Aflatoxin Control Research Articles

A review of agricultural aflatoxin management strategies and emerging innovations in sub-Saharan Africa

Aflatoxins are highly carcinogenic secondary metabolites produced by Aspergillus flavus , A. parasiticus and A. nomius . Aflatoxin contamination of food and animal feeds is, therefore, a major food security, food safety, trade, human and domestic animal health concern. Researchers worldwide have suggested various agriculture-based strategies to manage aflatoxigenic Aspergillus species and reduce contamination to safe levels. This paper reviews various agricultural strategies that could be employed to reduce contamination of aflatoxins in food crops and animal feeds, as well as the challenges faced by these reduction strategies. Among these strategies are innovations like Aflasafe TM and solar grain driers. It is hoped that this critique will stimulate refinement of the existing aflatoxin control approaches and innovations to maximize their efficacy. Keywords: aflatoxins, plant resistance, atoxigenic strains, drying instruments, aflatoxin control, food safety, mycotoxins, post-harvest losses

Inhibitory Activities of Alkyl Syringates and Related Compounds on Aflatoxin Production.

Inhibitors of aflatoxin production of aflatoxigenic fungi are useful for preventing aflatoxin contamination in crops. As methyl syringate weakly inhibits aflatoxin production, aflatoxin production inhibitory activities of additional alkyl syringates with alkyl chains from ethyl to octyl were examined. Inhibitory activity toward aflatoxin production of Aspergillus flavus became stronger as the length of the alkyl chains on the esters became longer. Pentyl, hexyl, heptyl, and octyl syringates showed strong activity at 0.05 mM. Heptyl and octyl parabens, and octyl gallate also inhibited aflatoxin production as strongly as octyl syringate. Alkyl parabens and alkyl gallates inhibit the complex II activity of the mitochondrial respiration chain; thus, whether alkyl syringates inhibit complex II activity was examined. Inhibitory activities of alkyl syringates toward complex II also became stronger as the length of the alkyl chains increased. The complex II inhibitory activity of octyl syringate was comparable to that of octyl paraben and octyl gallate. These results suggest that alkyl syringates, alkyl parabens, and alkyl gallates, including commonly used food additives, are useful for aflatoxin control.

Parents’ Practices Associated with Aflatoxin Contamination and Control of Complementary Foods in Central Tanzania

Parents’ practices that are associated with aflatoxin contamination and control in complementary foods were studied in Central Tanzania. A descriptive cross-sectional survey using interviewer-administered structured pretested questionnaire was conducted among 364 randomly selected parents of children aged between 6-23 months, and the mean age (SD) of the respondents was 30 (8.3) years old. The majority 33.2% of the participants harvest their crops in April followed by June which is 26.6%, March which is 21.2% and May which is 19% of all the participants. Most processing activities like dehulling, milling, drying, storage were analysed. The statistical packages SPSS (version 21) computer software packages were used to analyze the data. The results of logistic regression model for dehulling crops confirmed that respondents with less than or with US$ 22.8 as monthly income (OR=0.250, 95% CI: 0.111-0.564) were significantly 0.3 less likely to dehull crops (p<0.05) than respondents who earned more than US$. 22.8. On the other hand, petty trader participants (OR =3.712, 95% CI: 1.420-9.699) were significantly almost 4 times more with a tendency of dehulling the crops (p<0.05) than farmers. The study team recommends that parents should be trained on appropriate methods of drying, storage, and dehulling their crops after harvesting in order to control fungal and aflatoxin infestation. In addition, research on harvesting time, drying, storage, and dehulling practices of crops in Tanzanian is needed.

RNAi-mediated Control of Aflatoxins in Peanut: Method to Analyze Mycotoxin Production and Transgene Expression in the Peanut/&lt;em&gt;Aspergillus&lt;/em&gt; Pathosystem

