Recreational waterways are frequent leisure destinations for families and important economic resources for many coastal communities [1
]. Keeping visitors safe from harmful water pathogens and preventing adverse health outcomes is an important public health focus. Heavy rains can cause a significant rise in water level of creeks and streams, potentially carrying high levels of bacteria and other pollutants to the ocean, adding potential sources of fecal indicator bacteria (FIB) and surface runoff [2
]. These bacteria and pollutants may be present at recreational waterways and can cause recreational water illness (RWI). RWI is caused by swimmers swallowing water that is potentially contaminated by a variety of pathogens that can cause diarrhea, skin disease, or other potentially serious illness [3
]. In the United States and other countries, E. coli
(EC) and enterococci (ENT) bacteria, commonly called FIB, are often used to regulate beaches [4
]. The United States Environmental Protection Agency (USEPA) recently published new guidelines for EC and ENT that aim to keep the risk of gastrointestinal (GI) illness in swimmers below approximately 30 illnesses per 1000 swimmers (2012 RWQC) [5
]. In addition, there are epidemiology studies that link FIB to illnesses in swimmers at coastal beaches [6
]. Current protocols for monitoring and alerting the public of adverse water conditions to protect human health are limited, i.e., sample variability, no source determination, and 24 h for results. However, new laboratory methods are being developed and their use continues to show promise in adding depth to the current testing methodologies [7
There are many potential sources for fecal waste contamination in recreational waterways, from both human and non-human sources [9
]. The risks associated with these sources can fluctuate with environmental conditions such as rainfall. Storm water runoff can introduce animal fecal contamination. However, the risk to human health is often an order of magnitude lower than if it were from a human source [10
]. Gulls have also been identified as a potential source of fecal contamination, which can often be variable due to migration and the varied mobility of gull populations. The transient nature of gulls poses a difficult challenge as they have been shown to successfully transport fecal contamination from human waste sites and transport bacteria to beaches, thus increasing the risk to visitors [11
]. Dogs are also frequent visitors to beaches and contribute to fecal contamination [12
]. These studies show that fecal contamination can be complex and mixed with both human and non-human fecal sources, increasing potential risk of illness to visitors.
FIB such as EC and ENT are variable in monitoring recreational water and have limited capacity to discern human from non-human sources [4
]. Despite their limitations, detection of FIB remains the gold standard in recreational water monitoring. Determining the source of measured spikes of FIB and correlating them with an epidemiological study allow for better understanding of recreational waterways and possible links to adverse health outcomes [13
]. Microbial source tracking (MST) describes a suite of methods and investigative strategy to determine the source of fecal contamination in environmental water that depends on the association of certain fecal microorganisms with a particular host. MST field studies are being performed with more frequency to help identify the source of fecal contamination [12
]. These methods can detect host-associated Bacteroides
bacteria or other microorganisms from humans and other animals and link them to GI illness in recreational water [15
]. Inter-laboratory variation has been identified within MST studies; therefore, standardization of protocols and consumable reagents is recommended for repeatability and reproducibility [17
]. Few public health laboratories have fully adopted MST methods as part of routine beach management strategies, even though they show promise in helping control hazards to beachgoers.
The federal Clean Water Act (CWA) is the law that regulates pollution of United States waterways. Section 303(d) of the CWA requires states like California to identify surface waters where currently applied pollution control technologies cannot meet the water quality standards of the waterbody [18
]. Clam Beach is listed on the Section 303(d) list of impaired waterways for high bacteria levels. Clam Beach commonly exceeds the single sample maximum (SSM) for beach closures and advisories conducted by the Department of Health Services [19
]. Additionally, Clam Beach near Strawberry Creek was selected for this project because FIB levels have recently given this popular beach a grade of “F” on the Heal the Bay’s Annual Beach Report Card, a ranking of the 10 most polluted beaches in the State and earned the No. 2 spot on California’s Top Ten Beach Bummers List, up from the No. 4 spot the previous year [20
]. The greater Strawberry Creek watershed, comprised of Patrick Creek, Strawberry Creek, and Rose Creek, directly influences the density and potential source(s) of FIB levels at Clam Beach. This region has a variety of land uses that include agricultural, industrial, timberlands, public, and residential, all of which potentially influence fecal waste at Clam Beach. As part of a pilot project, during the dry weather beach monitoring season, which is April through October annually, a total of 91 FIB and 103 MST samples were collected and analyzed at Clam Beach from August 2015 to April 2018, which showed that birds were the most likely source of fecal contamination (Table S1
). The study presented here provides further insight into the potential source(s) and distribution of fecal waste at Clam Beach and Strawberry Creek watershed by conducting a beach study and watershed study. Additionally, it attempts to integrate these MST methods into the daily public health laboratory workflow and beach monitoring protocols.
