Non-toxigenic Aspergillus Research Articles

Aflatoxin Contamination, Mycoflora Diversity, and the Toxigenic Potential of Aspergillus spp. Associated with Maize Seeds and their Products

Aflatoxins are naturally occurring mycotoxins that are virtually impossible to eradicate from food and animal feed, and therefore contribute to the widespread contamination of maize products. These toxins are highly dangerous fungal substances that cause acute and chronic toxicity in humans and livestock. In the present study, 32 samples of different maize products were collected from different locations in Tamil Nadu, India, during 2021-2022 to assess aflatoxin contamination. Only two samples were free from aflatoxin contamination, whereas the other 30 maize samples were contaminated with different types of aflatoxins. In addition, maize seeds were collected from 15 locations covering 10 districts of Tamil Nadu, India, to study the mycoflora diversity. The results revealed the six main fungal genera, namely Aspergillus, Fusarium, Penicillium, Rhizopus, Alternaria, and Macrophomina, associated with maize seeds. Out of the 1152 mycoflora isolated from maize seeds, the genus Aspergillus contributed the most (82%), followed by Fusarium spp. (9%) and others (9%). Within the genus Aspergillus, A. niger, A. flavus, A. fumigatus, A. tamarii, and A. oryzae were recorded. Species composition studies revealed that A. niger, A. flavus and A. tamarii were present at all locations. Alpha diversity analysis of maize seed mycoflora and the genus Aspergillus using different indices revealed that abundance, dominance, and species richness were high at very few locations. Beta diversity analysis also revealed that there was no significant difference in the mycoflora between locations i.e. mycoflora distribution was even across all locations. Furthermore, the toxigenic potential of 24 Aspergillus isolates from different locations was assessed. The results revealed that the isolates Asp 16, Asp 17, Asp 20, and Asp 23 produced all four aflatoxins types, namely aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), and aflatoxin G2 (AFG2). Among the isolates, a maximum AFB1 of 10250.78 ppb/g of agar plug was recorded for the Asp 23 isolate, followed by Asp 20 (4246.584 ppb). Most isolates produced one or two aflatoxin types, and some non-toxigenic Aspergillus isolates were also recorded.

Application and antagonistic mechanisms of atoxigenic Aspergillus strains for the management of fungal plant diseases.

This review covers, for the first time, all methods based on the use of Aspergillus strains as biocontrol agents for the management of plant diseases caused by fungi and oomycetes. Atoxigenic Aspergillus strains have been screened in a variety of hosts, such as peanuts, maize kernels, and legumes, during the preharvest and postharvest stages. These strains have been screened against a wide range of pathogens, such as Fusarium, Phytophthora, and Pythium species, suggesting a broad applicability spectrum. The highest efficacies were generally observed when using non-toxigenic Aspergillus strains for the management of mycotoxin-producing Aspergillus strains. The modes of action included the synthesis of antifungal metabolites, such as kojic acid and volatile organic compounds (VOCs), secretion of hydrolytic enzymes, competition for space and nutrients, and induction of disease resistance. Aspergillus strains degraded Sclerotinia sclerotiorum sclerotia, showing high control efficacy against this pathogen. Collectively, although two Aspergillus strains have been commercialized for aflatoxin degradation, a new application of Aspergillus strains is emerging and needs to be optimized.

Discrimination of toxigenic and non-toxigenic Aspergillus flavus in wheat based on nanocomposite colorimetric sensor array

A novel method was developed for the early detection of wheat infected with Aspergillus flavus (A. flavus) using a nanocomposite colorimetric sensors array (CSA). LC-MS analysis revealed the presence of Aflatoxin B1 (AFB1) and Aflatoxin B2 (AFB2) on day seven, demonstrating mycotoxin variabilities in infected wheat. HS-SPME-GC–MS analysis detected 2-methylbutyral, a gas exclusively associated with toxigenic A. flavus. The CSA was modified using three nanoparticles of MOF and successfully used to detect the wheat infected with A. flavus. Discrimination of different types of infected wheat samples was achieved using the RGB difference map and Principal Component Analysis (PCA) model. Additionally, the Linear Discriminant Analysis (LDA) model accurately predicted the presence of toxigenic A. flavus at various stages of infection. These findings highlight the promising capabilities of nanocomposite CSA for early-stage detection of A. flavus infection in wheat.

