Assessing the Abundance and Diversity of Nitrite Ammonifiers in Manures.

Manures contain bacteria that can reduce nitrate/nitrite to ammonium through a process called dissimilatory nitrate reduction to ammonium, also known as nitrate/nitrite ammonification (NA). Unlike gaseous N₂O and N₂ from denitrification, the main product of NA (i.e. ammonium) can sorb to cation exchange sites or be assimilated, thereby slowing re-conversion to mobile nitrate. Thus, the ability to promote the NA pathway by adding manure to soils has potential to improve N retention under proper management. The molecular marker for bacterial NA is the nrfA gene, which encodes the penta-heme nitrite reductase enzyme, NrfA. The objective of this experiment is to characterize the poorly studied NA bacteria in raw and anaerobically-digested manures. Manure samples before and after anaerobic digestion were collected from 5 different biological manure treatment facilities in Pennsylvania during 2016-2017 for physico-chemical and molecular analysis. DNA was extracted from manure, and real-time PCR and amplicon sequencing on the Illumina MiSeq platform were used to evaluate nrfA gene abundance and diversity. Preliminary results suggest that manure handling techniques like anaerobic digestion lowered the C:N ratio by 36-56% in digested manure as compared to raw manure. The nrfA gene abundance ranged from 10²-10⁵ gene copy numbers ml^-1 manure, but was not significantly affected by the digestion process (P>0.05). Principal coordinate analysis revealed that anaerobic digestion of manure changed the nrfA community structure. Higher Shannon diversity of nrfA was observed in raw manure as compared to the digested manure. Our results indicate that the type of manure applied to soil can regulate nrfA gene abundance and community structure.