Use of microbial analysis to evaluate denitrification in the karstic aquifer of Okinawa, Japan

Abstract:

Denitrification, a microbial process in the nitrogen cycle, is a facultative respiratory pathway in which nitrate (NO₃^-), nitrite (NO₂^-), nitric oxide (NO), and nitrous oxide (N₂O), successively, are reduced to nitrogen gas (N₂). This study explores the use of microbial 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), this suggest that a new methods for detecting denitrification activity by comparing the gene dosage that has been detected by RT-PCR and the value of the δ¹⁵N_NO3- and δ¹⁸O_NO3-. This study focuses on a zone of significant NO₃^- attenuation occurring at underground dam catchment area in the karstic Ryukyu limestone aquifer, which is located 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, many bacteria related to denitrification have been extracted from suspended solids more than from groundwater in the aquifer. And, the correlation was high between nirK /nirS gene dosage that has been detected by RT-PCR and the value of the δ¹⁵N and δ¹⁸O; therefore, this study demonstrates the effectiveness of using Real-Time PCR analysis for providing insights into the processes affecting nitrate attenuation in ground water.