The Food and Agriculture Organization of the United Nations estimates that 25% of the food crops in the world are contaminated with aflatoxins. That represents 100 million tons of food being destroyed or diverted to non-human consumption each year. Aflatoxins are powerful carcinogens normally accumulated by the fungi Aspergillus flavus and A. parasiticus in cereals, nuts, root crops and other agricultural products. Silencing of five aflatoxin-synthesis genes by RNA interference (RNAi) in peanut plants was used to control aflatoxin accumulation following inoculation with A. flavus. Previously, no method existed to analyze the effectiveness of RNAi in individual peanut transgenic events, as these usually produce few seeds, and traditional methods of large field experiments under aflatoxin-conducive conditions were not an option. In the field, the probability of finding naturally contaminated seeds is often 1/100 to 1/1,000. In addition, aflatoxin contamination is not uniformly distributed. Our method uses few seeds per transgenic event, with small pieces processed for real-time PCR (RT-PCR) or small RNA sequencing, and for analysis of aflatoxin accumulation by ultra-performance liquid chromatography (UPLC). RNAi-expressing peanut lines 288-72 and 288-74, showed up to 100% reduction (p≤0.01) in aflatoxin B1 and B2 compared to the control that accumulated up to 14,000 ng.g-1 of aflatoxin B1 when inoculated with aflatoxigenic A. flavus. As reference, the maximum total of aflatoxins allowable for human consumption in the United States is 20 ng.g-1. This protocol describes the application of RNAi-mediated control of aflatoxins in transgenic peanut seeds and methods for its evaluation. We believe that its application in breeding of peanut and other crops will bring rapid advancement in this important area of science, medicine and human nutrition, and will significantly contribute to the international effort to control aflatoxins, and potentially other mycotoxins in major food crops.

RNAi-mediated Control of Aflatoxins in Peanut: Method to Analyze Mycotoxin Production and Transgene Expression in the Peanut/Aspergillus Pathosystem.

The Food and Agriculture Organization of the United Nations estimates that 25% of the food crops in the world are contaminated with aflatoxins. That represents 100 million tons of food being destroyed or diverted to non-human consumption each year. Aflatoxins are powerful carcinogens normally accumulated by the fungi Aspergillus flavus and A. parasiticus in cereals, nuts, root crops and other agricultural products. Silencing of five aflatoxin-synthesis genes by RNA interference (RNAi) in peanut plants was used to control aflatoxin accumulation following inoculation with A. flavus. Previously, no method existed to analyze the effectiveness of RNAi in individual peanut transgenic events, as these usually produce few seeds, and traditional methods of large field experiments under aflatoxin-conducive conditions were not an option. In the field, the probability of finding naturally contaminated seeds is often 1/100 to 1/1,000. In addition, aflatoxin contamination is not uniformly distributed. Our method uses few seeds per transgenic event, with small pieces processed for real-time PCR (RT-PCR) or small RNA sequencing, and for analysis of aflatoxin accumulation by ultra-performance liquid chromatography (UPLC). RNAi-expressing peanut lines 288-72 and 288-74, showed up to 100% reduction (p≤0.01) in aflatoxin B1 and B2 compared to the control that accumulated up to 14,000 ng.g-1 of aflatoxin B1 when inoculated with aflatoxigenic A. flavus. As reference, the maximum total of aflatoxins allowable for human consumption in the United States is 20 ng.g-1. This protocol describes the application of RNAi-mediated control of aflatoxins in transgenic peanut seeds and methods for its evaluation. We believe that its application in breeding of peanut and other crops will bring rapid advancement in this important area of science, medicine and human nutrition, and will significantly contribute to the international effort to control aflatoxins, and potentially other mycotoxins in major food crops.

Mitigating aflatoxin exposure to improve child growth in Eastern Kenya: study protocol for a randomized controlled trial.

BackgroundWhile the few studies that have looked at the association between stunting and aflatoxin exposure have found surprisingly large effects, the results remain inconclusive due to a lack of randomized controlled studies. This protocol describes a non-blinded, cluster-randomized controlled trial with the specific objective of testing the impact of reduced aflatoxin exposure on (individual) child linear growth.Methods/DesignParticipants were recruited from among households containing women in the last 5 months of pregnancy in 28 maize-growing villages within Meru and Tharaka-Nithi Counties in Kenya. Households in villages assigned to the intervention group are offered rapid testing of their stored maize for the presence of aflatoxin each month; any maize found to contain more than 10 ppb aflatoxin is replaced with an equal amount of maize that contains less than this concentration of the toxin. They are also offered the opportunity to buy maize that has been tested and found to contain less than 10 ppb aflatoxin at local shops. Clusters (villages) were allocated to the intervention group (28 villages containing 687 participating households) or control group (28 villages containing 536 participating households) using a random number generator. The trial, which is funded by United Kingdom (UK) aid from the UK government, the Global Food Security Portal, and the Ministry for Foreign Affairs of Finland, is currently ongoing.DiscussionThis study is the first randomized controlled trial (RCT) to test for a causal impact of aflatoxin exposure on child growth. Whether or not this relationship is found, its results will have implications for the prioritization of aflatoxin control efforts by governments in affected regions, as well as international donors.Trial registrationAmerican Economic Association RCT Registry # 0000105. Initial registration date: 6 November 2013, last updated 30 December 2014.Electronic supplementary materialThe online version of this article (doi:10.1186/s13063-015-1064-8) contains supplementary material, which is available to authorized users.

