Search

• Fecal and wastewater bacterial communities were at low proportions in surf zones. • Recycled water bacterial communities were absent in surf zone waters. • HF183 copies in surf zones were mainly from human feces and treated WWTP effluent. • Bather shedding contributed low levels of human fecal contamination in surf zones. • Sequencing based fecal source proportions appeared uncorrelated to qPCR results. Fecal sources to recreational surf zone waters should be identified to protect public health. While watershed origins of human and other fecal sources are often discoverable by quantitative polymerase chain reaction (qPCR) of fecal markers using spatially stratified samples, similarly assessing wastewater treatment plant (WWTP) outfall and other offshore contributions to surf zones is challenged by individual marker fate and transport. Here, bacterial communities were assessed for relatedness between all hypothesized fecal sources and surf zone waters for two urban California recreational beaches, by sequencing genes encoding 16S rRNA and analyzing data using SourceTracker and FEAST. Ambient marine bacterial communities dominated the surf zone, while fecal (human, dog, or gull) or wastewater (sewage or treated WWTP effluent) bacterial communities were present at low proportions and those from recycled water were absent. Based on the relative abundances of bacterial genera specifically associated with human feces, the abundances of HF183 in bacterial community sequences, and FEAST and SourceTracker results when benchmarked to HF183, the major sources of HF183 to surf zone waters were human feces and treated WWTP effluent. While surf zone sequence proportions from human sources (feces, sewage and treated WWTP effluent) appeared uncorrelated to previously obtained qPCR HF183 results, the proportions of human fecal and potential human pathogen sequences in surf zone waters were elevated when there were more swimmers (i.e. during weekday afternoons, holidays and busy weekends, and race events), thus confirming previously-published qPCR-based conclusions that bather shedding contributed low levels of human fecal contamination. Here, bacterial community sequencing also showed evidence that treated WWTP effluent from an offshore outfall was entering the surf zone, thereby resolving a prior uncertainty. Thus, bacterial community sequencing not only confirms qPCR HF183-based human marker detections, but further allows for confirming fecal sources for which individual marker quantification results can be equivocal. [Display omitted] [ABSTRACT FROM AUTHOR]

Abstract: The State of California has mandated the preparation of a guidance document on the application of fecal source identification methods for recreational water quality management. California contains the fifth highest population of cattle in the United States, making the inclusion of cow-associated methods a logical choice. Because the performance of these methods has been shown to change based on geography and/or local animal feeding practices, laboratory comparisons are needed to determine which assays are best suited for implementation. We describe the performance characterization of two end-point PCR assays (CF128 and CF193) and five real-time quantitative PCR (qPCR) assays (Rum2Bac, BacR, BacCow, CowM2, and CowM3) reported to be associated with either ruminant or cattle feces. Each assay was tested against a blinded set of 38 reference challenge filters (19 duplicate samples) containing fecal pollution from 12 different sources suspected to impact water quality. The abundance of each host-associated genetic marker was measured for qPCR-based assays in both target and non-target animals and compared to quantities of total DNA mass, wet mass of fecal material, as well as Bacteroidales, and enterococci determined by 16S rRNA qPCR and culture-based approaches (enterococci only). Ruminant- and cow-associated genetic markers were detected in all filters containing a cattle fecal source. However, some assays cross-reacted with non-target pollution sources. A large amount of variability was evident across laboratories when protocols were not fixed suggesting that protocol standardization will be necessary for widespread implementation. Finally, performance metrics indicate that the cattle-associated CowM2 qPCR method combined with either the BacR or Rum2Bac ruminant-associated methods are most suitable for implementation. [Copyright &y& Elsevier]

Abstract: A number of PCR-based methods for detecting human fecal material in environmental waters have been developed over the past decade, but these methods have rarely received independent comparative testing in large multi-laboratory studies. Here, we evaluated ten of these methods (BacH, BacHum-UCD, Bacteroides thetaiotaomicron (BtH), BsteriF1, gyrB, HF183 endpoint, HF183 SYBR, HF183 Taqman®, HumM2, and Methanobrevibacter smithii nifH (Mnif)) using 64 blind samples prepared in one laboratory. The blind samples contained either one or two fecal sources from human, wastewater or non-human sources. The assay results were assessed for presence/absence of the human markers and also quantitatively while varying the following: 1) classification of samples that were detected but not quantifiable (DNQ) as positive or negative; 2) reference fecal sample concentration unit of measure (such as culturable indicator bacteria, wet mass, total DNA, etc); and 3) human fecal source type (stool, sewage or septage). Assay performance using presence/absence metrics was found to depend on the classification of DNQ samples. The assays that performed best quantitatively varied based on the fecal concentration unit of measure and laboratory protocol. All methods were consistently more sensitive to human stools compared to sewage or septage in both the presence/absence and quantitative analysis. Overall, HF183 Taqman® was found to be the most effective marker of human fecal contamination in this California-based study. [Copyright &y& Elsevier]