Corrigendum to ‘Dietary aflatoxin B1 (AFB1) reduces growth performance, impacting growth axis, metabolism, and tissue integrity in juvenile gilthead sea bream (Sparus aurata)’. Aquaculture, volume 533, 25 February 2021, 736189

The authors regret the errors in a few table references within the text. Specifically, it should reads as follows within the following subsections/ page: 3.2. Blood analysis (page 5)info:eu-repo/semantics/publishedVersio

Evaluation of mycotoxin content in soybean (Glycine max l.) grown in Rwanda

Soybean is a critical food and nutritional security crop in Rwanda. Promoted by the Rwandan National Agricultural Research System for both adults and as an infant weaning food, soybean is grown by approximately 40% of households. Soybean may be susceptible to the growth of mycotoxin-producing moulds; however, data has been contradictory. Mycotoxin contamination is a food and feed safety issue for grains and other field crops. This study aimed to determine the extent of mycotoxin contamination in soybean, and to assess people’s awareness on mycotoxins. A farm-level survey was conducted in 2015 within three agro-ecological zones of Rwanda suitable for soybean production. Soybean samples were collected from farmers (n=300) who also completed questionnaires about pre-and post-harvest farm practices, and aflatoxin awareness. The concentration of total aflatoxin in individual soybean samples was tested by enzymelinked immunosorbent assay (ELISA) using a commercially-available kit. Other mycotoxins were analyzed using liquid chromatography-mass spectrometry (LCMS/ MS) on 10 selected sub samples. Only 7.3% of the respondents were aware of aflatoxin contamination in foods, but farmers observed good postharvest practices including harvesting the crop when the pods were dry. Using enzyme-linked immunosorbent assay (ELISA), only one sample had a concentration (11 μg/kg) above the most stringent EU maximum permitted limit of 4 μg/kg. Multi-mycotoxins liquid chromatography-mass spectrometry (LC-MS/MS) results confirmed that soybeans had low or undetectable contamination; only one sample contained 13μg/kg of sterigmatocystine. The soybean samples from Rwanda obtained acceptably low mycotoxin levels. Taken together with other studies that showed that soybean is less contaminated by mycotoxins, these results demonstrate that soybean can be promoted as a nutritious and safe food. However, there is a general need for educating farmers on mycotoxin contamination in food and feed to ensure better standards are adhered to safeguard the health of the consumers regarding these fungal secondary metabolites.Key words: soybean, safety, mould, aflatoxin, mycotoxins, sterigmatocystine, ELISA, LC-MS/MS, Rwand

Simultaneous determination of Deoxynivalenol, Deoxynivalenol-3-Glucoside and Nivalenol in wheat grains by HPLC-PDA with immunoaffinity column cleanup

Deoxynivalenol-3-glucoside (D3G) is a modified mycotoxin formed by the metabolism of plants through the conjugation of deoxynivalenol (DON) with glucose. Toxicology studies of D3G for human and animal health are still under investigation, and the development of practical and reliable methods for its direct determination, especially in cereal matrices, is of great importance. In the present study, a methodology for simultaneous determination of D3G, DON, and nivalenol (NIV) in wheat grains, using immunoaffinity column (IAC) cleanup, separation by C18 column and detection by ultraviolet (UV) absorption, was optimized and in-house validated. The results demonstrated adequate values of D3G recovery from IAC and spiked samples. Intraday precision, linearity, limit of detection and limit of quantification (LOQ) were also adequate for the determination of these mycotoxins. Range of applicability varied from 47.1 to 1000 g/kg for D3G and from 31.3 to 1000 g/kg for DON and NIV, with recovery ranging from 84.7±7.2 % to 112.3±8.1Felipe Trombete is grateful for a doctoral fellowship provided by the Brazilian Federal Agency for Support and Evaluation of Graduate Education (CAPES)

The Fusarium crown rot pathogen Fusarium pseudograminearum triggers a suite of transcriptional and metabolic changes in bread wheat (Triticum aestivum L.)

