Germán Barros

Sexuality Generates Diversity in the Aflatoxin Gene Cluster: Evidence on a Global Scale

Aflatoxins are produced by Aspergillus flavus and A. parasiticus in oil-rich seed and grain crops and are a serious problem in agriculture, with aflatoxin B1 being the most carcinogenic natural compound known. Sexual reproduction in these species occurs between individuals belonging to different vegetative compatibility groups (VCGs). We examined natural genetic variation in 758 isolates of A. flavus, A. parasiticus and A. minisclerotigenes sampled from single peanut fields in the United States (Georgia), Africa (Benin), Argentina (Córdoba), Australia (Queensland) and India (Karnataka). Analysis of DNA sequence variation across multiple intergenic regions in the aflatoxin gene clusters of A. flavus, A. parasiticus and A. minisclerotigenes revealed significant linkage disequilibrium (LD) organized into distinct blocks that are conserved across different localities, suggesting that genetic recombination is nonrandom and a global occurrence. To assess the contributions of asexual and sexual reproduction to fixation and maintenance of toxin chemotype diversity in populations from each locality/species, we tested the null hypothesis of an equal number of MAT1-1 and MAT1-2 mating-type individuals, which is indicative of a sexually recombining population. All samples were clone-corrected using multi-locus sequence typing which associates closely with VCG. For both A. flavus and A. parasiticus, when the proportions of MAT1-1 and MAT1-2 were significantly different, there was more extensive LD in the aflatoxin cluster and populations were fixed for specific toxin chemotype classes, either the non-aflatoxigenic class in A. flavus or the B1-dominant and G1-dominant classes in A. parasiticus. A mating type ratio close to 1∶1 in A. flavus, A. parasiticus and A. minisclerotigenes was associated with higher recombination rates in the aflatoxin cluster and less pronounced chemotype differences in populations. This work shows that the reproductive nature of the population (more sexual versus more asexual) is predictive of aflatoxin chemotype diversity in these agriculturally important fungi.

Influence of water activity and temperature on growth and mycotoxin production by Alternaria alternata on irradiated soya beans.

The aim of this study was to determine the effects of water activity (a(w)) (0.99-0.90), temperature (15, 25 and 30°C) and their interactions on growth and alternariol (AOH) and alternariol monomethyl ether (AME) production by Alternaria alternata on irradiated soya beans. Maximum growth rates were obtained at 0.980 a(w) and 25°C. Minimum a(w) level for growth was dependent on temperature. Both strains were able to grow at the lowest a(w) assayed (0.90). Maximum amount of AOH was produced at 0.98 a(w) but at different temperatures, 15 and 25°C, for the strains RC 21 and RC 39 respectively. Maximum AME production was obtained at 0.98 a(w) and 30°C for both strains. The concentration range of both toxins varied considerably depending on a(w) and temperature interactions. The two metabolites were produced over the temperature range 15 to 30°C and a(w) range 0.99 to 0.96. The limiting a(w) for detectable mycotoxin production is slightly greater than that for growth. Two-dimensional profiles of a(w)× temperature were developed from these data to identify areas where conditions indicate a significant risk from AOH and AME accumulation on soya bean. Knowledge of AOH and AME production under marginal or sub-optimal temperature and a(w) conditions for growth can be important since improper storage conditions accompanied by elevated temperature and moisture content in the grain can favour further mycotoxin production and lead to reduction in grain quality. This could present a hazard if the grain is used for human consumption or animal feedstuff.

Pathogenicity of phylogenetic species in the Fusarium graminearum complex on soybean seedlings in Argentina

