We investigated the diversity of planktonic Betaproteobacteria and the seasonal population changes of betaproteobacterial taxa in an oligo-mesotrophic lake (Piburger See, Austria). Focus was put on the vertical distribution of the investigated populations and on differences between their respective cell fractions with apparent amino acid incorporation. On average, 66% of betaproteobacterial cells and 73% of their diversity could be attributed to four clades within three lineages that were further analysed by fluorescence in situ hybridization. The numbers of bacteria from the R-BT subclade of the beta I lineage and from the PnecB subgroup of the beta II lineage were rather constant throughout the water column. In contrast, members of another subgroup of beta II (PnecC) and bacteria related to Methylophilus (beta IV) were particularly numerous in the oxygen-depleted zone. In general, only moderate seasonal changes in abundance were observed in the upper water layers, whereas there was a clear relationship between decreasing oxygen levels and the rise of bacteria from the PnecC and beta IV clades in deeper strata. On average, almost 80% of beta I bacteria, but < 15% of cells from the beta IV clade, showed amino acid incorporation. Our results suggest that the studied populations occupy distinct vertical and ecophysiological niches in Piburger See.