Male-specific RNA coliphages detected by plaque assay and RT-PCR in tropical river waters and animal fecal matter

Abstract

Male-specific RNA coliphages (FRNA) have been recommended as indicators of fecal contamination and of the virological quality of water. In this study, 16 river water and 183 animal fecal samples were examined for the presence of FRNA coliphages by a plaque assay using Salmonella typhimurium WG49 and WG25 to differentiate between male-specific and somatic phages, a RNase spot test to differentiate between DNA and RNA phages and a reverse transcriptase-polymerase chain reaction (RT-PCR) for the specific identification of FRNA phages. The overall recovery rate for F-specific coliphages was 8.0%. (4.4% from animal fecal matter and 50% from river water samples). Plaque counts were generally low (< 6×10² pfu per g feces or ml water), with FRNA (6.5%) and Male-specific DNA coliphages (FDNA) (7.0%) phages occurring at almost equal frequencies. The RT-PCR was positive in all FRNA plaques and was able to identify FRNA phages in mixed populations of FRNA, FDNA and somatic phages.

Keywords:

• FRNA coliphages
• river water
• chicken feces
• plaque assay
• RT-PCR

Acknowledgements

This research was supported by IRPA grant no. 060203-0771 and Monash University Malaysia research grant ES-16-02. For environmental sampling, we thank Mr C. K. Wong from University Malaya. The MS2, WG49 and WG45 were gifts from Prof. C. W. Wang, Department of Molecular Medicine, University of Malaya. The staining and microscopic observation was carried out with the assistance of Ms Azizah Ab Jalil in the Department of Microscopy and Microanalysis in University Putra Malaysia.