Molecular identification of Aspergillus parasiticus and flavus in forage samples by Duplex PCR method and comparison with culture and direct examination
Subject Areas : Journal of Comparative Pathobiologyناهید پدرام، منصور بیات، محمدحسن شاهحسینی، سعید بکائی، محمد قهری . 1
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Keywords: Aspergillus parasitcus, Aspergillus flavus, Duplex PCR, Forage,
Abstract :
Aspergillosis is one of infectious disease that it causes difficulties for human and poultry. The main aim of this study is the diagnosis of Aspergillus parasiticus and flavus in forage samples by Duplex PCR method and comparison with culture and direct examination. After extracting DNAs of standard strains of Aspergillus parasitcus and flavus along with primers of each fungus and PCR testing, genes of each fungus were amplified and PCR product was cloned using TA cloning by pTZ57R plasmid. After optimizing Duplex PCR (D-PCR) monoplex PCR tests, 50 forage samples by Duplex PCR method, culture and direct were tested.In the optimized PCR test, 343bp and 413bp products of Aspergillus parasiticus and flavus were respectively amplified. Fifty samples of forage by Duplex PCR method, culture and direct were tested. 13 samples were positive only by Duplex -PCR and 15 samples by both tests direct and culturing were positive and 26 samples were negative with three methods. Results of McNemar's test is equal about the three methods tests for diagnosing Aspergillus parasiticus and flavus (p=0.824). Molecular methods such as D-PCR are faster techniques than direct testing and culturing for diagnosis of Aspergillus parasiticus and flavus but there aren't difference between three methods for diagnosing Aspergillus parasiticus and flavus but with D-PCR we could diagnosis Aspergillus parasiticus.
