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Aflatoxin contamination of almond kernels, caused by Aspergillus flavus and A. parasiticus, is a severe concern for growers because of its high toxicity. In California, the global leader of almond production, aflatoxin can be managed by applying the biological control strain AF36 of A. flavus and selecting resistant cultivars. Here, we classified the almond genotypes by K-Means cluster analysis into three groups (susceptible [S], moderately susceptible [MS], or resistant [R]) based on aflatoxin content of inoculated kernels. The protective effects of the shell and seedcoat in preventing aflatoxin contamination were also examined. The presence of intact shells reduced aflatoxin contamination >100-fold. The seedcoat provided a layer of protection but not complete protection. In kernel inoculation assays, none of the studied almond genotypes showed a total resistance to the pathogen. However, nine traditional cultivars and four advanced selections were classified as R. Because these advanced selections contained germplasm derived from peach, we compared the kernel resistance of three peach cultivars to that shown by kernels of an R (Sonora) and an S (Carmel) almond cultivar and five pistachio cultivars. Overall, peach kernels were significantly more resistant to the pathogen than almond kernels, which were more resistant than pistachio kernels. Finally, we studied the combined effect of the cultivar resistance and the biocontrol strain AF36 in limiting aflatoxin contamination. For this, we coinoculated almond kernels of R Sonora and S Carmel with AF36 72 h before or 48 h after inoculating with an aflatoxin-producing strain of A. flavus. The percentage of aflatoxin reduction by AF36 strain was greater in kernels of Carmel (98%) than in those of Sonora (83%). Cultivar resistance also affected the kernel colonization by the biological control strain. AF36 strain limited aflatoxin contamination in almond kernels even when applied 48 h after the aflatoxin-producing strain. Our results show that biocontrol combined with the use of cultivars with resistance to aflatoxin contamination can result in a more robust protection strategy than the use of either practice in isolation.

The species Aspergillus flavus and A. parasiticus are commonly found in the soils of nut-growing areas in California. Several isolates can produce aflatoxins that occasionally contaminate nut kernels, conditioning their sale. Strain AF36 of A. flavus, which does not produce aflatoxins, is registered as a biocontrol agent for use in almond, pistachio, and fig crops in California. After application in orchards, AF36 displaces aflatoxin-producing Aspergillus spp. and thus reduces aflatoxin contamination. Vegetative compatibility assays (VCAs) have traditionally been used to track AF36 in soils and crops where it has been applied. However, VCAs are labor intensive and time consuming. Here, we developed a quantitative real-time PCR (qPCR) protocol to quantify proportions of AF36 accurately and efficiently in different substrates. Specific primers to target AF36 and toxigenic strains of A. flavus and A. parasiticus were designed based on the sequence of aflC, a gene essential for aflatoxin biosynthesis. Standard curves were generated to calculate proportions of AF36 based on threshold cycle values. Verification assays using pure DNA and conidial suspension mixtures demonstrated a significant relationship by regression analysis between known and qPCR-measured AF36 proportions in DNA (R2 = 0.974; P < 0.001) and conidia mixtures (R2 = 0.950; P < 0.001). Tests conducted by qPCR in pistachio leaves, nuts, and soil samples demonstrated the usefulness of the qPCR method to precisely quantify proportions of AF36 in diverse substrates, ensuring important time and cost savings. The outputs of this study will serve to design better aflatoxin management strategies for pistachio and other crops.

Aflatoxin contamination is caused by Aspergillus flavus and closely related fungi. In The Gambia, aflatoxin contamination of groundnut and maize, two staple and economically important crops, is common. Groundnut and maize consumers are chronically exposed to aflatoxins, sometimes at alarming levels, and this has severe consequences on their health and productivity. Aflatoxin contamination also impedes commercialization in local and international premium markets. In neighboring Senegal, an aflatoxin biocontrol product containing four atoxigenic isolates of A. flavus, Aflasafe SN01, has been registered and is approved for commercial use in groundnut and maize. We detected that the four genotypes composing Aflasafe SN01 are also native to The Gambia. The biocontrol product was tested during two years in 129 maize and groundnut fields and compared with corresponding untreated fields cropped by smallholder farmers in The Gambia. Treated crops contained up to 100% less aflatoxins than untreated crops. A large portion of the crops could have been commercialized in premium markets due to the low aflatoxin content (in many cases no detectable aflatoxins), both at harvest and after storage. Substantial aflatoxin reductions were also achieved when commercially produced groundnut received treatment. Here we report for the first time the use and effectiveness of an aflatoxin biocontrol product registered for use in two nations. With the current scale-out and -up efforts of Aflasafe SN01, a large number of farmers, consumers, and traders in The Gambia and Senegal will obtain health, income, and trade benefits. [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Aflatoxin contamination of staple crops, commonly occurring in warm areas, negatively impacts human and animal health, and hampers trade and economic development. The fungus Aspergillus flavus is the major aflatoxin producer. However, not all A. flavus genotypes produce aflatoxins. Effective aflatoxin control is achieved using biocontrol products containing spores of atoxigenic A. flavus. In Africa, various biocontrol products under the tradename Aflasafe are available. Private and public sector licensees manufacture Aflasafe using spores freshly produced in laboratories adjacent to their factories. BAMTAARE, the licensee in Senegal, had difficulties to obtain laboratory equipment during its first year of production. To overcome this, a process was developed in Ibadan, Nigeria, for producing high-quality dry spores. Viability and stability of the dry spores were tested and conformed to set standards. In 2019, BAMTAARE manufactured Aflasafe SN01 using dry spores produced in Ibadan and sent via courier and 19 000 ha of groundnut and maize in Senegal and The Gambia were treated. Biocontrol manufactured with dry spores was as effective as biocontrol manufactured with freshly produced spores. Treated crops contained safe and significantly (P < 0.05) less aflatoxin than untreated crops. The dry spore innovation will make biocontrol manufacturing cost-efficient in several African countries.

