Abstract
We propose an innovative, repeatable, and reliable experimental workflow to concentrate and detect environmental bacteria in drinking water using molecular techniques. We first concentrated bacteria in water samples using tangential flow filtration and then we evaluated two methods of environmental DNA extraction. We performed tests on both artificially contaminated water samples and real drinking water samples. The efficiency of the experimental workflow was measured through qPCR. The successful applicability of the high‐throughput DNA sequencing (HTS) approach was demonstrated on drinking water samples. Our results demonstrate the feasibility of our approach in high‐throughput‐based studies, and we suggest incorporating it in monitoring strategies to have a better representation of the microbial community. In the recent years, HTS techniques have become key tools in the study of microbial communities. To make the leap from academic laboratories to the routine monitoring (e.g., water treatment plants laboratories), we here propose an experimental workflow suitable for the introduction of HTS as a standard method for detecting environmental bacteria.
Highlights
Monitoring microbial contamination in drinking water, rinse solutions, juices, milk, and many other foodstuffs is a relevant topic of public health concern (Galimberti et al, 2015; Liu et al, 2012; Namvar, Haq, Shields, Amoako, & Warriner, 2013).The label “microbiologically pure” occurring on many food items means that no target microorganisms responsible for food spoilage and dangerous for human health (e.g., Escherichia coli, Legionella pneumophila, and Enterococci) were detected, but obviously it does not imply that there are no bacteria at all inside that product
Our results demonstrate that tangential flow filtration (TFF) coupled with DNA-based molecular techniques is the ideal tool for surveying microbial diversity in water samples and allows an unbiased and sensitive detection of microbes
We were able to concentrate 1 L of drinking water in
Summary
Monitoring microbial contamination in drinking water, rinse solutions, juices, milk, and many other foodstuffs is a relevant topic of public health concern (Galimberti et al, 2015; Liu et al, 2012; Namvar, Haq, Shields, Amoako, & Warriner, 2013). Culture-based methods are the classical techniques applied to detect indicator or sentinel microorganisms to monitor microbial contamination (Ashbolt, Grabow, & Snozzi, 2001; Boubetra, Nestour, Allaert, & Feinberg, 2011). They are biased by three relevant disadvantages: first, they require an enrichment step (typically incubation or filtration/volume reduction) that inevitably extends the analysis time. There is often the necessity of using selective media for specific bacteria, impeding the simultaneous detection and enumeration of different microorganisms which typically characterize complex matrices Molecular techniques, such as qPCR and high-throughput DNA sequencing (HTS), may provide a way to overcome these issues (Ercolini, 2013). Libraries for HTS sequencing were set up in order to demonstrate the feasibility of this pipeline in HTS-based studies and its potential application by a wide panel of stakeholders dealing with different aims (both theoretical and applicative/commercial)
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