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Article

Application of Same-Day Enterococcus qPCR-Based Analyses for Quality Assessment of Shorelines (Water and Sand) at Recreational Beaches

by
Faizan Saleem
,
Thomas A. Edge
and
Herb E. Schellhorn
*
Department of Biology, McMaster University, Hamilton, ON L8S 4K1, Canada
*
Author to whom correspondence should be addressed.
Water 2023, 15(13), 2338; https://doi.org/10.3390/w15132338
Submission received: 11 April 2023 / Revised: 11 June 2023 / Accepted: 19 June 2023 / Published: 24 June 2023
(This article belongs to the Special Issue Modern Methods for Analysis of Water and Related Environmental Samples)
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Abstract

:
Rapid water-quality monitoring methods for beach water and sand can be helpful for public health authorities to develop comprehensive beach monitoring programs. In this study, we evaluated the performance of the Enterococcus qPCR (USEPA 1609.1) method for quality monitoring of beach water and sand-porewater at two Niagara region beaches in Ontario, Canada (Lakeside and Sunset Beaches). While the USEPA 1609.1 method has been validated for beach water, its potential for assessing associated beach sands (which may function as a microbial reservoir) has not been fully explored. All beach water samples (n = 168) passed the qPCR quality control (QC). However, only 20 out of 48 (41.7%) sand-porewater samples passed the qPCR QC, potentially due to interference by soluble inhibitors. The proportion of the sand-porewater samples passing QC improved slightly to 63 out of 120 (52.5%) with a prefiltration step to remove sand and other large particles. The faecal indicator density in the sand-porewater, tested in parallel, did not correlate with the beach water faecal indicator density. Comparing beach water data for the same-day Enterococcus qPCR threshold with the previous-day E. coli culturing-based threshold across all beach days tested, Enterococcus qPCR analyses identified 3 (7%) and 7 (16%) false positive/lost beach days for Lakeside and Sunset Beaches, respectively. Additionally, of the total beach days tested, Enterococcus qPCR analyses identified 2 (5%) and 1 (2%) false negative/health-risk days for Lakeside and Sunset Beaches, respectively. Sand-porewater testing analyses identified days when faecal indicators (in the sand) exceeded beach water quality thresholds. Compared with conventional E. coli culturing, use of the same-day qPCR method would result in fewer beach postings and could identify several additional health-risk days (when the beaches may not be posted). Future studies could include additional prefiltration steps or modifications in the Enterococcus qPCR protocol to improve the method’s applicability for sand quality monitoring.

