Does Use of Atoxigenic Biocontrol Products to Mitigate Aflatoxin in Maize Increase Fumonisin Content in 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 (<1 part per million; ppm). Most untreated maize also harbored equally low fumonisin levels but contained higher aflatoxin levels. In addition, during one year, we detected considerably lower Fusarium spp. densities in treated maize than in untreated maize. Our results do not support the hypothesis that treating crops with atoxigenic isolates of A. flavus used in biocontrol formulations results in higher grain fumonisin levels.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Comparative Reliability of Screening Parameters for Anthracnose Resistance in Water Yam (Dioscorea alata).

Anthracnose, caused by Colletotrichum gloeosporioides, is one of the major constraints limiting water yam (Dioscorea alata) production in the tropics. In this region, yam anthracnose control is mostly achieved by the deployment of moderately resistant yam genotypes. Therefore, screening for new sources of anthracnose resistance is an important aspect of yam research in the tropics. The reliability and applicability of different yam anthracnose rating parameters has not been fully examined. Disease severity on detached leaves in the laboratory and leaf severity, lesion size, and spore production on whole plants in the greenhouse were used to screen an F1 yam population and correlate screening results with field evaluations. Anthracnose lesion size had the smallest predicted residual means but whole-plant severity and detached-leaf severity had the best variance homogeneity and relatively small predicted residual means. The concordance correlation coefficient (rc) and κ statistic were used to determine the agreement between anthracnose rating parameters and field evaluations. Detached-leaf (rc = 0.95, κ = 0.81) and whole-plant (rc = 0.96, κ = 0.86) evaluations had high positive agreement with field evaluation but spore production (κ = 0.69) and lesion size (κ = 0.57) had moderate positive agreement. These results suggest that all the evaluated rating parameters can be used to successfully screen yam germplasm for anthracnose resistance but lesion size and spore production data may need to be transformed.

Micro-climatic variations across Malawi have a greater influence on contamination of maize with aflatoxins than with fumonisins.

This study reports levels of aflatoxin and fumonisin in maize samples (n = 1294) from all agroecological zones (AEZs) in Malawi. Most maize samples (> 75%) were contaminated with aflatoxins and 45% with fumonisins, which co-occurred in 38% of the samples. Total aflatoxins varied across the AEZs, according to mean annual temperature (P < 0.05) of the AEZs. Samples from the lower Shire AEZ (median = 20.8 µg/kg) had higher levels of aflatoxins (P < 0.05) than those from the other AEZs (median = 3.0 µg/kg). Additionally, the majority (75%) of the positive samples from the lower Shire AEZ had aflatoxin levels exceeding the EU regulatory limit (4 µg/kg), whereas 25%, 37%, and 39% of positive samples exceeded the threshold in the mid-elevation, Lake Shore and upper and middle Shire, and highlands AEZs, respectively. The lower Shire AEZ is characterised by higher mean temperatures throughout the year and low erratic rainfall. However, total fumonisins did not show significant variation across AEZs, but all positive samples exceeded 150 µg/kg, required for tolerable daily intake of 1.0 µg/kg body weight per day, established by the European Food Safety Authority Panel on Contaminants in the Food Chain. Therefore, results of this study suggest that contamination of maize with aflatoxin responds to micro-climate more than with fumonisins. In addition, the data will be useful to public health policy-makers and stakeholders to articulate and implement monitoring and mitigation programs.

Aflatoxin contamination of maize and groundnut in Burundi: Distribution of contamination, identification of causal agents and potential biocontrol genotypes of Aspergillus flavus.

