Sulfide-driven denitrification: detecting active microorganisms in fed-batch enrichment cultures by DNA stable isotope probing

A microbial community was enriched in the anoxic compartment of a pilot-scale bioreactor that was operated for 180 days, fed with sewage and designed for organic matter, nitrogen and sulfide removal by coupling anaerobic digestion, nitrification and mixotrophic denitrification. Denitrification occurred with endogenous electron donors, mainly sulfide and residual organic matter, coming from the anaerobic compartment. The microorganisms involved in denitrification with sulfide as electron donor were identified by DNA-stable isotope probing with [U-C-13]-labelled CO2 and NaHCO3. Complete denitrification occurred every two days, and the applied NO3-/S2- ratio was 1.6. Bacteria belonging to the Sulfurimonas denitrificans was identified as a chemoautotrophic denitrifier, and those related to Georgfuchisa toluolica, Geothrix fermentans and Ferritrophicum radicicola were most probably associated with heterotrophic denitrification using endogenous cells and/or intermediate metabolites. This study showed that DNA-SIP was a suitable technique to identify the active microbiota involved in sulfide-driven denitrification in a complex environment, which may contribute to improve design and operation of bioreactors aiming for carbon-nitrogen-sulfur removal.