We investigated the presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in wastewater samples in southern Louisiana, USA. Untreated and treated wastewater samples were ...collected on five occasions over a four-month period from January to April 2020. The wastewater samples were concentrated via ultrafiltration (Method A), and an adsorption–elution method using electronegative membranes (Method B). SARS-CoV-2 RNA was detected in 2 out of 15 wastewater samples using two reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays (CDC N1 and N2). None of the secondary treated and final effluent samples tested positive for SARS-CoV-2 RNA. To our knowledge, this is the first study reporting the detection of SARS-CoV-2 RNA in wastewater in North America, including the USA. However, concentration methods and RT-qPCR assays need to be refined and validated to increase the sensitivity of SARS-CoV-2 RNA detection in wastewater.
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•First study in Louisiana, USA reporting the detection of SARS-CoV-2 RNA in wastewater using ultrafiltration.•Two out of seven untreated wastewater samples tested positive for SARS-CoV-2 RNA.•None of the secondary treated and final effluent samples tested positive.•Concentration methods and RT-qPCR assays applied for SARS-CoV-2 RNA detection need further refinement.
The occurrence of amoeba, Naegleria fowleri, in sediment samples from Lake Pontchartrain in Louisiana was investigated. This amoeba is pathogenic and can cause primary amoebic meningoencephalitis. In ...this study, quantitative polymerase chain reaction methods were used to test for the prevalence of Naegleria fowleri, HF183, and E. coli. N. fowleri was detected in 51.25% of our sediment samples. Illumina sequencing of sediment samples revealed ten different phyla, with Cyanobacteria being the most predominant at sites that generally presented with the highest median N. fowleri concentrations. N. fowleri was however strongly negatively correlated with HF183 (r = -0.859, p < 0.001). Whenever sediment E. coli concentrations were below 1.54 Log GC/g, there was only a 37.5% chance that N. fowleri would be detected in the same sample. When sediment E. coli concentrations exceeded 2.77 Log GC/g, the chances of detecting N. fowleri in the same sample increased to 90%, potentially suggesting predatory activity by the amoeba. The effect of temperature was observed to be different in relation to observed N. fowleri concentrations and detection rates. Although sediment samples collected during periods of higher temperatures had significantly lower mean N. fowleri concentrations (2.7 Log GC/g) compared to those collected at lower temperatures (3.7 Log GC/g, t(39) = 4.167, p < 0.001), higher N. fowleri detection rates in the overall samples were observed at higher temperatures (>19.1 °C) than at lower temperatures (<19.1 °C).
With a focus on five sites in an impaired, densely populated area in the New Orleans area, we investigated the temporal and spatial variability of standard FIB and a marker of human-associated ...pollution (Bacteroides HF183). With all sites combined, only a weak positive correlation (r = 0.345; p = 0.001) was observed between E. coli and HF183. Also, specific conductivity (r = - 0.374; p < 0.0001) and dissolved oxygen (r = - 0.390; p < 0.0001) were observed to show a weak moderate correlation with E. coli. These correlations increased to moderately negative when HF183 was correlated with specific conductivity (r = - 0.448; p < 0.0001) and dissolved oxygen (r = - 0.455; p < 0.0001). E. coli contamination was generally highest at the sites in the canal that are situated in the most densely populated part of the watershed while HF183 was frequently detected across all sites. E. coli concentrations were significantly higher (p < 0.05) when HF183 was present. HF183 was detected at significantly higher concentrations in samples that exceeded the EPA water quality standard (WQS) than those that did not (p < 0.05). Dissolved oxygen and specific conductivity were significantly lower when E. coli WQS was exceeded or when HF183 was present (p < 0.05). Rainfall impacted E. coli concentrations and HF183 differently at the study sites. While HF183 and E. coli concentrations levels were significantly higher (p < 0.05) if the days prior to sampling had been wet, the frequency of detection of HF183 was unimpacted, as comparable detection rates were recorded during wet and dry weather conditions. Without testing for HF183, it would have been assumed, based on testing for E. coli alone, that human fecal pollution was only associated with densely populated areas and rainfall events. E. coli alone may not be an effective indicator of sewage pollution at the study sites across all weather conditions and may need to be complemented with HF183 enumeration to optimize human fecal pollution identification and management at the watershed level.
