The Latest Practices in Culture-Free Detection of Bacteria in Water, From Sampling to Membrane Filtration and DNA...

The advancement of molecular biology in water research, combined with the lack of standardization in this area of research, exposed the need for presenting the different methodological approaches available to researchers. The aim of the article is to identify and critically discuss the water filtration methods for culture-free bacterial DNA extraction. A systematic review was conducted on PubMed and Web of Science, according to the 3Cochrane Handbook recommendations and PRISMA 2020 Checklist. The initial search retrieved 513 articles, and 53 were included with a multi-step approach screening (title, abstract, full text). Outcomes of interest involved details about sampling, filtration methods and DNA extraction. The most reported sampling methods were using containers, especially of 1000 mL. Filtration was performed using membrane filters, the majority of the studies using Polyethersulfone (PES) membranes with the pore size of 0.22 µm. Samples were prepared for extraction either with the help of an enzymatic pretreatment (protease, proteinase K, lysozyme, or lysostaphin), physical methods (bead-beating, centrifugation, vortexing, sonication, heating, freeze-thaw cycles), or detergent pretreatment. The preferred method for extraction was using commercially available kits, although some of the studies described in-house protocols. The methods must be adapted to the scientific scope. The review summarizes the existing methods and critically appraises their utility and promotes advancement in the field of environmental molecular biology.