In most sub-Saharan African countries, staple cereal grains harbor many fungi and some produce mycotoxins that negatively impact health and trade. Maize and three small grain cereals (sorghum, pearl millet, and finger millet) produced by smallholder farmers in Zimbabwe during 2016 and 2017 were examined for fungal community structure, and total aflatoxin (AF) and fumonisin (FM) content. A total of 800 maize and 180 small grain samples were collected at harvest and during storage from four agroecological zones. Fusarium spp. dominated the fungi associated with maize. Across crops, Aspergillusflavus constituted the main Aspergillus spp. Small grain cereals were less susceptible to both AF and FM. AF (52%) and FM (89%) prevalence was higher in maize than in small grains (13&ndash, 25% for AF and 0&ndash, 32% for FM). Less than 2% of small grain samples exceeded the EU regulatory limit for AF (4 &micro, g/kg), while &lt, 10% exceeded the EU regulatory limit for FM (1000 &micro, g/kg). For maize, 28% and 54% of samples exceeded AF and FM Codex guidance limits, respectively. Higher AF contamination occurred in the drier and hotter areas while more FM occurred in the wetter year. AF exposure risk assessment revealed that small grain consumption posed low health risks (&le, 0.02 liver cancer cases/100,000 persons/year) while maize consumption potentially caused higher liver cancer rates of up to 9.2 cases/100,000 persons/year depending on the locality. Additionally, FM hazard quotients from maize consumption among children and adults were high in both years, but more so in a wet year than a dry year. Adoption of AF and FM management practices throughout the maize value chain coupled with policies supporting dietary diversification are needed to protect maize consumers in Zimbabwe from AF- and FM-associated health effects. The higher risk of health burden from diseases associated with elevated concentration of mycotoxins in preferred maize during climate change events can be relieved by increased consumption of small grains.

In the tropics and subtropics, maize (Zea mays) and other crops are frequently contaminated with aflatoxins by Aspergillus flavus. Treatment of crops with atoxigenic isolates of A. flavus formulated into biocontrol products can significantly reduce aflatoxin contamination. Treated crops contain up to 100% fewer aflatoxins compared with untreated crops. However, there is the notion that protecting crops from aflatoxin contamination may result in increased accumulation of other toxins, particularly fumonisins produced by a few Fusarium species. The objective of this study was to determine if treatment of maize with aflatoxin biocontrol products increased fumonisin concentration and fumonisin-producing fungi in grains. Over 200 maize samples from fields treated with atoxigenic biocontrol products in Nigeria and Ghana were examined for fumonisin content and contrasted with maize from untreated fields. Apart from low aflatoxin levels, most treated maize also harbored fumonisin levels considered safe by the European Union ( [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Highlights • Efficacy of 2 aflatoxin biocontrol products reported across crops, years and zones. • Aflatoxin reduced by 99% in 800 maize and groundnut farmers’ fields in Ghana. • Results are the most consistent and highest aflatoxin reductions reported to date. • Displacement of toxigenic fungi by the active ingredients caused the reductions. • Having both products offers greater versatility to modulate Aspergillus communities., Biological control is one of the recommended methods for aflatoxin mitigation. Biocontrol products must be developed, and their efficacy demonstrated before widespread use. Efficacy of two aflatoxin biocontrol products, Aflasafe GH01 and Aflasafe GH02, were evaluated in 800 maize and groundnut farmers’ fields during 2015 and 2016 in the Ashanti, Brong Ahafo, Northern, Upper East, and Upper West regions of Ghana. Both products were developed after an extensive examination of fungi associated with maize and groundnut in Ghana. Each product contains as active ingredient fungi four Aspergillus flavus isolates belonging to atoxigenic African Aspergillus Vegetative Compatibility Groups (AAVs) widely distributed across Ghana. An untreated field was maintained for each treated field to determine product efficacy. Proportions of atoxigenic AAVs composing each product were assessed in soils before product application, and soils and grains at harvest. Significant (P

