Controlling mycotoxins in maize

One of the most important socioeconomic changes in the savannas of Africa has been the increasing production of maize in areas that previously were planted to millets and sorghum. The greater use of higher-yielding crop cultivars, increases in the availability and use of pesticides, and deregulation of the market for cereals also have changed these areas dramatically. Maize has essentially become a cash crop. Much of the increase in maize production has occurred in areas at significant risk of attack by pests of stored maize, e.g., fungi that produce mycotoxins.

Aflatoxin contamination is widespread in Africa: in Benin and Togo, aflatoxin levels in maize averaged five times the safe limit of 20 ng/g in up to 50% of the household grain stores surveyed (Egal et al., 2005; Gong et al., 2002). As a result, people, especially children (Gong et al., 2002), are being exposed to high levels of mycotoxins, often in mixtures, and the consequences have been largely ignored. For example, 99% of fully weaned children had ~2-fold higher aflatoxin-albumin adduct levels in their blood than do those receiving a mixture of breast milk and solid foods (Gong et al., 2003, 2004). Surveys also indicate that Fusarium infection is prevalent in field and stored maize at many African locations. Fusarium spp. are found in all agroecological zones of Benin, but their prevalence is higher in the South than the North. The incidence of Fusarium infection is higher in the field than in storage (K. Hell, unpublished). Fusarium infection usually is reduced during storage. The most common species found were F. verticillioides and F. proliferatum. Fu-monisins were found in the maize samples with levels often exceeding the limit of 4 ^g/g recommended by the U.S. Food and Drug Administration, especially in villages in southern Benin: Ye (12 ^g/g), Lainta (7 ^g/g), Adjohoun (6.7 ^g/g) and Kpome (4.7 ^g/g).

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Test cross to determine combining ability

Synthetic, Hybrids

Figure 2. A comprehensive breeding strategy for developing biotic stress resistant maize germplasm. RRS = reciprocal recurrent selection.

Similarly, nearly 35% of maize kernel samples collected from several villages during the rainy and post-rainy seasons of 2004-2005 in Andhra Pradesh (India) contained 1 to 20 ng/g af-latoxin (F. Waliyar, unpublished). Six percent of the rainy season samples and 7.6% of the post-rainy season samples contained > 20 ng/g aflatoxins soon after harvest, whereas 20% of the rainy season crop stored for up to 4 months had > 20 ng/g aflatoxins. This survey suggests that even though toxin levels can be low at the time of harvest, they probably increase during grain storage.

Many insect species, drought and other environment factors, e.g., nutrient stress, or pathogens are positively correlated with aflatoxin content in stored maize. These interactions between biotic and abiotic stresses not only reduce yield but also enable saprophytic fungi, such as Aspergillus flavus, to colonize the grain of stressed maize plants. In this regard, researchers at CIMMYT have been working to combine biotic and abiotic stress resistance, and to identify stress-tolerant lines or hybrids that have a reduced incidence of A. flavus. Line recycling, i.e., targeting crosses based on known traits in the parental lines, has used sources such as 'La Posta Sequia' for drought tolerance, lines extracted from the 'Multiple Insect Resistance Tropical' (MIRT) population for stem borer and armyworm resistance, stunt resistant populations (mainly 'P73' but also 'P76' and 'P79') and lines resistant to foliar diseases and ear rots (D. Bergvinson and D. Jeffers, unpublished). This broader approach to reducing the mycotoxin load has been successful and has led to efforts to breed source populations and synthetics against the maize weevil and larger grain borer, insects that serve as vectors for fungi and that breach the plant's external integrity allowing fungi to enter and colonize the plant. A comprehensive breeding strategy (Fig. 2) for developing biotic stress resistant maize germplasm has resulted in genetically enhanced lines with abiotic or biotic resis-

Figure 3. Aflatoxin accumulation in selected IITA maize inbred lines (i|) tested with a

kernel screening assay. A susceptible hybrid [after Brown et al. (2001)].

from the United States had 5300 ng/g tance to storage pests and aflatoxigenic fungi. Valuable sources for resistance to aflatoxin accumulation have been identified in several elite CIMMYT maize lines (Jeffers et al., 2005). These and other maize germplasm sources developed by IITA (Fig. 3) are being incorporated into both tropical and southern United States maize breeding materials (Brown et al, 2001).

Aflatoxin screening in maize kernels needs to be simple and cost-effective. Fluorescent screening uses a black light assay to observe fluorescence from kojic acid, a secondary metabolite observed in colonized grain. Under black light, uninfected kernels are opaque and those infected are bright. This mass preliminary screening of breeding materials, allows selection of parental sources for further population improvement of maize.

Competitive exclusion (one strain competing to exclude another) is a relatively new, but very promising, biological control strategy for aflatoxin management in Africa. This control option leads to the best-adapted fungal strain being dominant in a given environment, and is a promising strategy for replacing toxigenic strains of Aspergillus with atox-igenic forms of same fungus. Several strains of A. flavus have been isolated and are being tested in Nigeria and Benin, with the goal of using atoxigenic strains to reduce aflatoxin contamination. Systematic knowledge of pre- and postharvest practices can lead to complementary management of aflatoxins through cultural and storage practices (Turner et al., 2005). For example, lodged maize plants, drought predisposition, high grain moisture and grain damage at harvest all increase the risk of aflatoxin production in storage. Thus, farmers need to follow good management practices at harvest time, and in drying, including using an appropriate storage structure and controlling insects. Management options include preventing rain exposure of harvested cobs, storing maize in non-plastic bags, and sorting out kernels with insect damage and/or discoloration. Likewise, drying on black plastic sheets or cemented dry areas can reduce moisture content to safe levels after ~5 days, while drying maize cobs on the ground requires a minimum of 10 days.

Traditional maize processing also may help reduce mycotoxin levels; e.g., aflatoxin levels were reduced by 99% in Benin during the preparation of fermented mawe, and by 79% and 43% when preparing fermented ogi and owo (or "pate de mais"), respectively. Roasting and treatment with alkali also reduces the level of aflatoxin present, whereas boiling and soaking of maize grain in lime-water can eliminate or greatly reduce the levels of aflatoxin in the final product. Last but not least, selective removal or isolation of contaminated portions of the food commodity remains the most widely used physical method for aflatoxin decontamination. Awareness campaigns are needed to sensitize the population to the risks posed by aflatoxins and to popularize management options to minimize its effects. For example, due to civil society public action, more than 10 million people in Benin, Togo and Ghana became aware of the dangers posed by aflatoxin-contaminated feeds and foods.

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