Award: OPP-1643466

Overview. Chemoautotrophic production, the conversion of inorganic carbon to organic matter driven by energy from chemical reactions rather than by light, has been proposed to augment biomass production in polar regions, particularly during winter when water column photosynthesis is reduced by low light levels. The oxidation of ammonia to nitrite and then to nitrate, a process known as nitrification, is a major chemoautotrophic process known to take place in polar water columns, yet direct comparisons of the contribution of nitrification to chemoautotrophic production in polar waters have not been made. A major goal of this project was to generate a dataset from well-characterized waters that would allow direct comparison of these two processes. We collected samples from 3 or 4 depths chosen to represent different communities of bacterioplankton: Antarctic Surface Water (ASW, 0-35 m); Winter Water (WW, 36-174 m); Circumpolar Deep Water (CDW, 175-1000 m); and slope water (Slope, >1000 m), at stations occupied on a cruise surveying the continental shelf and slope west of the Antarctic Peninsula during the austral summer of 2018 (LMG 18-01). Concurrent sampling by the Palmer Long-Term Ecological Research program (PAL-LTER) provided ancillary data on bacterioplankton abundance and production, chlorophyll and nutrient concentrations, and physical and chemical variables that allowed us to put our measurements in a broader context. We provided them with data unique to our project (ammonium, nitrite and urea concentrations, measurements of total chemoautotrophy and nitrification rates, abundance of specific groups of microorganisms). Nitrification rates were determined by measuring the oxidation of 15N-labeled substrates. Nitrification rates were converted to carbon equivalents using factors derived from published work with pure cultures of ammonia- and nitrite- oxidizing archaea and bacteria. We also assessed the contribution of nitrogen (N) from urea and putrescine (1,4-diamino butane, a polyamine) to chemoautotrophy via nitrification. Total chemoautotrophic production was measured as the conversion of 14C-labeled dissolved inorganic carbon into organic matter during incubations of samples in the dark. The abundance of bacteria and ammonia- and nitrite-oxidizing organisms in plankton samples was measured by quantitative amplification by PCR of specific genes in DNA extracted from plankton samples. Significant Findings. Oxidation rates of N supplied as ammonium, nitrite or urea were negligible in ASW and were greatest in the WW and CDW water masses. Distributions of nitrifying organisms (ammonia and nitrite oxidizers) followed a similar pattern, though nitrification rates were not tightly correlated with nitrifier abundance. Mean rates (nmol L-1 d-1) across all samples (186-232 independent rate measurements) were 10.3, 3.0, 5.9 and 6.6 for ammonia, urea, nitrite and putrescine N, respectively. Mean abundance estimates over the same set of samples (31-92 independent measurements) were 7300, 1900, 660, 11 and 550 x 103 gene copies L-1 for Thaumarchaeota 16S rRNA, Water Column type B amoA, Thaumarchaeota ureC, beta-Proteobacteria amoA, and nitrite oxidizing bacteria 16S rRNA genes, respectively. Chemoautotrophic carbon fixation rates in a subset of 42 of samples from WW, CDW and Slope water masses averaged 1.9, 1.7 and 0.05 nmol C L-1 d-1. Our calculations indicate that ammonia oxidation supported 111% and 46%, oxidation of urea N supported 22% and 39%, while nitrite oxidation supported 51% and 24 % of the chemoautotrophic production in the WW and CDW water masses, respectively. Despite indications from previous work, oxidation rates of putrescine N were not highly correlated with ammonia oxidation rates. We had hypothesized that the oxidation of putrescine N is mediated by reactive oxygen species (ROS) produced by Thaumarchaeota during ammonia oxidation. We tested this by assessing the ability of pure cultures of two strains of Nitrospumilaceae (NOT from the Antarctic) to grow on a variety of organic nitrogen compounds. Neither isolate was able to grow on any of the polyamines tested. However, when cultures growing on unlabeled ammonia were supplied with 15N-labeled putrescine, 15NOx was produced, suggesting that the 15N supplied as putrescine was oxidized abiotically by ROS, or that Thaumarchaeota were oxidizing 15NH4 regenerated from PUT. Direct tests of abiotic, chemical oxidation of 15N-labeled putrescine by hydrogen peroxide or peroxynitrite, two ROS species known to be produced by Thaumarchaeota during growth on ammonia, were negative. These findings suggest that our field data resulted from regeneration of 15N from putrescine by heterotrophs, or that 15N from putrescine is oxidized by another ROS species, such as superoxide. Broader Impacts. This project resulted in training of a postdoctoral researcher and providing undergraduate students with opportunities to gain hands-on experience with research on microbial geochemistry. One of these students was an African-American woman who has gone on to graduate school in Environmental Studies and Public Policy at the University of Oregon. The LTER program, in general, is interested in ecosystem processes and how these change through space and, especially, time. Our work has contributed substantially to understanding an important aspect of nitrogen cycling and bacterioplankton production in the PAL-LTER study area. Last Modified: 11/03/2020 Submitted by: James T Hollibaugh