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It is not known which aflatoxigenic species are present in Zambia. Therefore, soil samples were collected during May to June 2012, at the end of the growing season in Eastern Province, from 399 farmers' fields that had been planted to groundnut ( Arachis hypogaea L.) in Nyimba, Petauke, Mambwe, and Chipata Districts. Population densities of Aspergillus spp. were estimated by plating 10−3 soil dilutions on modified dichloran rose Bengal (MDRB) media. To test for toxigenicity, colonies were randomly selected from MDRB dilution plates, single-spored, transferred to vials with yeast extract sucrose (YES) liquid media, and grown for a week at room temperature. Agra Strip ® lateral flow cards were then used to test the filtered extracts, from the YES cultures, for total aflatoxin at 4 and 20 parts per billion (ppb). We identified Aspergillus flavus (small and large sclerotia strains), A. parasiticus, A. niger, A. nomius, A. oryzae, A. tamarii, and A. terreus. 100% of the S-strain A. flavus isolates produced aflatoxin at 4 and 20 ppb, whereas 86% and 56% of the L-strain A. flavus isolates produced aflatoxins at 4 and 20 ppb, respectively. All the A. nomius isolates produced aflatoxins at 4 and 20 ppb. 79% and 64% the A. parasiticus isolates produced aflatoxins at 4 and 20 ppb. To our knowledge, this is the first peer reviewed report from Zambia documenting the population densities of A. flavus across different agroecologies. In addition, it is also the first report on the identification of different Aspergillus spp., such as A. nomius, A. terreus, A. oryzae, and A. tamarii from Zambia. This information, taken together with cropping practices, soil characteristics, agroecological and climatic data, can form a basis for developing holistic pre-harvest aflatoxin mitigation strategies. [ABSTRACT FROM AUTHOR]

Trichoderma species offer considerable potential for controlling aflatoxin contamination in groundnut and other crops. Initial classification of 48 Trichoderma isolates, derived from four different groundnut cultivation sites in India was based on alignment of 28S rDNA sequences to GenBank sequences of ex-type strains. This was found to be substantially more reliable than our routine morphological characterization, but did not provide a comprehensive diagnostic solution, as unique single nucleotide polymorphism (SNP) haplotypes could not be identified for all species. However, all the Trichoderma isolates could be readily distinguished by amplified fragment length polymorphism (AFLP) analysis, based on six primer pair combinations, which generated 234 polymorphic bands. In addition, individual AFLP bands were identified which differentiate closely related species. Similarly, AFLP bands were identified that correlated with different types of antagonism to Aspergillus flavus. The implications of these results for the development of simple polymerase chain reaction (PCR)-based diagnostic assays for antagonistic isolates of Trichoderma is discussed. [ABSTRACT FROM AUTHOR]

Sterility mosaic disease (SMD), an important biotic constraint on pigeonpea (Cajanus cajan) in the Indian subcontinent, is caused by Pigeonpea sterility mosaic virus (PPSMV) transmitted by the eriophyid mite,Aceria cajani.Distinct PPSMV isolates occur in different geographical regions and broad-based resistance to all these isolates is scarce in cultivated pigeonpea germplasm. Wild relatives of pigeonpea, which are known to possess resistance to several pests and diseases, were evaluated for broad-based SMD resistance. One hundred and fifteen wildCajanusaccessions from six species (C. albicans, C. platycarpus, C. cajanifolius, C. lineatus, C. scarabaeoidesandC. sericeus) were evaluated against three PPSMV isolates prevailing in peninsular India. Evaluations were done under greenhouse conditions in endemic locations of each isolate through mite-mediated virus inoculation. Fifteen accessions showed resistance to all three isolates: ICP 15614, 15615, 15626, 15684, 15688, 15700, 15701, 15725, 15734, 15736, 15737, 15740, 15924, 15925 and 15926. Most of the wild accessions did not support mite multiplication. The majority of the accessions resistant to PPSMV following inoculations with viruliferous mites were susceptible by graft inoculation, suggesting that vector resistance is conferring resistance to infection with PPSMV. The 15 accessions identified as being resistant to infection to all three virus isolates tested are cross compatible with pigeonpea by traditional breeding. They are therefore useful for exploitation in breeding programmes to increase both the level of SMD resistance and to diversify its genetic base in the cultivated pigeonpea gene pool. [ABSTRACT FROM AUTHOR]

