Partial and complete 16S rDNA sequences, their use in generation of 16S rDNA phylogenetic trees and their implications in molecular ecological studies

Abstract

When the first reports of 16S rRNA sequence analysis were published about 20 years ago, the potential of ribosomal RNA species for studies in evolution, phylogeny, systematics and identification [47] and ecology [28, 29] were far from being fully acknowledged. What started with short sequences from a limited range of pro- and eukaryotic species turned out to revolutionize our understanding of (i) the origin of life on the planet Earth, (ii) the development of unknown, ancient, ancestral forms into main lineages leading to extant organisms such as Bacteria, Archaea and Eukaryotes, (iii) the relatedness among recently evolved species, and (iv) the richness of microbial forms in the environment. The results obtained have not only resusciated interest in the work of systematists, but have stimulated the cooperative work of researchers from different disciplines. Both, microbial taxonomy and microbial ecology have revived from Sleeping Beauty-types, somewhat boring fields into exciting disciplines in which objectivity has replaced decades of subjective judgement of facts. Molecular biology, using techniques such as PCR amplification, cloning, sequencing, hybridization and probing has finally been fully accepted by these disciplines. Organisms known to exist as symbionts and parasites were suddenly available for further characterization, and the extent of living but uncultured microorganisms became obvious without, however, allowing an estimation of the order of magnitude with which these dormant forms exceed the culturable organisms. In the past, uncultured strains have been observed in natural samples solely by light and electron microscopy. The determination of phylogenetic relationships of some these obligately inter- and intracellular organisms has been one of the scientific highlights in the eighties. Using the cataloguing approach, the task was easy when the symbiont occurred abundantly as an almost pure culture, as in the case of Prochloron species within their ascidian (Didemnid) hosts [38]. The reverse transcriptase technique of 16S rRNA sequencing was used to determine the phylogeny of the extremely slow growing pathogenic organism Mycobacterium leprae [39], but the main breakthrough occurred recently when the 16S rDNA of small numbers of cells could be amplified by the PCR approach and sequenced, either directly or following a cloning step. Examples are symbionts of shipworms [8], Holospora [1], the nematode lysing Pasteuria penetrans [Stackebrandt, Dorsch and Paschke, unpublished], the giant fish bacterium [2], Buchnera species which are symbions of aphids [43] or the methanogenic symbionts of anaerobic ciliates [11].