Primers: Functional Genes for Nitrogen-Cycling Microbes in Oil Reservoirs

Microbial communities found in the subsurface are important in the biogeochemical cycling of nitrogen (N) both in the oxidative and reductive processes, and changes in their functional structure might affect the stability of a petroleum reservoir. In petroleum reservoirs, where in situ conditions are predominantly anoxic, denitrification involving the stepwise reduction of nitrate (NO3 −) via nitrite (NO2 −) and nitric oxide (NO) to nitrous oxide (N2O) or dinitrogen gas (N2) is a major process. Microorganisms may also decompose organic N to ammonium (NH4 +) by ammonification, which can subsequently be oxidised to NO3 − via NO2 − by the process of nitrification. Autotrophic ammonia oxidation is known in three groups of microorganisms: aerobic autotrophic ammonia-oxidising bacteria (AOB) and Archaea (AOA) and anaerobic ammonia-oxidising bacteria (anammox). Since the microorganisms involved in many of these N transformations are taxonomically diverse, 16S rRNA-based methods are generally not suitable. Instead, a common approach has been to target the protein-encoding genes involved in the transformation of N as biomarkers. This chapter describes the common PCR primers that have been used to target the major functional genes involved in the cycling of N, with the key N transformations likely to occur in petroleum reservoirs highlighted throughout.