1. Introduction

Water quality for Ontario freshwater beaches is currently determined using conventional E. coli culturing-based enumeration methods to mark the beach water quality as safe or unsafe for recreational purposes [1]. Two major practical issues associated with beach monitoring programs are (1) Delayed results associated with culturing-based methods and (2) Lack of routine sand quality monitoring [2,3]. Faecal indicator levels in freshwater beaches can change overnight [4], and thus the faecal indicator bacteria (FIB) counts based on samples collected on the previous day may not be a good indicator of the next day’s beach quality [5,6]. Additionally, beachgoers’ recreational use may involve direct contact with beach water and sand, whereas public health agencies currently actively test beach water quality only [7,8]. If a beach is posted to prevent swimming, this may result in more beachgoer time spent onshore and more human exposure to beach sand.
Beach sand can be a reservoir for faecal indicators and pathogenic bacteria [9,10,11,12]; bacterial accumulation and long-term persistence are possible because of the sand’s ultraviolet and temperature shielding effects [3]. Epidemiology studies [13,14] also indicate that the direct contact of beachgoers with shore sand positively correlates with gastrointestinal disease morbidity. In addition, the wave action in beach water may mobilize sand-particle-attached bacteria, which can be transported to the adjacent shoreline, resulting in lower beach water quality [4]. However, beach sand monitoring is not typically performed, and when it is, it mainly relies on culturing-based 24 h-delayed methods such as Colilert® and Enterolert® (IDEXX, Westbrook, ME, USA) [15]. Therefore, incorporating sand quality assessment methods may result in more effective, improved beach quality monitoring strategies.
Culturing-based methods for faecal indicator bacteria have inherent limitations for assessing beach water quality and may not be suitable for dormant or non-culturable enterococci [16]. Compared with culturing-based beach monitoring, qPCR-based methods, such as the USEPA 1609.1 Enterococcus testing protocol, are much more robust and can provide results to the beach managers within 3–4.5 h of sample collection [5,17]. In addition, qPCR-based methods can more comprehensively estimate faecal indicator densities by quantifying the culturable and non-culturable indicators [15]. National Environmental Epidemiological Assessment of Recreational Water (NEEAR) studies indicate that qPCR-based enterococci measures are a potentially more reliable indicator of gastrointestinal diseases among beachgoers [18,19]. While the applicability of the qPCR-based method has been tested for beach water, it is not generally used for monitoring beach sands (FIB may be transported onto beaches by wave action or rainwater runoff). Compared with beach water, sand can potentially harbour higher levels of qPCR inhibitory compounds, such as humic acids, that can inhibit PCR reactions [2]. Testing the applicability of the qPCR-based method for more rapid monitoring of both beach water and beach sand may inform public health agencies in revising beach monitoring regimens.
Despite the growing adoption of rapid beach monitoring methods [4,20], many public health agencies still depend on 24 h-delayed E. coli culturing from water samples, and sand-porewater is not routinely monitored. Our study builds on previous studies on the microbial analysis of important freshwater beaches [5,6,10] and focuses on the applicability/comparative analysis of the Enterococcus qPCR-based method for both beach water and sand environments at Niagara beaches. Compared with the Toronto beaches (Marie Curtis Park East and Sunnyside Beaches) [5,6], the Niagara beaches (Lakeside and Sunset Beaches) are posted less frequently and are not as heavily contaminated by industrial/faecal contamination sources. For this study, we selected the Niagara beaches as part of our goal to extend testing of the applicability of an Enterococcus qPCR-based beach water testing method for different geographical locations (with different sources or levels of contamination) and different environments (shore water and sand). Objectives for our study were: (1) Testing the applicability of the USEPA 1609.1 Enterococcus protocol for assessing beach water and sand quality at two different Niagara region beaches, (2) Comparative analysis of the FIB densities in beach water and sand between beach open and posted days, (3) Analysis of beach posting outcomes using Enterococcus qPCR compared with Public Health’s E. coli culturing data, and (4) Testing the correlation between the FIB densities (as measured by culture and qPCR) and environmental variables such as temperature and wave height.

2. Materials and Methods

2.1. Study Plan

Our study design included beach water and sand-porewater sampling of two Niagara beaches on Lake Ontario in Ontario, Canada (Lakeside and Sunset Beaches). The sample collection plan included the 2022 summer season from 31 May 2022 to 1 September 2022. Figure 1 and Supplementary Table S1 show the geographical locations/coordinates of the sampling sites corresponding to Lakeside and Sunset Beaches. Water and sand samples were collected from two sampling sites for each beach (LK3 and LK5 for Lakeside Beach; SS3 and SS5 for Sunset Beach). Samples were collected in the morning between 8 a.m. and 9 a.m. for three consecutive days a week and were delivered to the lab for processing before 10 a.m. For each sampling day, eight samples were collected, including four samples from beach water and four from beach sand-porewater. The beach season for summer 2022 consisted of 94 beach days from 31 May 2022 to 1 September 2022. In total, 336 samples (168 beach water samples and 168 sand-porewater samples) corresponding to 42 beach days were collected and processed for the 2022 summer season.

2.2. Water Sample Collection and Filtration

Beach water samples were collected in sterile 1000 mL polyethene terephthalate (PET) water bottles 30 cm below the water surface at a water depth of 1.0 m. For sand-porewater samples, a 30 cm × 30 cm × 30 cm hole was dug 1 m inland from the lake in the foreshore sand region of each sampling site, followed by water collection (500 mL) by allowing surrounding groundwater to seep into the bottom of the hole (Figure 2) [10]. After the sample receipt in the lab, a 100 mL beach water sample was processed through a 0.45 µm polycarbonate membrane filter (Millipore Corp., Bedford, MA, USA). For beach sand-porewater, initially (31 May 2022 to 30 June 2022), a 50 mL water sample was directly passed through a 0.45 µm membrane filter. For the samples from July 01 onwards, 50 mL sand-porewater samples were first passed through a prefiltration assembly made by enclosing a 50 µm nylon filter (Dynamic Aqua Supply, Surrey, BC, Canada) inside a plastic garden mesh (Supplementary Figure S1) to remove larger sand and dirt particles, followed by passing the filtrate through a 0.45 µm filter.