Aflatoxin contamination of the staples maize and groundnut is a concern for health and economic impacts across sub-Saharan Africa. The current study (i) determined aflatoxin levels in maize and groundnut collected at harvest in Burundi, (ii) characterized populations of Aspergillus section Flavi associated with the two crops, and (iii) assessed aflatoxin-producing potentials among the recovered fungi. A total of 120 groundnut and 380 maize samples were collected at harvest from eight and 16 provinces, respectively. Most of the groundnut (93%) and maize (87%) contained aflatoxin below the European Union threshold, 4 µg/kg. Morphological characterization of the recovered Aspergillus section Flavi fungi revealed that the L-morphotype of A. flavus was the predominant species. Aflatoxin production potentials of the L-morphotype isolates were evaluated in maize fermentations. Some isolates produced over 137,000 µg/kg aflatoxin B1. Thus, despite the relatively low aflatoxin levels at harvest, the association of both crops with highly toxigenic fungi poses significant risk of post-harvest aflatoxin contamination and suggests measures to mitigate aflatoxin contamination in Burundi should be developed. Over 55% of the L-morphotype A. flavus did not produce aflatoxins. These atoxigenic L-morphotype fungi were characterized using molecular markers. Several atoxigenic genotypes were detected across the country and could be used as biocontrol agents. The results from the current study hold promise for developing aflatoxin management strategies centered on biocontrol for use in Burundi to reduce aflatoxin contamination throughout the value chain.

Impact of frequency of application on the long-term efficacy of the biocontrol product Aflasafe in reducing aflatoxin contamination in maize.

Aflatoxins, produced by several Aspergillus section Flavi species in various crops, are a significant public health risk and a barrier to trade and development. In sub-Saharan Africa, maize and groundnut are particularly vulnerable to aflatoxin contamination. Aflasafe, a registered aflatoxin biocontrol product, utilizes atoxigenic A. flavus genotypes native to Nigeria to displace aflatoxin producers and mitigate aflatoxin contamination. Aflasafe was evaluated in farmers' fields for 3 years, under various regimens, to quantify carry-over of the biocontrol active ingredient genotypes. Nine maize fields were each treated either continuously for 3 years, the first two successive years, in year 1 and year 3, or once during the first year. For each treated field, a nearby untreated field was monitored. Aflatoxins were quantified in grain at harvest and after simulated poor storage. Biocontrol efficacy and frequencies of the active ingredient genotypes decreased in the absence of annual treatment. Maize treated consecutively for 2 or 3 years had significantly (p < 0.05) less aflatoxin (92% less) in grain at harvest than untreated maize. Maize grain from treated fields subjected to simulated poor storage had significantly less (p < 0.05) aflatoxin than grain from untreated fields, regardless of application regimen. Active ingredients occurred at higher frequencies in soil and grain from treated fields than from untreated fields. The incidence of active ingredients recovered in soil was significantly correlated (r = 0.898; p < 0.001) with the incidence of active ingredients in grain, which in turn was also significantly correlated (r = -0.621, p = 0.02) with aflatoxin concentration. Although there were carry-over effects, caution should be taken when drawing recommendations about discontinuing biocontrol use. Cost-benefit analyses of single season and carry-over influences are needed to optimize use by communities of smallholder farmers in sub-Saharan Africa.

Aflatoxin Contamination of Maize, Groundnut, and Sorghum Grown in Burkina Faso, Mali, and Niger and Aflatoxin Exposure Assessment.

Aflatoxin contamination of staple crops by Aspergillus flavus and closely related fungi is common across the Sahel region of Africa. Aflatoxins in maize, groundnut, and sorghum collected at harvest or from farmers' stores within two weeks of harvest from Burkina Faso, Mali, and Niger were quantified. Thereafter, aflatoxin exposure values were assessed using per capita consumption rates of those crops. Mean aflatoxin concentrations in maize were high, 128, 517, and 659 µg/kg in Mali, Burkina Faso, and Niger, respectively. The estimated probable daily intake (PDI) of aflatoxins from maize ranged from 6 to 69, 29 to 432, and 310 to 2100 ng/kg bw/day in Mali, Burkina Faso, and Niger, respectively. Similarly, mean aflatoxin concentrations in sorghum were high, 76 and 259 µg/kg in Mali and Niger, respectively, with an estimated PDI of 2-133 and 706-2221. For groundnut, mean aflatoxin concentrations were 115, 277, and 628 µg/kg in Mali, Burkina Faso, and Niger, respectively. Aflatoxin exposure values were high with an estimated 9, 28, and 126 liver cancer cases/100,000 persons/year in Mali, Burkina Faso, and Niger, respectively. Several samples were extremely unsafe, exceeding manyfold regulatory levels of diverse countries (up to 2000 times more). Urgent attention is needed across the Sahel for integrated aflatoxin management for public health protection, food and nutrition security, and access to trade opportunities.