Background and Aims: Fusarium crown rot caused by the fungal pathogen Fusarium pseudograminearum is a disease of wheat and barley, bearing significant economic cost. Efforts to develop effective resistance to this disease have been hampered by the quantitative nature of resistance and a lack of understanding of the factors associated with resistance and susceptibility. Here, we aimed to dissect transcriptional responses triggered in wheat by F. pseudograminearum infection. Methods: We used an RNA-seq approach to analyse host responses during a compatible interaction and identified >2700 wheat genes differentially regulated after inoculation with F. pseudograminearum. The production of a few key metabolites and plant hormones in the host during the interaction was also analysed. Key Results: Analysis of gene ontology enrichment showed that a disproportionate number of genes involved in primary and secondary metabolism, signalling and transport were differentially expressed in infected seedlings. A number of genes encoding pathogen-responsive uridine-diphosphate glycosyltransferases (UGTs) potentially involved in detoxification of the Fusarium mycotoxin deoxynivalenol (DON) were differentially expressed. Using a F. pseudograminearum DON-non-producing mutant, DON was shown to play an important role in virulence during Fusarium crown rot. An over-representation of genes involved in the phenylalanine, tryptophan and tyrosine biosynthesis pathways was observed. This was confirmed through metabolite analyses that demonstrated tryptamine and serotonin levels are induced after F. pseudograminearum inoculation. Conclusions: Overall, the observed host response in bread wheat to F. pseudograminearum during early infection exhibited enrichment of processes related to pathogen perception, defence signalling, transport and metabolism and deployment of chemical and enzymatic defences. Additional functional analyses of candidate genes should reveal their roles in disease resistance or susceptibility. Better understanding of host responses contributing to resistance and/or susceptibility will aid the development of future disease improvement strategies against this important plant pathogen

Risk assessment of chronic dietary exposure to the conjugated mycotoxin deoxynivalenol-3-β-glucoside in the Dutch population

In this study, a risk assessment of dietary exposure to the conjugated mycotoxin deoxynivalenol-3-β-glucoside (DON-3G) in the Dutch population was conducted. Data on DON-3G levels in food products available in the Netherlands are scarce. Therefore, data on co-occurring levels of DON-3G and deoxynivalenol (DON), its parent compound, were used to estimate the DON-3G/DON ratio for several food product categories. This resulted in a DON-3G/DON ratio of 0.2(90% confidence interval (CI):0.04-0.9) in grains & grain-milling products, 0.3 (90% CI: 0.03-2.8) in grain-based products and 0.8 (90% CI: 0.4-1.8) in beer. These ratios were applied to the Dutch monitoring data of DON to estimate the DON-3G concentrations in food products available in the Netherlands. DON and DON-3G concentrations were combined with food consumption data of two Dutch National Food Consumption Surveys to assess chronic exposure in young children (2-6 years), children (7-16 years) and adults (17-69 years) using the Monte Carlo Risk Assessment program. The chronic exposure levels of DON, DON-3G and the sum of both compounds (DON+DON-3G) were compared to the tolerable daily intake (TDI) of 1 μg/kg body weight/day which is based on the most critical effect of DON, namely decreased body weight gain. The assumption was made that DON-3G is deconjugated and then fully absorbed as DON in the gastro-intestinal tract. Exposure (P97.5) of the population aged 7-16 years and 17-69 years to DON or DON-3G separately, did not exceed the TDI. However, exposure to upper bound levels of DON+DON-3G (i.e. worst-case scenario) in the same age categories (P97.5) exceeded the TDI with a maximum factor of 1.3. Exposure (P97.5) of the 2-6 year-olds to DON was close to the TDI. Within this group, exposure (P97.5) to upper bound levels of DON+DON-3G exceeded the TDI with not more than a factor 2.</p

Biofilm mode of cultivation leads to an improvement of the entomotoxic patterns of two aspergillus species

