High-throughput sequencing techniques have dramatically improved our capabilities to better understand the diversity, ecology, and evolution of soil bacterial communities in comparison with the traditional culture-based approaches. However, in the last few years, as genome sequencing data accumulate, there has been a new interest in culturing techniques to obtain as-yet-uncultured soil bacteria since isolates are still required to fully discover physiology and ecology of bacteria and improve reference databases for sequence annotation. Although new approaches are available, culture plating methods are still commonly used to describe culture-dependent bacterial diversity of soils and to isolate novel bacteria species. So far, little is known about the impact of using different culture media on the taxonomic diversity of heterotrophic bacteria recovered by plating. Therefore, we aimed to address the following questions here: (i) how different are bacteria isolated from a specific soil sample depending on the culture media used? (ii) do oligotrophic (versus eutrophic) culture media yield a higher culturable bacterial diversity?
To answer these questions, total bacteria (targeted by using extracted soil DNA) and bacteria growing in four culture media differing in their nutrient contents (TSA (trypticase soy agar) > R2A (full-strength Reasoner's 2A agar) > SEM (soil extract medium) > 0.05 R2A (20-fold dilution of R2A)) from agricultural, redwood forest, and petroleum hydrocarbon-contaminated soils were identified and compared by 16S rRNA gene metabarcoding.
Results/Conclusions
On average, considering the results of the four media used, only 15, 13, and 26 % of the total bacterial OTUs were culturable in the agricultural, redwood forest, and petroleum hydrocarbon-contaminated soils, respectively. The highest number of culturable OTUs was collected using SEM, while TSA (the richest medium) yielded the lowest amount of culturable OTUs in the three soils. Taxonomic composition of the culturable bacterial communities in the three soils was significantly dependent on the culture medium used. The growth of Gammaproteobacteria and Bacilli was favored by richer medium composition. Instead, Actinobacteria, Alphaproteobacteria, and Sphingobacteriia were recovered to a higher extent using the oligotrophic media (i.e., SEM and 0.05 R2A). In general, SEM yielded the most diverse bacterial communities at OTU and genus levels in the three soils. Therefore, according to our results, the use of culture media based on soil extracts seems to be a good option to obtain more diverse communities of soil culturable bacteria and potential as-yet-uncultured isolates.