The article discusses the effect of aflatoxin B1 (AFB1) contaminated food on human beings and development of a technique for its detection. As stated, biomarkers in blood, urine, and tissues have been used to monitor the AFB1exposure. It further informs that indirect competitive enzyme linked immunosorbent assay (IC-ELISA) has the potential for epidemiological studies in order to minimize aflatoxin contamination in the AFB1 exposed individuals.

Highlights: [•] Groundnut germplasm was assessed across varying drought intensities. [•] Drought intensity increased aflatoxin concentration in seeds. [•] Across trials aflatoxin concentration and grain yield reduction were highly correlated. [•] However within trial, aflatoxin and grain yield reduction were unrelated, showing no direct relationship between drought tolerance and resistance to aflatoxin contamination. [•] Mechanisms of drought tolerance and aflatoxin contamination are likely not common. [ABSTRACT FROM AUTHOR]

Abstract: Soil moisture and soil temperature affect pre-harvest infection with Aspergillus flavus and production of aflatoxin. The objectives of our field research in Niger, West Africa, were to: (i) examine the effects of sowing date and irrigation treatments on pod yield, infection with A. flavus and aflatoxin concentration; and (ii) to quantify relations between infection, aflatoxin concentration and soil moisture stress. Seed of an aflatoxin susceptible peanut cv. JL24 was sown at two to four different sowing dates under four irrigation treatments (rainfed and irrigation at 7, 14 and 21 days intervals) between 1991 and 1994, giving 40 different ‘environments’. Average air and soil temperatures of 28–34°C were favourable for aflatoxin contamination. CROPGRO-peanut model was used to simulate the occurrence of moisture stress. The model was able to simulate yields of peanut well over the 40 environments (r 2 =0.67). In general, early sowing produced greater pod yields, as well as less infection and lower aflatoxin concentration. There were negative linear relations between infection (r 2 =0.62) and the average simulated fraction of extractable soil water (FESW) between flowering and harvest, and between aflatoxin concentration (r 2 =0.54) and FESW in the last 25 days of pod-filling. This field study confirms that infection and aflatoxin concentration in peanut can be related to the occurrence of soil moisture stress during pod-filling when soil temperatures are near optimal for A. flavus. These relations could form the basis of a decision-support system to predict the risk of aflatoxin contamination in peanuts in similar environments. [Copyright &y& Elsevier]

Stem necrosis disease caused by Tobacco streak virus (TSV), first recognized in 2000, has emerged as a potential threat to peanut (Arachis hypogaea) in southern states of India. The virus induces severe necrosis of shoots leading to death of the plant, and plants that survive are malformed, with severe reduction in pod yield. All the currently grown peanut cultivars in India are highly susceptible to the virus. Therefore, wild relatives of peanut were evaluated to identify potential sources of resistance to TSV infection. In all, 56 germplasm accessions from 20 wild Arachis spp. in four sections (Arachis, Erectoides, Procumbenie, and Rhizomatosae), along with susceptible peanut cultivars (JL 24 and K 1375), were evaluated for resistance to TSV under greenhouse conditions using mechanical sap inoculations. Systemic virus infection, determined by enzyme-linked immunosorbent assay (ELISA), in the test accessions ranged between 0 and 100%. Twenty-four accessions in section Arachis that had 0 to 35% systemically infected plants were retested, and systemic infection was not detected in eight of these accessions in repeated trials in the greenhouse. These are International Crops Research Institute for the Semi-Arid Tropics groundnut (ICO) accession nos. 8139, 8195, 8200, 8203, 8205, and 11550 belonging to A. duranensis; ICG 8144 belonging to A. villosa; and ICG 13210 belonging to A. stenospernia. Even though the resistant accessions had 0 to 100% ISV infection in inoculated leaves, ISV was not detected in the subsequently emerged leaves. This is the first report of TSV resistance in Arachis spp. The eight TSV resistant accessions are cross compatible with A. hypogaea for utilization in breeding for stem necrosis disease resistance. [ABSTRACT FROM AUTHOR]

