A non-radioactive assay of bacterial productivity optimized for oligotrophic pelagic environments
Limnol. Oceanogr. Methods 3:211-220 (2005) | DOI: 10.4319/lom.2005.3.211
ABSTRACT: The measurement of bacterial production is a cornerstone of research in aquatic biogeochemistry and microbial ecology, yet the standard radioisotope-based techniques are threatened by increasing logistical obstacles. The two primary goals of this study were (1) the development of a reliable nonradioactive assay of bacterial productivity interpretable in the context of previous 3H-thymidine (3H-TdR)based metrics, and (2) maximizing the sensitivity of the assay to allow routine measurement of bacterial productivity in open-ocean and lowproductivity environments. We optimize existing methods for immunochemical detection of the incorporation of the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) to present a functional assay of community bacterial productivity comparable to radioisotope-based assays in economy and field implementation and capable of consistently detecting bacterial production rates within the range of 0.1 to 10 pmol BrdU L1 h1. A standardization procedure provides complete separation of radioisotopes from the working methodology and gives statistically consistent results between calibrations with no degradation of the labeled DNA standards. Immunoassayed BrdU incorporation rates were highly correlated with parallel measurements of 3H-TdR incorporation rates spanning a range of 0.1 to 10 pmol L1 h1 (r = 0.96, n = 20, P < 0.001). Model II linear regression (BrdU = 0.65[3H-TdR] + 0.12) yielded a conversion factor not significantly different from that reported previously comparing incorporation rates measured using radioisotope protocols (3H-BrdU = 0.69[3H-TdR] 0.81). The assay is interpretable in the context of 3H-TdRbased measures of production, providing continuity as a nonradioactive measure of bacterial production in low-productivity aquatic ecosystems.