16S ribosomal DNA analysis of microbial populations associated with hydrocarbon reservoirs
The sulphate-reducing bacteria (SRB) are a diverse group of organisms which use sulphate as a terminal electron acceptor and produce the highly toxic gas, hydrogen sulphide. The deleterious effects of this include hydrocarbon reservoir souring, formation damage and microbial corrosion. The SRB are of major economic importance to the oil industry. However, knowledge of the microbial ecology of the deep subsurface remains limited. The aim of this project was to investigate whether organisms are indigenous to the hydrocarbon formation and/or are introduced during drilling operations. A range of molecular techniques such as 16S rDNA sequence analysis, probing with labelled oligonucleotides, and denaturing gradient gel electrophoresis (DGGE) were employed to investigate the microbial diversity in oil field samples. A wide range of bacterial 16S rDNA sequences were identified using these molecular methods. An analysis of drilling mud samples revealed a diverse range of bacterial 16S rDNA sequences confirming that bacteria, including SRB, can be introduced to the reservoir during drilling operations. A number of bacterial 16S rDNA sequences were recovered from a geological core sample taken from a depth of 9,770 feet. The microbial diversity was remarkable in such a high temperature, high pressure environment. This lends credence to the theory that certain bacteria may be indigenous to the subsurface environment. Scanning electron micrographs of core which had been incubated in growth medium indicated the presence of 'nannobacteria'. These tiny coccoids, with a diameter of only 0.1 μm are far smaller than the generally accepted minimum size for cellular life forms. The nannobacteria grew in regular colony shaped structures and were seen only in sections taken from inside the rock. This study indicates that hydrocarbon reservoirs provide an environment in which bacteria, if introduced during drill operations, may become established. However, the subsurface also contains complex indigenous microbial populations that demonstrate considerable species diversity and may include unrecognised life forms.