Molecular detection of Bacillus anthracis: evaluation of the efficiency of DNA extraction and a novel dry PCR

Abstract

Abstract Background Due to recent increase in mailings of anthrax spores, the detection of bioweapons has gained a great deal of interest. This study aimed to investigate the yield and purity of DNA obtained from spores and vegetative forms of Bacillus anthracis for detection by conventional (wet) and dry (lyophilized) PCR methods. Materials and methods Biosamples from stock solution were reconstituted to a concentration 108 cfu/mL followed by the spectrophotometric measurement of the yield and purity of acquired DNA. Twelve wet and 12 dry PCR studies of four various DNA dilution samples were performed for each three target gene (cap, pag, sap) of B. anthracis. Results Significant differences for both DNA yields and purity were found between liquid-agar and liquid-spore samples. No significant difference was observed between wet and dry PCR in concentration of 2.5 ng/μL DNA for all gene regions. PCR results of sap gene region with DNA concentrations of 1.5 ng/μL and 0.9 ng/μL was found statistically significant in favor of conventional wet PCR method (p = 0.047 and p ≤ 0.001, respectively). Conclusion It is concluded that B. anthracis cultivated in liquid medium is more usable than vegetative or spore form obtained on plate agar for proper DNA extraction.