Detection of bacteria in their natural habitats

The laboratory and isolation cultivation of a particular Bacterium from a biotope was at one time the sole evidence that that species was present. However, many studies have shown that there are many more prokaryotes in the environment than can be cultured using laboratory media and nucleic acid analysis has complemented classical microbial ecology.  Individual cells can be labeled with a dye attached to an oligonucleotide that hybridizes to ribosomal RNA. The organisms that bind the oligonucleotide will then show up under a fluorescence microscope. Careful choice of oligonucleotide can mean that these fluorescent probes can be species- or even strain-specific, allowing the microbial ecologist to examine and even count various sorts of microorganism in a particular niche. Various developments of this FISH technique have allowed the differentiation between metabolically active and inactive microorganisms.

The development of PCR means that the total ribosomal RNA in a microbial community can be examined. By sequencing a library of ribosomal cDNA from a particular biotope the ecologist has some idea of the diversity of active microorganisms. Extending this idea of amplification of DNA to other genes has developed into the field of environmental genomics.

Both FISH and PCR of total genomic DNA suggest that there is a far greater diversity of Bacteria and Archaea than first thought, to the extent that the idea of discrete species barriers is beginning to break down. Although these new organisms are often held  to be VBNC new techniques and  laboratory practices are finally yielding pure and  mixed  cultures suitable for physiological and  biochemical studies.