Fungal Contamination and Characterization of Aspergillus flavus on Poultry Feeds and their Ingredients in North Sumatera

Poultry feeds mainly consist of cereal grains and crops that are vulnerable to fungal infection. Improper handling of the feeds increases spoilage and contamination by aflatoxigenic fungi. The aim of this study was to determine fungal contamination including characterization of toxigenic and non-toxigenic Aspergillus flavus strains in poultry feed and its ingredients. We collected 200 composite samples of compound poultry feeds and their ingredients sold by retailers at traditional markets. The fungal population was enumerated using a dilution followed by the pour plate in dichloran 18% glycerol agar medium. All isolated A. flavus were identified by culture technique and molecular approach. Aflatoxin production was determined in 10% coconut agar medium with thin layer chromatography. Regulatory (aflR) and structural (nor-1, ver-1, omt-1) genes of A. flavus strains were isolated using four sets of primers (aflR, nor-1, ver-1, omt-1). We found that all feeds were infected by fungi. Aspergillus chevalieri was the most found in all feeds, particularl on chick starter. Of the 200 samples of compound and poultry feed ingredients, 57 isolates of A. flavus were examined and 16 of them (28%) were toxigenic, each strain producing aflatoxin B1 from <3.01 to 35.50 ppb. Among feeds, layer finisher, pellet starter and grain corn were the most infected by toxigenic A. flavus, respectively. The contamination occurred on feeds that contained corn as the main compound. Routine analysis of compound poultry feeds and their ingredients for aflatoxigenic fungi particularly A. flavus is compulsory

Complete mitochondrial genome sequence of Afla-Guard®, commercially available non-toxigenic Aspergillus flavus

Afla-Guard® is a commercial non-toxigenic Aspergillus flavus strain used to decrease aflatoxin contamination level in field. Its mitochondrial genome was sequenced, showing that its length is 29,208 bp with typical configuration of Aspergillus mitochondrial genome. 17 SNPs and 27 INDELs were identified by comparing with previous A. flavus mitochondrial genome. Phylogenetic trees present that A. flavus of Afla-Guard® was clustered with the previous A. flavus mitochondrial genome.

A non-toxigenic Aspergillus flavus strain prevents the spreading of Fusarium verticillioides and fumonisins in maize

We evaluated Fusarium verticillioides and fumonisin contamination in maize samples after application of a non-aflatoxigenic Aspergillus flavus strain in the field. The sampling was performed 150 days after planting. The results showed a reduction in F. verticillioides frequency, as well as in fumonisin levels when samples were obtained from field areas treated with non-aflatoxigenic A. flavus strain. These results suggested a competition for substrate or space between fungi reducing the frequency of F. verticillioides and, consequently, fumonisin production.

Thermosensitive Hydrogel for Encapsulation and Controlled Release of Biocontrol Agents to Prevent Peanut Aflatoxin Contamination.

