Impact of reduced sampling frequency of illicit drug wastewater monitoring in the Netherlands.

Drug consumption estimates are traditionally based on surveys or information from police seizures. Alternatively, residues of illicit drugs in untreated wastewater (influent) can be used to calculate mass loads and subsequently estimate drug consumption in the community throughout the week. For this purpose, wastewater is commonly sampled for seven consecutive days within the Sewage analysis CORe group Europe (SCORE), while other sampling schemes may be implemented in long-term studies outside this consortium. The current study demonstrates how sampling frequency of illicit drug residues in the influent of wastewater treatment plants (WWTPs) affects the derived weekly average. Thirty WWTPs were sampled over the course of 12 years and influents were analyzed for five drugs (metabolites): 3,4-methylenedioxymethamphetamine (MDMA), methamphetamine, amphetamine, benzoylecgonine (a metabolite of cocaine), and 11-nor-9-Carboxy tetrahydrocannabinol (THC-COOH). Subsequently, small and large WWTPs were grouped with a threshold of 100,000 inhabitants. After data curation, standardized loads were calculated (mg/d per 1000 inhabitants). Weekly averages of loads of the drug residues were calculated based on six scenarios (sampling one to six weekdays) and compared to the weekly average in the control situation (sampling seven weekdays) in a Monte Carlo simulation. Results indicate that drug residues with more dynamic loads over a week require more frequent sampling. The analysis illustrates that a decreased sampling frequency (4 or 5 days per week) still leads to a representative weekly average for all drugs tested when a deviation up to a factor of 1.25 is deemed acceptable. However, knowledge on typical levels is necessary to define outliers. We therefore recommend to study dynamics in drug residue loads for WWTPs before reducing sampling frequency in long term monitoring programs.

Mapping consumptions and market size of cocaine, amphetamine and MDMA through wastewater analysis: A Dutch case study.

BACKGROUND AND AIMS: Illicit drug consumption is associated with public health effects and criminal activities. This study aimed to estimate Illicit drug consumption and annual market in the Netherlands from wastewater analysis of drug residues. METHODS: Residues of cocaine, amphetamine and 3,4-methylene dioxymethamphetamine (MDMA) were measured between 2015 and 2022 in 30 Dutch wastewater treatment plants serving both rural and urban populations. These wastewater treatment plants covered 20% of the total Dutch population. The Dutch annual retail market was estimated by extrapolating consumption to the total Dutch population, back-calculating consumption volume, correcting for drug purity and street price collected in voluntary checking services, and accounting for the correlation of consumption and urbanity. RESULTS: The per capita MDMA and cocaine consumption correlated positively with the urbanity of the wastewater treatment plant catchments with r2 of 31% and 64%, respectively. Amphetamine did not show a significant correlation with urbanity. The three studied drugs were conservatively estimated to cover an average annual market value of 903 (95% prediction interval 829 to 987) million Euro for the studied period. Market estimations from prevalence figures and interceptions of international trade were similar. CONCLUSIONS: Illicit drug consumption in the Netherlands appears to correlate positively with urban (in contrast to rural) areas. Wastewater analysis can be used to estimate the volume and monetary value of illicit drug markets as a proof of concept.

Estimating alcohol and tobacco consumption of university students and urban population in Ho Chi Minh City by wastewater analysis.

The quantitative measurement of urinary biomarkers in wastewater has emerged as a robust tool for estimating alcohol and tobacco consumption in populations. In this study, we applied the wastewater-based epidemiology (WBE) approach to compare alcohol and tobacco use between university students and urban inhabitants in Ho Chi Minh City, Vietnam. Ethyl sulfate and cotinine serve as markers for alcohol and tobacco use, respectively. Our findings reveal that urban inhabitants aged 15 and above consume 1.56 ± 0.23 mL of pure ethanol and 2.8 ± 0.33 mg of nicotine per day, while university students consume 0.69 ± 0.13 mL of pure alcohol and 1.2 ± 0.2 mg of nicotine per day. This indicates that, on average, students consume less alcohol and tobacco compared with urban adults. A Monte Carlo simulation indicated that, on average, university students in our study smoke 1.5 cigarettes per day, while urban residents aged 15 and above smoke 4.3 cigarettes per day. Considering the smoking prevalence, a student smoker in this study consumes 6.5 cigarettes per day, a level high enough to establish addiction. On the other hand, alcohol use estimation is significantly lower than previous survey-based reports, likely due to degradation within on-site septic tanks. Future research should aim to extend the sampling period to capture seasonal variations and improve the understanding of tobacco and alcohol consumption patterns. The results from this study are crucial for decision-makers in Ho Chi Minh City to develop effective public health strategies and interventions. PRACTITIONER POINTS: Wastewater-based approach is applicable to estimate the tobacco consumption in Ho Chi Minh City. Each current smoker in the urban area of Ho Chi Minh City smokes nearly a package a day. The estimated consumption for student smokers in U-town is 6.5 cigarettes per day, a level high enough to establish addiction. The existence of septic tanks within Vietnam's drainage systems prevents reliable estimation of alcohol consumption for the entire population.