2.3. E. coli Culture and Enterococcus qPCR

For E. coli culture enumeration from sand-porewater samples, 1:10 dilutions were prepared as required before filtration. Each sand-porewater sample was filtered in triplicate, and membrane filters were placed on differential coliform agar plates (OxoidTM), followed by incubation for 24 h at 44.5 °C. Enterococcus qPCR was performed as described in the USEPA 1609.1 method: Enterococci in Water by TaqManTM Quantitative Polymerase Chain Reaction (qPCR) with Internal Amplification Control (IAC) Assay [17], as recently validated [5,6]. In brief, the qPCR assay included calibrator positive control, internal amplification control (to check for qPCR inhibition), sample processing control (salmon sperm DNA qPCR to check for extraction efficiency), water matrix spike (to assess for interference effects from the water matrix), and non-template control (to check for contamination) as quality controls. DNA extraction for the standard curve was performed using Norgen soil plus DNA extraction kit (Norgen Biotek, Ontario, Canada) while, for routine sample analysis, the standard DNA extraction protocol was used as described in the USEPA 1609.1 method [17]. Four individual standard curves ranging from 40,000 to 10 target sequence copies (TSC)/5 µL were analysed using Enterococcus faecalis (ATCC 2921) cell suspension (109 cells) to obtain a composite standard curve. For each sampling day, two calibrator positive controls, two method blanks, and one non-template control were analysed along with the beach water and sand-porewater samples. All the samples and quality controls were analysed in duplicate on the Bio-Rad CFX96 Touch Real-Time PCR Detection System thermocycler (Bio-Rad Inc. Mississauga, ON, Canada).

2.4. Data Analysis

Calculations for qPCR inhibition, DNA recovery, and calibrator cell equivalents (CCEs/100mL) were performed using a USEPA 1609.1 Excel sheet (https://www.epa.gov/cwa-methods/other-clean-water-act-test-methods-microbiological#file-183743, accessed on 1 June 2022). The arithmetic means of the density of Enterococcus calibrator cell equivalents were calculated using counts from both sampling sites for each beach. The E. coli culturing counts from the sand-porewater samples included calculating the arithmetic mean of three replicates for each sample and two sampling sites for each beach (3 replicates/sample × 2 sampling sites/beach). The current Health Canada and Ontario quality threshold for E. coli culturing (beach water) is 200 CFUs/100 mL with a single-sample max of 400 CFUs/100 mL [8]. For Enterococcus qPCR and E. coli culturing, the Beach Action Value and water quality thresholds used for data interpretation were 1000 calibrator cell equivalents (CCEs)/100 mL and 200 colony-forming units (CFUs)/100 mL, respectively. For the sand quality comparison, we used faecal indicator beach water quality thresholds due to the unavailability of quality thresholds for beach sand. Culturing and qPCR data were log-transformed for correlation analysis and analysed for normality distribution using the Shapiro–Wilk test (http://www.sthda.com/english/wiki/correlation-test-between-two-variables-in-r, accessed on 15 September 2022). The Pearson correlation with a 95% confidence interval and p < 0.01 was used to evaluate the degree of correlation between the environmental variables and faecal indicator counts. We also obtained E. coli data and environmental variable data such as wave height, air/water temperature, and wind speed from Niagara Public Health (referred to as Public Health in the later sections). The beach postings comparison analysis was performed using Public Health’s beach posting decisions for the summer of 2022 based on an E. coli geomean of samples at a beach. Retrospective same-day E. coli data from Public Health enabled an assessment of the hypothetical possibility of obtaining culturing results on the same day as sample collection. For the beach postings comparison, beach days were categorized as false negative days when the beaches were not posted according to E. coli culturing and the same-day Enterococcus qPCR results identified Beach Action Value threshold exceedances. Beach days were categorized as false positive beach days when the beaches were posted according to E. coli culturing and the same-day Enterococcus qPCR results did not identify Beach Action Value threshold exceedances. Due to common qPCR quality control failures, beach posting comparisons and correlations using sand-porewater qPCR results often included enterococci calibrator cell equivalents from a single sampling site for each beach (no sampling site replicate). All the plots were generated using the ggplot2 package in R [21].

