Correlating the relative abundance of ammonia- and nitrite-oxidizing bacteria with the nitrification performance using Hierarchical Oligonucleotide Primer Extension (HOPE)

Establishing an optimal ratio of ammonia-oxidizing bacteria (AOB) to nitrite-oxidizing bacteria (NOB) is important in ensuring the functionality of the nitrification process. A high-throughput method based on hierarchical oligonucleotide primer extension (HOPE) to quantify for the abundance of the AOB and NOB was proposed in this study to track the proportional ratio between these two bacterial groups. The method was tested against wastewater samples collected from a full-scale wastewater treatment plant and biomass obtained from a laboratory-scale biofilm-based trickling reactor. Our findings indicate a significant increase in the abundance of AOB, specifically Nitrosomonas, with increasing ammonium content and shock loading (t-test, p < 0.001). In contrast, Nitrosospira remained stable in its relative abundance against the total community throughout the operational phases. There was a corresponding significant decrease in the relative abundance of NOB group, specifically Nitrospira. Although Nitrobacter increased in its relative abundance despite the shock loading, nitrite accumulation in the effluent suggested that Nitrobacter may not be playing an active role in the nitrification process within the wastewater environment compared to Nitrospira. It was determined that the optimal ratio of AOB against NOB ranged from 0.7 to 2.5 during stable reactor performance. Multivariate analysis from both HOPE and next generation sequencing datasets showed good correlation at a significant confidence level. The HOPE method allows for the simultaneous monitoring of relative abundance of AOB and NOB groups, and would provide indicative measurements of the reactor performance and nitrification functionality.