Several approaches were used to help identify the potential source(s) of fecal contamination to better understand Clam Beach near Strawberry Creek. For the pilot project, the 3 years’ worth of weekly sample data indicated that birds were the most frequent source consistently detected throughout the year. As a result, we designed a short-term, targeted study at Clam Beach that showed frequent bird marker detections with no detectable human or ruminant sources. This suggests that failing septic systems or pastured cattle, wild deer, and elk are not likely to be a major contributor to fecal contamination. Nearly half (50%) of the FIB samples collected at Clam Beach near Strawberry Creek during the ocean beach study period exceeded the SSM objectives, which would have prompted posting a beach advisory and subsequent re-testing until levels returned to acceptable levels. This location is meeting the TC objective. However, there were resamples due to exceedances of EC and ENT. Only two (17%) of the samples collected at Clam Beach near Mad River (site 1) would have resulted in a resample, which was due to exceedance of the TC SSM objective. Strawberry Creek had a higher density of EC and ENT bacteria density when compared to Mad River, which could be attributed to the size of the streamflow. It should be noted that during the watershed study, no sample exceeded any FIB SSM objective at site 1. In addition, site 2a had only two (17%) exceedances due to ENT, which would have resulted in a resample. Mad River’s flow is significantly larger than Strawberry Creek, which is perhaps flushing the bacteria levels further out into the ocean.
In the beach study, the second most-abundant MST marker detected was the dog marker (9%) but none of the detections were quantifiable. Throughout the day several dogs were observed, which was consistent with the low level of dog MST marker detections, none of which appeared to pose any human health risk since all six samples detected were BLOQ. The most abundant MST marker detected was the bird marker (65%), with a little less than half of the samples in the quantifiable range. In addition, no sample had co-occurring bird and human marker, which would have lowered the potential risk threshold [7
]. Despite the high frequency of bird marker detections, no sample exceeded the QMRA derived criteria of 4 × 106
copies per 100 mL, which correlates to the USEPA benchmark of 30 GI illnesses per 1000 swimmers [5
]. Despite the strong presence of bird marker there was no association with any of the FIB, which is consistent with previous studies [27
]. The day of this study, both Strawberry Creek and Mad River beaches showed striking visible influence from birds.