Maize is a staple for billions across the globe. However, in tropical and sub-tropical regions, maize is frequently contaminated with aflatoxins by Aspergillus section Flavi fungi. There is an ongoing search for sources of aflatoxin resistance in maize to reduce continuous exposures of human populations to those dangerous mycotoxins. Large variability in susceptibility to aflatoxin contamination exists within maize germplasm. In Mexico, several maize landrace (MLR) accessions possess superior resistance to both Aspergillus infection and aflatoxin contamination but their mechanisms of resistance have not been reported. Influences of kernel integrity on resistance of four resistant and four susceptible MLR accessions were evaluated in laboratory assays. Wounds significantly (P < 0.05) increased susceptibility to aflatoxin contamination even when kernel viability was unaffected. Treatments supporting greater A. flavus reproduction did not (P > 0.05) proportionally support higher aflatoxin accumulation suggesting differential influences by some resistance factors between sporulation and aflatoxin biosynthesis. Physical barriers (i.e., wax and cuticle) prevented both aflatoxin accumulation and A. flavus sporulation in a highly resistant MLR accession. In addition, influence of temperature on aflatoxin contamination was evaluated in both viable and non-viable kernels of a resistant and a susceptible MLR accession, and a commercial hybrid. Both temperature and living embryo status influenced (P < 0.05) resistance to both aflatoxin accumulation and A. flavus sporulation. Lower sporulation on MLR accessions suggests their utilization would result in reduced speed of propagation and associated epidemic increases in disease both in the field and throughout storage. Results from the current study should encourage researchers across the globe to exploit the large potential that MLRs offer to breed for aflatoxin resistant maize. Furthermore, the studies provide support to the importance of resistance based on the living host and maintaining living status to reducing episodes of post-harvest contamination.

Aflatoxin contamination of important food and feed crops occurs frequently in warm tropical and subtropical regions. The contamination is caused mainly by Aspergillus flavus and A. parasiticus. Aflatoxin contamination negatively affects health and trade sectors and causes economic losses to agricultural industries. Many pre- and post-harvest technologies can limit aflatoxin contamination but may not always reduce aflatoxin concentrations below tolerance thresholds. However, the use of atoxigenic (non-toxin producing) isolates of A. flavus to competitively displace aflatoxin producers is a practical strategy that effectively limits aflatoxin contamination in crops from field to plate. Biocontrol products formulated with atoxigenic isolates as active ingredients have been registered for use in the US, several African nations, and one such product is in final stages of registration in Italy. Many other nations are seeking to develop biocontrol products to protect their crops. In this review article we present an overview of the biocontrol technology, explain the basis to select atoxigenic isolates as active ingredients, describe how formulations are developed and tested, and describe how a biocontrol product is used commercially. Future perspectives on formulations of aflatoxin biocontrol products, along with other important topics related to the aflatoxin biocontrol technology are also discussed. Fil: Moral, Juan. Universidad de Córdoba; España Fil: Garcia-Lopez, Maria Teresa. Universidad de Córdoba; España Fil: Camiletti, Boris Xavier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Agropecuarias. Departamento de Recursos Naturales. Cátedra de Microbiología Agrícola; Argentina Fil: Jaime, Ramon. University of California at Davis; Estados Unidos Fil: Michailides, Themis J.. University of California at Davis; Estados Unidos Fil: Bandyopadhyay, Ranajit. International Institute of Tropical Agriculture; Nigeria Fil: Ortega-Beltran, Alejandro. International Institute of Tropical Agriculture; Nigeria

Aflatoxins are secondary metabolites produced by soilborne saprophytic fungus Aspergillus flavus and closely related species that infect several agricultural commodities including groundnut and maize. The consumption of contaminated commodities adversely affects the health of humans and livestock. Aflatoxin contamination also causes significant economic and financial losses to producers. Research efforts and significant progress have been made in the past three decades to understand the genetic behavior, molecular mechanisms, as well as the detailed biology of host-pathogen interactions. A range of omics approaches have facilitated better understanding of the resistance mechanisms and identified pathways involved during host-pathogen interactions. Most of such studies were however undertaken in groundnut and maize. Current efforts are geared toward harnessing knowledge on host-pathogen interactions and crop resistant factors that control aflatoxin contamination. This study provides a summary of the recent progress made in enhancing the understanding of the functional biology and molecular mechanisms associated with host-pathogen interactions during aflatoxin contamination in groundnut and maize.

