To culture or not to culture: a snapshot of culture-dependent and culture-independent bacterial diversity from peanut rhizosphere.

Abstract

BackgroundSequencing driven metagenomics studies have been instrumental in various aspects of microbiology including identification of newer taxa. While this culture-independent approach has its own merits and demerits, several studies have focussed on comparing it with traditional culture-dependent (CD) approach. However, most of these comparative studies rely on Sanger sequencing of complete 16S rRNA gene from pure culture colonies to determine the culturable bacterial diversity. This approach undercounts culturable diversity as only fewer isolates are selected, sequenced, and identified.MethodsIn this study, we have used an Illumina based partial 16S sequencing to identify all the microbes growing on the media and directly comparing with its culture-independent (CI) counterpart. Eight different media were used to target different organisms from soil. Diversity on these media were compared with their CI counterpart. The NGS data was analysed using DADA2 to provide more resolution to the data.ResultsIn line with studies of similar nature, current study presented higher bacterial diversity in CI approach. However, the current study reflected that a greater number of sequence variants were missed out in CI approach as compared to number of sequence variants shared with CD approach. We observed around 322 (5.98%) ASVs (Amplicon Sequence Variants) exclusively present in CD samples while, 234 (4.35%) ASVs were shared between both approaches. Most of these 322 CD exclusive ASVs were classified as Enterobacteriaceae family and Bacillus genus, with several ASVs annotated at the species level as well, and these organisms are more commonly observed in soil and were also detected in CI approach. Furthermore, 22 genera were exclusively detected in CD samples, most of which were reported from soil and water.