Evaluating the Performance of Lateral Flow Devices for Total Aflatoxins with Special Emphasis on Their Robustness under Sub-Saharan Conditions.

As aflatoxins are a global risk for humans and animals, testing methods for rapid on-site screening are increasingly needed alongside the standard analytical laboratory tools. In the presented study, lateral flow devices (LFDs) for rapid total aflatoxin screening were thoroughly investigated with respect to their matrix effects, cross-reactivity, their performance under harsh conditions in Sub-Saharan Africa (SSA), and their stability, as well as when compared with liquid chromatography-tandem mass spectrometry (LC-MS/MS). To analyze the matrix effects, qualitative test kits offering a certain cutoff level were used to screen different nut samples. In addition, these tests were challenged on their cross-reactivity with 230 fungal toxins and metabolites. Furthermore, the resulting measurements performed under harsh tropical conditions (up to 38.4 °C and 91% relative humidity) in SSA, specifically Burkina Faso and Mozambique, were compared with the results from a well-established and validated LC-MS/MS-based reference method. The comparison of the on-site LFD results with the reference method showed a good agreement: 86.4% agreement, 11.8% non-agreement, and 1.8% invalid test results. To test the robustness of the cutoff tests, short- and long-term stability testing was carried out in Mozambique and Nigeria. For both experiments, no loss of test performance could be determined. Finally, a subset of African corn samples was shipped to Austria and analyzed under laboratory conditions using semiquantitative aflatoxin tests. A good correlation was found between the rapid strip tests and the LC-MS/MS reference method. Overall, the evaluated LFDs showed satisfying results regarding their cross-reactivity, matrix effects, stability, and robustness.

Temperature Influences on Interactions Among Aflatoxigenic Species of Aspergillus Section Flavi During Maize Colonization.

Fungal species within Aspergillus section Flavi contaminate food and feed with aflatoxins. These toxic fungal metabolites compromise human and animal health and disrupt trade. Genotypically and phenotypically diverse species co-infect crops, but temporal and spatial variation in frequencies of different lineages suggests that environmental factors such as temperature may influence structure of aflatoxin-producing fungal communities. Furthermore, though most species within Aspergillus section Flavi produce sclerotia, divergent sclerotial morphologies (small or S-type sclerotia vs. large or L-type sclerotia) and differences in types and quantities of aflatoxins produced suggest lineages are adapted to different life strategies. Temperature is a key parameter influencing pre- and post-harvest aflatoxin contamination of crops. We tested the hypothesis that species of aflatoxin-producing fungi that differ in sclerotial morphology will vary in competitive ability and that outcomes of competition and aflatoxin production will be modulated by temperature. Paired competition experiments between highly aflatoxigenic S-type species (A. aflatoxiformans and Lethal Aflatoxicosis Fungus) and L-type species (A. flavus L morphotype and A. parasiticus) were conducted on maize kernels at 25 and 30°C. Proportions of each isolate growing within and sporulating on kernels were measured using quantitative pyrosequencing. At 30°C, S-type fungi were more effective at host colonization compared to L-type isolates. Total aflatoxins and the proportion of B vs. G aflatoxins were greater at 30°C compared to 25°C. Sporulation by L-type isolates was reduced during competition with S-type fungi at 30°C, while relative quantities of conidia produced by S-type species either increased or did not change during competition. Results indicate that both species interactions and temperature can shape population structure of Aspergillus section Flavi, with warmer temperatures favoring growth and dispersal of highly toxigenic species with S-type sclerotia.

