|Microdiversity of Rhodopirellula genes in coastal sediments|
Biswas, B. (2011). Microdiversity of Rhodopirellula genes in coastal sediments. MSc Thesis. Max Planck Institute for Marine Microbiology: Bremen. 69 pp.
Rhodopirellula Schlesner, Rendsmann, Tindall, Gade, Rabus, Pfeiffer & Hirsch, 2004 [WoRMS]
ANE, Waddenzee [Marine Regions]
Rhodopirellula; microdiversity; functional genes; phylogenetic diversity
Most of the microbial diversity seems to be hiding in the large amount of uncultured bacteria. With the increasing development of molecular tools, the microbial diversity analysis in culture-independent manner is exploring insights into the question “how diverse are the bacteria?” Rhodopirellula baltica is one of the members of the unusual, lacking of peptidoglycan, but widely distributed bacterial phylum Planctomycetes. Many strains of the genus have been recently isolated. However, isolates are too few to reveal a comprehensive microbial biodiversity view. Analysis of 16S rRNA gene has become the primary approach though the microdiversity analysis is skewed due to the limited resolution of the 16S rRNA gene, which is conserved to identify species unambiguously. However, housekeeping genes that are not facing such a strong functional selective pressure as the 16S rRNA vary significantly and can resolve species. Whole genome analyses indicated a species border at 6% gene diversity for housekeeping genes. Rhodopirellula has recently revealed a biogeography in European sediments. In this thesis, three candidates of housekeeping genes, namely fumC, guaA and carB, were applied to analysis the microdiversity of Rhodopirellula in coarse, sandy sediments of the Wadden Sea in Sylt. PCR amplification was successful for the carB gene. In total 137 carB sequences were obtained after TOPO® TA cloning of the carB amplicon. These were 68 unique sequences of 693 base pairs. Reference sequences of isolates have shown that strains of one species differ up to 3% dissimilarity, whereas the next closely related species had a difference of 8% dissimilarity. We applied a threshold of 10% dissimilarity. This reduced the diversity to 18 operational taxonomic units (OTUs) of the clones. Rarefaction analysis suggested the presence of up to 25 OTUs. In maximum-likelihood phylogenetic trees, most of the clones (84%, 14 OTUs) were closely affiliated with different members of Rhodopirellula though others (4 OTUs) were branched distantly from Rhodopirellula. The phylogenetic branching and clustering of clones were similar in different phylogenetic methods except a few branching in maximum parsimony. This initial insight has revealed a high diversity of Rhodopirellula species in coarse, sandy surface sediment of the Wadden Sea.