Community Structure of Aspergillus flavus and A. parasiticus in Major Almond-Producing Areas of California, United States.

Several nut crops, including almond, pistachio, and walnut, can become contaminated with mycotoxins. Of greatest economic significance are aflatoxins, which are mainly produced by members of Aspergillus section Flavi. The distribution of the two sclerotial-size morphotypes of Aspergillus flavus (i.e., S and L strains) and A. parasiticus, the main species responsible for aflatoxin production among section Flavi, was monitored in the soil of almond orchards in California over a 5-year period from 2007 to 2011, excluding 2009. In total, 4,349 Aspergillus isolates were collected from 28 almond orchards located in the northern, central, and southern Central Valley in California. Overall, A. flavus L strain was the most frequent, followed by A. parasiticus and A. flavus S strain. However, variations in the spatial distribution of these three taxa were found between the three regions. Over the 5-year period, higher frequencies of L strain were more often observed in the southern region (79.9 to 95.1%, depending on year) compared with the northern region (21.4 to 47.1%). In the north, A. parasiticus was the most common strain, with frequencies of 28.5 to 61% for the various years. In addition, the frequency of aflatoxin-producing isolates among L strains fluctuated from year to year. A significant increase (P = 0.0001) was observed from 2008 (75% of the isolates produced aflatoxins) to 2007 (59%), and a decrease was observed from 2010 (61%) to 2011 (53%). Aflatoxin-producing L strain isolates were significantly more prevalent than atoxigenic isolates in each region during the 5-year survey, except in 2011 in the north, where more isolates were atoxigenic (56%) than aflatoxin-producing (44%). Our results indicate that the structure of A. flavus and A. parasiticus communities in the soil and the proportion of toxigenic isolates vary across regions and years. Such knowledge may help devise appropriate aflatoxin control strategies, including the use of atoxigenic isolates as biological control agents adapted to the soil environments in each region.

Biological control of aflatoxin is effective and economical in Mississippi field trials

Aflatoxin contamination of corn reduces grain quality and can negatively impact the economic well-being of Mississippi's corn farmers. Mitigation of aflatoxin contamination in corn through pre-harvest field application of non-aflatoxigenic strains of Aspergillus flavus holds promise as an effective management strategy. In this study, field trials were conducted at fourteen sites over three years, in experimental plots and within commercial fields, to evaluate the efficacy of two biocontrol products at 11 or 22 kg ha−1 application rates and at various timings, including two vegetative-stages (V5 and V10), tasseling (VT), a reproductive application made at blister (R2) and two split-applications. The greatest reduction in aflatoxin generally occurred with applications at the earliest developmental stages. Furthermore, six treatments proved more cost-effective across multiple fields and years. These observations support the use of atoxigenic strains of A. flavus to promote the safety and marketability of corn grown in areas susceptible to aflatoxin contamination.

Aflatoxin control in maize by Trametes versicolor.

Aspergillus flavus is a well-known ubiquitous fungus able to contaminate both in pre- and postharvest period different feed and food commodities. During their growth, these fungi can synthesise aflatoxins, secondary metabolites highly hazardous for animal and human health. The requirement of products with low impact on the environment and on human health, able to control aflatoxin production, has increased. In this work the effect of the basidiomycete Trametes versicolor on the aflatoxin production by A. flavus both in vitro and in maize, was investigated. The goal was to propose an environmental loyal tool for a significant control of aflatoxin production, in order to obtain feedstuffs and feed with a high standard of quality and safety to enhance the wellbeing of dairy cows. The presence of T. versicolor, grown on sugar beet pulp, inhibited the production of aflatoxin B1 in maize by A. flavus. Furthermore, treatment of contaminated maize with culture filtrates of T. versicolor containing ligninolytic enzymes, showed a significant reduction of the content of aflatoxin B1.

