Phylogenetic analysis of bacterial diversity using ribosomal RNA gene sequences

Abstract

Broad-range PCR amplification of ribosomal RNA (rRNA) gene sequences and community profiling methods were applied in studies of the microbiology of three different environmental habitats. Length heterogeneity analysis of PCR-amplified 16S rRNA gene sequences (LH-PCR) was developed for use in profiling bacterial diversity and for optimisation of the broad-range bacterial PCR procedure. A bioremediation system of polychlorophenol-contaminated groundwater in Karkola, Finland, was investigated over a six-year period. It was dependent on a stable bacterial community with low species diversity. One of the dominant organisms was isolated and named Novosphingobium sp. MT1. Molecular analysis of polychlorophenol-degrading bacteria isolated from the groundwater showed that they represented a wide range of phylogenetic groups. Comparative analysis of 16S rRNA and pcpB gene sequences suggested that the pcpB gene, involved in the initiation of chlorophenol degradation, had been achieved in Karkola sphingomonads by a recent horizontal transfer. Bacterial diversity profiles in a thermophilic aerobic biofilm process treating paper mill process water showed that the bacterial community was completely different in the biofilm and in the suspended biomass. The biofilm community was sensitive to pH alterations, but recovered after disturbances. The study showed that LH-PCR was a fast and reliable method for screening biotechnological processes, but the separation resolution between bacterial groups was hampered by overlapping sequence lengths. A direct diagnostic procedure based on broad-range bacterial PCR and sequencing was applied for the first time for the fish diseases. Direct molecular analysis was shown to be a potential tool for improving and expediting the diagnosis of flavobacteriosis, which is a serious salmonid disease in Finland.