Microbial community analysis of groundwater related to the denitrification in Ryukyu limestone aquifer

Abstract

This study explores the use of microbial community analysis to evaluate the processes involved in nitrate attenuation in groundwater. Polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) is used to identify denitrifiers based only on their 16SrRNA gene sequences, and Real-Time PCR analysis is used to quantify nitrite reducing genes (nirK and nirS). The latter suggest a new method to be used for detecting denitrification activity by comparing the gene dosage that has been detected by Real-Time PCR and the value of the nitrate (NO 3-) δ 15 N and δ 18 O. This study focuses on a zone of significant NO 3-attenuation occurring at underground dam catchment area in the karstic Ryukyu limestone aquifer, which is located in the southern part of Okinawa, Japan. As a result of microbial analysis, the bacteria were detected at all observation points which have been reported to have denitrification ability. And it has been confirmed that the bacteria has a gene nirS which is related to denitrification. In addition, the correlation was high between nirK /nirS gene dosage that has been detected by Real-Time PCR and the value of the nitrate δ 15 N and δ 18 O; therefore, this study demonstrates the effectiveness of using Real-Time PCR analysis for providing insights into the processes affecting nitrate attenuation in groundwater.