3. Results

3.1. qPCR Quality Control Analysis

The coefficient of determination (R2) for the composite standard curve was 0.998, while the slope and intercept values were observed as −3.28 and 39.55, respectively (Table 1). In total, 336 water samples were collected/processed, including 168 samples each for beach water and sand-porewater, corresponding to 42 beach sampling days. All the beach water samples passed the quality control criteria recommended in the USEPA 1609.1 protocol [17]. For sand-porewater samples processed without nylon mesh prefiltration, the quality control (QC) qPCR pass rate was only 20 of 48 (41.7%). The pass rate improved slightly with nylon mesh prefiltration in 63 out of 120 (52.5%) (Table 1). In comparison with beach water and sand-porewater samples that passed the QC criteria, most (74/85; 87%) of the QC failed samples showed a highly significant (p < 0.01) increase in the sample processing control Ct value (Supplementary Figure S2).

3.2. Association between Sand-Porewater and Beach Water Faecal Indicator Densities

No significant correlation (p > 0.05) was observed between the sand-porewater and beach water Enterococcus calibrator cell equivalents (CCEs) or between the sand-porewater E. coli colony-forming units (CFUs) and beach water Enterococcus CCE (Supplementary Figures S3 and S4). However, there was a significant positive correlation (p < 0.01, R = 0.5–0.6) between the sand-porewater Enterococcus CCEs and E. coli CFUs (Supplementary Figure S5). Among the environmental variables, there was a significant positive correlation between Enterococcus CCE and wave height (Supplementary Figure S6), while there was no significant correlation between either air/water temperature or wind speed and the faecal indicator counts.

3.3. Comparison of Same-Day qPCR-Based Monitoring Method with 24 h-Delayed E. coli Enumeration by Culture for Beach Posting Outcomes

According to the E. coli CFUs data for summer 2022 (Public Health’s data), out of our 42 tested beach days, Lakeside and Sunset Beaches were posted for 3 and 13 beach days, respectively. Compared with Public Health’s data, our same-day Enterococcus qPCR identified considerable differences in the beach postings. For Lakeside Beach (Figure 2), of 42 tested beach days, our same-day Enterococcus qPCR results identified three (7%) false positives, or lost beach days, and two (5%) false negatives, or health-risk days (Figure 3). For Sunset Beach (Figure 4), our same-day Enterococcus qPCR results identified seven (16%) false positives, or lost beach days, and one (2%) false negative, or health-risk day (Figure 5).
Same-day Enterococcus qPCR provided results for 27 and 32 beach days for sand-porewater samples from Lakeside and Sunset Beaches, respectively. Of the 27 beach days for Lakeside Beach, same-day qPCR identified 7 (26%) beach days when Enterococcus density in beach sand was higher than the USEPA Beach Action Value for beach water quality (BAV; 1000 CCEs/100 mL) and the beach remained open for recreational activities (Figure 6 and Supplementary Table S2). For Sunset Beach, same-day Enterococcus qPCR identified 13 (41%) beach days when the beach was not posted and sand Enterococcus levels exceeded the USEPA BAV for beach water quality (Figure 7 and Supplementary Table S2). Enterococcus qPCR for sand-porewater also identified two (7%) and three (9%) beach days for Lakeside and Sunset sand-porewater, respectively, when the beaches were posted (while Enterococcus densities remained below BAV) (Figure 6 and Figure 7).
E. coli culturing results were obtained for all 42 tested beach days. For sand-porewater from Lakeside Beach (Figure 8 and Supplementary Table S2), 25 (60%) beach days were identified with retrospective same-day E. coli density in sand higher than the Ontario Public Health posting threshold for beach water (200 CFUs/100 mL), and the beach remained open to the public. For sand-porewater samples from Sunset Beach (Figure 9 and Supplementary Table S2), we observed 22 (53%) beach days when retrospective same-day E. coli density in the sand was higher than the Ontario Public Health posting threshold for beach water and the beach remained open to the public. E. coli culturing also identified one (2%) and three (7%) beach days for Lakeside and Sunset Beaches, respectively, when beaches were posted while sand-porewater did not show BAV exceedance.