Prevalence of Aflatoxin- and Fumonisin-Producing Fungi Associated with Cereal Crops Grown in Zimbabwe and Their Associated Risks in a Climate Change Scenario.

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 (13-25% for AF and 0-32% for FM). Less than 2% of small grain samples exceeded the EU regulatory limit for AF (4 µg/kg), while <10% exceeded the EU regulatory limit for FM (1000 µg/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 (≤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.

Field efficacy of two atoxigenic biocontrol products for mitigation of aflatoxin contamination in maize and groundnut in Ghana.

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 < 0.05) displacement of toxigenic fungi occurred in both crops during both years, in all five regions. Biocontrol-treated crops consistently had significantly (P < 0.05) less aflatoxins (range = 76% to 100% less; average = 99% less) than untreated crops. Results indicate that both biocontrol products are highly efficient, cost-effective, environmentally safe tools for aflatoxin mitigation. Most crops from treated fields could have been sold in both local and international food and feed premium markets. Adoption and use of biocontrol products have the potential to improve the health of Ghanaians, and both income and trade opportunities of farmers, aggregators, distributors, and traders.

Performance of Broilers Fed with Maize Colonized by Either Toxigenic or Atoxigenic Strains of Aspergillus flavus with and without an Aflatoxin-Sequestering Agent.

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® 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 < 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.

"Ground-Truthing" Efficacy of Biological Control for Aflatoxin Mitigation in Farmers' Fields in Nigeria: From Field Trials to Commercial Usage, a 10-Year Study.

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 <20 ppb total aflatoxins, and a significant proportion (>90%) contained <4 ppb total aflatoxins. Grains from treated plots had preponderantly >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.

Potential of Atoxigenic Aspergillus flavus Vegetative Compatibility Groups Associated With Maize and Groundnut in Ghana as Biocontrol Agents for Aflatoxin Management.

Increasing knowledge of the deleterious health and economic impacts of aflatoxin in crop commodities has stimulated global interest in aflatoxin mitigation. Current evidence of the incidence of Aspergillus flavus isolates belonging to vegetative compatibility groups (VCGs) lacking the ability to produce aflatoxins (i.e., atoxigenic) in Ghana may lead to the development of an aflatoxin biocontrol strategy to mitigate crop aflatoxin content. In this study, 12 genetically diverse atoxigenic African A. flavus VCGs (AAVs) were identified from fungal communities associated with maize and groundnut grown in Ghana. Representative isolates of the 12 AAVs were assessed for their ability to inhibit aflatoxin contamination by an aflatoxin-producing isolate in laboratory assays. Then, the 12 isolates were evaluated for their potential as biocontrol agents for aflatoxin mitigation when included in three experimental products (each containing four atoxigenic isolates). The three experimental products were evaluated in 50 maize and 50 groundnut farmers' fields across three agroecological zones (AEZs) in Ghana during the 2014 cropping season. In laboratory assays, the atoxigenic isolates reduced aflatoxin biosynthesis by 87-98% compared to grains inoculated with the aflatoxin-producing isolate alone. In field trials, the applied isolates moved to the crops and had higher (P < 0.05) frequencies than other A. flavus genotypes. In addition, although at lower frequencies, most atoxigenic genotypes were repeatedly found in untreated crops. Aflatoxin levels in treated crops were lower by 70-100% in groundnut and by 50-100% in maize (P < 0.05) than in untreated crops. Results from the current study indicate that combined use of appropriate, well-adapted isolates of atoxigenic AAVs as active ingredients of biocontrol products effectively displace aflatoxin producers and in so doing limit aflatoxin contamination. A member each of eight atoxigenic AAVs with superior competitive potential and wide adaptation across AEZs were selected for further field efficacy trials in Ghana. A major criterion for selection was the atoxigenic isolate's ability to colonize soils and grains after release in crop field soils. Use of isolates belonging to atoxigenic AAVs in biocontrol management strategies has the potential to improve food safety, productivity, and income opportunities for smallholder farmers in Ghana.

