Evaluation of molecular- and culture-dependent MST markers to detect fecal contamination and indicate viral presence in good quality groundwater.

Microbial contamination of groundwater represents a significant health risk to resource users. Culture-dependent Bacteroides phage and molecular-dependent Bacteroidales 16S rRNA assays are employed in microbial source tracking (MST) studies globally, however little is known regarding how these important groups relate to each other in the environment and which is more suitable to indicate the presence of waterborne fecal pollution and human enteric viruses. This study addresses this knowledge gap by examining 64 groundwater samples from sites with varying hydrogeological properties using a MST toolbox containing two bacteriophage groups (phage infecting GB-124 and ARABA-84), and two Bacteroidales 16S rRNA markers (Hf183 and BacR); those were compared to fecal indicator bacteria, somatic coliphage, Bacteroidales 16S rRNA marker AllBac, four human enteric viruses (norovirus GI and II, enterovirus and group A rotavirus) and supplementary hydrogeological/chemical data. Bacteroidales 16S rRNA indicators offered a more sensitive assessment of both human-specific and general fecal contamination than phage indicators, but may overestimate the risk from enteric viral pathogens. Comparison with hydrogeological and land use site characteristics as well as auxiliary microbiological and chemical data proved the plausibility of the MST findings. Sites representing karst aquifers were of significantly worse microbial quality than those with unconsolidated or fissured aquifers, highlighting the vulnerability of these hydrogeological settings.