In Zambia, groundnut products (milled groundnut powder, groundnut kernels) are mostly sold in under-regulated markets. Coupled with the lack of quality enforcement in such markets, consumers may be at risk to aflatoxin exposure. However, the level of aflatoxin contamination in these products is not known. Compared to groundnut kernels, milled groundnut powder obscures visual indicators of aflatoxin contamination in groundnuts such as moldiness, discoloration, insect damage or kernel damage. A survey was therefore conducted from 2012 to 2014, to estimate and compare aflatoxin levels in these products (n = 202), purchased from markets in important groundnut growing districts and in urban areas. Samples of whole groundnut kernels (n = 163) and milled groundnut powder (n = 39) were analysed for aflatoxin B 1 (AFB 1 ) by competitive enzyme-linked immunosorbent assay (cELISA). Results showed substantial AFB 1 contamination levels in both types of groundnut products with maximum AFB 1 levels of 11,100 μg/kg (groundnut kernels) and 3000 μg/kg (milled groundnut powder). However, paired t test analysis showed that AFB 1 contamination levels in milled groundnut powder were not always significantly higher (P > 0.05) than those in groundnut kernels. Even for products from the same vendor, AFB 1 levels were not consistently higher in milled groundnut powder than in whole groundnut kernels. This suggests that vendors do not systematically sort out whole groundnut kernels of visually poor quality for milling. However, the overall contamination levels of groundnut products with AFB 1 were found to be alarmingly high in all years and locations. Therefore, solutions are needed to reduce aflatoxin levels in such under-regulated markets.

A 3-year comprehensive analysis of aflatoxin contamination in peanut butter was conducted in Zambia, sub-Saharan Africa. The study analyzed 954 containers of 24 local and imported peanut butter brands collected from shops in Chipata, Mambwe, Petauke, Katete, and Nyimba districts and also in Lusaka from 2012 to 2014. For analysis, a sample included six containers of a single brand, from the same processing batch number and the same shop. Each container was quantitatively analyzed for aflatoxin B1 (AFB1) in six replicates by using competitive enzyme-linked immunosorbent assay; thus, aflatoxin contamination level of a given sample was derived from an average of 36 test values. Results showed that 73% of the brands tested in 2012 were contaminated with AFB1 levels >20 μg/kg and ranged up to 130 μg/kg. In 2013, 80% of the brands were contaminated with AFB1 levels >20 μg/kg and ranged up to 10,740 μg/kg. Compared with brand data from 2012 and 2013, fewer brands in 2014, i.e., 53%, had aflatoxin B1 levels >20 μg/kg and ranged up to 1,000 μg/kg. Of the eight brands tested repeatedly across the 3-year period, none consistently averaged ≤20 μg/kg. Our survey clearly demonstrates the regular occurrence of high levels of AF B1 in peanut butter in Zambia. Considering that some of the brands tested originated from neighboring countries such as Malawi, Zimbabwe, and South Africa, the current findings provide a sub-Saharan regional perspective regarding the safety of peanut butter.

