The population of ammonia-oxidizing bacteria in a temperate oligotrophic freshwater lake was analyzed by
recovering 16S ribosomal DNA (rDNA) from lakewater and sediment samples taken throughout a seasonal
cycle. Nitrosospira and Nitrosomonas 16S rRNA genes were amplified in a nested PCR, and the identity of the
products was confirmed by oligonucleotide hybridization. Nitrosospira DNA was readily identified in all
samples, and nitrosomonad DNA of the Nitrosomonas europaea-Nitrosomonas eutropha lineage was also directly
detected, but during the summer months only. Phylogenetic delineation with partial (345 bp) 16S rRNA gene
sequences of clones obtained from sediments confirmed the fidelity of the amplified nitrosomonad DNA and
identified two sequence clusters closely related to either N. europaea or N. eutropha that were equated with the
littoral and profundal sediment sites, respectively. Determination of 701-bp sequences for 16S rDNA clones
representing each cluster confirmed this delineation. A PCR-restriction fragment length polymorphism
(RFLP) system was developed that enabled identification of clones containing N. europaea and N. eutropha 16S
rDNA sequences, including subclasses therein. It proved possible to analyze 16S rDNA amplified directly from
sediment samples to determine the relative abundance of each species compared with that of the other. N.
europaea and N. eutropha are very closely related, and direct evidence for their presence in lake systems is
limited. The correlation of each species with a distinct spatial location in sediment is an unusual example of
niche adaptation by two genotypically similar bacteria. Their occurrence and relative distribution can now be
routinely monitored in relation to environmental variation by the application of PCR-RFLP analysis.