Impact of pH and salinity fluctuations on oxidation of Fe(II) by nitrate-reducing microorganisms enriched from the reduced tidal sediment of an extreme acidic river (Río Tinto, Spain)

Nitrate reduction coupled to Fe(II) oxidation (NRFeOx) contributes to Fe cycling in the estuarian sediments of the Río Tinto river (Huelva, Spain). However, it is not yet known (i) whether and which NRFeOx microorganisms can be enriched from the reduced sediment layer and (ii) how in-situ pH and salinity fluctuations affect NRFeOx. Therefore, we (i) used two different approaches such as microcosm experiments (sediment amended with either NO3−/Fe2+aq or acetate/NO3−/Fe2+aq) and enrichment cultures (medium amended with acetate/NO3−/Fe2+aq) to enrich NRFeOx microorganisms to (ii) test their salinity and pH tolerance under simulated high tide and low tide conditions. We found that different microorganisms such as Thiobacillus (up to 9.7±5.8% DNA-based 16S rRNA gene abundance) and Denitromonas (83.6% DNA-based 16S rRNA gene abundance) were contributing to NRFeOx in the microcosm experiments and enrichment culture approach, respectively. The strong buffering capacity of the native sediment and the presence of additional organic carbon as acetate can favour NRFeOx microorganisms during acidic water influx (low tide) events. The ⁓100% conversion of NO3⁻ to NO₂⁻ under high tide conditions was observed both in the enrichment cultures and microcosm experiment when acetate was added suggesting the chemodenitrification may be the primary Fe(II) oxidation pathway under salty conditions.