Aflatoxin contamination of groundnut and maize infected by Aspergillus section Flavi fungi is common throughout Senegal. The use of biocontrol products containing atoxigenic Aspergillus flavus strains to reduce crop aflatoxin content has been successful in several regions, but no such products are available in Senegal. The biocontrol product Aflasafe SN01 was developed for use in Senegal. The four active ingredients of Aflasafe SN01 are atoxigenic A. flavus genotypes native to Senegal and distinct from active ingredients used in other biocontrol products. Efficacy tests on groundnut and maize in farmers’ fields were carried out in Senegal during the course of 5 years. Active ingredients were monitored with vegetative compatibility analyses. Significant (P < 0.05) displacement of aflatoxin producers occurred in all years, districts, and crops. In addition, crops from Aflasafe SN01-treated fields contained significantly (P < 0.05) fewer aflatoxins both at harvest and after storage. Most crops from treated fields contained aflatoxin concentrations permissible in both local and international markets. Results suggest that Aflasafe SN01 is an effective tool for aflatoxin mitigation in groundnut and maize. Large-scale use of Aflasafe SN01 should provide health, trade, and economic benefits for Senegal. [Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Aflatoxin contamination in maize and groundnut is perennial in Ghana with substantial health and economic burden on the population. The present study examined for the first time the prevalence of aflatoxin contamination in maize and groundnut in major producing regions across three agroecological zones (AEZs) in Ghana. Furthermore, the distribution and aflatoxin-producing potential of Aspergillus species associated with both crops were studied. Out of 509 samples (326 of maize and 183 of groundnut), 35% had detectable levels of aflatoxins. Over 15% of maize and 11% of groundnut samples exceeded the aflatoxin threshold limits set by the Ghana Standards Authority of 15 and 20 ppb, respectively. Mycoflora analyses revealed various species and morphotypes within the Aspergillus section Flavi. A total of 5,083 isolates were recovered from both crops. The L morphotype of Aspergillus flavus dominated communities with 93.3% of the population, followed by Aspergillus spp. with S morphotype (6%), A. tamarii (0.4%), and A. parasiticus (0.3%). Within the L morphotype, the proportion of toxigenic members was significantly (P < 0.05) higher than that of atoxigenic members across AEZs. Observed and potential aflatoxin concentrations indicate that on-field aflatoxin management strategies need to be implemented throughout Ghana. The recovered atoxigenic L morphotype fungi are genetic resources that can be employed as biocontrol agents to limit aflatoxin contamination of maize and groundnut in Ghana. [Formula: see text] Copyright © 2018 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

In 2015, all United Nations Member States adopted the 2030 Agenda for Sustainable Development to achieve peace and prosperity for all people in the planet. Meeting that ambitious agenda depends on fulfilling all objectives of 17 Sustainable Development Goals (SDGs). Multiple approaches by diverse actors, many of them interconnected, will allow achieving each SDG. However, with compromised food security and food safety, many SDGs will not be realized. In sub-Saharan Africa (SSA), maize and groundnut are two staple crops frequently contaminated with aflatoxins, which threaten food security and food safety. Aflatoxins are extremely dangerous compounds produced primarily by the fungus Aspergillus flavus. Even at minute concentrations, aflatoxins negatively influence health, income, and trade sectors. Farmers, traders, industries, and consumers become affected. However, practical solutions exist. Non-aflatoxin producing isolates (referred to as atoxigenic) of A. flavus can decrease crop aflatoxin content when used in biocontrol formulations to competitively displace aflatoxin producers during crop development. Typically, treated crops contain 80%–100% less aflatoxin than non-treated crops. The technology was developed by USDA-ARS for use in the US and has been adapted and improved for use in SSA where several products under the tradename Aflasafe are available. There are biocontrol products registered for use in 10 SSA countries and more are being developed. On the other hand, although highly effective, biocontrol is not a panacea. Less aflatoxin occurs across value chains when biocontrol is combined with other practices. In this review, we discuss how i) aflatoxin biocontrol products are developed, manufactured, licensed, and commercialized, ii) aflatoxin management strategies are designed, and iii) integrated aflatoxin management is or will soon be contributing to achieve, in several countries, many targets of most SDGs. We present integrated aflatoxin management as a model intervention contributing to tackle several challenges impeding prosperity and peace in SSA.

