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Ammonia-oxidizers’ diversity in wastewater treatment processes

ABSTRACT The diversity of ammonia-oxidizing bacteria (AOB) within the beta-subclass of Proteobacteria was investigated by genus- and family-specific real-time quantitative PCR (qPCR) assays on samples drawn from wastewater treatment systems. The 16S rRNA gene copy numbers ranged from 7.0 × 103 to 6.8 × 106, 1.1 × 107 to 1.8 × 107, and 2.9 × 105 to 1.5 × 107 copies/mL, respectively. Volumetric ammonium load (VAL) in the wastewater treatment systems calculated using the AOB numbers was in the range of 2.1–12.6 mM/d. Distribution patterns of eutrophic (i.e. Nitrosomonas europaea and Nitrosomonas nitrosa clusters) and oligotrophic (i.e. Nitrosomonas cryotolerans cluster) AOB groups were correlated with the VAL values. A high possibility of potential false-positive detection by family-specific qPCR assays was established by evaluating theoretical specificity in in silico and experimental investigations. The specificities of genus-specific qPCR assays were confirmed by amoA PCR, followed by cloning and sequencing. VAL must be the factor influencing the inclusion of AOB species. However, there was no significant correlation between the volatile suspended solid concentration representing chemical oxygen demand and N. europaea’s community population, indicating that the degree of ammonia oxidation influenced the community cluster of Nitrosomonas relatively more.