Monitoring Aspergillus flavus Genotypes in a Multi-Genotype Aflatoxin Biocontrol Product With Quantitative Pyrosequencing.

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.

Aflatoxin in Chili Peppers in Nigeria: Extent of Contamination and Control Using Atoxigenic Aspergillus flavus Genotypes as Biocontrol Agents.

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' 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.

Distribution and toxigenicity of Aspergillus species isolated from maize kernels from three agro-ecological zones in Nigeria.

Maize samples were collected during a survey in three agro-ecological zones in Nigeria to determine the distribution and aflatoxin-producing potential of members of Aspergillus section Flavi. The three agro-ecological zones were, Derived Savannah (DS) and Southern Guinea Savannah (SGS) in the humid south and North Guinea Savannah (NGS) in the drier north. Across agro-ecological zones, Aspergillus was the most predominant fungal genera identified followed by Fusarium with mean incidences of 70 and 24%, respectively. Among Aspergillus, A. flavus was the most predominant and L-strains constituted >90% of the species identified, while the frequency of the unnamed taxon S(BG) was <3%. The incidence of atoxigenic strains of A. flavus was higher in all the districts surveyed except in the Ogbomosho and Mokwa districts in DS and SGS zones, respectively, where frequency of toxigenic strains were significantly (P<0.05) higher than that of atoxigenic strains. The highest and lowest incidence of aflatoxin positive samples was recorded in the SGS (72%) and NGS (20%), respectively. Aflatoxin contamination in grain also followed a similar trend and the highest mean levels of B-aflatoxins were detected in maize samples obtained from Bida (612 ng g(-1)) and Mokwa (169 ng g(-1)) districts, respectively, in the SGS. Similarly, the highest concentrations of G-aflatoxins were detected in samples from Akwanga district in the SGS with a mean of 193 and 60 ng g(-1), respectively. When agro-ecological zones were compared, B-aflatoxins were significantly (P<0.05) higher in SGS than in NGS, and intermediate in maize samples from the DS agro-ecological zone.

Occurrence of aflatoxin in agricultural produce from local markets in Burundi and Eastern Democratic Republic of Congo.

Aflatoxins are noxious secondary metabolites, of certain fungal species, found in food and feed. Contamination of a commodity with aflatoxins is associated with production and storage losses, and subsequently less food availability. Aflatoxins can also pose human health risks and represent a barrier to the development of trade, in both domestic and international markets. In this study, samples of cassava, maize, groundnut, beans, soybean, sorghum and milk, and their processed products were collected from local markets in Burundi and Eastern DRC. In order to investigate the levels of aflatoxin, crop samples were analyzed using a single step lateral flow immunochromatographic assay (Reveal Q+), while enzyme-linked immune-sorbent assay (ELISA) was used to analyze aflatoxin-M1 in milk, yogurt, and cheese samples. The results revealed the presence of aflatoxins in all samples from both countries, with levels ranging from 1.3 to 2,410 µg/kg. Samples collected from Burundi contained relatively higher (p > 0.0.5) levels of aflatoxins. In 51% of all the crops samples, total aflatoxin contamination was above the EU maximum tolerable level of 4 µg/kg. Processed products, particularly from groundnut, maize, and sorghum, had the highest levels of aflatoxin contamination when compared to unprocessed grain. With regard to milk and dairy products, the level of aflatoxin-M1 ranged from 4.8 to 261.1 ng/kg. Approximately 29% of milk and yogurt samples had aflatoxin-M1 higher than the EU regulatory limit of 50 ng/kg, whereas 20% of cheese samples were found to be contaminated at levels higher than the maximum limit of 250 ng/kg. These results can serve as the basis for pre- and postharvest approaches to reduce aflatoxin contamination in agricultural commodities in Burundi and Eastern DRC in order to reduce health risk, avoid reduced production in livestock, and open up export markets.

