An assessment of three methods for extracting bacterial DNA from beach sand.

Autor: Gallard-Gongora J; Department of Integrative Biology, University of South Florida, Tampa, Florida, USA., Lobos A; Department of Integrative Biology, University of South Florida, Tampa, Florida, USA., Conrad JW; Department of Integrative Biology, University of South Florida, Tampa, Florida, USA., Peraud J; Department of Integrative Biology, University of South Florida, Tampa, Florida, USA., Harwood VJ; Department of Integrative Biology, University of South Florida, Tampa, Florida, USA.
Jazyk: angličtina
Zdroj: Journal of applied microbiology [J Appl Microbiol] 2022 Apr; Vol. 132 (4), pp. 2990-3000. Date of Electronic Publication: 2021 Dec 26.
DOI: 10.1111/jam.15423
Abstrakt: Aims: Beach water quality is regulated by faecal indicator bacteria levels, sand is not, despite known human health risk from exposure to beach sand. We compared the performance of three methods to extract bacterial DNA from beach sand as a step toward a standard method.
Methods and Results: The analytical sensitivity of quantitative polymerase chain reaction (qPCR) for Enterococcus was compared for the slurry (suspension, agitation, membrane filtration of supernatant), versus direct extraction using PowerSoil™ or PowerMax Soil™ kits. The slurry method had the lowest limit of detection at 20-80 gene copies g -1 , recovered significantly more DNA, and the only method that detected Enterococcus by qPCR in all samples; therefore, the only method used in subsequent experiments. The slurry method reflected the spatial variability of Enterococcus in individual transect samples. Mean recovery efficiency of the microbial source tracking marker HF183 from wastewater spiked marine and freshwater beach sand was 100.8% and 64.1%, respectively, but varied, indicating that the mixing protocol needs improvement.
Conclusions: Among the three methods, the slurry method had the best analytical sensitivity and produced extracts that were useful for culture or molecular analysis.
Significance and Impact of Study: Standardization of methods for extraction of bacterial DNA from sand facilitates comparisons among studies, and ultimately contributes to the safety of recreational beaches.
(© 2021 The Society for Applied Microbiology.)
Databáze: MEDLINE
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