Next-generation sequencing identification of pathogenic bacterial genes and their relationship with fecal indicator bacteria in different water sources in the Kathmandu Valley, Nepal

Abstract

Bacteriological analysis of drinking water leads to detection of only conventional fecal indicator bacteria. This study aimed to explore and characterize bacterial diversity, to understand the extent of pathogenic bacterial contamination, and to examine the relationship between pathogenic bacteria and fecal indicator bacteria in different water sources in the Kathmandu Valley, Nepal. Sixteen water samples were collected from shallow dug wells (n=12), a deep tube well (n=1), a spring (n=1), and rivers (n=2) in September 2014 for 16S rRNA gene next-generation sequencing. A total of 525 genera were identified, of which 81 genera were classified as possible pathogenic bacteria. Acinetobacter, Arcobacter, and Clostridium were detected with a relatively higher abundance (>0.1% of total bacterial genes) in 16, 13, and 5 of the 16 samples, respectively, and the highest abundance ratio of Acinetobacter (85.14%) was obtained in the deep tube well sample. Furthermore, the blaOXA23-like genes of Acinetobacter were detected using SYBR Green-based quantitative PCR in 13 (35%) of 37 water samples, including the 16 samples that were analyzed for next-generation sequencing, with concentrations ranging 5.3–7.5logcopies/100mL. There was no sufficient correlation found between fecal indicator bacteria, such as Escherichia coli and total coliforms, and potential pathogenic bacteria, as well as the blaOXA23-like gene of Acinetobacter. These results suggest the limitation of using conventional fecal indicator bacteria in evaluating the pathogenic bacteria contamination of different water sources in the Kathmandu Valley.