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•A weak positive correlation between E. coli and HF183•Higher E.coli WQS exceedances associated with densely populated areas.•Wet weather had weaker mobilization effect on HF183 than E. coli.•Wet and dry weather HF183 detection suggest chronic human fecal pollution.•E. coli alone may not be an effective indicator of sewage pollution.
Prison populations are unlikely to have access to prompt, effective medical care as the general population. Therefore, vaccination and effective surveillance systems have been recommended to mitigate ...coronavirus disease 2019 (COVID-19) transmission in prison settings. This pilot study aimed to assess the application of wastewater-based epidemiology (WBE) in a prison to act as an early warning tool for COVID-19 transmission. In this study, weekly wastewater samples (n = 21) were collected for 21 weeks from a prison facility in New Orleans, LA, USA, and analyzed for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and the results were compared with the number of confirmed cases during the same period. SARS-CoV-2 was concentrated using two methods and quantified via RT-qPCR using CDC N1 and N2 assays. Overall, SARS-CoV-2 was detected in eight samples (38%). An equal number of samples tested positive for SARS-CoV-2 using the two concentrating methods, indicating the effectiveness of both methods for building-scale WBE. Despite limited clinical testing in the studied prison facility, instances of SARS-CoV-2 detection in wastewater prior to the diagnosis of COVID-19 depict the potential use of wastewater surveillance in detecting the presence of early and averting outbreaks in asymptomatic COVID-19 patients.
Abstract
Concerns over fecal contamination in stormwater canals have promoted the need for pollution control strategies, including the use of microbial source tracking, to identify fecal ...contamination in the Greater New Orleans Area. Surface water samples were collected over a 12-month period at five canal locations within Jefferson Parish, Louisiana. Quantitative polymerase chain reaction and the IDEXX method were used to assess the concentrations of coliforms, Escherichia coli (E. coli) and human fecal 183 bacteroides (HF183) in stormwater samples. A 100% positive detection rate of total coliforms and E. coli was observed across all tested sites. Despite the closeness of the five sites, when averaged across all sampling time points, Kruskal–Wallis tests indicated that E. coli was present at significantly different concentrations in these locations (χ2(5) = 19.8, p = 0.0005). HF183 was detected in 62% of the water samples collected during the stormwater sampling. Without further testing for HF183 markers, the conclusion from this study would have been that fecal contamination from an unknown source was always present at varying levels during the study period. Analysis of HF183 markers therefore adds another layer of conclusions to the results deductible from E. coli concentrations. A 100% E. coli detection rate, high E. coli concentrations coupled with low rates of HF183 detection particularly at the Esplanade, Poplar Street, and Bonnabel Boat Launch sites, the sites closest to the lake outlet, throughout the study period, indicate that fecal contamination at these stormwater canal sites comes primarily from non-human sources. However, the Metairie Road and Napoleon Avenue sites, which have the highest HF183 detection rates, on top of chronic pollution by other non-human sources, are also influenced by human fecal pollution, possibly because of human development and faulty infrastructure. This study highlights the advantages of the use of microbial source-tracking methods to complement traditional indicator bacteria.
Wastewater treatment plants (WWTPs) represent all known types of antibiotic resistance mechanisms and are considered as the critical points for the spread of antibiotic resistance genes (ARGs). The ...purpose of this study is to investigate the removal of a Class 1 integrase gene (intI1) and a selected set of ARGs (blaTEM, ermF, mecA, and tetA) at two conventional WWTPs by using chlorination in Louisiana, USA. We collected 69 wastewater samples (23 each from influent, secondary effluent, and final effluent) and determined the concentrations of ARGs by using quantitative polymerase chain reaction. All tested ARGs, except for mecA, were detected in 83–96% and 30–65% of influent and final effluent samples, respectively. Although the ARGs underwent approximately 3-log10 reduction, two WWTPs on an average still released 3.3 ± 1.7 log10 copies/mL of total ARGs studied in the effluents. Chlorination was found to be critical in the significant reduction of total ARGs (p < 0.05). Correlation analysis and the ability of intI1 to persist through the treatment processes recommend the use of intI1 as a marker of ARGs in effluents to monitor the spread of antibiotic resistance in effluents. Our study suggests that conventional WWTPs using chlorination do not favor the proliferation of antibiotic resistance bacteria and ARGs during wastewater treatment.
