Better water quality indicators for understanding microbial health risks.

Abstract

The aims of the research were to evaluate existing microbial water quality indicators, and refine and/or develop alternative, improved indicators for determining the source of faecal contamination in urban and rural surface waters. There has been concern that because E. coli is capable of long term persistence in the environment in temperate climates that it is no longer a valid frontline tool for water quality monitoring. This research explored urban (untreated human sewage) and rural (cow faeces) impacts on water quality, and investigated relationships between faecal source tracking (FST) markers, faecal ageing determinants, microbial indicators and pathogens. The variables measured were FST markers (quantitative Polymerase Chain Reaction (qPCR), and faecal steroids), the faecal ageing ratio of atypical colonies/total coliforms (AC/TC), and the indicator microorganism, Escherichia coli. In the urban river study, additional determinants were indicator microorganisms, Clostridium perfringens and F-RNA phage; potential pathogens belonging to the genera of Campylobacter, Giardia and Cryptosporidium, and the FST marker, fluorescent whitening agents (FWA). In the urban study, a river had been impacted by major discharges of untreated human sewage. Variables were monitored in the river water and underlying sediment at three locations both during discharge, and up to eighteen months post-discharge. Relationships between E. coli and potential pathogens in water demonstrated that E. coli was a reliable indicator of public health risk. As a signal of a recent human faecal input, F-RNA phage were identified as suitable, cost-effective indicators to be measured in conjunction with E. coli. In contrast, the ubiquitous C. perfringens was observed to accumulate in sediments, confounding its ability as an indicator in water. PCR markers and faecal steroids in water were similar and even superior to E. coli as predictors of protozoan pathogen presence, and hence indicative of human health risk. The faecal ageing ratio, AC/TC in water, was significantly, negatively correlated with increasing pathogen detection. Campylobacter had the weakest associations with all microbial and FST indicators. It was observed, however, that where elevated E. coli levels were detected in water, identification of the HumM3 PCR marker in conjunction with F-RNA phage and a low AC/TC ratio <1.5 was indicative of fresh pollution and an associated health risk from Campylobacter. River sediments appeared to be a reservoir for steroids and FWA, Cryptosporidium and Giardia but not Campylobacter or F-RNA phage. FST PCR markers were not assayed in the sediments. There was no relationship observed between chemical FST markers in sediments and the overlying water, and few correlations between chemical FST markers and target microorganisms in sediment. In the rural study, the decomposition of cowpats was investigated to determine the mobilisation rates of water quality determinants when irrigated and non-irrigated cowpats were subjected to simulated flood and rainfall runoff events. It was observed that decomposing cowpats harboured concentrations of E. coli, which were available for mobilisation after flood and rainfall events for at least five and a half months post-deposition under flood conditions, and for at least two and a half months after lighter rainfall. Persistent levels of total coliforms in ageing cowpats showed that AC/TC ratios would indicate fresh sources of faecal contamination in a waterway after flood conditions up to four months post-deposition. An amplicon–based metagenomic study of the ageing cowpat investigated shifts in microbial populations as the cowpat decomposed. Major bacterial community shifts were observed over 161 days in the mobilised fraction from decomposing cowpats. Dominant bacteria that inhabited the cow rumen and fresh faeces, such as a Ruminococcus species, were displaced by bacterial groups that could be utilised as potential PCR targets of aged bovine faecal sources. Faecal steroid ratios were observed to be reliable and stable FST markers during the ageing process. The PCR marker ratio of BacR/TotalBac (ruminant (BacR)/Total Bacteroidetes) has potential as an indicator of 100% contribution from fresh bovine sources. Recommendations for water managers are outlined for the cost-effective application of FST tools based on findings from this current research. The differential fate and transport of microbial and FST markers noted in this research supported the use of multiple lines of evidence through application of a cohort of indicators for tracking the source(s) of faecal contamination and indicating the associated public health risk. In the urban river study, strong to moderate correlations between PCR and steroid markers suggested they could be used individually or combined for greater confidence in the result. Some of the FST host-associated PCR markers (HumM3 and CowM2) were shown to be useful indicators of recent faecal inputs to a waterbody. The lack of correlation between chemical FST markers and microorganisms in sediment suggested that chemical markers in sediment were indicative of historical faecal sources, and restricted their predictive value for health risks. Due to the persistence of potential pathogens, re-suspension of sediment has the potential to increase risk to human health for those who participate in recreational and work activities in the river environment. It is suggested that where runoff from non-flood conditions may confound water quality monitoring, application of the Bacteroidales host-associated PCR markers would be preferable to the more persistent E. coli. In addition, AC/TC testing should only be performed during baseflow conditions. The sequence information generated from the cowpat metagenomic study could be used for development of a metagenomic FST library of bacteria. Mobilisation rates of FST markers from cowpat runoff determined in this rural study can contribute to models designed to apportion contamination from agricultural sources.