Microbial source tracking in highly vulnerable karst drinking water resources.

Water resources situated in areas with underlying karst geology are particularly vulnerable to fecal pollution. In such vulnerable systems, microbial source tracking (MST) methods are useful tools to elucidate the pathways of both animal and human fecal pollution, leading to more accurate water use risk assessments. Here, we describe the application of a MST toolbox using both culture-dependent bacteriophage and molecular-dependent 16S rRNA assays at spring and well sites in the karstic St Imier Valley, Switzerland. Culture-dependent and molecular-dependent marker performance varied significantly, with the 16S rRNA assays displaying greater sensitivity than their phage counterpart; HF183 was the best performing human wastewater-associated marker while Rum2Bac was the best performing ruminant marker. Differences were observed in pollution regimes between the well and spring sampling sites, with the spring water being more degraded than the well site. Our results inform the choice of marker selection for MST studies and highlight differences in microbial water quality between well and spring karst sites.

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.

The effect of UV-C radiation (254 nm) on candidate microbial source tracking phages infecting a human-specific strain of Bacteroides fragilis (GB-124).

The enumeration of phages infecting host-specific strains of Bacteroides has been widely recognised as an effective and low-cost method of microbial source tracking (MST). A recently described human-specific Bacteroides host strain (GB-124) has been shown to detect bacteriophages exclusively in human-impacted waters and is emerging as a useful MST tool. However, a better understanding of the morphology and ecological behaviour of the phages, especially in wastewater disinfection processes, is now required in order to validate their role as MST markers. Bacteriophages infecting Bacteroides fragilis GB-124 (n = 21) were isolated from wastewater effluent and irradiated using laboratory-based UV-C (254 nm) collimated beam experiments. Bacteriophages were found to be both a morphologically and ecologically homogeneous group, with all specimens showing highly similar first order log-linear inactivation profiles (mean fluence required to inactivate phages by 4-log(10) was 36 mJ/cm(2)). These findings present the first evidence that phages infecting GB-124 are inactivated by the levels of UV-C radiation routinely delivered during tertiary wastewater treatment processes. More importantly, comparison with previously published inactivation data suggests that their response to UV-C radiation makes GB-124 phages more suitable surrogates for selected enteric viruses in UV disinfection processes than traditional faecal indicator bacteria or human-specific molecular markers.