BCIG-SMAC medium and PMA-qPCR for differential detection of viable Escherichia coli in potable water

Rehan Deshmukh; Sunil Bhand; Utpal Roy

doi:10.18502/ijm.v13i5.7427

Rehan Deshmukh Department of Biological Sciences, Birla Institute of Technology and Sciences, Pilani, Goa, India
Sunil Bhand Department of Biology, Faculty of Science, Dr. Vishwanath Karad MIT World Peace University, Pune, India
Utpal Roy Department of Biological Sciences, Birla Institute of Technology and Sciences, Pilani, Goa, India

DOI: https://doi.org/10.18502/ijm.v13i5.7427

Keywords: Coliforms; Detection; Escherichia coli; Pathogens; Propidium monoazide; Public health

Abstract

Background and Objectives: Public health protection requires timely evaluation of pathogens in potable water to minimize outbreaks caused by microbial contaminations. The present study was aimed at assessing the microbiological quality of water obtained from Shantinagar (a rural area in the South Goa region of Goa, India) using 5-Bromo-4-Chloro-3-Indoxyl β-D-glu- curonide-Sorbitol MacConkey agar (BCIG-SMAC) medium and, propidium monoazide-quantitative polymerase chain reac- tion (PMA-qPCR) assay for differential detection and quantification of viable Escherichia coli cells in water samples.

Materials and Methods: Membrane filtration method was used for both BCIG-SMAC medium and PMA-qPCR methods. To determine the efficiency of detection of viable cells, we first evaluated the PMA treatment protocol and established the standard calibration curves using previously reported primers.

Results: PMA-qPCR detected as low as 7 femtograms of DNA of E. coli per qPCR reaction whereas the limit of detection (LOD) of BCIG-SMAC medium was 1.8 CFU/100mL. A total of 71 water samples spanning 2017-2018 have been analyzed using BCIG-SMAC medium and PMA-qPCR, of which 95.77% (68/71) and 7.04% (5/71) were found to be total E. coli and E. coli O157:H7, respectively. PMA-qPCR study showed the viable counts of total viable E. coli cells ranging from 3CFU/100mL to 8.2×102 CFU/100mL. The total E. coli CFU/100mL quantified by PMA-qPCR significantly exceeded (paired t-test; P<0.05) the number on BCIG-SMAC medium.

Conclusion: The present study indicates that the microbiological quality of environmental water samples analyzed do not comply with the regulatory standard. Therefore, special attention is warranted to improve the overall portable quality of water in the perspective of public health.