Study on the Identification Methods for Effective Microorganisms in Commercially Available Organic Agriculture Materials.

The identification of microorganisms in closely related groups is challenging. The present work focused on the different molecular methodology for the accurate microbial identification in the five commercially available organic agriculture materials enriched with effective microorganisms. From the tested five organic agricultural materials, a total of seven distinct bacterial colonies (A-1, B-1, C-1, D-1, E-1, E-2, and E-3) were isolated and processed for sequential identification utilizing HiCrome™ agar, biochemical tests with API CHB50, 16S rRNA gene analysis, random amplified polymorphic DNA (RAPD), and species-specific PCR analysis. All the isolated microorganisms were Gram-positive rods and spore former belonging to group and appeared as a differential characteristic feature on HiCrome™ agar. All isolates showed high-percentage similarities with the different members of species in biochemical testing and 16S rRNA gene analysis. The collective identification results revealed isolates, A-1, B-1, and C-1, close to . Further RAPD-PCR and species-specific PCR discriminated and provided confirmatory evidence for D-1 as and E-1, E-2, and E-3 as , respectively. In addition, presence of was also confirmed by toxin crystal protein staining. In conclusion, the species-specific primers could be used as a rapid and accurate identification tool to discriminate closely related species such as , , and .