Starch, alginate, and poly(N-isopropylacrylamide) (PNIPAAm) were combined to prepare a semi-interpenetrating network (IPN) hydrogel with temperature sensitivity. Calcium chloride was used as cross-linking agent, the non-toxigenic Aspergillus flavus spores were successfully encapsulated as biocontrol agents by the method of ionic gelation. Characterization of the hydrogel was performed by Fourier-transform infrared spectroscopy (FTIR), scanning electron micrograph (SEM), and thermogravimetry analysis (TGA). Formulation characteristics, such as entrapment efficiency, beads size, swelling behavior, and rheological properties were evaluated. The optical and rheological measurements indicated that the lower critical solution temperature (LCST) of the samples was about 29–30 °C. TGA results demonstrated that the addition of kaolin could improve the thermal stability of the semi-IPN hydrogel. Morphological analysis showed a porous honeycomb structure on the surface of the beads. According to the release properties of the beads, the semi-IPN hydrogel beads containing kaolin not only have the effect of slow release before peanut flowering, but they also can rapidly release biocontrol agents after flowering begins. The early flowering stage of the peanut is the critical moment to apply biocontrol agents. Temperature-sensitive hydrogel beads containing kaolin could be considered as carriers of biocontrol agents for the control of aflatoxin in peanuts.

BIOLOGICAL CONTROL OF AFLATOXIGENIC FUNGI ON PEANUT: FOR THE PRE-HARVEST APPROACH

This study was carried out to determine the efficacy of different applications of a biopesticide for reduction of aflatoxin contamination in peanut. The biopesticide, afla-guard, delivers a nontoxigenic Aspergillus flavus to the field where it competes with naturally occurring toxigenic fungus. Biocontrol treatments included: (ı) soil application during sowing, (ıı) multiple application during sowing and 40 days after planting, (ııı) foliar application at 60 days after planting (ıv) control (untreated plots). Biopesiticide was applied to peanut plots in 2015 and 2016 in Randomized Complete Block Design with four replications. Peanuts were collected from control and treated plots at harvest-drying-pre-storage periods and analysed for aflatoxins. Aflatoxin concentrations were generally quite low in 2015, also the aflatoxin concentration in treated samples (from 0.04 to 0.71 μg/kg) was reduced by 97.38 to 99.82% compared with controls (from 21.84 to 27.12 μg/kg). In 2016, reductions were also noted for all biocontrol treatments (from 89.07 to 92.39%) compared with controls. In conjunction with the reductions in aflatoxin contamination, biocontrol treatments produced significant reductions with biopesticide in peanut. Therefore, it can be said that a biological control method is a promising approach for controlling aflatoxin.

Controlled Release of Biological Control Agents for Preventing Aflatoxin Contamination from Starch⁻Alginate Beads.

For the wise use of fungal biocontrol and metalaxyl fungicide, starch–alginate-based formulations have been developed by encapsulating metalaxyl and non-toxigenic Aspergillus flavus spores simultaneously in the form of microspheres using calcium chloride as a cross-linking agent. The formulations were characterized by Fourier transform infrared spectroscopy (FTIR), a scanning electron micrograph (SEM), and thermogravimetry (TGA). Formulation characteristics, including the bead size, entrapment efficiency, swelling ratio of the beads, and rheological properties, were analyzed. The release behavior of beads with different formulations was evaluated. The addition of kaolin and rice husk powder in starch–alginate beads retarded the release profile of spores and metalaxyl. The release of the active ingredient from starch–alginate–kaolin beads and starch–alginate–rice husk powder beads occurred in both a controlled and sustained manner. Additionally, the release rate decreased with the increase of kaolin or rice husk powder content. The beads added with kaolin were slower than the release of rice husk powder. In comparison, spores released slower and lasted longer than metalaxyl. The starch–alginate–kaolin formulations could be used as controlled release material in the field of biocontrol and reduce the harm of fungicides to the environment.

Glyphosate in vitro removal and tolerance by Aspergillus oryzae in soil microcosms

