Habitat Segregation of a Functional Gene Encoding Nitrate Ammonification in Estuarine Sediments

Distribution and diversity of nrfA gene encoding dissimilatory nitrite reduction to ammonium (DNRA) in the sediments of the Colne River, North Essex, UK, were investigated. Sequencing cloned nrfA fragments amplified from environmental DNA enabled structure analysis of the bacterial community responsible for this pathway. The DNA was extracted from the sediment samples at different depths from the estuary ranging from freshwater to seawater regions, and amplified using specific PCR primer pairs targeting for the nrfA gene. Analysis of the nrfA clones showed two distinct clusters corresponding to their origins, namely, divided into the stable sites (marine and freshwater regions) and the unstable sites (brackish water region), where the tidal rise and fall constantly disturbs the environmental conditions. In addition, the nrfA clones from the deeper layer of the sediment formed a more homogenous community than those from the surface layer of the sediment. This may be due to more isolated and anaerobic conditions kept in the deeper sediment less influenced by the overlying water and other environmental factors. Most of the nrfA clones from the Colne estuarine sediments formed several distinct clusters including known nitrate ammonifiers such as Aeromonas, Shewanella, Desulfovibrio and Sulfurospillum. One of which was, however, related to Bacteroides but still quite divergent (∼70% identity) and the rest forming unknown clusters of supposedly uncultured members of bacteria. This is the first trial to describe the nrfA partial sequences derived from a natural environment, with reference to their habitat-specific community structure.

Keywords:

• DNRA
• estuarine sediment
• environmental gradient
• distribution
• nitrate ammonification
• nrfA

Acknowledgments

This study was carried out as a part of self-funded projects at the University of Essex. The author thanks the past members of molecular and environmental microbiology laboratories for their technical assistance. Thanks are also due to his colleagues at the University of Wales Bangor for giving him helpful comments on the manuscript

Notes

^a Lab stocks at Essex;

^b environmental isolates in this study;

^c gift from Dr Kondo;

^d gifts from Dr Sass;

^e negative control; +++, strongly positive; ++, positive; +, weakly positive

^f semi-nested PCR primer set developed in this study;

^g touchdown PCR primer set (Mohan et al. 2004).

^a : this study,

^b : Mohan et al. (2004).