Farmer perceptions of aflatoxin management strategies in lower Eastern Kenya

Aflatoxins pose a serious problem to maize grains, rendering them unsuitable for human and livestock consumption. This study assessed farmers’ attitude toward, and knowledge and perceptions of, the nature, causes and use of biological technologies in aflatoxin control in Embu, Machakos, Makueni and Kitui Counties of Kenya. A total of 480 households were randomly selected from two districts per county using a GPS system. The tools used in the survey included pre-tested semi-structured questionnaires, focused group discussions, key informant interviews and secondary data exploration. The information collected included crop production constraints, existing aflatoxin management strategies, farmers’ experiences with use of atoxigenic strains of Aspergillus and other household farm production activities. The study found that farmers were largely aware of the aflatoxin problem and its negative effects, factors that contribute to its occurrence, and available options to address it. The main pre-harvest aflatoxin control technologies used were crop rotation, irrigation, use of resistant crop varieties and pest control. The main post-harvest aflatoxin control technologies used were proper storage, drying, sorting, and use of pesticides to manage pests. Modern aflatoxin control technologies such as ammoniation and use of hydrogels were not in use mainly due to unavailability, high cost and safety concerns (especially regarding grains meant for human consumption). Effective management of aflatoxin contamination requires that farmers are aware of the problem and use a combination of strategies targeting the crop before and after harvesting, and understanding farmers’ perceptions and knowledge is a vital step to finding a local solution for aflatoxin management. The study recommends the strengthening of existing public extension service system to enable it deliver up-to-date information, through a variety of channels, on aflatoxin and its management to farmers in a more effective and timely manner. Key words: Aflatoxin, Aflasafe, farmers, perceptions, Kenya.

Study on the Key Environmental Factors &lt;i&gt;Aspergillus flavus&lt;/i&gt; Growth and Aflatoxin Production

Single factor and Plackett-Burman design were used to determine the key environmental factors for Aspergillus flavus growth and aflatoxin production. The results showed that the key factors were culture time, moisture content. So Controling of moisture was crucial for Aflatoxin contamination prevention and control.

Aflatoxin and Dairy Products: A Review

Aflatoxins are toxic secondary metabolites produced by Aspergillus fungus. Continuous monitoring of aflatoxins in the entire food chain, from the field to milk and milk products is very important, as it grows readily in favourable condition. It causes major economic losses and adversely affects the animal and human health including infants and children. Public should be aware towards ill effects of aflatoxin. In all countries, there should be regulation of AFM 1 in milk and its products to protect consumers specially children. This toxin cannot be inactivated by thermal processing used in dairy industry. HACCP is considered as an effective method for control of aflatoxins in food supply. Feed given to cattle should be free from aflatoxin. Many environmentally sustainable pre or post-harvest methods like proper irrigation, genetically resistant crop strains and bio-pesticide management can be applied to reduce the contamination of aflatoxins in food chain.

Potential of essential oils for protection of grains contaminated by aflatoxin produced by Aspergillus flavus.

Aflatoxin B1 (AFB1) is a highly toxic and carcinogenic metabolite produced by Aspergillus species on food and agricultural commodities. Inhibitory effects of essential oils of Ageratum conyzoides (mentrasto) and Origanum vulgare (oregano) on the mycelial growth and aflatoxin B1 production by Aspergillus flavus have been studied previously in culture medium. The aim of this study was to evaluate aflatoxin B1 production by Aspergillus flavus in real food systems (corn and soybean) treated with Ageratum conyzoides (mentrasto) and Origanum vulgare (oregano) essential oils. Samples with 60 g of the grains were treated with different volumes of essential oils, 200, 100, 50, and 10 μL for oregano and 50, 30, 15, and 10 μL for mentrasto. Fungal growth was evaluated by disk diffusion method. Aflatoxin B1 production was evaluated inoculating suspensions of A. flavus containing 1.3 × 105 spores/mL in 60 g of grains (corn and soybeans) after adjusting the water activity at 0.94. Aflatoxin was quantified by photodensitometry. Fungal growth and aflatoxin production were inhibited by essential oils, but the mentrasto oil was more effective in soybeans than that of oregano. On the other hand, in corn samples, the oregano essential oil was more effective than that of mentrasto. Chemical compositions of the essential oils were also investigated. The GC/MS oils analysis showed that the main component of mentrasto essential oil is precocene I and of the main component of oregano essential oil is 4-terpineol. The results indicate that both essential oils can become an alternative for the control of aflatoxins in corn and soybeans.