Argentinian agriculture is based on a technological package that includes the application of significant amounts of pesticides. Glyphosate is the active ingredient of several glyphosate herbicides which are frequently used for weed control. Fungi can use glyphosate both as a nutrient and an energetic source. Therefore, the aim of the present study was to evaluate the in vitro glyphosate removal as the only source of phosphorus or nitrogen by two non-toxigenic Aspergillus oryzae strains. A. oryzae AM1 and AM2 were successfully able to use the herbicide both as phosphorus or nitrogen source. The degradation percentages were higher than 50% at the end of the incubation period (15 days) with glyphosate concentrations of 1 and 1.5 mM. In addition, AM1 strain was able to remove 57% of 10 mM of glyphosate at 0.99 of aW. Soil microcosm assay was performed in order to study the GP tolerance, permanence and competitiveness of A. oryzae AM1 in the presence of native mycota. A. oryzae count remained constant along the incubation period and was not significantly affected by the different glyphosate concentrations tested (10, 20 and 50 mM). This strain showed great in vitro removal ability, and it was also able to remain viable on soil microcosms contaminated with glyphosate. Therefore, these results encourage future studies in order to evaluate the ability of these strains to degrade glyphosate on soil and then promote them as potential bioremediation agents.

Development and removal ability of non-toxigenic Aspergillus section Flavi in presence of atrazine, chlorpyrifos and endosulfan

This study evaluated the in vitro effect of three concentrations of atrazine, chlorpyrifos and endosulfan on the growth parameters of four non-toxigenic Aspergillus section Flavi strains. The ability of the strains to remove these pesticides in a synthetic medium was also determined. Growth parameters were measured on soil extract solid medium supplied with 5, 10 and 20mg/l of each pesticide, and conditioned to −0.70, −2.78, −7.06 and −10.0 water potential (MPa). Removal assays were performed in Czapek Doc medium (CZD) supplied with 20mg/l of each pesticide under optimal environmental conditions (−2.78 of MPa and 25°C). The residual levels of each pesticide were detected by the reversed-phase HPLC/fluorescence detection system. The lag phases of the strains significantly decreased in the presence of the pesticides with respect to the control media. This result indicates a fast adaptation to the conditions assayed. Similarly, the mycelial growth rates in the different treatments increased depending on pesticide concentrations. Aspergillus oryzae AM 1 and AM 2 strains showed high percentages of atrazine degradation (above 90%), followed by endosulfan (56 and 76%) and chlorpyrifos (50 and 73%) after 30 days of incubation. A significant (p<0.001) correlation (r=0.974) between removal percentages and growth rate was found. This study shows that non-toxigenic Aspergillus section Flavi strains from agricultural soils are able to effectively grow in the presence of high concentrations of atrazine, chlorpyrifos and endosulfan under a wide range of MPa conditions. Moreover, these strains have the ability to remove high levels of these pesticides in vitro in a short time.

Use of microsatellite markers to assess the competitive ability of nontoxigenic Aspergillus flavus strains in studies on biocontrol of aflatoxins in maize in Thailand

ABSTRACTMany nontoxigenic strains of Aspergillus flavus have been used in studies on biocontrol by competitive exclusion, but assessing their competitive ability is difficult. This paper reports on the use of a microsatellite marker technique for assessing competitiveness. The chosen microsatellite markers were able to differentiate, at an individual level, between the four biocontrol strains used in a study on the biocontrol of aflatoxins in maize in Thailand. The microsatellite markers were then used to determine which of the four biocontrol strains used were identical with 86 nontoxigenic strains of A. flavus taken from dried maize samples produced in that study. Fifty-one of the 86 strains (59%) were identified as one of the four biocontrol strains, with another four likely to be so. Analysis of microsatellites in A. flavus strains taken from dried samples at the conclusion of a field trial was shown to be of value in understanding the competitive ability of the specific strains used for biocontrol. This method provides an objective assessment of the competitiveness of biocontrol strains.

Identification and Quantification of a Toxigenic and Non-Toxigenic Aspergillus flavus Strain in Contaminated Maize Using Quantitative Real-Time PCR.