Two fungi, i.e., Aspergillus flavus Link and Aspergillus oryzae (Ahlb.) E. Cohn, were cultivated according to two methodologies, namely submerged and biofilm cultures with the primary aim to use their secondary metabolites the supernatant CL50, and CL90 varied between 1.3% (v/v) to 12.7% (v/v) for incubation times from 24 to 72 h. While the A. flavus supernatant entomotoxicity was higher than this of A. oryzae, the biofilm culture application increased the efficiency of the former. Proteomic analysis of the supernatants revealed discrepancies among the two species and modes of cultivation. Furthermore, the secondary metabolite profiles of both Aspergillus cultures were verified. Aspergillic acid, beta-cyclopiazonic acid, cyclopiazonic acid, ferrineospergillin, flavacol, and spermadin A were most predominant. Generally, these secondary metabolites were present in higher concentrations in the supernatants of A. flavus and biofilm cultures. These molecular identifications correlated positively with entomotoxic activity. Noteworthy, the absence of carcinogenic aflatoxins was remarkable, and it will allow further valorization to produce A. flavus to develop potential biopesticides. © 2020 by the authors. Licensee MDPI, Basel, Switzerland

Incidence of toxigenic Aspergillus and Fusarium species occurring in maize kernels from Kenyan households

Aspergillus and Fusarium are fungal genera that include toxigenic and pathogenic species, able to suffuse farmers' crops and secrete an array of small molecular weight secondary metabolites which can cause health complications to humans and animals when ingested. In sub-Sahara Africa, contamination and persistence of these fungi is increased by the tropical climatic conditions which are ideal for the fungi to thrive. This study evaluated the incidence, regional distribution and toxigenic potential of Aspergillus and Fusarium species occurring in maize kernels from Eastern, Western, Coastal and the Lake Victoria agro-ecological zones of Kenya. Maize kernels were collected from 16 households in each agro-ecological zone. Single spore technique was used to isolate pure cultures of Aspergillus and Fusarium which were identified morphologically. Further, molecular analysis was done using the internal transcribed spacer 1 (ITS 1) region of the ribosomal DNA for Aspergillus and the translation elongation factor-1 alpha (TEF-1 alpha) for Fusarium. The potential of the isolated fungi to produce mycotoxins was probed by polymerase chain reaction (PCR) based on the aflatoxin regulatory aflaR gene in Aspergillus, and the fumonisin backbone structure gene FUM1 in Fusarium. Among the potentially aflatoxigenic A. flavus species isolated, 55% were from Eastern, 27% from the Coastal zone, 13% from Lake Victoria zone and 5% from Western Kenya. Among the potentially fumonisin producing F. verticillioides isolated, 45% were from the Lake Victoria agro-ecological zone, 30% were from Western, 15% from Eastern Kenya and 10% from the Coastal agro-ecological zone. This study adds data on potential mycotoxin hotspots in Kenya useful in employing targeted and regional mycotoxin mitigation strategies in efforts to avert future mycotoxicoses outbreaks in Kenya

Volatile organic compounds emitted by Aspergillus flavus strains producing or not aflatoxin B1

Aspergillus flavus is a phytopathogenic fungus able to produce aflatoxin B1 (AFB1), a carcinogenic mycotoxin that can contaminate several crops and food commodities. In A. flavus, two different kinds of strains can co-exist: toxigenic and non-toxigenic strains. Microbial-derived volatile organic compounds (mVOCs) emitted by toxigenic and non-toxigenic strains of A. flavus were analyzed by solid phase microextraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS) in a time-lapse experiment after inoculation. Among the 84 mVOCs emitted, 44 were previously listed in the scientific literature as specific to A. flavus, namely alcohols (2-methylbutan-1-ol, 3-methylbutan-1-ol, 2-methylpropan-1-ol), aldehydes (2-methylbutanal, 3-methylbutanal), hydrocarbons (toluene, styrene), furans (2,5-dimethylfuran), esters (ethyl 2-methylpropanoate, ethyl 2-methylbutyrate), and terpenes (epizonaren, trans-caryophyllene, valencene, alpha-copaene, beta-himachalene, gamma-cadinene, gamma-muurolene, delta-cadinene). For the first time, other identified volatile compounds such as alpha-cadinol, cis-muurola-3,5-diene, alpha-isocomene, and beta-selinene were identified as new mVOCs specific to the toxigenic A. flavus strain. Partial Least Square Analysis (PLSDA) showed a distinct pattern between mVOCs emitted by toxigenic and non-toxigenic A. flavus strains, mostly linked to the diversity of terpenes emitted by the toxigenic strains. In addition, the comparison between mVOCs of the toxigenic strain and its non-AFB1-producing mutant, coupled with a semi-quantification of the mVOCs, revealed a relationship between emitted terpenes (beta-chamigrene, alpha-corocalene) and AFB1 production. This study provides evidence for the first time of mVOCs being linked to the toxigenic character of A. flavus strains, as well as terpenes being able to be correlated to the production of AFB1 due to the study of the mutant. This study could lead to the development of new techniques for the early detection and identification of toxigenic fungi

