Identification of bacterial populations in drinking water using 16S rRNA-based sequence analyses

Intracellular RNA is rapidly degraded in stressed cells and is more unstable outside of the cell than DNA. As a result, RNA-based methods have been suggested to study the active microbial fraction in environmental matrices. The aim of this study was to identify bacterial populations in drinking water by analyzing 16S rRNA-based clone libraries. Hollow-fiber ultrafiltration was used to concentrate bacterial communities from 40 l of tap water collected at 12 different times during three different summer months from a single point-of-use. Total RNA was extracted from the microbial concentrates and used to develop 16S rRNA-based clone libraries. Phylogenetic analyses of 1231 partial 16S rRNA gene sequences showed that difficult-to-classify bacterial sequences were the most predominant clones, representing 57.6% of the sequences analyzed. Within these unclassified clades, most sequences were closely related to sequences retrieved from previous DNA- and RNA-based drinking water studies. Other bacterial groups represented in this study included Proteobacteria, cyanobacteria, Actinobacteria, Bacteroidetes, and Planctomycetes. Overall, the results suggest that these bacterial groups are amongst potentially active bacteria in drinking water. Diversity analyses of clones generated show that while overall diversity is similar amongst the different months, membership changes with respect to time. The results from this study further improve our understanding of the molecular diversity and bacterial population dynamics of drinking water microbial communities. Moreover, these results provide the sequence foundation for the development of molecular assays that target active drinking water bacteria.