Environmental factors play an important role in FIB concentration and can influence the density throughout the entire day, which was shown in this study by the fluctuating detected FIB concentrations [4
]. The laboratory data produced were consistent with the literature on known spatial and tidal patterns of fecal contamination [28
]. The detection of significantly higher TC density in the early morning was consistent with the known effects of sunlight on FIB concentration in water [30
]. Tides also play a critical role in FIB levels, making it extremely challenging to evaluate true health risk. As with many other studies, TC and ENT densities were higher at low tide when compared to high tide [31
]. Additionally, TC density was higher during outgoing tide when compared to incoming tide, highlighting the complex nature of beach sampling and monitoring. Birds influence fecal contamination at Clam Beach, but sample location plays an important factor in bird marker detections. The samples taken in the mixing zone (site 2a) had much higher bird marker density when compared to the sample taken in Strawberry Creek (site 2b), which was freshwater. While sampling, thousands of birds were observed bathing between the two sample locations: upstream of 2a and downstream from 2b. In addition, there was one dead bird found between the sampling locations, which may have contributed to the findings of this study. It also highlights the importance of taking a strategic approach to sampling, especially if health risk is substantial or regulatory decisions are being based on these data [32
The watershed study was used to determine if the Strawberry Creek watershed was contributing directly to the increased FIB levels found at Clam Beach. Potential sources varied by sample location. The Mad River site 1 was the only sample location that showed any influence from birds, which is consistent with the Beach Study. The Strawberry Creek watershed did not show any influence from birds. The only site that showed influence from dogs was the footbridge (site 5). Interestingly, one sample collected at this site exceeded all three FIB markers and had 4200 copies/100 mL of the dog marker detected. There were two quantifiable human detections: one at site 8 (Patrick Creek) and one at site 11 (the pond). Both detections were at the water quality threshold of 525 copies/100 mL, indicating potential health risk [7
]. However, these two sites are distant from Clam Beach and did not appear to affect this location. Furthermore, Patrick Creek showed higher ENT when compared to the other creeks, suggesting further study of this creek is necessary. The most important finding was the consistent ruminant detections throughout the Strawberry Creek watershed and the Clam Beach near Strawberry Creek site 2a. Ruminants appear to be more influential than originally thought, which is not surprising due to the land use in this region. MST methods are still evolving. In this study, we were successful in automating the extraction process, which was key in being able to integrate these methods in a public health laboratory setting. Moreover, future research could focus on improving the methodology, making it easier to adapt in a variety of laboratory settings. First, bridging extraction and PCR platforms to these methods will make adoption more palatable if labs can add these methods without significant capital investment for equipment. Using familiar equipment and similar workflows is attractive to labs considering adding to their testing menu. Second, improving the extraction control by blending the human, ruminant, dog, and bird marker into one calibrator sample with known quantities to serve as a single extraction control is needed. It would improve quality control by minimizing sample controls while lowering cost. Lastly, advancing these molecular assays into a multiplex option would greatly improve these methods and cut the work and time by 25%. Results in real time are needed to make this a practical solution to water quality management. The way the assays are designed now, multiple thermocyclers are needed to acquire data that are actionable. In the future, the use of MST multiplex assays with similar sensitivity would significantly reduce both reagent costs and technical time expenses. This would make it easier for labs to integrate these protocols into existing workflows.
Clam Beach near Strawberry Creek has been consistently identified for having high FIB detections and has been publicly named as one of the top 10 “bad beaches” in California. Routine sampling often leads to beach postings and warnings to the general public. In addition, these SSM exceedances lead to resamples and follow-up laboratory testing until the levels return to acceptable limits. This approach to beach monitoring can result in the use of more resources and often unclear answers to actual human health risk. With MST added to the current analysis we can provide a better assessment of human health risk and potential sources of fecal contamination. However, they often do not correlate to FIB levels. Our data indicated that failing septic systems are not likely to be the source of fecal contamination and demonstrated that the high concentrations of bird fecal matter at Clam Beach is likely due to a natural population of nearshore birds regularly observed along Clam Beach.
MST methods were used, which identified birds as a potential vector for human pathogens, potentially increasing human risk for illness [11
]. This is important because the median risk of illness using HF183 (human fecal marker) is three orders of magnitude less than similar CAT (bird fecal marker) concentrations [15
]. Understanding the source of FIB exceedances is invaluable information for recreational water managers to more accurately evaluate the human health risks. In this study, we showed that the FIB exceedances were most likely due to bird sources. Therefore, the probability of illness is lower than if the source were human. Since the potential source was determined to be bird, it is reasonable to conclude that risks to human health are within the regulatory illness risk benchmark of approximately 30 illnesses per 1000 recreators [5
]. Clam Beach is frequented by birds, which adds to the allure of its location and should be celebrated not feared. Perhaps in the future, the grading methodology used to create the “Beach Bummer List” could take into account MST data when evaluating human health risk.