Aflatoxins pose significant food security and public health risks, decrease productivity and profitability of animal industries, and hamper trade. To minimize aflatoxin contamination in several crops, a biocontrol technology based on atoxigenic strains of Aspergillus flavus is commercially used in the United States and some African countries. Significant efforts are underway to popularize the use of biocontrol in Africa by various means including incentives. The purpose of this study was to develop quantitative pyrosequencing assays for rapid, simultaneous quantification of proportions of four A. flavus biocontrol genotypes within complex populations of A. flavus associated with maize crops in Nigeria to facilitate payment of farmer incentives for Aflasafe (a biocontrol product) use. Protocols were developed to confirm use of Aflasafe by small scale farmers in Nigeria. Nested PCR amplifications followed by sequence by synthesis pyrosequencing assays were required to quantify frequencies of the active ingredients and, in so doing, confirm successful use of biocontrol by participating farmers. The entire verification process could be completed in 3–4 days proving a savings over other monitoring methods in both time and costs and providing data in a time frame that could work with the commercial agriculture scheme. Quantitative pyrosequencing assays represent a reliable tool for rapid detection, quantification, and monitoring of multiple A. flavus genotypes within complex fungal communities, satisfying the requirements of the regulatory community and crop end-users that wish to determine which purchased crops were treated with the biocontrol product. Techniques developed in the current study can be modified for monitoring other crop-associated fungi.

In sub-Saharan Africa (SSA), diverse fungi belonging to Aspergillus section Flavi frequently contaminate staple crops with aflatoxins. Aflatoxins negatively impact health, income, trade, food security, and development sectors. Aspergillus flavus is the most common causal agent of contamination. However, certain A. flavus genotypes do not produce aflatoxins (i.e., are atoxigenic). An aflatoxin biocontrol technology employing atoxigenic genotypes to limit crop contamination was developed in the United States. The technology was adapted and improved for use in maize and groundnut in SSA under the trademark Aflasafe. Nigeria was the first African nation for which an aflatoxin biocontrol product was developed. The current study includes tests to assess biocontrol performance across Nigeria over the past decade. The presented data on efficacy spans years in which a relatively small number of maize and groundnut fields (8–51 per year) were treated through use on circa 36,000 ha in commercially-produced maize in 2018. During the testing phase (2009–2012), fields treated during one year were not treated in the other years while during commercial usage (2013–2019), many fields were treated in multiple years. This is the first report of a large-scale, long-term efficacy study of any biocontrol product developed to date for a field crop. Most (>95%) of 213,406 tons of maize grains harvested from treated fields contained 90%) contained 80% less aflatoxin content than untreated crops. The frequency of the biocontrol active ingredient atoxigenic genotypes in grains from treated fields was significantly higher than in grains from control fields. A higher proportion of grains from treated fields met various aflatoxin standards compared to grains from untreated fields. Results indicate that efficacy of the biocontrol product in limiting aflatoxin contamination is stable regardless of environment and cropping system. In summary, the biocontrol technology allows farmers across Nigeria to produce safer crops for consumption and increases potential for access to premium markets that require aflatoxin-compliant crops.

In warm agricultural areas across the globe, maize, groundnut, and other crops become frequently contaminated with aflatoxins produced primarily by the fungus Aspergillus flavus. Crop contamination with those highly toxic and carcinogenic compounds impacts both human and animal health, as well as the income of farmers and trade. In Nigeria, poultry productivity is hindered by high prevalence of aflatoxins in feeds. A practical solution to decrease crop aflatoxin content is to use aflatoxin biocontrol products based on non-toxin-producing strains of A. flavus. The biocontrol product Aflasafe&reg, was registered in 2014 for use in maize and groundnut grown in Nigeria. Its use allows the production of aflatoxin-safe maize and groundnut. A portion of the maize treated with Aflasafe in Nigeria is being used to manufacture feeds used by the poultry industry, and productivity is improving. One of the conditions to register Aflasafe with the national regulator was to demonstrate both the safety of Aflasafe-treated maize to avian species and the impact of Aflasafe as a public good. Results presented here demonstrate that the use of maize colonized by an atoxigenic strain of Aflasafe resulted in superior (p &lt, 0.05) broiler performance in all evaluated parameters in comparison to broilers fed with toxigenic maize. Use of an aflatoxin-sequestering agent (ASA) was not sufficient to counteract the harmful effects of aflatoxins. Both the safety and public good value of Aflasafe were demonstrated during our study. In Nigeria, the availability of aflatoxin-safe crops as a result of using Aflasafe allows poultry producers to improve their productivity, their income, and the health of consumers of poultry products.

