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A molecular tool for detection and tracking of a potential indigenous Beauveria bassiana strain for managing emerald ash borer populations in Canada

Johny, Shajahan, Kyei-Poku, George
Journal of invertebrate pathology 2014 v.122 pp. 16-21
Agrilus planipennis, Beauveria bassiana, DNA, alleles, cross reaction, genetic variation, invasive species, monitoring, pathogens, polymerase chain reaction, single nucleotide polymorphism, trapping, Asia, Canada
Emerald ash borer is an invasive species from Asia. Beauveria bassiana strain L49–1AA is being tested for the control of emerald ash borer in Canada, using an autocontamination trapping system. We have developed a simplified allele discrimination polymerase chain reaction (PCR) assay to screen B. bassiana strain, L49–1AA from other Beauveria species by targeting the inter-strain genetic differences in 5′ end of EF1-α gene of the genus Beauveria. A single nucleotide polymorphism (SNP) site, T→C was identified only in L49–1AA and was used to develop a simplified allele discrimination polymerase chain reaction (PCR) assay based on a modified allelic inhibition of displacement activity (AIDA) approach for distinguishing B. bassiana L49–1AA from all background Beauveria isolates. The SNP site was employed to design inner primers but with a deliberate mismatch introduced at the 3′ antepenultimate from the mutation site in order to maximize specificity and detection efficiency. Amplification was specific to L49–1AA without cross-reaction with DNA from other Beauveria strains. In addition, the designed primers were also tested against environmental samples in L49–1AA released plots and observed to be highly efficient in detecting and discriminating the target strain, L49–1AA from both pure and crude DNA samples. This new method can potentially allow for more discriminatory tracking and monitoring of released L49–1AA in our autocontamination and dissemination projects for managing EAB populations. Additionally, the modified-AIDA format has potential as a tool for simultaneously identifying and differentiating closely related Beauveria species, strains/isolates as well as general classification of other pathogens or organisms.