Exposure to environmental pollutants: A mini-review on the application of wastewater-based epidemiology approach.

Wastewater-based epidemiology (WBE) is considered an innovative and promising tool for estimating community exposure to a wide range of chemical and biological compounds by analyzing wastewater. Despite scholars' interest in WBE studies, there are uncertainties and limitations associated with this approach. This current review focuses on the feasibility of the WBE approach in assessing environmental pollutants, including pesticides, heavy metals, phthalates, bisphenols, and personal care products (PCPs). Limitations and challenges of WBE studies are initially discussed, and then future perspectives, gaps, and recommendations are presented in this review. One of the key limitations of this approach is the selection and identification of appropriate biomarkers in studies. Selecting biomarkers considering the basic requirements of a human exposure biomarker is the most important criterion for validating this new approach. Assessing the stability of biomarkers in wastewater is crucial for reliable comparisons of substance consumption in the population. However, directly analyzing wastewater does not provide a clear picture of biomarker stability. This uncertainty affects the reliability of temporal and spatial comparisons. Various uncertainties also arise from different steps involved in WBE. These uncertainties include sewage sampling, exogenous sources, analytical measurements, back-calculation, and estimation of the population under investigation. Further research is necessary to ensure that measured pollutant levels accurately reflect human excretion. Utilizing data from WBE can support healthcare policy in assessing exposure to environmental pollutants in the general population. Moreover, WBE seems to be a valuable tool for biomarkers that indicate healthy conditions, lifestyle, disease identification, and exposure to pollutants. Although this approach has the potential to serve as a biomonitoring tool in large communities, it is necessary to monitor more metabolites from wastewater to enhance future studies.

Leveraging wastewater-based epidemiology to monitor the spread of neglected tropical diseases in African communities.

Neglected tropical diseases continue to cause a significant burden worldwide, with Africa accounting for more than one-third of the global burden. Over the past decade, progress has been made in eliminating, controlling, and eradicating these diseases in Africa. By December 2022, 47 out of 54 African countries had eliminated at least one neglected tropical disease, and more countries were close to achieving this milestone. Between 2020 and 2021, there was an 80 million reduction in people requiring intervention. However, continued efforts are needed to manage neglected tropical diseases and address their social and economic burden, as they deepen marginalisation and stigmatisation. Wastewater-based epidemiology involves analyzing wastewater to detect and quantify biomarkers of disease-causing pathogens. This approach can complement current disease surveillance systems in Africa and provide an additional layer of information for monitoring disease spread and detecting outbreaks. This is particularly important in Africa due to limited traditional surveillance methods. Wastewater-based epidemiology also provides a tsunami-like warning system for neglected tropical disease outbreaks and can facilitate timely intervention and optimised resource allocation, providing an unbiased reflection of the community's health compared to traditional surveillance systems. In this review, we highlight the potential of wastewater-based epidemiology as an innovative approach for monitoring neglected tropical disease transmission within African communities and improving existing surveillance systems. Our analysis shows that wastewater-based epidemiology can enhance surveillance of neglected tropical diseases in Africa, improving early detection and management of Buruli ulcers, hookworm infections, ascariasis, schistosomiasis, dengue, chikungunya, echinococcosis, rabies, and cysticercosis for better disease control.

Campus source to sink wastewater surveillance of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).

