Stable nitrogen isotope measurements of marine bacterial proteins and nucleic acids: tracers of microbial activity

Abstract

Stable nitrogen isotopes ([]15N) can trace elemental cycling in aquatic ecosystems if the enzyme mediated fractionations associated with nutrient uptake and assimilation are negligible or consistently predictable. In this study, bacterial proteins and nucleic acids from cultures and environmental samples were analyzed to determine if protein []15N reliably reflects whole cell []15N. In batch culture experiments, the net fractionation between Vibrio harveyi and bacterial protein extracts, varied as a function of growth rate and substrate. Exponential cell growth resulted in a reduction of discrimination during both organic and inorganic nitrogen assimilation. Organic nitrogen assimilation resulted in protein extracts that averaged 3%[] to 4% ([]1.0%[]) heavier than the whole cells, confirming previously reported values. Protein extracts from cultures assimilating NH4+, however, exhibited more variable discrimination but also averaged 3%[] to 4%[] (+1.0%[]) during exponential and stationary growth. Large volume batch culture reduced the variation in measured discrimination and resulted in protein extracts that averaged 0.8%[] heavier than whole cells. Nucleic acid extracts prepared using a separate, previously established procedure averaged 1.6%[] lighter than cells. Mass balance of protein and nucleic acid accurately reflected the than cells. Mass balance of protein and nucleic acid accurately reflected the proportions of these two intracellular nitrogen pools. Both protein and nucleic acid 515N were determined to reflect whole cell values. Estimates of intracellular discrimination were strongly linear for large volume cultures with proteins being []2.8 [] 0.8%[] (n=l0 ) heavier than nucleic acids. Protein and nucleic acid extracts from estuarine and marine water samples were measured and bacterial population []15N values were estimated. lntracellular discrimination in natural samples averaged less than half that observed in culture (n=10). When the []15N of suspended particulate material (SPM), was compared with that calculated for whole cells, marine and estuarine samples were clearly distinguished, as were waters influenced by allochthonous Mississippi River input.