Identification and Quantification of a Toxigenic and Non-Toxigenic Aspergillus flavus Strain in Contaminated Maize Using Quantitative Real-Time PCR

Toxins

J. Erik Mylroie³  Seval Ozkan¹  Renuka Shivaji²  Gary L. Windham³  Michael N. Alpe³  W. Paul Williams³ 

[1] Department of Plant and Soil Sciences, Mississippi State University, Mississippi State City, MS 39762, USA;University of North Carolina at Greensboro Molecular Core Lab, University of North Carolina at Greensboro, Greensboro, NC 27412, USA;United States Department of Agriculture, Agricultural Research Service, Corn Host Plant Resistance Research Unit, Mississippi State City, MS 39762, USA;

关键词: aflatoxin;    Aspergillus flavus;    PCR;    quantification;    maize;    corn;   

DOI  :  10.3390/toxins8010015

来源: mdpi

【 摘 要 】

Aflatoxins, which are produced by Aspergillus flavus, are toxic to humans, livestock, and pets. The value of maize (Zea mays) grain is markedly reduced when contaminated with aflatoxin. Plant resistance and biological control using non-toxin producing strains are considered effective strategies for reducing aflatoxin accumulation in maize grain. Distinguishing between the toxin and non-toxin producing strains is important in determining the effectiveness of bio-control strategies and understanding inter-strain interactions. Using polymorphisms found in the fungal rRNA intergenic spacer region (IGS) between a toxigenic strain of A. flavus (NRRL 3357) and the non-toxigenic strain used in the biological control agent Afla-Guard^® (NRRL 21882), we developed a set of primers that allows for the identification and quantification of the two strains using quantitative PCR. This primer set has been used to screen maize grain that was inoculated with the two strains individually and co-inoculated with both strains, and it has been shown to be effective in both the identification and quantification of both strains. Screening of co-inoculated ears from multiple resistant and susceptible genotypic crosses revealed no significant differences in fungal biomass accumulation of either strain in the field tests from 2010 and 2011 when compared across the means of all genotypes. Only one genotype/year combination showed significant differences in strain accumulation. Aflatoxin accumulation analysis showed that, as expected, genotypes inoculated with the toxigenic strain accumulated more aflatoxin than when co-inoculated with both strains or inoculated with only the non-toxigenic strain. Furthermore, accumulation of toxigenic fungal mass was significantly correlated with aflatoxin accumulation while non-toxigenic fungal accumulation was not. This primer set will allow researchers to better determine how the two fungal strains compete on the maize ear and investigate the interaction between different maize lines and these A. flavus strains.

【 授权许可】

CC BY   
© 2016 by the authors; licensee MDPI, Basel, Switzerland.