4. Discussion

Current beach monitoring practices rely heavily on culturing methodology, which has the well-known limitation of delayed data availability [9]. However, faecal indicator densities change within 24 h [4], and results obtained from the previous day’s samples may not reflect the change in water quality. Rapid molecular methods, including Enterococcus qPCR, can provide results for beach quality monitoring within 3.5–4 h and can overcome the delay associated with culturing-based methods. This study continues our goal to evaluate the use and applicability of rapid molecular methods, including Enterococcus qPCR (USEPA 1609.1), for rapid quality assessment of different environments (beach water and sand) and geographical locations. Prior use of the Enterococcus qPCR-based method identified additional beach open/health-risk days for the Toronto beaches (Marie Curtis Park East and Sunnyside Beaches) and that the beach sands are potential reservoirs of bacterial pathogens [5,6,10]. To further demonstrate the adaptability and applications of the Enterococcus qPCR-based method, in this study, we expanded our sampling regime to the Niagara beaches (Lakeside and Sunset Beaches), which have less impact from high-density human activity compared with the Toronto beaches. We systematically analysed sand-porewater as often as beach water for the same beach days to provide a comparative estimate of the faecal indicator densities between the two environments.
The Enterococcus qPCR analyses for the Niagara beaches revealed that all the beach water samples passed the USEPA 1609.1 qPCR quality control (QC) parameters (no significant qPCR inhibition or poor DNA recovery), and the results were reported to Public Health within 3.5 h after sample receipt in the lab. This was similar to our validation study at two Toronto beaches [5,6], further supporting the applicability of the same-day qPCR-based method for water quality monitoring at Ontario beaches. However, there remain challenges in adapting USEPA method 1609.1 for assessing sand-porewater quality. For sand-porewater samples, with the inclusion of the prefiltration step using 50 µm nylon mesh, we observed a higher proportion of samples passing the qPCR QC. Most sand-porewater samples that failed the qPCR QC criteria showed significantly higher Ct values for the sample processing control than the beach water samples, possibly due to DNA loss during the extraction (bead beating) step. Coral sands have been identified to interfere with DNA extraction using rapid DNA extraction protocols [2]. Similar to the coral sands, our results indicate that using the rapid DNA extraction protocol (USEPA 1609.1) in the presence of sand particles can result in poor DNA recovery. However, the Enterococcus qPCR method can be modified further for sand quality monitoring by acidifying the water samples before filtration [2] or including additional prefiltration steps before sample processing.
Although beach sand can be a reservoir for faecal indicators and pathogenic bacteria [10,11,12], beach monitoring protocols of public health agencies currently rely on beach water quality analysis. In addition, factors such as erosion, rainwater runoff, and wave swash allow the bacterial communities in beach sand to enter beach water. Thus, beach sand can serve as a non-point source of faecal contamination [22,23]. Our results indicate little correlation between the samples from the different environments (beach water vs. sand faecal indicator densities). Similar to our study on Toronto beach waters [5], a significant positive correlation was observed between retrospective same-day E. coli culturing and Enterococcus qPCR for beach water samples taken from the two Niagara beaches. In this Niagara study, these two beach monitoring methods were also correlated with sand-porewater samples. However, additional research is needed to understand better the potential for health risks associated with faecal indicator bacteria in beach sand, as epidemiology studies have largely focused on beach water. Additional research is also needed to further advance the methods and safety thresholds for assessing faecal indicator bacteria levels in beach sand, as there are no standard field sampling and lab processing methods for beach sand.
Enterococcus qPCR analyses identified additional beach loss days and health-risk days for both Lakeside and Sunset Beaches. In addition, the E. coli culturing method posted the beaches more frequently than the Enterococcus qPCR. Our results concur with a Chicago beaches (U.S.) study [4], which concluded that there is no better than a 50% chance of agreement between the previous-day’s (first sampling day in two consecutive beach days) and next-day’s (second sampling day in two consecutive beach days) faecal indicator densities, and that culturing-based methods can generate about three times more beach postings. Faecal indicator densities in recreational waters can change markedly within hours [4,24], leading to the potential for less reliable beach posting decision-making if the 24 h-delayed culturing-based methods are used alone for beach monitoring.
Comparing beach sand-porewater faecal indicator densities (Enterococcus qPCR and E. coli culturing) with Public Health’s beach postings revealed that a substantial number of our beach testing days had higher faecal indicator densities in the beach sand than the shoreline water. Noticeably higher faecal indicator densities in sand-porewater than in beach water suggest that faecal contamination sources may significantly impact the sand but have less or no contact with neighbouring beach water [25]. Similarly, studies performed from beaches in the U.S. [26,27], Australia [28], and France [29] have demonstrated the pattern of comparatively higher faecal indicator densities in beach sand than in shoreline water. Alternatively, wave-induced resuspension of sand bacteria into beach water can deteriorate the water quality near the beach shoreline [30,31] and, for some beaches, can serve as a source of enterococci dissemination into beach water [7,32]. Therefore, standard water quality testing at a distance out from the shoreline may not provide real-time or comprehensive information about faecal bacteria dynamics for a complex beach water/sand interface.
This study identified differences in beach posting interpretations using the same-day Enterococcus qPCR method compared with Public Health’s 24 h-delayed E. coli culturing-based results. Differences in beach posting outcomes might be particularly due to changing water conditions over the 24 h delay associated with the culturing-based E. coli method. The significance of these differences could have been much higher if our sampling had been performed for the whole summer and extended to all 19 Niagara region beaches (www.niagararegion.ca/living/water/beaches/default.aspx, accessed on 15 September 2022) Our results provide further support for public health agencies to consider adopting the same-day USEPA Enterococcus qPCR-based method for beaches.

