Rapid detection of coliforms in drinking water of Arak city using multiplex PCR method in comparison with the standard method of culture (Most Probably Number)

Dehghan Fatemeh; Zolfaghari Mohammad Reza; Arjomandzadegan Mohammad; Kalantari Salomeh; Ahmari Gholam Reza; Sarmadian Hossein; Sadrnia Maryam; Ahmadi Azam; Shojapoor Mana; Najarian Negin; Kasravi Alii Reza; Falahat Saeed

doi:10.12980/APJTB.4.2014C896

Rapid detection of coliforms in drinking water of Arak city using multiplex PCR method in comparison with the standard method of culture (Most Probably Number)

Asian Pac J Trop Biomed. 2014 May;4(5):404-9. doi: 10.12980/APJTB.4.2014C896.

Affiliations

¹ Department of Microbiology, Qom Branch, Islamic Azad University, Qom, Iran.
² Tuberculosis and Infectious Research Center and Department of Microbiology, Arak University of Medical Sciences, Iran.
³ Hygiene and Quality Control Office of Markazi, Province Water and Wastewater Company, Iran.
⁴ Department of Biology, Payame Noor University, P.O.Box 19395-4697, Tehran, I.R. of Iran.
⁵ Molecular and Medicine Research Center, Arak University of Medical Sciences, Iran.
⁶ Department of Microbiology, Arak University of Medical Sciences, Iran.
⁷ Department of Microbiology, Islamic Azad University, Sciences and Research Branch, Arak, Iran.

Abstract

Objective: To analyse molecular detection of coliforms and shorten the time of PCR.

Methods: Rapid detection of coliforms by amplification of lacZ and uidA genes in a multiplex PCR reaction was designed and performed in comparison with most probably number (MPN) method for 16 artificial and 101 field samples. The molecular method was also conducted on isolated coliforms from positive MPN samples; standard sample for verification of microbial method certificated reference material; isolated strains from certificated reference material and standard bacteria. The PCR and electrophoresis parameters were changed for reducing the operation time.

Results: Results of PCR for lacZ and uidA genes were similar in all of standard, operational and artificial samples and showed the 876 bp and 147 bp bands of lacZ and uidA genes by multiplex PCR. PCR results were confirmed by MPN culture method by sensitivity 86% (95% CI: 0.71-0.93). Also the total execution time, with a successful change of factors, was reduced to less than two and a half hour.

Conclusions: Multiplex PCR method with shortened operation time was used for the simultaneous detection of total coliforms and Escherichia coli in distribution system of Arak city. It's recommended to be used at least as an initial screening test, and then the positive samples could be randomly tested by MPN.

Keywords: Coliforms; PCR; Rapid detection; Water.