Drinking water, water used in food production and for irrigation, water for fish farming, waste water, surface water, and recreational water have been recently recognized as a vector for the transm...

Numerous quantitative PCR assays for microbial fecal source tracking (MST) have been developed and evaluated in recent years. Widespread application has been hindered by a lack of knowledge regarding the geographical stability and hence applicability of such methods beyond the regional level. This study assessed the performance of five previously reported quantitative PCR assays targeting human-, cattle-, or ruminant-associated Bacteroidetes populations on 280 human and animal fecal samples from 16 countries across six continents. The tested cattle-associated markers were shown to be ruminant-associated. The quantitative distributions of marker concentrations in target and nontarget samples proved to be essential for the assessment of assay performance and were used to establish a new metric for quantitative source-specificity. In general, this study demonstrates that stable target populations required for marker-based MST occur around the globe. Ruminant-associated marker concentrations were strongly correlated with total intestinal Bacteroidetes populations and with each other, indicating that the detected ruminant-associated populations seem to be part of the intestinal core microbiome of ruminants worldwide. Consequently tested ruminant-targeted assays appear to be suitable quantitative MST tools beyond the regional level while the targeted human-associated populations seem to be less prevalent and stable, suggesting potential for improvements in human-targeted methods.

Noroviruses (previously Norwalk-like viruses) are the most common viral agents associated with food- and waterborne outbreaks of gastroenteritis. In the absence of culture methods for noroviruses, animal caliciviruses were used as model viruses to study inactivation by nonionizing (253.7-nm-wavelength [UV]) and ionizing (gamma) radiation. Here, we studied the respiratory feline calicivirus (FeCV) and the presumed enteric canine calicivirus (CaCV) and compared them with the well-studied bacteriophage MS2. When UV irradiation was used, a 3-log ₁₀ reduction was observed at a fluence of 120 J/m ² in the FeCV suspension and at a fluence of 200 J/m ² for CaCV; for the more resistant phage MS2 there was a 3-log ₁₀ reduction at a fluence of 650 J/m ² . Few or no differences were observed between levels of UV inactivation in high- and low-protein-content virus stocks. In contrast, ionizing radiation could readily inactivate MS2 in water, and there was a 3-log ₁₀ reduction at a dose of 100 Gy, although this did not occur when the phage was diluted in high-protein-content stocks of CaCV or FeCV. The low-protein-content stocks showed 3-log ₁₀ reductions at a dose of 500 Gy for FeCV and at a dose of 300 for CaCV. The inactivation rates for both caliciviruses with ionizing and nonionizing radiation were comparable but different from the inactivation rates for MS2. Although most FeCV and CaCV characteristics, such as overall particle and genome size and structure, are similar, the capsid sequences differ significantly, making it difficult to predict human norovirus inactivation. Adequate management of UV and gamma radiation processes for virus inactivation should limit public health risks.

Legionella infections are among the most important waterborne infections with constantly increasing numbers of cases in industrialized countries, as a result of aging populations, rising numbers of immunocompromised individuals and increased need for conditioned water due to climate change. Surveillance of water systems is based on microbiological culture-based techniques; however, it has been shown that high percentages of the Legionella populations in water systems are not culturable. In the past two decades, the relevance of such viable but non-culturable (VBNC) legionellae has been controversially discussed, and whether VBNC legionellae can directly infect human macrophages, the primary targets of Legionella infections, remains unclear. In this study, it was demonstrated for the first time that several starved VBNC Legionella strains (four L. pneumophila serogroup 1 strains, a serogroup 6 strain and a L. micdadei strain) can directly infect different types of human macrophages and amoebae even after one year of starvation in ultrapure water. However, under these conditions, the strains caused infection with reduced efficacy, as represented by the lower percentages of infected cells, prolonged time in co-culture and higher multiplicities of infection required. Interestingly, the VBNC cells remained mostly non-culturable even after multiplication within the host cells. Amoebal infection by starved VBNC Legionella, which likely occurs in oligotrophic biofilms, would result in an increase in the bacterial concentration in drinking-water systems. If cells remain in the VBNC state, the real number of active legionellae will be underestimated by the use of culture-based standard techniques. Thus, further quantitative research is needed in order to determine, whether and how many starved VBNC Legionella cells are able to cause disease in humans.

Numerous bacterial genetic markers are available for the molecular detection of human sources of fecal pollution in environmental waters. However, widespread application is hindered by a lack of knowledge regarding geographical stability, limiting implementation to a small number of well-characterized regions. This study investigates the geographic distribution of five human-associated genetic markers (HF183/BFDrev, HF183/BacR287, BacHum-UCD, BacH, and Lachno2) in municipal wastewaters (raw and treated) from 29 urban and rural wastewater treatment plants (750 – 4,400,000 population equivalents) from 13 countries spanning six continents. In addition, genetic markers were tested against 280 human and non-human fecal samples from domesticated, agricultural and wild animal sources. Findings revealed that all genetic markers are present in consistently high concentrations in raw (median log10 7.2 - 8.0 marker equivalents (ME) 100 ml-1) and biologically treated wastewater samples (median log10 4.6 - 6.0 ME 100 ml-1) regardless of location and population. The false positive rates of the various markers in non-human fecal samples ranged from 5% to 47%. Results suggest that several genetic markers have considerable potential for measuring human-associated contamination in polluted environmental waters. This will be helpful in water quality management and pollution modeling and health risk assessment across the globe (as demonstrated by QMRAcatch).