A somatic coliphage threshold approach to improve the management of activated sludge wastewater treatment plant effluents in resource-limited regions

Versión aceptada para publicaciónEffective wastewater management is crucial to ensure the safety of water reuse projects and 29 effluent discharge into surface waters. Multiple studies have demonstrated that municipal 30 wastewater treatment with conventional activated sludge processes is inefficient for the removal 31 of the wide spectrum of viruses in sewage. In this study, a well-accepted statistical approach was 32 used to investigate the relationship between viral indicators and human enteric viruses during 33 wastewater treatment in a resource-limited region. Influent and effluent samples from five urban 34 wastewater treatment plants (WWTP) in Costa Rica were analyzed for somatic coliphage and 35 human enterovirus, hepatitis A virus, norovirus genotype I and II, and rotavirus. All WWTP 36 provide primary treatment followed by conventional activated sludge treatment prior to 37 discharge into surface waters that are indirectly used for agricultural irrigation. The results 38 revealed a statistically significant relationship between the detection of at least one of the five 39 human enteric viruses and somatic coliphage. Multiple logistic regression and Receiver Operating Characteristic curve analysis identified a threshold of 3.0 ×103 40 (3.5-log10) somatic 41 coliphage plaque forming unit per 100 mL, which corresponded to an increased likelihood of encountering enteric viruses above the limit of detection (>1.83×102 42 virus target/100 mL). 43 Additionally, quantitative microbial risk assessment was executed for famers indirectly reusing 44 WWTP effluent that met the proposed threshold. The resulting estimated median cumulative 45 annual disease burden complied with World Health Organization recommendations. Future 46 studies are needed to validate the proposed threshold for use in Costa Rica and other regions.Universidad de Costa Rica/[]/UCR/Costa RicaNational Science Foundation/[OCE-1566562]/NSF/Estados UnidosUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto de Investigaciones en Salud (INISA)UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiologí

An efficient strategy for evaluating new non-invasive screening tests for colorectal cancer: the guiding principles

Objective: New screening tests for colorectal cancer (CRC) are rapidly emerging. Conducting trials with mortality reduction as the end point supporting their adoption is challenging. We re-examined the principles underlying evaluation of new non-invasive tests in view of technological developments and identification of new biomarkers. Design: A formal consensus approach involving a multidisciplinary expert panel revised eight previously established principles. Results: Twelve newly stated principles emerged. Effectiveness of a new test can be evaluated by comparison with a proven comparator non-invasive test. The faecal immunochemical test is now considered the appropriate comparator, while colonoscopy remains the diagnostic standard. For a new test to be able to meet differing screening goals and regulatory requirements, flexibility to adjust its positivity threshold is desirable. A rigorous and efficient four-phased approach is proposed, commencing with small studies assessing the test’s ability to discriminate between CRC and non-cancer states (phase I), followed by prospective estimation of accuracy across the continuum of neoplastic lesions in neoplasia-enriched populations (phase II). If these show promise, a provisional test positivity threshold is set before evaluation in typical screening populations. Phase III prospective studies determine single round intention-to-screen programme outcomes and confirm the test positivity threshold. Phase IV studies involve evaluation over repeated screening rounds with monitoring for missed lesions. Phases III and IV findings will provide the real-world data required to model test impact on CRC mortality and incidence. Conclusion: New non-invasive tests can be efficiently evaluated by a rigorous phased comparative approach, generating data from unbiased populations that inform predictions of their health impact

Viruses Found in Raw Sewage and Their Potential to Indicate Fecal Pollution in Coastal Environments

The presence of pathogenic viruses in coastal environments is an important tool in evaluating water quality and health risks. Millions of viruses are excreted in fecal matter and bacterial indicators do not correlate with the presence of pathogenic viruses. Enteroviruses have been used to identify fecal pollution in the environment; however, other viruses shed in fecal matter could be used to indicate fecal pollution. The purpose of this research is to develop a baseline understanding of the diversity of viruses found in raw sewage and to assess their presence in the marine environment. PCR was used to detect adenoviruses, herpesviruses, hepatitis B viruses, morbilliviruses, noroviruses, papillomaviruses, pepper mild mottle viruses, picobirnaviruses, reoviruses, rotaviruses, and sapporoviruses in raw sewage collected from throughout the United States and from five marine environments ranging in their proximity to dense human populations. Adenoviruses, noroviruses, pepper mild mottle viruses, and picobirnaviruses were detected in raw sewage but absent in the marine environment, making these viruses potential indicators of fecal pollution in marine environments. These viruses were also found in many of the final effluent samples. Pepper mild mottle viruses may be useful for source tracking fecal contamination since it was consistently found in human sewage and is not expected in the feces of other animals due to its dietary origin. Furthermore, this research uncovered previously unknown sequence diversity in human picobirnaviruses. This baseline understanding of viruses in raw sewage and the marine environment will enable educated decisions to be made regarding the use of viruses in water quality assessments

