Contamination Of Recreational Waters Research Articles

Rapid QPCR-based assay for fecal Bacteroides spp. as a tool for assessing fecal contamination in recreational waters

Concentrations of fecal indicator bacteria (FIB; e.g. Escherichia coli, and Enterococcus sp.) can only be used in limited ways for determining the source of fecal contamination in recreational waters because they cannot distinguish human from non-human fecal contamination. Several Bacteroides spp. have been suggested as potential alternative indicators. We have developed a rapid, culture-independent method for quantifying fecal Bacteroides spp. using quantitative PCR (QPCR) targeting the 16S rRNA gene. The assay specifically targets and quantifies the most common human Bacteroides spp. The details of the method are presented, including analyses of a wide range of fecal samples from different organisms. Specificity and performance of the QPCR assay were also tested via a laboratory experiment where human sewage and gull guano were inoculated into a range of environmental water samples. Concentrations of fecal Bacteroides spp., total Enterococcus sp., Enterococcus faecium, Enterococcus faecalis, and Enterococcus casseliflavus were measured using QPCR, and total Enterococcus sp. and E. coli were quantified by membrane filtration (MF). Samples spiked with gull guano were highly concentrated with total Enterococcus sp., E. coli, E. faecalis, and E. casseliflavus, demonstrating that these indicators are prominent in animal feces. On the other hand, fecal Bacteroides spp. concentrations were high in samples containing sewage and were relatively low in samples spiked with gull guano. Sensitivity and specificity results suggest that the rapid fecal Bacteroides spp. QPCR assay may be a useful tool to effectively predict the presence and concentration of human-specific fecal pollution.

Investigation of transcription-mediated amplification as a rapid test method for Enterococci in recreational water

Growth-based methods for detecting faecal contamination of recreational water require at least 24 hours to yield results, which can delay detection, action and remediation, if required. Such delays can put human health at risk while closing beaches inappropriately can cause unnecessary economic losses. There is a requirement for more rapid methods to facilitate the management of pollution events. In this study, undertaken in our respective laboratories, a molecular assay based on transcription-mediated amplification (TMA) technology was compared with established growth-based methods. The assays were used to quantitate enterococcal faecal indicator bacteria (relative to the legislative threshold of 104 colony forming units/100 mL) in 138 water samples collected from 41 different locations, representative of different recreational water types routinely sampled in Southern California. The results demonstrate the efficacy of the TMA assay for the detection of Enterococcus species in recreational water as a rapid alternative to traditional methods.

Genetic variability and pathogenicity potential of Escherichia coli isolated from recreational water reservoirs

Contamination of recreational waters and public water supplies by Escherichia coli represents a risk for public health, since some strains can be pathogenic or propagated with other pathogenic microorganisms. In this study, two reservoirs, Billings and Guarapiranga (São Paulo metropolitan area, Brazil), were investigated in order to assess E. coli diversity. Genetic typing using rep-PCR completely differentiated all strains and enabled the determination of their genetic variability. Although the same level of genetic variability was observed for strains originating from both reservoirs, randomization procedures showed that isolates from the same reservoir were more closely related to each other. Phylogenetic group frequencies in each reservoir suggested that contamination in the Billings reservoir was mostly from humans, whereas contamination in the Guarapiranga reservoir was mostly from animals. Colony blot experiments using probes from several virulence factor genes showed that both reservoirs contained potential pathogenic strains and may represent a risk to recreational or household usage of these water resources.

Rapid tests for detection and quantitation of Enterococcus contamination in recreational waters