Wastewater-based surveillance (WBS) offers an aggregate, and cost-effective approach for tracking infectious disease outbreak prevalence within communities, that provides data on community health complementary to individual clinical testing. This study reports on a 16-month WBS initiative on a university campus in England, UK, assessing the presence of SARS-CoV-2 in sewers from large buildings, downstream sewer locations, raw wastewater, partially treated and treated effluents. Key findings include the detection of the Alpha (B.1.1.7) variant in wastewater, with 70 % of confirmed campus cases correlating with positive wastewater samples. Notably, ammonium nitrogen (NH4-N) levels showed a positive correlation (ρ = 0.543, p < 0.01) with virus levels at the large building scale, a relationship not observed at the sewer or wastewater treatment works (WWTW) levels due to dilution. The WWTW was compliant to wastewater standards, but the secondary treatment processes were not efficient for virus removal as SARS-CoV-2 was consistently detected in treated discharges. Tools developed through WBS can also be used to enhance traditional environmental monitoring of aquatic systems. This study provides a detailed source-to-sink evaluation, emphasizing the critical need for the widespread application and improvement of WBS. It showcases WBS utility and reinforces the ongoing challenges posed by viruses to receiving water quality.

Evaluation of spatial and temporal changes in illicit drug use in the Taipei metropolitan area via wastewater-based epidemiology.

Taiwan, identified as pivotal in the Asian drug trafficking chain, has been experiencing a surge in illicit drug-related issues. Wastewater-based epidemiology (WBE) has emerged as a promising approach for comprehensive evaluation of actual illicit drug usage. This study presents the first WBE investigation of illicit drug consumption in Taiwan based on the analysis of wastewater from four wastewater treatment plants (WWTPs) in the Taipei metropolitan area. Additionally, it demonstrates a high correlation between the amounts of illicit drugs seized and influent concentrations over an extended period of time. The reliability of solid-phase extraction and analysis via high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was validated for 16 illicit drugs (methamphetamine, ketamine, cocaine, codeine, methadone, morphine, meperidine, fentanyl, sufentanil, para-methoxyamphetamine (PMA), para-methoxymethamphetamine (PMMA), 3,4-methylenedioxymethamphetamine (MDMA), cathinone, methcathinone, mephedrone (MEPH), and 4-methylethcathinone (4-MEC)). Methamphetamine, ketamine, and 4-MEC were consistently detected in all wastewater samples, underscoring their prevalence in the Taipei metropolitan area. Biochemical oxygen demand (BOD) and ammonia nitrogen (ammonia N) were employed to reduce uncertainty in estimations of population size during back-calculation of illicit drug consumption. The results indicate that methamphetamine was the most consumed drug (175-740 mg day-1 1000 people-1), followed by ketamine (22-280 mg day-1 1000 people-1). In addition, urban-related WWTPs exhibited higher consumption of methamphetamine and ketamine than did the suburban-related WWTP, indicating distinct illicit drug usage patterns between suburban and urban regions. Moreover, an examination of temporal trends in wastewater from the Dihua WWTP revealed a persistent predominance of ketamine and methamphetamine, consistent with statistical data pertaining to seizure quantities and urine test results. The study provides encouraging insight into spatial and temporal variations in illicit drug usage in the Taipei metropolitan area, emphasizing the complementary role of WBE in understanding trends in illicit drug abuse.

Understanding treatment of pain during SARS-CoV-2 pandemic in a two-year intercity longitudinal study using wastewater-based epidemiology.

SARS-CoV-2 pandemic had a significant impact on the society, economy, and health of people around the world with consequences that need to be better understood for future pandemic preparedness. This manuscript provides insights into the usage of pharmaceuticals for pain treatment management throughout SARS-CoV-2 pandemic. Four towns and cities with a total population of > 1 million people covering an area of 2000 km2 in South West England were monitored for twenty-four months. Results showed different patterns in pain pharma usage, with small towns having higher population normalised daily loads (PNDLs) than big cities for majority of pain killers studied. This is likely due to demographics of these cities with smaller cities having older population. Per capita consumption of non-steroidal anti-inflammatory drugs (NSAIDs) increased compared to pre-pandemic usage in line with SARS-CoV-2 infections (ibuprofen and acetaminophen), while body pain drugs (diclofenac and naproxen) decreased in line with restrictions and closure of sports facilities. Changes in population normalised daily intake (PNDI) of pain killers were particularly apparent during the 1st and 3rd national lockdown. Comparison of PNDIs with prescriptions highlighted differences related to medication availability (OTC drugs) and patients' nonadherence (prescribed drugs). In addition, several instances of direct disposal events across the catchments were observed which raises an issue of lack of pharma compliance and general understanding of potential environmental impacts from pharma usage.

