefore, it was selected for the isolation of potential biocontrol agents. In this context, Hanseniaspora uvarum U1 was selected for biocontrol in vitro assays. The results showed that this yeast is able to reduce the growth rate of the main ochratoxigenic and aflatoxigenic Aspergillus spp. occurring on grapes. Moreover, H. uvarum U1 seems to be an effective detoxifying agent for aflatoxin B1 and ochratoxin A, probably mediated by the mechanisms of adsorption to the cell wall and other active mechanisms. Therefore, H. uvarum U1 should be considered in an integrated approach to preventing AFB1 and OTA in grapes due to its potential as a biocontrol and detoxifying agent.

Melinacidin-Producing Acrostalagmus luteoalbus, a Major Constituent of Mixed Mycobiota Contaminating Insulation Material in an Outdoor Wall.

Occupants may complain about indoor air quality in closed spaces where the officially approved standard methods for indoor air quality risk assessment fail to reveal the cause of the problem. This study describes a rare genus not previously detected in Finnish buildings, Acrostalagmus, and its species A. luteoalbus as the major constituents of the mixed microbiota in the wet cork liner from an outdoor wall. Representatives of the genus were also present in the settled dust in offices where occupants suffered from symptoms related to the indoor air. One strain, POB8, was identified as A. luteoalbus by ITS sequencing. The strain produced the immunosuppressive and cytotoxic melinacidins II, III, and IV, as evidenced by mass spectrometry analysis. In addition, the classical toxigenic species indicating water damage, mycoparasitic Trichoderma, Aspergillus section Versicolores, Aspergillus section Circumdati, Aspergillus section Nigri, and Chaetomium spp., were detected in the wet outdoor wall and settled dust from the problematic rooms. The offices exhibited no visible signs of microbial growth, and the airborne load of microbial conidia was too low to explain the reported symptoms. In conclusion, we suggest the possible migration of microbial bioactive metabolites from the wet outdoor wall into indoor spaces as a plausible explanation for the reported complaints.

Ochratoxin A and related fungi in Brazilian black pepper (Piper nigrum L.)

Ochratoxin A (OTA) is a mycotoxin with nephrotoxic, genotoxic, teratogenic and carcinogenic properties, produced by several species of Aspergillus, mainly those belonging to the A. section Circumdati and A. section Nigri. Although this toxin has been detected in spices and condiments, in black pepper (Piper nigrum L.) few studies have investigated the mycobiota (based on a molecular approach) and the presence of OTA in this food. The aim of this study was to investigate the presence of potentially ochratoxigenic species and ochratoxin A in black pepper marketed in Brazil, one of the largest producers in the world. A total of 60 samples of black pepper (29 in powder and 31 in grain) were collected in markets. The presence of OTA was investigated in black pepper samples using High-Performance Liquid Chromatography (HPLC), OTA was detected in 55% of the samples, with levels ranging from 0.05 to 13.15 μg/kg, all of which were below the Brazilian legal tolerances. A. section Nigri and A. section Circumdati were found in 80% of the samples, but the species of A. section Nigri were significantly more frequent than those of A. section Circumdati. The potential for OTA production by fungal isolates was tested using the agar plug technique and confirmed by HPLC. Among the isolates belonging to A. section Nigri (n = 1,083) and A. section Circumdati (n = 129), 3.7% and 3.8%, respectively, were able to produce OTA in Yeast Extract Sucrose Agar (YESA). A total of 25 strains from A. section Circumdati and 64 from A. section Nigri were identified using molecular data. The following potentially ochratoxigenic species were found in black pepper: A. niger, A. welwitschiae, A. carbonarius, A. westerdijkiae and A. ochraceus. The occurrence of these species denotes the need for continuous monitoring of black pepper by regulatory bodies in order to safeguard consumers' health.