Presently, growth-based tests are used for the detection and quantitation of microbiological contaminants in the environment. These tests take a minimum of 24 h to generate a result, which compromises the ability to take the most appropriate action. This report describes a rapid test for Enterococcus in recreational water as an indicator of faecal contamination. This method involves (1) isolation and lysis of the target organism, (2) purification of ribosomal RNA (rRNA) from the lysate and (3) amplification and detection of the purified rRNA. rRNA is used as the target since, in contrast to DNA, there are hundreds to thousands of copies in the cell. The rRNA is purified from the lysate by target capture onto magnetic microspheres, which removes interfering substances present in the sample. The rRNA is then quantitated using transcription-mediated amplification (TMA) with real-time homogeneous detection of amplicon using a fluorescent oligonucleotide probe. Compared to polymerase chain reaction (PCR) amplification, TMA is isothermal, more rapid, and ideally suited to RNA detection. The test described here demonstrates sensitive detection and quantitation of enterococci over a wide dynamic range with a high level of analytical specificity. The latter is particularly important for accurate and relevant monitoring both for protecting public health and for source tracking. Many conventional microbiological tests are time-consuming, exhibit limited dynamic range and are known to lack specificity. This assay demonstrates the advantages achievable by the application of TMA of rRNA targets to current environmental testing challenges.

Bacteroides spp. as reliable marker of sewage contamination in Hawaii's environmental waters using molecular techniques

Standard PCR (SPCR) and quantitative PCR (QPCR) assays using primers for general and for human-specific Bacteroides 16S rRNA markers were selected as the molecular tests to assess sewage contamination in recreational waters of Hawaii and these same water samples were assayed for culturable concentrations of selected faecal microbial indicators. The results of this study showed that the general primer for Bacteroides was not useful because ambient and polluted water samples were positive for this marker. However, use of human-specific primers reliably detected sewage contamination. The human-specific Bacteroides detection data supported previously reported conclusions that concentrations of alternative faecal indicators (C. perfringens, FRNA coliphages) but not traditional faecal indicators (faecal coliform, E. coli, enterococci) are reliable indicators of faecal contamination in Hawaii's environmental waters. The QPCR assay for the human-specific Bacteroides 16S rRNA marker was faster, more sensitive and more reliable than comparable SPCR assay because OPCR assay provided additional information such as melting temperatures, which confirmed that the right amplicons were being measured and Ct values, which indicated the relative level of faecal contamination.

Evaluation of Beach Grooming Techniques on Escherichia coli Density in Foreshore Sand at North Beach, Racine, WI

ABSTRACT Elevated levels of Escherichia coli(E. coli) in bathing waters at North Beach, a popular recreational site in Racine, Wisconsin, have been a persistent problem often resulting in the issuance of poor water quality advisories. Moreover, waterfowl (mostly Larus delawarensis and L. argentatus) in nearshore and offshore areas are common and may serve as non-point sources for bacterial contamination of recreational waters. Current beach management practice involves daily mechanical grooming of the nearshore sand for aesthetics and removal of hazardous debris. However, this practice has not been evaluated in terms of its effects on E. coli loading to beach sand and potential introduction to contiguous swimming water. In this study, we tested E. coli responses to three treatments: mechanical groomer, daily and twice weekly hand raking, and a control (no raking/grooming). A randomized block design consisted of replicated treatments and one control (10 each), for a total of 40 blocks sampled daily for 10 days. Foreshore sand samples were collected by hand coring to an average depth of 10 cm. Median E. coli recovered were 73 (mechanically groomed), 27 (hand-raked daily), 32 (hand-raked twice weekly), and 22 (control) colony-forming units (CFU) per gram dry weight sand. E. coli counts in sand that was groomed were significantly higher than hand rakings and control (p <0.0001), and there was no significant difference between control and raking treatments (p<0.01). This study demonstrates the beach management implications related to grooming efficacy and the importance of understanding non-point sources of bacterial contamination.

Detection of &lt;I&gt;Bacteroides&lt;/I&gt; species by Polymerase Chain Reaction (PCR) for Identifying Sewage Contamination in Recreational Water

Detection of &lt;I&gt;Bacteroides&lt;/I&gt; species by Polymerase Chain Reaction (PCR) for Identifying Sewage Contamination in Recreational Water

Diseases Associated with the use of Recreational Waters: Assessing Their Relationship to Microbiological Parameters

The evidence for the link presumed to exist between feecal contamination of recreational waters and the diseases suffered by users of such waters is reassessed. For minor diseases the causes of the symptoms are not clearly established and it is concluded that more research is needed before preventative measures can be realistically devised.