Assessment of restriction measures implemented during COVID pandemics on community lifestyle choices via wastewater-based epidemiology.

SARS-CoV-2 pandemic affected lifestyle habits, and the mental health and wellbeing of people around the world. In this manuscript, two towns (Paulton and Radstock) and two cities (Bath and Bristol) in Southwest England (> 1 million people) were monitored for two years using Wastewater-Based Epidemiology to assess impacts of COVID (including management measures such as lockdowns and movement restrictions) on community lifestyle choices: illicit drugs, legal stimulants, abused pharmaceuticals and pain pharma usage. Results were triangulated with key dates captured during the pandemic (national lockdowns, restrictions and social distancing measures, etc.). This highlighted a reduction in cocaine intake (as benzoylecgonine) (community average: -36 %) during the first lockdown and an increase in illicit drugs usage after the 3rd national lockdown (community averages for amphetamine: +8 %, cocaine/benzoylecgonine: +39 %, ketamine:+70 %) when restrictions were removed, and social interaction and recreational activities increased. There was a reduction in the intake of caffeine (as 1,7-dimethylxanthine) (community average:-39 %) after 3rd national lockdown coinciding with pubs reopening while nicotine intake (via cotinine) remained stable indicating lack of impact of COVID on smoking habits. Pain pharma often used in pain management resulting from injuries linked with sport activities (naproxen and diclofenac) showed decrease in usage due to lockdown restrictions in physical exercise and access to gyms/sport facilities.

Fate-and-transport modeling of SARS-CoV-2 for rural wastewater-based epidemiology application benefit.

Wastewater-based epidemiology (WBE) for the detection of agents of concern such as severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has been prevalent in literature since 2020. The majority of reported research focuses on large urban centers with few references to rural communities. In this research the EPA-Storm Water Management Model (EPA-SWMM) software was used to describe a small sewershed and identify the effects of temperature, temperature-affected decay rate, flow rate, flush time, fecal shedding rate, and historical infection rates during the spread of the Omicron variant of the SARS-CoV-2 virus within the sewershed. Due to the sewershed's relative isolation from the rest of the city, its wastewater quality behavior is similar to a rural sewershed. The model was used to assess city wastewater sampling campaigns to best appropriate field and or lab equipment when sampling wastewater. An important aspect of the assessment was the comparison of SARS-CoV-2 quantification methods with specifically between a traditional microbiological lab (practical quantitation limit, PQL, 1 GC/mL) versus what can be known from a field method (PQL 10 GC/mL). Understanding these monitoring choices will help rural communities make decisions on how to best implement the collection and testing for WBE agents of concern. An important outcome of this work is the knowledge that it is possible to simulate a WBE agent of concern with reasonable precision, if uncertainties are incorporated into model sensitivity. These ideas could form the basis for future mixed monitoring-modeling studies that will enhance its application and therefore adoption of WBE techniques in communities of many sizes and financial means.

Real-Time On-Site Monitoring of Viruses in Wastewater Using Nanotrap® Particles and RICCA Technologies.

Wastewater-based epidemiology (WBE) is an effective and efficient tool for the early detection of infectious disease outbreaks in a community. However, currently available methods are laborious, costly, and time-consuming due to the low concentration of viruses and the presence of matrix chemicals in wastewater that may interfere with molecular analyses. In the present study, we designed a highly sensitive "Quick Poop (wastewater with fecal waste) Sensor" (termed, QPsor) using a joint approach of Nanotrap microbiome particles and RICCA (RNA Isothermal Co-Assisted and Coupled Amplification). Using QPsor, the WBE study showed a strong correlation with standard PEG concentrations and the qPCR technique. Using a closed format for a paper-based lateral flow assay, we were able to demonstrate the potential of our assay as a real-time, point-of-care test by detecting the heat-inactivated SARS-CoV-2 virus in wastewater at concentrations of 100 copies/mL and within one hour. As a proof-of-concept demonstration, we analyzed the presence of viral RNA of the SARS-CoV-2 virus and PMMoV in raw wastewater samples from wastewater treatment plants on-site and within 60 min. The results show that the QPsor method can be an effective tool for disease outbreak detection by combining an AI-enabled case detection model with real-time on-site viral RNA extraction and amplification, especially in the absence of intensive clinical laboratory facilities. The lab-free, lab-quality test capabilities of QPsor for viral prevalence and transmission in the community can contribute to the efficient management of pandemic situations.

