Evaluation of the Heterotrophic Plate Count Test for Drinking Water Safety: Comparing Culture-based vs. Molecular Methods for Identifying Bacteria

Undergraduate Honors Thesis

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Citeable URL: https://scholar.colorado.edu/concern/undergraduate_honors_theses/0k225b57q

Abstract

Federal regulations of drinking-water quality regarding microbial contamination require the use of the culture-based heterotrophic plate count (HPC). This study compares HPC to culture-independent techniques for enumeration and identification of bacteria in environmental samples. Drinking water samples were collected from four different sites at two different times of year and analyzed using HPC, bacterial identification from 16s ribosomal RNA gene sequences, and direct cell counts from epifluorescence microscopy. No significant correlation was found between HPC measurements and either direct cell counts or measures of sample biodiversity from sequence analysis. 16s rRNA gene sequences from bulk DNA extractions reveal microbial communities in drinking water to comprise a broad array of bacterial diversity, including microbes of potential concern to human health such as mycobacteria. Conversely, HPC consistently selected for members of the Alphaproteobacteria (Sphingomonas, 45.8%; Methylobacteria, 33.8%; Porphyrobacter, 11.7%). These organisms comprised 25.0, 2.8, and 0.8 percent espectively of all 16s rRNA gene libraries from bulk DNA extractions. This result suggests that the heterotrophic plate count is not a relevant measure of drinking water quality.

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