Affordable Enteric Virus Detection Techniques Are Needed to Support Changing Paradigms in Water Quality Management

In light of water quality monitoring paradigms shifting to a more holistic approach, it is essential that environmental microbiologists embrace new methodological developments in clinical virology to create rapid, laboratory‐free methods for the identification of wastewater pollution. It is widely accepted that routine monitoring of fecal indicator bacteria (FIB) does not adequately reflect human health risks associated with fecal pollution, especially risks posed by viruses. Enteric viruses are typically more resistant to wastewater treatment and persist longer in the environment than FIB. Furthermore, enteric viruses often have extremely low infectious doses. Currently, the incorporation of sanitary surveys, short‐term monitoring of reference pathogens, exploratory quantitative microbial risk assessments, and predictive ecological models is being championed as the preferred approach to water management. In addition to improved virus concentration methods, simple, point‐of‐use tests for enteric viruses and/or improved viral indicators are needed to complement this emerging paradigm and ensure microbial safety worldwide

Eukaryotic Viruses in Wastewater Samples from the United States▿

Human fecal matter contains a large number of viruses, and current bacterial indicators used for monitoring water quality do not correlate with the presence of pathogenic viruses. Adenoviruses and enteroviruses have often been used to identify fecal pollution in the environment; however, other viruses shed in fecal matter may more accurately detect fecal pollution. The purpose of this study was to develop a baseline understanding of the types of viruses found in raw sewage. PCR was used to detect adenoviruses, enteroviruses, hepatitis B viruses, herpesviruses, morbilliviruses, noroviruses, papillomaviruses, picobirnaviruses, reoviruses, and rotaviruses in raw sewage collected throughout the United States. Adenoviruses and picobirnaviruses were detected in 100% of raw sewage samples and 25% and 33% of final effluent samples, respectively. Enteroviruses and noroviruses were detected in 75% and 58% of raw sewage samples, respectively, and both viral groups were found in 8% of final effluent samples. This study showed that adenoviruses, enteroviruses, noroviruses, and picobirnaviruses are widespread in raw sewage. Since adenoviruses and picobirnaviruses were detected in 100% of raw sewage samples, they are potential markers of fecal contamination. Additionally, this research uncovered previously unknown sequence diversity in human picobirnaviruses. This baseline understanding of viruses in raw sewage will enable educated decisions to be made regarding the use of different viruses in water quality assessments

Pepper Mild Mottle Virus as an Indicator of Fecal Pollution

Accurate indicators of fecal pollution are needed in order to minimize public health risks associated with wastewater contamination in recreational waters. However, the bacterial indicators currently used for monitoring water quality do not correlate with the presence of pathogens. Here we demonstrate that the plant pathogen Pepper mild mottle virus (PMMoV) is widespread and abundant in wastewater from the United States, suggesting the utility of this virus as an indicator of human fecal pollution. Quantitative PCR was used to determine the abundance of PMMoV in raw sewage, treated wastewater, seawater exposed to wastewater, and fecal samples and/or intestinal homogenates from a wide variety of animals. PMMoV was present in all wastewater samples at concentrations greater than 1 million copies per milliliter of raw sewage. Despite the ubiquity of PMMoV in human feces, this virus was not detected in the majority of animal fecal samples tested, with the exception of chicken and seagull samples. PMMoV was detected in four out of six seawater samples collected near point sources of secondary treated wastewater off southeastern Florida, where it co-occurred with several other pathogens and indicators of fecal pollution. Since PMMoV was not found in nonpolluted seawater samples and could be detected in surface seawater for approximately 1 week after its initial introduction, the presence of PMMoV in the marine environment reflects a recent contamination event. Together, these data demonstrate that PMMoV is a promising new indicator of fecal pollution in coastal environments

Removal of Six Estrogenic Endocrine-Disrupting Compounds (EDCs) from Municipal Wastewater Using Aluminum Electrocoagulation

Conventional wastewater treatment plant (WWTP) processes are primarily designed to reduce the amount of organic matter, pathogens, and nutrients from the incoming influent. However, these processes are not as effective in reducing the concentrations of micropollutants, including endocrine-disrupting compounds (EDCs), which notoriously evade traditional wastewater treatment technologies and are found even in tertiary-treated effluent. For WWTPs practicing deep-well injection or surface-water discharge, EDCs in the treated effluent are discharged into groundwater or the aquatic environment where humans and wildlife may potentially suffer the effects of chemical exposure. In the current laboratory-scale study, we tested a bench-top electrocoagulation (EC) unit utilizing aluminum blades for the removal of six estrogenic EDCs [estrone (E1), 17β-estradiol (E2), estriol (E3), 17α-ethinylestradiol (EE2), bisphenol-A (BPA), and nonylphenol (NP)]. Samples of municipal wastewater influent and tertiary-treated effluent were spiked with the six EDCs in order to test the removal efficiency of the EC unit. The mean concentration of each EDC component was statistically lower after EC treatment (removal range = 42%–98%). To our knowledge, this is the first study to investigate aluminum electrocoagulation for removal of these specific EDCs, including nonylphenol (without the ethoxylate chain), as well as natural and synthetic estrogens

