The depth specific significiance, relative abundance and phylogeography of anaerobic ammonium oxidation (anammox) in marine and estuarine sediments

Abstract

The availability of fixed forms of nitrogen is critical to the regulation of primary production. Until recently, denitrification (the sequential reduction of NO3 -, through NO2 - , to di-nitrogen gas) was recognised as the only significant pathway facilitating N removal. The discovery of anaerobic ammonium oxidation (a process whereby NH4 + is anaerobically oxidised with NO2 - to form N2 gas), however, has redefined this concept. Environmental studies clearly indicate that anammox is a globally significant sink for N, yet the factors that govern variations in the potential for anaerobic ammonium oxidation (anammox), the abundance or the natural diversity of these organisms are poorly understood. The purpose of this investigation was to identify the organisms responsible for anammox across a gradient of the Medway estuary, Irish Sea and North Atlantic. DNA amplification was performed using the Planctomycete forward primer ‘S-P-Planc-0046-a- A-18’ in combination with either ‘S-G-Sca-1309-a-A-21’ (targeting members of the genus ‘Scalindua’) or ‘S-*-Amx-0368-a-A-18’reverse. Analysis of 16S rRNA gene fragments indicated that the majority of sequences shared large phylogenetic distances with the ‘candidate’ species ‘Scalindua sorokinni’ (!93% sequence similarity). A number of the sequences extracted from both marine and estuarine sediments, however, cluster into 2 sub-groups that share common origins with the anammox lineage. In addition, the zone of potential anammox activity was characterised using a combination of 15N isotope labelling experiments, pore water oxygen profiles and depth specific rates sediment metabolism (CO2 production). This was performed in combination with fluorescence in situ hybridisation (FISH), to map shifts in the abundance of anammox organisms with depth, thus potentially linking the depth integrated capacity for anammox to deviations in population size. The potential for anammox activity and positive FISH signals confirm the presence of anammox at all sites investigated. The contribution of anammox to total N2 production (ra%) varied, on average, between 4- 35% in estuarine and 13-49% in marine sediments relative to denitrification. This was linked to a small population of anammox organisms constituting <1-3% of total bacteria in the estuarine sediments and <1-5% in marine samples. Whilst the depth specific values of ra correlate with the relative abundance of anammox organisms in continental shelf (r2=0.86, P=0.024) and slope sediments (r2=0.84, P=0.011), no such relationship was observed in the Medway estuary. The overall capacity for therefore appears to be dependant upon the depth integrated potential for anammox and is not inherent to differences in population size.