Molecular characterization of atoxigenic Aspergillus flavus isolates collected in China

Aspergillus flavus strains were isolated from peanut fields of Liaoning, Shandong, Hubei and Guangdong Provinces in China, and identified through phenotypic and molecular approaches. Of the 323 A. flavus strains isolated, 76 strains did not produce aflatoxins detectable by UPLC. The incidence of atoxigenic A. flavus strains decreased with increase in temperature and increased with increase in latitude in different geographical locations. Amplification of all the aflatoxin genes in the aflatoxin gene cluster in the atoxigenic isolates showed that there were 25 deletion patterns (A-Y), with 22 deletion patterns identified for the first time. Most of the atoxigenic A. flavus isolates with gene deletions (97%) had deletions in at least one of the four genes (aflT, nor-1, aflR, and hypB), indicating that these four genes could be targeted for rapid identification of atoxigenic strains. The atoxigenic isolates with gene deletions, especially the isolates with large deletions, are potential candidates for aflatoxin control.

Inhibitory effects of respiration inhibitors on aflatoxin production.

Aflatoxin production inhibitors, which do not inhibit the growth of aflatoxigenic fungi, may be used to control aflatoxin without incurring a rapid spread of resistant strains. A respiration inhibitor that inhibits aflatoxin production was identified during a screening process for natural, aflatoxin-production inhibitors. This prompted us to evaluate respiration inhibitors as potential aflatoxin control agents. The inhibitory activities of four natural inhibitors, seven synthetic miticides, and nine synthetic fungicides were evaluated on aflatoxin production in Aspergillus parasiticus. All of the natural inhibitors (rotenone, siccanin, aptenin A5, and antimycin A) inhibited fungal aflatoxin production with IC50 values around 10 µM. Among the synthetic miticides, pyridaben, fluacrypyrim, and tolfenpyrad exhibited strong inhibitory activities with IC50 values less than 0.2 µM, whereas cyflumetofen did not show significant inhibitory activity. Of the synthetic fungicides, boscalid, pyribencarb, azoxystrobin, pyraclostrobin, and kresoxim-methyl demonstrated strong inhibitory activities, with IC50 values less than 0.5 µM. Fungal growth was not significantly affected by any of the inhibitors tested at concentrations used. There was no correlation observed between the targets of respiration inhibitors (complexes I, II, and III) and their IC50 values for aflatoxin-production inhibitory activity. This study suggests that respiration inhibitors, including commonly used pesticides, are useful for aflatoxin control.

Kıbrıs Geleneksel Peynir (Hellim) Örneklerinde Aflatoksin M1(AFM1)Oluşumunun Belirlenmesi: Kesitsel Bir Çalışma

Summary This Cross-Sectional present study was conducted to determine the occurence of aflatoxin M1 (AFM1) in samples of Turkish Republic of North Cyprus (TRNC) traditional cheese (halloumi). In the current study a total of 128 halloumi cheese samples including 36 industrial made and 92 home made were selected by using cluster sampling method and analyzed for AFM1 with the competitive ELISA. The percent of AFM1 contamination in halloumi cheese was found to be low, since 28.8% (0-16.66 ng/kg) of the samples were positive in industrial made and 21.7% (0-4.63 ng/kg) in home made. The mean, standard error of mean (SEM), 95% Confidence Interval (95% CI) values of aflatoxin M1 in halloumi cheese with industrial and home made samples were 0.84±0.24, (95% CI :0.35-1.35) and 1.28±0.32, (95% CI: 0.63-1.93) respectively. Both means were not significantly difference (P=0.422) and found very low from the limits of European Commission (EC) (250 ng/kg) and Turkish Food Codex (TFC) (500 ng/kg) (P<0.001). In order to prevent from introduction of aflatoxin M1 into cheese industry cycle, hygienic conditions, appropriate storage and control of livestock feed at all stages of planting and requires system that makes aflatoxin control are necessary.

An Assessment on Aflatoxin Control in Pistachio‐Processing Units from Raw Material Reception to Packaging Based on ISO22000:2005 Model

AbstractBecause of the importance and usage of pistachios, it is necessary to improve their agricultural situation, particularly by establishing good processing equipment and packaging units near pistachio farms. It is possible to supply a high‐quality product in large quantities for foreign and domestic markets. In this research, International Organization for Standardization 22000:2005 requirements were implemented in a pistachio‐processing unit. Aflatoxin B1 content was measured by high‐performance liquid chromatography before and after washing, immersing pool, wet sorting, drying, temporary storage and dry sorting. After analysis of measured data, it was found that the most reducing stage in aflatoxin content was wet sorting. Washing stage had no significant effect on aflatoxin B1 reduction. It is concluded that operational prerequisite program and critical control point have no differences important to hazard control, but they are different from a type of control measure aspect.Practical ApplicationIn this study, the authors employed food safety management system based on International Organization for Standardization (ISO) 22000:2005 model to prepare safe pistachios with less aflatoxin content. All process units (pistachio reception, de‐hulling, washing, immersing pool, dehumidifier, goo separator, wet sorting, drying, temporary storage, dry sorting, packaging) evaluated upon ISO22000:2005 requirements. Then the stages that had more significant effect on aflatoxin B1 content identified. The quantification of aflatoxin B1 in accordance with standard methods was also explained in the paper. The authors also investigated operational prerequisite programs and critical control points in hazard control.

