In situ Analyses of the Distributions and Phylogeny of Cultivatable and Non-cultivatable Planctomycetales Using Phylogenetic Group-Specific RNA Probes

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The majority of directly countable bacteria in microbial ecosystems cannot be identified or cultivated using traditional microbiological techniques thus, cultured bacteria may not be representative of actual microbial diversity. The Planctomycetales are one of the ten major eubacterial divisions defined by 16S ribosomal RNA. the unusual ribosomal RNA signatures of this group make them exceptionally well-suited for microbial ecology studies using ribosomal RNA probe technologies. The recent development of phylogenetic group-specific DNA probes has made it possible to study the distribution and diversity of the Planctomycetales in natural habitats without the biases of cultivation requirements. Two probes for the Planctomycetales (PL1R and PL2R) and a "universal" rRNA probe will be used to measure the fraction of Planctomycetales ribosomal RNA genes in DNA extracted from mixed microbial populations. Flourescently labeled DNA probes will be used to measure the contribution of the Planctomycetales to attached microbial populations adhering to contact slides. Phylogenetic group-specific gene amplification primers will be used to identify particular constituents of the Planctomycetales population by the cloning and sequencing of rRNA genes. These results will be compared to a data base of known Planctomycetales sequences to determine whether previously uncultivated species are present in the population.