An Assessment of Willingness To Pay by Maize and Groundnut Farmers for Aflatoxin Biocontrol Product in Northern Nigeria.

In Nigeria, Aflasafe is a registered biological product for reducing aflatoxin infestation of crops from the field to storage, making the crops safer for consumption. The important questions are whether farmers will purchase and apply this product to reduce aflatoxin contamination of crops, and if so under what conditions. A study was carried out to address these questions and assess determinants of willingness to pay (WTP) for the product among maize and groundnut farmers in Kano and Kaduna states in Nigeria. A multistage sampling technique was used to collect primary data from 492 farmers. The majority of farmers who had direct experience with Aflasafe (experienced farmers) in Kano (80.7%) and Kaduna (84.3%) had a WTP bid value equal to or greater than the threshold price ($10) at which Aflasafe was to be sold. The mean WTP estimates for Aflasafe for experienced farmers in Kano and Kaduna were statistically the same. However, values of $3.56 and $7.46 were offered in Kano and Kaduna states, respectively, by farmers who had never applied Aflasafe (inexperienced farmers), and the difference here was significant (P < 0.01). Regression results indicate that contact with extension agents (P < 0.01) and access to credit (P < 0.05) positively and significantly influenced the probability that a farmer would be willing to pay more for Aflasafe than the threshold price. Lack of awareness of the importance of Aflasafe was the major reason cited by inexperienced farmers (64% in Kano state and 21% in Kaduna state) for not using the product. A market strategy promoting a premium price for aflatoxin-safe produce and creating awareness and explaining the availability of Aflasafe to potential users should increase Aflasafe usage.

Environmental distribution and genetic diversity of vegetative compatibility groups determine biocontrol strategies to mitigate aflatoxin contamination of maize by Aspergillus flavus.

Maize infected by aflatoxin-producing Aspergillus flavus may become contaminated with aflatoxins, and as a result, threaten human health, food security and farmers' income in developing countries where maize is a staple. Environmental distribution and genetic diversity of A. flavus can influence the effectiveness of atoxigenic isolates in mitigating aflatoxin contamination. However, such information has not been used to facilitate selection and deployment of atoxigenic isolates. A total of 35 isolates of A. flavus isolated from maize samples collected from three agro-ecological zones of Nigeria were used in this study. Ecophysiological characteristics, distribution and genetic diversity of the isolates were determined to identify vegetative compatibility groups (VCGs). The generated data were used to inform selection and deployment of native atoxigenic isolates to mitigate aflatoxin contamination in maize. In co-inoculation with toxigenic isolates, atoxigenic isolates reduced aflatoxin contamination in grain by > 96%. A total of 25 VCGs were inferred from the collected isolates based on complementation tests involving nitrate non-utilizing (nit(-)) mutants. To determine genetic diversity and distribution of VCGs across agro-ecological zones, 832 nit(-) mutants from 52 locations in 11 administrative districts were paired with one self-complementary nitrate auxotroph tester-pair for each VCG. Atoxigenic VCGs accounted for 81.1% of the 153 positive complementations recorded. Genetic diversity of VCGs was highest in the derived savannah agro-ecological zone (H = 2.61) compared with the southern Guinea savannah (H = 1.90) and northern Guinea savannah (H = 0.94) zones. Genetic richness (H = 2.60) and evenness (E5 = 0.96) of VCGs were high across all agro-ecological zones. Ten VCGs (40%) had members restricted to the original location of isolation, whereas 15 VCGs (60%) had members located between the original source of isolation and a distance > 400 km away. The present study identified widely distributed VCGs in Nigeria such as AV0222, AV3279, AV3304 and AV16127, whose atoxigenic members can be deployed for a region-wide biocontrol of toxigenic isolates to reduce aflatoxin contamination in maize.