Aflatoxins pose significant food security and public health risks, decrease productivity and profitability of animal industries, and hamper trade. To minimize aflatoxin contamination in several crops, a biocontrol technology based on atoxigenic strains of Aspergillus flavus is commercially used in the United States and some African countries. Significant efforts are underway to popularize the use of biocontrol in Africa by various means including incentives. The purpose of this study was to develop quantitative pyrosequencing assays for rapid, simultaneous quantification of proportions of four A. flavus biocontrol genotypes within complex populations of A. flavus associated with maize crops in Nigeria to facilitate payment of farmer incentives for Aflasafe (a biocontrol product) use. Protocols were developed to confirm use of Aflasafe by small scale farmers in Nigeria. Nested PCR amplifications followed by sequence by synthesis pyrosequencing assays were required to quantify frequencies of the active ingredients and, in so doing, confirm successful use of biocontrol by participating farmers. The entire verification process could be completed in 3–4 days proving a savings over other monitoring methods in both time and costs and providing data in a time frame that could work with the commercial agriculture scheme. Quantitative pyrosequencing assays represent a reliable tool for rapid detection, quantification, and monitoring of multiple A. flavus genotypes within complex fungal communities, satisfying the requirements of the regulatory community and crop end-users that wish to determine which purchased crops were treated with the biocontrol product. Techniques developed in the current study can be modified for monitoring other crop-associated fungi.

Across sub-Saharan Africa, chili peppers are fundamental ingredients of many traditional dishes. However, chili peppers may contain unsafe aflatoxin concentrations produced by Aspergillus section Flavi fungi. Aflatoxin levels were determined in chili peppers from three states in Nigeria. A total of 70 samples were collected from farmers&rsquo, stores and local markets. Over 25% of the samples contained unsafe aflatoxin concentrations. The chili peppers were associated with both aflatoxin producers and atoxigenic Aspergillus flavus genotypes. Efficacy of an atoxigenic biocontrol product, Aflasafe, registered in Nigeria for use on maize and groundnut, was tested for chili peppers grown in three states. Chili peppers treated with Aflasafe accumulated significantly less aflatoxins than nontreated chili peppers. The results suggest that Aflasafe is a valuable tool for the production of safe chili peppers. Use of Aflasafe in chili peppers could reduce human exposure to aflatoxins and increase chances to commercialize chili peppers in premium local and international markets. This is the first report of the efficacy of any atoxigenic biocontrol product for controlling aflatoxin in a spice crop.

Maize is an important staple crop for the majority of the population in Uganda. However, in tropical and subtropical climates, maize is frequently contaminated with aflatoxins, a group of cancer-causing and immuno-suppressive mycotoxins produced by Aspergillus section Flavi fungi. In Uganda, there is limited knowledge about the causal agents of aflatoxin contamination. The current study determined both the aflatoxin levels in pre-harvest maize across Uganda and the structures of communities of aflatoxin-producing fungi associated with the maize. A total of 256 pre-harvest maize samples were collected from 23 major maize-growing districts in eight agro-ecological zones (AEZ). Maize aflatoxin content ranged from 0 to 3760 ng/g although only around 5% for Ugandan thresholds. For EU it is about 16% of the samples contained aflatoxin concentrations above tolerance thresholds. A total of 3105 Aspergillus section Flavi isolates were recovered and these were dominated by the A. flavus L morphotype (89.4%). Densities of aflatoxin-producing fungi were negatively correlated with elevation. Farming systems and climatic conditions of the AEZ are thought to have influenced communities' structure composition. Fungi from different AEZ varied significantly in aflatoxin-producing abilities and several atoxigenic genotypes were identified. The extremely high aflatoxin concentrations detected in some of the studied regions indicate that management strategies should be urgently designed for use at the pre-harvest stage. Atoxigenic genotypes detected across Uganda could serve as aflatoxin biocontrol agents to reduce crop contamination from fields conditions and throughout the maize value chain.

Aflatoxin accumulation in maize (Zea mays L.) kernels is a serious economic and health problem that reduces grain quality and nutritional values and causes death to livestock and humans. Understanding the genetic parameters and heterotic responses of exotic maize inbred lines can facilitate their use for developing aflatoxin resistant parents of hybrids in Africa. This study was designed to (1) determine the heterotic affinities of aflatoxin resistant exotic lines, (2) identify exotic inbreds with good combining ability, and (3) determine the mode of inheritance of resistance to aflatoxin contamination in these lines. A line × tester mating design was used to determine combining ability of 12 yellow and 13 white inbreds and classify them into heterotic groups. The inbreds were crossed to two adapted testers representing two African heterotic groups and the resulting testcrosses along with hybrid checks were evaluated in separate trials at two locations for 2 years in Nigeria. General combining ability (GCA) effects were more important than specific combining ability effects for aflatoxin and grain yield. Among 15 exotic inbred lines having negative GCA effects for aflatoxin and 13 with positive GCA effects for grain yield, six combined the two desired traits. Five white and six yellow endosperm testcrosses were found to be good specific combiners for the two desired traits. The exotic lines with negative GCA effects for aflatoxin accumulation will be used as donor parents to develop backcross populations for generating new inbred lines with much higher levels of resistance to aflatoxin accumulation.