Aflatoxins, which are produced by Aspergillus flavus, are toxic to humans, livestock, and pets. The value of maize (Zea mays) grain is markedly reduced when contaminated with aflatoxin. Plant resistance and biological control using non-toxin producing strains are considered effective strategies for reducing aflatoxin accumulation in maize grain. Distinguishing between the toxin and non-toxin producing strains is important in determining the effectiveness of bio-control strategies and understanding inter-strain interactions. Using polymorphisms found in the fungal rRNA intergenic spacer region (IGS) between a toxigenic strain of A. flavus (NRRL 3357) and the non-toxigenic strain used in the biological control agent Afla-Guard® (NRRL 21882), we developed a set of primers that allows for the identification and quantification of the two strains using quantitative PCR. This primer set has been used to screen maize grain that was inoculated with the two strains individually and co-inoculated with both strains, and it has been shown to be effective in both the identification and quantification of both strains. Screening of co-inoculated ears from multiple resistant and susceptible genotypic crosses revealed no significant differences in fungal biomass accumulation of either strain in the field tests from 2010 and 2011 when compared across the means of all genotypes. Only one genotype/year combination showed significant differences in strain accumulation. Aflatoxin accumulation analysis showed that, as expected, genotypes inoculated with the toxigenic strain accumulated more aflatoxin than when co-inoculated with both strains or inoculated with only the non-toxigenic strain. Furthermore, accumulation of toxigenic fungal mass was significantly correlated with aflatoxin accumulation while non-toxigenic fungal accumulation was not. This primer set will allow researchers to better determine how the two fungal strains compete on the maize ear and investigate the interaction between different maize lines and these A. flavus strains.

Isolation of culturable mycobiota from agricultural soils and determination of tolerance to glyphosate of nontoxigenic Aspergillus section Flavi strains

ABSTRACTGlyphosate-based herbicides are extensively used in Argentina's agricultural system to control undesirable weeds. This study was conducted to evaluate the culturable mycobiota [colony forming units (CFU) g−1 and frequency of fungal genera or species] from an agricultural field exposed to pesticides. In addition, we evaluated the tolerance of A. oryzae and nontoxigenic A. flavus strains to high concentrations (100 to 500 mM – 17,000 to 84,500 ppm) of a glyphosate commercial formulation. The analysis of the mycobiota showed that the frequency of the main fungal genera varied according to the analyzed sampling period. Aspergillus spp. or Aspergillus section Flavi strains were isolated from 20 to 100% of the soil samples. Sterilia spp. were also observed throughout the sampling (50 to 100%). Aspergillus section Flavi tolerance assays showed that all of the tested strains were able to develop at the highest glyphosate concentration tested regardless of the water availability conditions. In general, significant reductions in growth rates were observed with increasing concentrations of the herbicide. However, a complete inhibition of fungal growth was not observed with the concentrations assayed. This study contributes to the knowledge of culturable mycobiota from agricultural soils exposed to pesticides and provides evidence on the effective growth ability of A. oryzae and nontoxigenic A. flavus strains exposed to high glyphosate concentrations in vitro.

Diversity of black Aspergilli isolated from raisins in Argentina: Polyphasic approach to species identification and development of SCAR markers for Aspergillus ibericus

Aspergillus section Nigri is a heterogeneous fungal group including some ochratoxin A producer species that usually contaminate raisins. The section contains the Series Carbonaria which includes the toxigenic species Aspergillus carbonarius and nontoxigenic Aspergillus ibericus that are phenotypically undistinguishable. The aim of this study was to examine the diversity of black aspergilli isolated from raisins and to develop a specific genetic marker to distinguish A. ibericus from A. carbonarius. The species most frequently found in raisins in this study were Aspergillus tubingensis (35.4%) and A. carbonarius (32.3%), followed by Aspergillus luchuensis (10.7%), Aspergillus japonicus (7.7%), Aspergillus niger (6.2%), Aspergillus welwitschiae (4.6%) and A. ibericus (3.1%). Based on inter-simple sequence repeat (ISSR) fingerprinting profiles of major Aspergillus section Nigri members, a sequence-characterized amplified region (SCAR) marker was identified. Primers were designed based on the conserved regions of the SCAR marker and were utilized in a PCR for simultaneous identification of A. carbonarius and A. ibericus. The detection level of the SCAR-PCR was found to be 0.01ng of purified DNA. The present SCAR-PCR is rapid and less cumbersome than conventional identification techniques and could be a supplementary strategy and a reliable tool for high-throughput sample analysis.