Dietary aflatoxin B1 (AFB1) reduces growth performance, impacting growth axis, metabolism, and tissue integrity in juvenile gilthead sea bream (Sparus aurata)

Mycotoxins are an increasing threat to all the related commodities from agriculture. Its occurrence is expected to increase due to climate change. Here, we examined the impacts of dietary toxicity of aflatoxin B1 (AFB1) in gilthead sea bream (Sparus aurata) at levels of 1 or 2 mg AFB1 kg(-1)- fish feed. Inclusion of AFB1 in the diet resulted in 80% inhibition of the total weight gain during the 85-day trial. Carbohydrate and lipid energetic metabolites, both in plasma and liver, were depleted. Moreover, the histopathological analysis revealed several tissue anomalies in the liver, kidney, and spleen. Furthermore, the relative expression of gene transcripts for growth regulation was affected by AFB1. Adenohypophyseal gh and hepatic igf1 were inversely correlated due to AFB1 effects. Relative expression levels of gene transcripts as stress indicators were increased at AFB1 highest doses, such as hypothalamic trh, crh, and crhbp, as well as star in head kidney. Interestingly circulating levels of cortisol were unaffected. Overall, our results showed that aquafeeds with AFB1 impaired growth, alter metabolism, tissue integrity, and transcriptomic responses. However, we did find AFB1 residue neither in the liver nor muscle.Spanish Ministry of Economy and Business-MINECO [AGL2016-76069-C2-1-R]; Agencia Estatal de InvestigaciOn (MINECO, Spanish Government) [AGL2016-81808-REDT]; University of Cadiz Ph.D. scholarship [PIF UCA/RECO2VIT/2014]; Foundation for Science and Technology (FCT)Portuguese Foundation for Science and TechnologyEuropean Commission [UIDB/04326/2020

Genetic and toxigenic variability within Aspergillus flavus population isolated from maize in two diverse environments in Kenya

Aspergillus flavus is the main producer of carcinogenic aflatoxins in agricultural commodities such as maize. This fungus occurs naturally on crops, and produces aflatoxins when environmental conditions are favorable. The aim of this study is to analyse the genetic variability among 109 A. flavus isolates previously recovered from maize sampled from a known aflatoxin-hotspot (Eastern region, Kenya) and the major maize-growing area in the Rift Valley (Kenya), and to determine their toxigenic potential. DNA analyses of internal transcribed spacer (ITS) regions of ribosomal DNA, partial β-tubulin gene (benA) and calmodulin gene (CaM) sequences were used. The strains were further analyzed for the presence of four aflatoxin-biosynthesis genes in relation to their capability to produce aflatoxins and other metabolites, targeting the regulatory gene aflR and the structural genes aflP, aflD, and aflQ. In addition, the metabolic profile of the fungal strains was unraveled using state-of-the-art LC-MS/MS instrumentation. The three gene-sequence data grouped the isolates into two major clades, A. minisclerotigenes and A. flavus. A. minisclerotigenes was most prevalent in Eastern Kenya, while A. flavus was common in both regions. A. parasiticus was represented by a single isolate collected from Rift Valley. Diversity existed within the A. flavus population, which formed several subclades. An inconsistency in identification of some isolates using the three markers was observed. The calmodulin gene sequences showed wider variation of polymorphisms. The aflatoxin production pattern was not consistent with the presence of aflatoxigenic genes, suggesting an inability of the primers to always detect the genes or presence of genetic mutations. Significant variation was observed in toxin profiles of the isolates. This is the first time that a profound metabolic profiling of A. flavus isolates was done in Kenya. Positive associations were evident for some metabolites, while for others no associations were found and for a few metabolite-pairs negative associations were seen. Additionally, the growth medium influenced the mycotoxin metabolite production. These results confirm the wide variation that exists among the group A. flavus and the need for more insight in clustering the group