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Field-based evaluation of a male-specific (F+) RNA coliphage concentration method
Citation:
Perez-Mendez, A., J. C. Chandler, J. Paar, M. Doolittle, B. Bisha, AND L. D. Goodridge. Field-based evaluation of a male-specific (F+) RNA coliphage concentration method. JOURNAL OF VIROLOGICAL METHODS. Elsevier Science BV, Amsterdam, Netherlands, 239(January):9-16, (2017).
Impact/Purpose:
Validate a new, low-volume, resin-based, viral concentration method through the analyses of fecal contamination in water samples by RT-PCR.
Description:
Fecal contamination of water poses a significant risk to public health due to the potential presence of pathogens, including enteric viruses. Thus, sensitive, reliable and easy to use methods for the detection of microorganisms are needed to evaluate water quality. In this study, we performed a field evaluation of an anion-exchange resin based platform to concentrate F-RNA coliphages (fecal/enteric virus indicators) from diverse fecally impacted environmental waters. In this platform, F-RNA coliphages are adsorbed to anion-exchange resin and direct nucleic acid isolation is performed, yielding a sample amenable to real-time reverse transcriptase PCR detection. Matrix-dependent inhibition was evaluated using known quantities of spiked F-RNA coliphage genogroups GI, GII, GII and GIV. Detection was successful in 97%, 72%, 85% and 98% of the samples for spiked F-RNA coliphage GI, GII, GIII and GIV, respectively, and was differentially affected by inhibitory properties specific to each water sample. No association between inhibition and the water samples’ physicochemical properties was apparent. Parallel evaluations of the spiked samples with internal amplification control (IAC) reactions (a widely used control to assess inhibition) demonstrated that IAC reaction inhibition was not agreement with that observed for spiked samples, suggesting that testing of spiked samples allows for better assessments of matrix-dependent inhibition. Additionally, the anion-exchange resin platform was able to detect F-RNA coliphages in 40 out 65 unspiked samples, demonstrating this system is a viable alternative to current strategies to detect F-RNA coliphages from water.