Effect of Water Activity, Temperature, and Mixed Fungal Spore Interactions on Ochratoxin A Production by Aspergillus carbonarius

The purpose of this work was to investigate the potential of two nontoxigenic Aspergillus section Nigri species (Aspergillus tubingensis and Aspergillus japonicus) to influence the in vitro ochratoxin A (OTA) production of three toxigenic Aspergillus carbonarius isolates (Ac-28, Ac-29, and Ac-33) from Greek vineyards of different geographical areas. OTA accumulation was evaluated by inoculation of 0:100, 25:75, 50:50, 75:25, and 100:0 ratios of mixed spore suspensions on a synthetic grape juice medium for up to 28 days at different temperatures (15, 20, and 25°C), water activity (aw) levels (0.95 and 0.98 aw) and incubation time (7, 14, 21, and 28 days). Results confirmed that environmental factors and fungal species had a significant effect on OTA production. Specifically, maximum OTA concentration for Ac-28 (3.21 μg g−1) and Ac-29 (7.69 μg g−1) was observed at 20°C/0.98 aw and for Ac-33 (9.13 μg g−1) at 15°C/0.95 aw, regardless of incubation time. Moreover, A. tubingensis had no significant influence on OTA concentration of all toxigenic isolates assayed, regardless of temperature, aw, and incubation time. On the other hand, the presence of A. japonicus slightly inhibited OTA production of Ac-29 and Ac-33, while for Ac-28, stimulation of OTA was observed in some cases. Overall, lower aw levels reduced OTA accumulation for Ac-28 and Ac-29, regardless of temperature, inoculum ratio, and time. On the contrary, for Ac-33, low aw increased OTA production, regardless of the investigated parameters. The importance of this study concerns the understanding of interspecific interactions on OTA diffusion by A. carbonarius in an attempt to find ways to prevent the presence of toxins in grapes and their derivatives.

Prevention of toxigenic fungal growth in stored grains by carbon dioxide detection

The growth of toxigenic fungi can adversely affect grain quality and even produce mycotoxins of food safety concern, which should be sensitively monitored and controlled during grain storage. To establish the relationship between the growth of toxigenic fungi and their carbon dioxide (CO2) production, the pattern of CO2 concentration changes was studied during the fungal growth in grain. The results showed the CO2 concentrations increased exponentially (r ≥ 0.96) during the growth of toxigenic fungi Aspergillus flavus, Penicillium sp. and Aspergillus ochraceus, which was different from the linear increase of CO2 concentration produced by the non-toxigenic xerophilic fungi Aspergillus glaucus and Aspergillus restrictus. The acceleration of CO2 concentration was found much earlier than the growth of toxigenic fungi, which would be useful for the prevention of grain spoilage. In addition, the CO2 concentration changes were also determined in storage containers loaded with grain of different moisture content and significant correlation (p < 0.05) was found between changes of CO2 concentration and fungal growth as well as mycotoxin production. The nonlinear increase of CO2 concentration in stored grains could be considered as an indication of the rapid growth of toxigenic fungi and greater risk of microbial spoilage of grains. The results can provide a valid foundation for the prevention of toxigenic fungi and mycotoxin production in stored grains through monitoring the CO2 concentration changes.