5. Conclusions

  • The Enterococcus qPCR-based beach monitoring method provided same-day results within 3.5 h of sample processing for the Niagara beaches (Lakeside and Sunset Beaches).
  • There was no correlation between the sand-porewater and beach water faecal indicator densities for either the culturing- or qPCR-based methods.
  • Of the 33 beach days tested, Enterococcus qPCR analyses identified 3 (7%) and 7 (16%) false positive/lost beach days for Lakeside and Sunset Beaches, respectively.
  • Sand-porewater testing indicated that up to 60% of the tested days exceeded the beach water quality posting thresholds. However, beach water quality thresholds may not be applicable for sand-porewater quality, and therefore further evaluation is required to identify suitable safety thresholds for sand-porewater.
  • While a prefiltering screen improved the Enterococcus qPCR method by ~10% for assessing Enterococcus levels in beach sand-porewater, additional research is required to advance the application of this method for evaluating beach sands.
  • The Enterococcus qPCR-based method provides rapid/robust beach water quality assessment and can augment current beach monitoring methods.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/w15132338/s1. Figure S1: Collection of sand-porewater samples from foreshore sand regions of sampling sites. (a) Hole for sand-porewater collection, (b) Seeped groundwater inside sand-porewater collection hole, (c) Sand-porewater collected in 500 mL sterile PET bottles, and (d) Prefiltration assembly with 50 µ nylon filter enclosed in plastic mesh for sand-porewater samples. Figure S2: Comparison between sample processing control (SPC; salmon sperm DNA) Ct values from beach water, QC-passed sand-porewater, and QC-failed sand-porewater samples. Figure S3: Correlation between sand-porewater log E. coli CFUs/100 mL and beach water enterococci calibrator cell equivalents (CCEs/100 mL) for Lakeside and Sunset Beaches. Correlation analysis was performed using Pearson correlation at a 95% confidence interval. Figure S4: Correlation between enterococci calibrator cell equivalents (CCEs/100mL) of beach water and sand-porewater from Lakeside and Sunset Beaches. Correlation analysis was performed using Pearson correlation at a 95% confidence interval. Figure S5: Correlation between sand-porewater log E. coli CFUs/100 mL and enterococci calibrator cell equivalents (CCEs/100 mL) for Lakeside and Sunset Beaches. Correlation analysis was performed using Pearson correlation at a 95% confidence interval. Figure S6: Correlation between wave height and log enterococci calibrator cell equivalents (CCEs/100 mL) for sand-porewater and beach water from Lakeside and Sunset Beaches. Correlation analysis was performed using Pearson correlation at a 95% confidence interval. Wave height data were obtained from Niagara Public Health for analysis. Figure S7: Correlation between wave height and sand porewater log E. coli CFUs/100 mL for Lakeside and Sunset Beaches. Correlation analysis was performed using Pearson correlation at 95% confidence interval. Wave height data were obtained from Niagara Public Health for analysis. Table S1: Geographical coordinates for sampling sites from Lakeside and Sunset Beaches. Table S2: Beach posting status in comparison with E. coli culturing beach water threshold exceedances for sand-porewater samples from Lakeside and Sunset Beaches.

