Biotechnology Investigations—Ocean Margins Program (BI-OMP)

Molecular approaches for in situ study of nitrate use by marine bacteria

Funded by DOE. December 2003 to November 2006. Bronk, Co-PI. Bronk part $186,160.
Lead PI Marc Frischer (SkIO)

Abstract

Traditionally, the importance of inorganic nitrogen (N) for the nutrition and growth of marine phytoplankton has been recognized, while inorganic N utilization by bacteria has received less attention. Likewise, organic N has been thought to be important for heterotrophic organisms but not for phytoplankton. However, accumulating evidence suggests that bacteria compete with phytoplankton for nitrate and other N species. The consequences of this competition may have a profound effect on the flux of N, and therefore carbon (C), in ocean margins. Because it has been difficult to differentiate between N uptake by heterotrophic bacterioplankton versus autotrophic phytoplankton; the processes that control N utilization, and the consequences of these competitive interactions, have traditionally been difficult to study. Significant bacterial utilization of DIN may have a profound effect on the flux of N and C in the water column because sinks for dissolved N that do not incorporate inorganic C represent mechanisms that reduce the atmospheric CO₂ drawdown via the “biological pump” and limit the flux of POC from the euphotic zone.

Since 1998 with the support of the DOE, we have developed a tool kit of molecular methods (PCR, RT-PCR, Q-PCR, QRT-PCR, and TRFLP) that allow selective isolation, characterization, and study of the diversity and genetic expression (mRNA) of the structural gene responsible for the assimilation of nitrate by heterotrophic bacteria (nasA). To date, our studies have revealed that bacteria capable of assimilating nitrate are ubiquitous in marine waters, that the nasA gene is expressed in these environments, that heterotrophic bacteria can account for a significant fraction of total DIN uptake in different ocean margin systems, that the expression of nasA is differentially regulated in genetically distinct nitrate assimilating bacteria, and that the best predictors of nasA gene expression are either nitrate concentration or nitrate uptake rates (Allen et al. 1999; Allen et al. in press; Booth et al. in prep). These studies provide convincing evidence of the importance of bacterial utilization of nitrate, insight into controlling processes, and provide a rich dataset that can be used to develop linked C and N modeling components necessary to evaluate the significance of bacterial DIN utilization to global C cycling.