Survey of Aspergillus section Flavi presence in agricultural soils and effect of glyphosate on nontoxigenic A. flavus growth on soil-based medium

To evaluate the cultivable mycobiota from agricultural soils exposed to pesticides, the aflatoxigenic capacity of Aspergillus section Flavi strains and the effect of glyphosate on lag phase and growth rates of native nontoxigenic Aspergillus flavus under different water potential (MPa) conditions on soil-based medium. Culturable mycobiota analysis from different agricultural soils was performed by the surface spread method. The effect of glyphosate (0-20 mmol l(-1)) on the growth of A. flavus strains was evaluated on a soil extract solid medium. Mycobiota analysis of crop soils showed the presence of twenty-one genera of filamentous fungi. Aspergillus flavus and Aspergillus niger aggregate strains were isolated from the three soil types. Ninety-two per cent of A. flavus strains were toxigenic. In vitro assay results showed that at -0·70 MPa, a significant increase in growth rate in all strains was recorded at 5 and 20 mmol l(-1) of glyphosate. At -2·78 MPa, this parameter remained constant at all glyphosate concentrations, except in GM4 strain where an increase in growth rate was recorded with increasing pesticide concentrations. At -7·06 MPa, a significant increase in growth rate has also been observed in GM 3 strain with 5 mmol l(-1) and in GM 4 strain with 10 and 20 mmol l(-1). This study showed that the imperfecti fungi Aspergillus spp., Penicillium spp., Trichoderma spp., Cladosporium spp. and Paecilomyces spp. are isolated as prevalent groups in agricultural soil exposed to pesticides, and the capacity of nontoxigenic A. flavus strains to tolerate different glyphosate concentrations under different water potential (MPa) conditions. This manuscript makes a contribution to the knowledge of cultivable fungal populations from agricultural soils exposed to pesticides and the glyphosate tolerance of A. flavus strains.

Genetic variability of Aspergillus flavus isolates from a Mississippi corn field.

A nontoxigenic Aspergillus flavus strain, K49, is currently being tested as a biological control agent in corn fields in the Mississippi Delta. However, little is known about the overall genetic diversity of A. flavus from year to year in corn fields and specifically in Mississippi. Our objective was to assess the genetic variability of A. flavus isolates from different seasons, inoculum sources, and years, from a no-till corn field. Of the 175 A. flavus isolates examined, 74 and 97 had the typical norB-cypA type I (1.5 kb) and type II (1.0 kb) deletion patterns, respectively. Variability in the sequence of the omtA gene of the majority of the field isolates (n = 118) was compared to strain K49. High levels of haplotypic diversity (24 omtA haplotypes; Hd = 0.61 ± 0.04) were found. Among the 24 haplotypes, two were predominant, H1 (n = 71), which consists of mostly toxigenic isolates, and H49 (n = 18), which consists of mostly atoxigenic isolates including K49. Toxigenic isolates were prevalent (60%) in this natural population. Nonetheless, about 15% of the population likely shared the same ancestral origin with K49. This study provides valuable information on the diversity of A. flavus. This knowledge can be further used to develop additional biological control strains.

Implications of European corn borer, Ostrinia nubilalis, infestation in an Aspergillus flavus‐biocontrolled corn agroecosystem

A novel biocontrol strategy consisting of field application of bioplastic-based granules inoculated with a non-toxigenic Aspergillus flavus L. strain has recently been shown to be effective for reducing aflatoxin contamination in corn. This study focused on other factors that may affect the feasibility of this biocontrol technique, and more specifically the role of the European corn borer (ECB), Ostrinia nubilalis H., in the dispersal and infestation of A. flavus in corn and its impact on crop yield. In spite of the high percentage of corn ears showing larval feeding damage, ECB-bored kernels accounted for only 3 and 4% in 2009 and 2010 respectively. Most of the damaged kernels were localised in the ear tip or immediately below. More precisely, the average incidence of ECB-bored kernels in the upper end of the ear was 32%. However, less than 5% of kernels from the central body of the ear, which includes the majority of kernels, were injured by ECB. Although ECB larvae showed a high tolerance to aflatoxin B1 and thus had the potential to serve as vectors of the mould, fungal infection of kernels was poorly associated with insect damage. ECB infestation resulted in grain yield losses not exceeding 2.5%.