Author Contributions

Validation, F.S.; methodology, F.S.; writing—original draft preparation, F.S.; writing—review and editing F.S., T.A.E., and H.E.S.; supervision, H.E.S. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Government of Ontario (Canada-Ontario Agreement grant agreement #2607). Such support does not indicate endorsement by the Government of Ontario of the contents or conclusions of this contribution.

Data Availability Statement

Data is contained within the article or supplementary material.

Acknowledgments

We would like to thank Niagara Public Health, who provided guidance and support in data acquisition. Albert Simhon, Ontario Ministry of Environment, Conservation, and Parks (MECP), provided support and suggestions throughout the project.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. Geographical map of sampling sites from Lakeside and Sunset Beaches (by Ontario Geohub).
Figure 1. Geographical map of sampling sites from Lakeside and Sunset Beaches (by Ontario Geohub).
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Figure 2. Comparison between Public Health’s beach posting decisions and same-day enterococci qPCR quantification for Lakeside Beach. The dashed line represents the USEPA Beach Action Value for Enterococcus qPCR quantification method (≥1000 CCEs/100 mL), while the grey highlighted days represent beach days posted by Public Health based on their 24 h-delayed E. coli geometric mean. Error bars represent the standard deviation of log-transformed Enterococcus calibrator cell equivalents.
Figure 2. Comparison between Public Health’s beach posting decisions and same-day enterococci qPCR quantification for Lakeside Beach. The dashed line represents the USEPA Beach Action Value for Enterococcus qPCR quantification method (≥1000 CCEs/100 mL), while the grey highlighted days represent beach days posted by Public Health based on their 24 h-delayed E. coli geometric mean. Error bars represent the standard deviation of log-transformed Enterococcus calibrator cell equivalents.
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Figure 3. Analysis of impacts on Lakeside Beach postings using the same-day Enterococcus qPCR and compared with Public Health’s reported 24 h-delayed E. coli geometric mean for our summer 2022 beach sampling days (n = 42).
Figure 3. Analysis of impacts on Lakeside Beach postings using the same-day Enterococcus qPCR and compared with Public Health’s reported 24 h-delayed E. coli geometric mean for our summer 2022 beach sampling days (n = 42).
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Figure 4. Comparison between Public Health’s beach posting decisions and same-day Enterococcus qPCR quantification for Sunset Beach. The dashed line represents the USEPA Beach Action Value for the Enterococcus qPCR quantification method (≥1000 CCEs/100 mL), while the grey highlighted days represent beach days posted by Public Health based on their 24 h-delayed E. coli geometric mean. Error bars represent the standard deviation of log-transformed Enterococcus calibrator cell equivalents.
Figure 4. Comparison between Public Health’s beach posting decisions and same-day Enterococcus qPCR quantification for Sunset Beach. The dashed line represents the USEPA Beach Action Value for the Enterococcus qPCR quantification method (≥1000 CCEs/100 mL), while the grey highlighted days represent beach days posted by Public Health based on their 24 h-delayed E. coli geometric mean. Error bars represent the standard deviation of log-transformed Enterococcus calibrator cell equivalents.
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Figure 5. Analysis of impacts on Sunset Beach postings using the same-day Enterococcus qPCR and compared with Public Health’s reported 24 h-delayed E. coli geometric mean for our summer 2022 beach sampling days (n = 42).
Figure 5. Analysis of impacts on Sunset Beach postings using the same-day Enterococcus qPCR and compared with Public Health’s reported 24 h-delayed E. coli geometric mean for our summer 2022 beach sampling days (n = 42).