In this study, we investigated the occurrence of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) RNA in primary influent (
= 42), secondary effluent (
= 24) and tertiary treated effluent ...(
= 34) collected from six wastewater treatment plants (WWTPs A-F) in Virginia (WWTP A), Florida (WWTPs B, C, and D), and Georgia (WWTPs E and F) in the United States during April-July 2020. Of the 100 wastewater samples analyzed, eight (19%) untreated wastewater samples collected from the primary influents contained SARS-CoV-2 RNA as measured by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) assays. SARS-CoV-2 RNA were detected in influent wastewater samples collected from WWTP A (Virginia), WWTPs E and F (Georgia) and WWTP D (Florida). Secondary and tertiary effluent samples were not positive for SARS-CoV-2 RNA indicating the treatment processes in these WWTPs potentially removed SARS-CoV-2 RNA during the secondary and tertiary treatment processes. However, further studies are needed to understand the log removal values (LRVs) and transmission risks of SARS-CoV-2 RNA through analyzing wastewater samples from a wider range of WWTPs.
Deterioration of surface water microbial quality, primarily attributed to increased anthropogenic activity and aging infrastructure, poses a significant threat. Human fecal contamination, stemming ...from various sources, heavily impacts water quality. While indicator bacteria like fecal coliforms, E. coli, and enterococci are commonly used to assess water quality due to their simplicity and cost-effectiveness, they do not identify contamination sources. In our first objective, we employed microbial source tracking methods and found a positive correlation between E. coli and HF183 levels and physicochemical parameters, highlighting the intricate dynamics of water quality. E. coli contamination was most pronounced in densely populated watershed regions, whereas HF183 persisted consistently across all sites, even during dry weather. Rainfall impacted E. coli levels, but HF183 concentrations remained steady, indicating enduring contamination sources. This study underscores the need to supplement E. coli testing with HF183 to manage fecal pollution in urban water bodies effectively. In our second objective, quantitive polymerase chain reaction methods were used to explore the prevalence of the pathogenic amoeba Naegleria fowleri in sediment samples from Louisiana's Lake Pontchartrain. N. fowleri was detected in over half of the sediment samples, with an intriguing negative correlation with HF183, hinting at possible predatory behavior. The concentration of E. coli in sediment samples influenced the likelihood of N. fowleri detection, suggesting a potential relationship. Temperature variations also affected N. fowleri concentrations and detection rates, with higher temperatures associated with increased detection. The emergence of the global SARS-CoV-2 pandemic, initially a respiratory virus, led to the detection of viral RNA in feces. This prompted the use of wastewater-based surveillance to understand SARS-CoV-2's epidemiology. Our third objective was to examine the presence of SARS-CoV-2 RNA in wastewater samples from six U.S. wastewater treatment plants using the reverse transcriptase quantitative polymerase chain reactions. It detected viral RNA in untreated wastewater samples from select plants, while secondary and tertiary effluent samples were free of the virus, indicating potential removal during treatment. The study underscores the importance of further research to comprehend log removal values and transmission risks of SARS-CoV-2 RNA in wastewater treatment plants.This dissertation unites its various studies under the overarching mission of pinpointing the origins of human fecal contamination and quantifying the emergence of pathogens within natural and engineered water systems. It underscores the vital role of ongoing wastewater surveillance as a cornerstone for improving water quality and shaping public health policies, ultimately serving the welfare of communities. The significance of these findings becomes evident in their contribution to the multifaceted challenges associated with water quality, especially in densely populated urban areas. They emphasize the necessity for custom-tailored interventions and establishing region-specific standards to protect public health and the environment.
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