Aspergillus flavus and Aspergillus parasiticus are ubiquitous in natural environments and have the potential to produce aflatoxins that can cause both chronic and acute diseases in humans and animals. The near-infrared (800–2500 nm) reflectance spectra of almonds from the 2012 and 2013 harvest seasons inoculated with one selected isolate of either A. flavus or A. parasiticus were analysed to determine if the spectra could be used to distinguish infected almonds from uninfected control almonds. A canonical classifier was developed that could discriminate infected almonds from the uninfected control kernels with a total cross-validation error rate of 0.26% and zero false negative errors. Additionally, a follow-up canonical classifier was developed to evaluate the potential to discriminate between infections by the two Aspergillus species, once infection was detected. Spectral analysis indicated that the NIR wavebands associated with lipid content were the most indicative factors in both identifying the infected almonds as well as discriminating between almonds infected by the A. flavus or the A. parasiticus isolate. Further comparison of the canonical discriminant analysis results from the 2nd derivative spectra and the original spectra also suggested that the chemical differences caused by the fungal metabolism had a much greater contribution to the discriminant function than the structural changes caused by the fungal invasion. The wavelengths identified in this study could be helpful in developing non-destructive multispectral sensing systems to effectively scan and detect almonds with fungal infection.

Almonds can be contaminated with aflatoxins, produced mainly by Aspergillus flavus and A. parasiticus. Infection can be facilitated by insect injuries during hull split, which begins four to six weeks before harvest. Within this period, it is unknown which kernel stages are most susceptible to aflatoxin contamination. Developing almonds of the Nonpareil cultivar were inoculated weekly with a spore suspension of A. flavus or A. parasiticus for five weeks after hull split in 2013. The almonds were infested with eggs of the lepidopteron navel orangeworm (NOW) (Amyelois transitella) before each spore inoculation. Aflatoxin levels were quantified at harvest using HPLC. Aflatoxin contamination was consistently higher in NOW-damaged kernels, although aflatoxins were also detected in undamaged kernels at each inoculation date. Insect injury is not required for kernel infection but it is a key risk factor for high aflatoxin contamination. Laboratory inoculations were also performed on Nonpareil almond kernels collected during the summers of 2013 and 2015. Aflatoxin levels were significantly lower on dried almonds but the ability to produce aflatoxins was restored when almonds were incubated with high humidity or when the Aspergillus species were inoculated on almond meal agar amended with ground kernels. Therefore, aflatoxins can accumulate in kernels with low aw, should sufficient moisture favors aflatoxin production. In our field experiment, the orchard micro-climate had sufficient humidity to enable aflatoxin production in both damaged and undamaged dried kernels.

Maize, the critical staple food for billions of people, was domesticated in Mexico about 9,000 YBP. Today, a great array of maize landraces (MLRs) across rural Mexico is harbored in a living library that has been passed among generations since before the establishment of the modern state. MLRs have been selected over hundreds of generations by ethnic groups for adaptation to diverse environmental settings. The genetic diversity of MLRs in Mexico is an outstanding resource for development of maize cultivars with beneficial traits. Maize is frequently contaminated with aflatoxins by Aspergillus flavus, and resistance to accumulation of these potent carcinogens has been sought for over three decades. However, MLRs from Mexico have not been evaluated as potential sources of resistance. Variation in susceptibility to both A. flavus reproduction and aflatoxin contamination was evaluated on viable maize kernels in laboratory experiments that included 74 MLR accessions collected from 2006 to 2008 in the central west and northwest regions of Mexico. Resistant and susceptible MLR accessions were detected in both regions. The most resistant accessions accumulated over 99 % less aflatoxin B1 than did the commercial hybrid control Pioneer P33B50. Accessions supporting lower aflatoxin accumulation also supported reduced A. flavus sporulation. Sporulation on the MLRs was positively correlated with aflatoxin accumulation (R = 0.5336, P0.0001), suggesting that resistance to fungal reproduction is associated with MLR aflatoxin resistance. Results of the current study indicate that MLRs from Mexico are potentially important sources of aflatoxin resistance that may contribute to the breeding of commercially acceptable and safe maize hybrids and/or open pollinated cultivars for human and animal consumption.

Humans and animals are exposed to aflatoxins, toxic carcinogenic fungal metabolites, through consumption of contaminated food and feed. Aspergillus flavus, the primary causal agent of crop aflatoxin contamination, is composed of phenotypically and genotypically diverse vegetative compatibility groups (VCGs). Molecular data suggest that VCGs largely behave as clones with certain VCGs exhibiting niche preference. VCGs vary in aflatoxin-producing ability, ranging from highly aflatoxigenic to atoxigenic. The prevalence of individual VCGs is dictated by competition during growth and reproduction under variable biotic and abiotic conditions. Agronomic practices influence structures and average aflatoxin-producing potentials of A. flavus populations and, as a result, incidences and severities of crop contamination. Application of atoxigenic strains has successfully reduced crop aflatoxin contamination across large areas in the United States. This strategy uses components of the endemic diversity to alter structures of A. flavus populations and improve safety of food, feed, and the overall environment.