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Figure 6. Comparison of Lakeside Beach sand Enterococcus exceedances of beach water Beach Action Value with Niagara Public Health’s reported 24 h-delayed E. coli posting results for our summer 2022 beach sampling days (n = 27).
Figure 6. Comparison of Lakeside Beach sand Enterococcus exceedances of beach water Beach Action Value with Niagara Public Health’s reported 24 h-delayed E. coli posting results for our summer 2022 beach sampling days (n = 27).
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Figure 7. Comparison of Sunset Beach sand Enterococcus exceedances of beach water Beach Action Value with Niagara Public Health’s reported 24 h-delayed E. coli posting results for our summer 2022 beach sampling days (n = 32).
Figure 7. Comparison of Sunset Beach sand Enterococcus exceedances of beach water Beach Action Value with Niagara Public Health’s reported 24 h-delayed E. coli posting results for our summer 2022 beach sampling days (n = 32).
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Figure 8. Comparison of Lakeside Beach sand retrospective same-day E. coli exceedances of Niagara Public Health’s reported 24 h-delayed E. coli beach water postings for our summer 2022 beach sampling days (n = 42). Error bars represent the standard deviation of log-transformed E. coli colony-forming units.
Figure 8. Comparison of Lakeside Beach sand retrospective same-day E. coli exceedances of Niagara Public Health’s reported 24 h-delayed E. coli beach water postings for our summer 2022 beach sampling days (n = 42). Error bars represent the standard deviation of log-transformed E. coli colony-forming units.
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Figure 9. Comparison of Sunset Beach sand retrospective same-day E. coli exceedances of Niagara Public Health’s reported 24 h-delayed E. coli beach water postings for our summer 2022 beach sampling days (n = 42). Error bars represent the standard deviation of log-transformed E. coli colony-forming units.
Figure 9. Comparison of Sunset Beach sand retrospective same-day E. coli exceedances of Niagara Public Health’s reported 24 h-delayed E. coli beach water postings for our summer 2022 beach sampling days (n = 42). Error bars represent the standard deviation of log-transformed E. coli colony-forming units.
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Table
Table 1. qPCR quality control analytics for beach and sand-porewater samples from Lakeside and Sunset Beaches.
Table 1. qPCR quality control analytics for beach and sand-porewater samples from Lakeside and Sunset Beaches.
Data Quality Parameters
Standard Curve
Number of Standard Curves4
Coefficient of Determination0.998
Amplification Efficiency2.02
Slope−3.28
Intercept39.55
Beach Water Samples
Number of Samples168
qPCR Quality Pass 168 (100%)
Sand-Porewater Samples
Before Nylon MeshAfter Nylon Mesh
Number of Samples48Number of Samples120
qPCR Quality Pass20 (41.7%)qPCR Quality Pass63 (52.5%)
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MDPI and ACS Style

Saleem, F.; Edge, T.A.; Schellhorn, H.E. Application of Same-Day Enterococcus qPCR-Based Analyses for Quality Assessment of Shorelines (Water and Sand) at Recreational Beaches. Water 2023, 15, 2338. https://doi.org/10.3390/w15132338

AMA Style

Saleem F, Edge TA, Schellhorn HE. Application of Same-Day Enterococcus qPCR-Based Analyses for Quality Assessment of Shorelines (Water and Sand) at Recreational Beaches. Water. 2023; 15(13):2338. https://doi.org/10.3390/w15132338

Chicago/Turabian Style

Saleem, Faizan, Thomas A. Edge, and Herb E. Schellhorn. 2023. "Application of Same-Day Enterococcus qPCR-Based Analyses for Quality Assessment of Shorelines (Water and Sand) at Recreational Beaches" Water 15, no. 13: 2338. https://doi.org/10.3390/w15132338

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