Identification of dominant microbial community in aerophilic biofilm reactors by fluorescence in situ hybridization and PCR-denaturing gradient gel electrophoresis

Abstract

This study was conducted by combining fluorescence in situ hybridization (FISH) performed on 16S rRNA and polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) with 16S CTOs primers to characterize the nitrifying microbial communities in biofilm processes, which were tested to retrofit the S municipal wastewater treatment plant in Busan, Korea. Four aerophilic biofilm reactors were operated with hydraulic retention times of 2 to 8 h and biofilms were grown on ceramic media. The same low COD/ NH ⁺₄ -N ratio (100 mg/L of COD over 40 mg/L of NH ⁺₄ -N) with the S plant was used. The average relative population ratios of Nitrosomonas spp. to ammonia oxidizing bacteria (AOB) as measured by specific FISH probes (%/Nso190) were 75.0%, 80.0%, 73.0% and 73.5%, respectively, while those of Nitrosospira spp. to AOB were 21.0%, 14.7%, 24.6% and 24.1% after 180 days of operation. The microbial composition of Nitrobacter spp. detected by using a Nit3 probe was below 10% in each reactor. In contrast, Nitrospira genus detected with an Ntspa662 probe was around 20%. When CTOs primer was applied in PCR-DGGE analysis to define the nitrifying bacteria, the bands of group B in the R-1 reactor with the highest hydraulic retention time (HRT) had the strongest light intensity compared with two other reactors with lower HRTs after day 64. The bands of the groups were responsible for nitrification with the major dominant population in each reactor depending on the change of ammonia removal rate. These results would directly lead to an understanding of the reactor performance in relation to the ammonia removal, when conventional municipal wastewater treatment plants are retrofitted or upgraded to biological nitrogen removal processes using biofilm.