Microbial Source Tracking in Shellfish Harvesting Waters in the Gulf of Nicoya, Costa Rica

Current microbial water quality monitoring is generally limited to culture-based measurements of fecal indicator bacteria (FIB). Given the many possible sources of fecal pollution within a watershed and extra-intestinal FIB reservoirs, it is important to determine source(s) of fecal pollution as a means to improve water quality and protect public health. The principal objective of this investigation was to characterize the microbial water quality of shellfish harvesting areas in the Gulf of Nicoya, Costa Rica during 2015. In order to achieve this objective, the specificity and sensitivity of 11 existing microbial source tracking (MST) PCR assays, associated with cows (BacCow), dogs (BacCan, DogBac), domestic wastewater (PMMoV), general avian (GFD), gulls (Gull2), horses (HorseBac, HoF), humans (HF183, HPyV), and pigs (PF), were evaluated using domestic wastewater and animal fecal samples collected from the region. The sensitivity of animal-associated assays ranged from 13 to 100%, while assay specificity ranged from 38 to 100%. The specificity of pepper mild mottle virus (PMMoV) and human polyomavirus (HPyV) was 100% for domestic wastewater, as compared to 94% specificity of the HF183 Bacteroidales marker. PMMoV was identified as a useful domestic wastewater-associated marker, with concentrations as high as 1.1 × 105 copies/ml and 100% sensitivity and specificity. Monthly surface water samples collected from four shellfish harvesting areas were analyzed using culture-based methods for Escherichia coli as well as molecular methods for FIB and a suite of MST markers, which were selected for their specificity in the region. While culturable E. coli results suggested possible fecal pollution during the monitoring period, the absence of human/domestic wastewater-associated markers and low FIB concentrations determined using molecular methods indicated sufficient microbial water quality for shellfish harvesting. This is the first study to our knowledge to test the performance of MST markers in Costa Rica as well as in Central America. Given the lack of wastewater treatment and the presence of secondary sources of FIB, this study highlights the importance of an MST toolbox approach to characterize water quality in tropical regions. Furthermore, it confirms and extends the geographic range of PMMoV as an effective tool for monitoring domestic wastewater pollution

Evaluating the Role of Methylated Circulating Tumor DNA in Combination With Pathological Prognostic Factors for Predicting Recurrence of Colorectal Cancer

Background: Colorectal cancer (CRC) has a high rate of recurrence, in particular for advanced disease, but prognosis based on staging and pathology at surgery can have limited efficacy. The presence of circulating tumor DNA (ctDNA) at diagnosis could be used to improve the prediction for disease recurrence. Objectives: To assess the impact of detecting methylated BCAT1/IKZF1 ctDNA at diagnosis in combination with demographic, lifestyle, clinical factors and tumor pathology, to assess predictive value for recurrence. Design: A retrospective cohort study. Methods: The cohort included 180 patients (36 with recurrent CRC), who had undergone complete treatment and surveillance for a minimum of 3 years. Participant clinical details and ctDNA methylated BCAT1 / IKZF1 results were compared between those with and without recurrence, and cox regression analysis assessed each factor on disease-free survival. Results: Clinical factors independently associated with reduced disease-free survival included nodal involvement (HR = 3.83, 95% CI 1.56-9.43, P  = .003), M1 stage (HR = 4.41, 95% CI 1.18-16.45, P  = .027), a resection margin less than 2 mm (HR = 4.60, 95% CI 1.19-17.76, P  = .027), perineural involvement (HR = 2.50, 95% CI 1.01-6.17, P  = .047) and distal tumors (HR = 3.13, 95% CI 1.07-9.18, P  = .037). Methylated BCAT1 / IKZF1 was detected in 51.7% (93/180) of pre-treatment plasma samples. When a positive ctDNA finding was considered in combination with these clinical prognostic factors, there was improved predictive power of recurrence for patients with perineural involvement (HR = 4.44, 95% CI 1.92-10.26, P  < .001), and it marginally improved the predictive factor for M1 stage (HR = 7.59, 95% CI 2.30-25.07, P  = .001) and distal tumors (HR = 5.04, 95% CI 1.88-13.49, P  = .001). Conclusions: Nodal invasion, metastatic disease, distal tumor site, low resection margins and perineural invasion were associated with disease recurrence. Pre-treatment methylated ctDNA measurement can improve the predictive value for recurrence in a subset of patients, particularly those with perineural involvement. Registration: Australian and New Zealand Clinical Trials Registry #12611000318987