Overestimation of prokaryotic production by leucine incorporation—and how to avoid it

Prokaryotes play a central role in aquatic ecosystems by consuming approximately half of the organic matter produced by aquatic primary production, of which a fraction is used for growth. Accurately measuring this prokaryotic biomass production is key to understanding aquatic carbon and nutrient cycles, since it is instrumental in driving biogeochemical processes that control parameters such as atmospheric carbon content. Aquatic prokaryotic biomass production is typically estimated from incorporation rates of the amino acid leucine during radiotracer experiments—a method widely used since the 1980s. Here we evaluate the underlying assumptions of the method with a focus on the associated conversion factors and review them in the context of empirical data. We demonstrate that the commonly used theoretical conversion factors fail to account for leucine's use as precursor for de novo protein synthesis and its respiration. As a consequence, prokaryotic biomass production is likely considerably overestimated when applying the standard conversion factors. Most severely affected are open-ocean, mesopelagic and benthic environments, where 25% of the estimates are likely to be overestimated by at least a factor of 6.1, 4.9, and 6.5, respectively. We propose a refined carbon-to-leucine conversion factor and make recommendations for improving and selecting appropriate experimental protocols.