Comprehensive Detection of Pathogenic Bacteria in Jar Water, Community Well Groundwater, and Environmental Water in the Kathmandu Valley, Nepal

Abstract

Microbial contamination of water sources remains a serious problem in the Kathmandu Valley, Nepal. This study was conducted to comprehensively investigate the occurrence of pathogenic bacteria in water samples collected from the valley, most of which were used for drinking and other domestic purposes. The water samples consisted of 10 samples of commercial jar water, 10 of groundwater from shallow dug wells, 3 of groundwater from deep tube wells, 1 of stone spout water, 2 of spring water, and 2 of river water. DNA microarray analysis targeting 941 different pathogenic bacterial species/groups showed the prevalence of Acinetobacter, Arthrobacter, Brevibacterium, Pseudomonas, and Legionella and the presence of biosafety level 3 pathogens in some samples, which were partially confirmed by genus-specific PCR quantification for Legionella and Brucella. Additionally, the occurrence of most pathogens detected via DNA microarray analysis was not correlated with that of fecal indicator bacteria (total coliforms and Escherichia coli). These results will aid in understanding the overall picture of pathogenic bacterial contamination and in screening important pathogens that should be investigated in detail.

Comparison of Pathogenic Bacteria in Water and Fecal-Source Samples in the Kathmandu Valley, Nepal, Using High-Throughput DNA Microarray

“…'%' indicates presence of bacteria in samples divided by total number of samples collected.A classification at the genus level revealed that 12 generaPseudomonas, and Xanthomonas) were present in 15 (94%) of the 16 samples tested. This result was comparable to that of a previous study, where genera such as Acinetobacter, Arthrobacter, Brevibacterium, Legionella, and Pseudomonas, were detected in 82% (23/28) of water samples, which had been collected in 2014[27]. Waterborne pathogenic bacteria that could be transmitted through the fecal-oral route were identified, whereby Vibrio cholerae, Klebsiella spp., and Salmonella enterica serovars were detected in 63%, 38%, and 31% of the water samples tested, respectively.…”

“…People use 12% and 22% of groundwater for drinking and cooking purpose [1]. A previous study of groundwater and river water revealed 5 BSL 3 pathogens, of which the Anaplasma marginale/centrale group (39%) was detected most frequently [27]. In this study, 6 BSL 3 pathogens were detected from different water samples, and the detection ratio of the A. marginale/centrale group (94%) was the highest (Table 1).…”

“…The applicability of the microarray to environmental samples had been assured by the developers and previously applied to landfill leachets, river water samples, and shallow well groundwater [5,24,25]. In brief, Cy3-labeled polymerase chain reaction (PCR) products were prepared by using a PCR mixture (50 µL) containing 25 µL of SapphireAmp Fast PCR Master Mix (Takara Bio, Kusatsu, Japan), 0.25 µL each of 15 pmol/µL forward and reverse primers [5,[26][27][28], 1 µL of DNA template, and 23.5 µL of ultrapure water.…”