Environmental Hygiene, Food Safety and Growth in less than Five Year Old Children in Zimbabwe and Ethiopia

Childhood stunting remains a significant public health challenge with adverse developmental and health outcomes in life and over generations. This study explored novel causes of poor growth in infants and young children. Using in‐depth observation methods, soil and chicken feces ingestion were identified as key pathways for fecal‐oral transmission of bacteria in infants and young children in rural Zimbabwe. In a cross‐sectional survey, poor environmental hygiene was associated with linear growth faltering (p =0.032), independent of socioeconomic status, infant feeding and recent morbidity, in 24–59 month old Ethiopian children. Significant aflatoxin exposure in mothers was associated with severe stunting in 6 to 59 months old Zimbabwean children in a dose response manner (p=0.035). We identified poor environmental hygiene and aflatoxin exposure as two novel causes of stunting in African children. Existing WASH interventions are not sufficient to protect infants and young children from fecal bacteria ingestion through soil and poultry feces. In addition, existing hygiene interventions do not directly address improving household environmental hygiene. Our findings raise the need for low cost strategies for aflatoxin control and a holistic approach in designing context‐ and age‐specific hygiene interventions to prevent childhood stunting.Grant Funding Source: Cornell International Institute for Food, Agriculture and Development (CIIFAD); the Department for International Development (DFID), UK and Borlaug Leadership in Agriculture Enhancement Program (LEAP), UC, Davis, USA; Bill &amp; Melinda Gates Foundation.

Pengendalian aflatoksin pada pascapanen jagung melalui penerapan HACCP

In order to support the self-sufficiency of corn, aflatoxin control system in the supply chains have to be very important attention. Good handling and drying will make longer storage life of corn. Also scheduling in cultivation system, handling, storage and distribution are very important in order to fulfill the need of corn along the year either for food or feed. Due to its problems, the research is conducted on Creating HACCP guidance on handling and storage with follow the Guidance of Compilation Plan HACCP (BSN-Pedoman 1004-1999). Identification of critical point dealing with contamination on corn from harvesting until storage has been conducted. The processing step which is critical on post harvest of corn such as harvesting moment (CCP1), grading (CCP 2), drying moment (CCP3), grading of quality (CCP 4) and storage moment (CCP 5). On the steps above, it needed extra control from the farmer and the seller for controlling the quality of corn in order to avoid the physical contaminant, chemist and microbiology such as fungi and insect which can produce aflatoxins, so it can reduce the weight of corn that is very harming.

Prevention of aflatoxin contamination by a soil bacterium of Stenotrophomonas sp. that produces aflatoxin production inhibitors

A soil bacterium, designated strain no. 27, was found to produce aflatoxin-production inhibitors. The strain was identified as a species of the genus Stenotrophomonas, and was found to be closely related to Stenotrophomonas rhizophila. Two diketopiperazines, cyclo(L-Ala-L-Pro) and cyclo(L-Val-L-Pro), were isolated from the bacterial culture filtrate as main active components. These compounds inhibited aflatoxin production of Aspergillus parasiticus and Aspergillus flavus in liquid medium at concentrations of several hundred µM without affecting fungal growth. Both inhibitors inhibited production of norsorolinic acid, a biosynthetic intermediate involved in an early step of the aflatoxin biosynthetic pathway, and reduced the mRNA level of aflR, which is a gene encoding a key regulatory protein necessary for the expression of aflatoxin-biosynthetic enzymes. These results indicated that the inhibitors targets are present in early regulatory steps leading to AflR expression. Co-culture of strain no. 27 with aflatoxigenic fungi in liquid medium effectively suppressed aflatoxin production of the fungus without affecting fungal growth. Furthermore, application of the bacterial cells to peanuts in laboratory experiments and at a farmer's warehouse in Thailand by dipping peanuts in the bacterial cell suspension strongly inhibited aflatoxin accumulation. The inhibitory effect was dependent on bacterial cell numbers. These results indicated that strain no. 27 may be a practically effective biocontrol agent for aflatoxin control.