Aflatoxins are toxic, carcinogenic, mutagenic, teratogenic and immunosuppressive byproducts of Aspergillus spp. that contaminate a wide range of crops such as maize, peanut, and cotton. Aflatoxin not only affects crop production but renders the produce unfit for consumption and harmful to human and livestock health, with stringent threshold limits of acceptability. In many crops, breeding for resistance is not a reliable option because of the limited availability of genotypes with durable resistance to Aspergillus. Understanding the fungal/crop/environment interactions involved in aflatoxin contamination is therefore essential in designing measures for its prevention and control. For a sustainable solution to aflatoxin contamination, research must be focused on identifying and improving knowledge of host-plant resistance factors to aflatoxin accumulation. Current advances in genetic transformation, proteomics, RNAi technology, and marker-assisted selection offer great potential in minimizing pre-harvest aflatoxin contamination in cultivated crop species. Moreover, developing effective phenotyping strategies for transgenic as well as precision breeding of resistance genes into commercial varieties is critical. While appropriate storage practices can generally minimize post-harvest aflatoxin contamination in crops, the use of biotechnology to interrupt the probability of pre-harvest infection and contamination has the potential to provide sustainable solution., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Recombinant genes conferring resistance to antibiotics or herbicides are widely used as selectable markers in plant transformation for selecting the primary transgenic events. However, these become redundant once the transgenic plants have been developed and identified. Although, there is no evidence that the selectable marker genes are unsafe for consumers and the environment, it would be desirable if the marker genes can be eliminated from the final transgenic events. The availability of efficient transformation methods can enable the possibility of developing transgenic events that are devoid of the marker gene/s upfront. Taking advantage of the high and consistent transformation potential of peanut, we report a technique for developing its transgenics without the use of any selectable marker gene. Marker-free binary vectors harboring either the phytoene synthase gene from maize (Zmpsy1) or the chitinase gene from rice (Rchit) were constructed and used for Agrobacterium tumefaciens-mediated transformation of peanut. The putative transgenic events growing in vitro were initially identified by PCR and further confirmed for gene integration and expression by dot blots assays, Southern blots, and RT-PCR where they showed a transformation frequency of over 75%. This system is simple, efficient, rapid, and does not require the complex segregation steps and analysis for selection of the transgenic events. This approach for generation of marker-free transgenic plants minimizes the risk of introducing unwanted genetic changes, allows stacking of multiple genes and can be applicable to other plant species that have high shoot regeneration efficiencies.

High-titer rabbit polyclonal antibodies to aflatoxin M(1) (AFM1) were produced by utilizing AFM1-bovine serum albumin (BSA) conjugate as an immunogen. An indirect competitive enzyme-linked immunosorbent assay was standardized for estimating AFM1 in milk and milk products. To avoid the influence of interfering substances present in the milk samples, it was necessary to prepare AFM1 standards in methanol extracts of certified reference material (CRM) not containing detectable AFM1 (< 0.05 ng/g). The reliability of the procedure was assessed by using CRM with AFM1 concentrations of < 0.5 and 0.76 ng/g. Also, assays of milk samples mixed with AFM1 ranging in concentration between 0.5 and 50 ng/L gave recoveries of > 93%. The relative cross-reactivity with aflatoxins (AF) and ochratoxin A, assessed as the amount of AFM1 necessary to cause 50% inhibition of binding, was 5% for AFB1 and much less for AFB2, AFG1, and AFG2; there was no reaction with ochratoxin A. AFM1 contamination was measured in retail milk and milk products collected from rural and periurban areas in Andhra Pradesh, India. Of 280 milk samples tested, 146 were found to contain < 0.5 ng/mL of AFM1; in 80 samples it varied from 0.6 to 15 ng/mL, in 42 samples from 16 to 30 ng/mL, and in 12 samples from 31 to 48 ng/mL. Most of the milk samples that contained high AFM1 concentrations were obtained from periurban locations. The results revealed a significant exposure of humans to AFM1 levels in India and thus highlight the need for awareness of risk among milk producers and consumers.

Foliar diseases caused by Cercospora arachidicola, Cercosporium personatum, and Puccinia arachidis are major constraints to peanut production in the world. Fungicides are among the most efficient available control methods. Field trials were conducted in 1991 and 1992 in Benin and Niger, West Africa, to evaluate the cost effectiveness of fungicide application timings and frequencies on four peanut cultivars. A combination of four timings (40, 55, 70, and 85 days after sowing) was scheduled. Early (causal organism, C. arachidicola) and late (caused by C. personatum) leaf spot were prevalent in both years, but late leaf spot was the more economically important disease as shown by high values of area under the disease progress curve. Application of fungicide reduced late leaf spot incidence and increased pod yield. Pod yield responded to an interaction of number and timing of fungicide applications. With appropriate timing two or three fungicide applications were enough to significantly increase pod yield. Properly timed fungicide sprays can result in substantial monetary gains for peanut farmers in West Africa.