Soybean (Glycine max L.) is one of the main crops in Argentina. Most of the studies of pathogenicity in the Fusarium graminearum complex have focused on strains isolated from wheat and maize, and there is little information on strains isolated from soybean. Our objective in the present study was to compare the pathogenicity among soybean isolates of different phylogenetic species within the Fusarium graminearum complex on soybean seedlings under controlled conditions. Six strains representing three different phylogenetic species (F. graminearum, F. meridionale and F. cortaderiae) were identified by partial sequencing of the Translation Elongation Factor -1α gene (TEF-1) and evaluated for pathogenicity. All six strains reduced emergence, mainly by causing pre-emergence damping-off, seedling height and root dry weight and produced abnormal seedlings. The mean disease severity averaged across all isolates was approximately 3.0 in a 0–4 rating scale where 0 = healthy seedling and 4 = dead seedling. Significant differences in pathogenicity were observed among F. graminearum, F. meridionale and F. cortaderiae. These results are consistent with the hypothesis that different phylogenetic species within the Fusarium graminearum complex isolated from soybean are pathogenic under controlled conditions to soybean seedlings in Argentina. The present study demonstrates for the first time the pathogenic effect of F. meridionale on soybean in Argentina.

Impact of cycling temperatures on Fusarium verticillioides and Fusarium graminearum growth and mycotoxins production in soybean

BACKGROUND Fusarium graminearum and F. verticillioides are two very important mycotoxigenic species as they cause diverse diseases in crops. The effects of constant and cycling temperatures on growth and mycotoxin production of these species were studied on soybean based medium and on irradiated soya beans. RESULTS F. graminearum grew better when was incubated at 15, 20 and 15-20 °C (isothermal or cycling temperature) during 21 days of incubation. Maximum levels of zearalenone and deoxynivalenol (39.25 and 1040.4 µg g(-1), respectively) were detected on soya beans after 15 days of incubation and the optimal temperature for mycotoxin production was 15 °C for zearalenone and 20 °C for deoxynivalenol. F. verticillioides grew better at 25 °C in culture medium and at 15/20 °C and 15/25 °C on soybean seeds. Fumonisin B(1) was produced only in culture medium, and the maximum level (7.38 µg g(-1)) was found at 15 °C after 7 days of incubation. CONCLUSION When growth and mycotoxin production under cycling temperatures were predicted from the results under constant conditions, observed values were different from calculated for both species and substrate medium. Therefore, care should be taken if data at constant temperature conditions are to be extrapolated to real field conditions.

Safety Aspects in Soybean Food and Feed Chains: Fungal and Mycotoxins Contamination

Soybean (Glycine max L. Merr.) is an Asiatic leguminous plant cultivated in many parts of the world for its oil and proteins, which are extensively used in the manufacture of animal and human foodstuffs (FAO, 2004a; Hepperly, 1985). The production reached 47.5 million tons during the 2006/2007 harvest season ranking Argentina third as soybean producer in the world. In Argentina, during the last quarter of the century, soybean production has increased at an unprecedented rate from a cultivated area of 38.000 hectares in 1970 to 16 million hectares today. Around 70% of the soybean harvested is processed, providing 81% and 36% of the worlds exported soybean oil and meal, respectively (SAGPyA, 2010). Soybean is often attacked by fungal infections during cultivation, or post-harvest (in transit or in storage), significantly affecting its productivity. Seeds and infected harvest debris are the main sources of primary infections, and the level of seed damage depends on environmental conditions such as high relative humidity, dew, and temperatures above 25 oC. These species can be potential mycotoxin producers. Mycotoxins (from “myco” fungus and toxin) are relatively low-molecular weight, fungal secondary metabolic products that may affect exposed vertebrates such as animals in a variety of ways. Mycotoxins are considered secondary metabolites because they are not necessary for fungal growth and are simply a product of primary metabolic processes. The functions of mycotoxins have not been clearly established, but they are believed to play a role in eliminating other microorganisms competing in the same environment. They are also believed to help parasitic fungi invade host tissues. The amount of toxins needed to produce adverse health effects varies widely among toxins, as well as within each person’s immune system (Brase et al., 2009). Some mycotoxins are carcinogenic, some are vasoactive, and some cause central nervous system damage. The mycotoxins can be acutely or chronically toxic, or both, depending on the kind of toxin, the dose, the health, the age and nutritional status of the exposed individual or animal, and the possible synergistic effects between mycotoxins. The most frequently studied mycotoxins are produced by species of Aspergillus, Penicillium, Fusarium and Alternaria. There is an increasing world consumer demand for high quality and inocuous food and drink products with the lowest possible level of contaminants such as mycotoxins. As a result, the food industry in the developed world demands raw ingredients of the best