Interactions between pathogenic and nonpathogenic fungal species in the tree canopy are complex and can determine if disease will manifest in the plant and in other organisms such as honey bees. Seasonal dynamics of fungi were studied in an almond orchard in California where experimental release of the atoxigenic biopesticide Aspergillus flavus AF36 to displace toxigenic Aspergillus strains has been conducted for five years. The presence of the vegetative compatibility group (VCG) YV36, to which AF36 belongs, in the blossoms, and the honey bees that attend these blossoms, was assessed. In blossoms, A. flavus frequencies ranged from 0 to 4.5%, depending on the year of study. Frequencies of honey bees carrying A. flavus ranged from 6.5 to 10%. Only one A. flavus isolate recovered from a blossom in 2016 belonged to YV36, while members of the VCG were not detected contaminating honey bees. Exposure of pollinator honey bees to AF36 was detected to be very low. The density of several Aspergillus species was found to increase during almond hull split and throughout the final stages of maturation; this also occurred in pistachio orchards during the maturation period. Additionally, we found that AF36 effectively limited almond aflatoxin contamination in laboratory assays. This study provides knowledge and understanding of the seasonal dynamics of Aspergillus fungi and will help design aflatoxin management strategies for almond. The evidence of the low levels of VCG YV36 encountered on almond blossoms and bees during pollination and AF36's effectiveness in limiting aflatoxin contamination in almond provided additional support for the registration of AF36 with USEPA to use in almond in California.

AIMS To assess frequencies of the Aspergillus flavus atoxigenic vegetative compatibility group (VCG) YV36, to which the biocontrol agent AF36 belongs, in maize-growing regions of Mexico. METHODS AND RESULTS Over 3500 A. flavus isolates recovered from maize agroecosystems in four states of Mexico during 2005 through 2008 were subjected to vegetative compatibility analyses based on nitrate nonutilizing mutants. Results revealed that 59 (1·6%) isolates belong to VCG YV36. All 59 isolates had the MAT1-2 idiomorph at the mating-type locus and the single nucleotide polymorphism in the polyketide synthase gene that confers atoxigenicity. Additional degradation of the aflatoxin gene cluster was detected in three isolates. Microsatellite loci analyses revealed low levels of genetic diversity and no linkage disequilibrium within VCG YV36. CONCLUSIONS The VCG to which the biocontrol agent AF36 belongs, YV36, is also native to Mexico. The North American Free Trade Agreement should facilitate adoption of AF36 for use by Mexico in aflatoxin prevention programs. SIGNIFICANCE AND IMPACT OF THE STUDY An USEPA registered biocontrol agent effective at preventing aflatoxin contamination of crops in the US, is also native to Mexico. This should facilitate the path to registration of AF36 as the first biopesticide for aflatoxin mitigation of maize in Mexico. Economic and health benefits to the population of Mexico should result once aflatoxin mitigation programs based on AF36 applications are implemented.

In California, aflatoxin contamination of almond, fig, and pistachio has become a serious problem in recent years due to long periods of drought and probably other climatic changes. The atoxigenic biocontrol product Aspergillus flavus AF36 has been registered for use to limit aflatoxin contamination of pistachio since 2012 and for use in almond and fig since 2017. New biocontrol technologies employ multiple atoxigenic genotypes because those provide greater benefits than using a single genotype. Almond, fig, and pistachio industries would benefit from a multi-strain biocontrol technology for use in these three crops. Several A. flavus vegetative compatibility groups (VCGs) associated with almond, fig, and pistachio composed exclusively of atoxigenic isolates, including the VCG to which AF36 belongs to, YV36, were previously characterized in California. Here, we report additional VCGs associated with either two or all three crops. Representative isolates of 12 atoxigenic VCGs significantly (P < 0.001) reduced (>80%) aflatoxin accumulation in almond and pistachio when challenged with highly toxigenic isolates of A. flavus and A. parasiticus under laboratory conditions. Isolates of the evaluated VCGs, including AF36, constitute valuable endemic, well-adapted, and efficient germplasm to design a multi-crop, multi-strain biocontrol strategy for use in tree crops in California. Availability of such a strategy would favor long-term atoxigenic A. flavus communities across the affected areas of California, and this would result in securing domestic and export markets for the nut crop and fig farmer industries and, most importantly, health benefits to consumers.