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Enrichment of DNRA bacteria in a continuous culture.

ABSTRACT: Denitrification and dissimilatory nitrate reduction to ammonium (DNRA) are competing microbial nitrate-reduction processes. The occurrence of DNRA has been shown to be effected qualitatively by various parameters in the environment. A more quantitative understanding can be obtained using enrichment cultures in a laboratory reactor, yet no successful DNRA enrichment culture has been described. We showed that a stable DNRA-dominated enrichment culture can be obtained in a chemostat system. The enrichment was based on the hypothesis that nitrate limitation is the dominant factor in selecting for DNRA. First, a conventional denitrifying culture was enriched from activated sludge, with acetate and nitrate as substrates. Next, the acetate concentration in the medium was increased to obtain nitrate-limiting conditions. As a result, conversions shifted from denitrification to DNRA. In this selection of a DNRA culture, two important factors were the nitrate limitation and a relatively low dilution rate (0.026 h(-1)). The culture was a highly enriched population of Deltaproteobacteria most closely related to Geobacter lovleyi, based on 16S rRNA gene sequencing (97% similarity). We established a stable and reproducible cultivation method for the enrichment of DNRA bacteria in a continuously operated reactor system. This enrichment method allows to further investigate the DNRA process and address the factors for competition between DNRA and denitrification, or other N-conversion pathways.

SUBMITTER: van den Berg EM

PROVIDER: S-EPMC4579468 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

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Project description:Denitrification and dissimilatory reduction to ammonium (DNRA) are competing nitrate-reduction processes that entail important biogeochemical consequences for nitrogen retention/removal in natural and man-made ecosystems. The nature of the available carbon source and electron donor have been suggested to play an important role on the outcome of this microbial competition. In this study, the influence of lactate as fermentable carbon source on the competition for nitrate was investigated for varying ratios of lactate and nitrate in the influent (Lac/N ratio). The study was conducted in an open chemostat culture, enriched from activated sludge, under strict anoxia. The mechanistic explanation of the conversions observed was based on integration of results from specific batch tests with biomass from the chemostat, molecular analysis of the biomass enriched, and a computational model. At high Lac/N ratio (2.97 mol/mol) both fermentative and respiratory nitrate reduction to ammonium occurred, coupled to partial oxidation of lactate to acetate, and to acetate oxidation respectively. Remaining lactate was fermented to propionate and acetate. At a decreased Lac/N ratio (1.15 mol/mol), the molar percentage of nitrate reduced to ammonium decreased to 58%, even though lactate was supplied in adequate amounts for full ammonification and nitrate remained the growth limiting compound. Data evaluation at this Lac/N ratio suggested conversions were comparable to the higher Lac/N ratio, except for lactate oxidation to acetate that was coupled to denitrification instead of ammonification. Respiratory DNRA on acetate was likely catalyzed by two Geobacter species related to G. luticola and G. lovleyi. Two Clostridiales members were likely responsible for lactate fermentation and partial lactate fermentation to acetate coupled to fermentative DNRA. An organism related to Propionivibrio militaris was identified as the organism likely responsible for denitrification. The results of this study clearly show that not only the ratio of available substrates, but also the nature of the electron donor influences the outcome of competition between DNRA and denitrification. Apparently, fermentative bacteria are competitive for the electron donor and thereby alter the ratio of available substrates for nitrate reduction.

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Enrichment of DNRA bacteria in a continuous culture.

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