Influent wastewater analysis to investigate emerging trends of new psychoactive substances use in Europe.

Wastewater-based epidemiology (WBE) can provide objective and timely information on the use of new psychoactive substances (NPS), originally designed as legal alternatives of internationally controlled drugs. NPS have rapidly emerged on the global drug market, posing a challenge to drug policy and constituting a risk to public health. In this study, a WBE approach was applied to monitor the use of more than 300 NPS, together with fentanyl and its main metabolite norfentanyl, in influent wastewater collected from 12 European cities during March-June 2021. Quantitative and qualitative analysis of NPS in composite 24 h influent wastewater samples were based on solid phase extraction and liquid chromatography-mass spectrometry. In-sample stability tests demonstrated the suitability of most investigated biomarkers, except for a few synthetic opioids, synthetic cannabinoids and phenetylamines. Fentanyl, norfentanyl and eight NPS were quantified in influent wastewater and at least three substances were found in each city, demonstrating their use in Europe. N,N-dimethyltryptamine and 3-methylmethcathinone (3-MMC) were the most common NPS found, with the latter having the highest mass loads (up to 24.8 mg/day/1000 inhabitants). Seven additional substances, belonging to five categories of NPS, were identified in different cities. Spatial trends of NPS use were observed between cities and countries, and a changing weekly profile of use was observed for 3-MMC. WBE is a useful tool to rapidly evaluate emerging trends of NPS use, complementing common indicators (i.e. population surveys, seizures) and helping to establish measures for public health protection.

Urban wastewater-based epidemiology for multi-viral pathogen surveillance in the Valencian region, Spain.

Wastewater-based epidemiology (WBE) has lately arised as a promising tool for monitoring and tracking viral pathogens in communities. In this study, we analysed WBE's role as a multi-pathogen surveillance strategy to detect the presence of several viral illness causative agents. Thus, an epidemiological study was conducted from October 2021 to February 2023 to estimate the weekly levels of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), Respiratory Syncytial virus (RSV), and Influenza A virus (IAV) in influent wastewater samples (n = 69). In parallel, a one-year study (October 2021 to October 2022) was performed to assess the presence of pathogenic human enteric viruses. Besides, monitoring of proposed viral fecal contamination indicators crAssphage and Pepper mild mottle virus (PMMoV) was also assessed, along with plaque counting of somatic coliphages. Genetic material of rotavirus (RV), human astrovirus (HAStV), and norovirus genogroup I (GI) and GII was found in almost all samples, while hepatitis A and E viruses (HAV and HEV) only tested positive in 3.77 % and 22.64 % of the samples, respectively. No seasonal patterns were overall found for enteric viruses, although RVs had a peak prevalence in the winter months. All samples tested positive for SARS-CoV-2 RNA, with a mean concentration of 5.43 log genome copies per liter (log GC/L). The tracking of the circulating SARS-CoV-2 variants of concern (VOCs) was performed by both duplex RT-qPCR and next generation sequencing (NGS). Both techniques reliably showed how the dominant VOC transitioned from Delta to Omicron during two weeks in Spain in December 2021. RSV and IAV viruses peaked in winter months with mean concentrations 6.40 and 4.10 log GC/L, respectively. Moreover, the three selected respiratory viruses strongly correlated with reported clinical data when normalised by wastewater physico-chemical parameters and presented weaker correlations when normalising sewage concentration levels with crAssphage or somatic coliphages titers. Finally, predictive models were generated for each respiratory virus, confirming high reliability on WBE data as an early-warning system and communities illness monitoring system. Overall, this study presents WBE as an optimal tool for multi-pathogen tracking reflecting viral circulation and diseases trends within a selected area, its value as a multi-pathogen early-warning tool stands out due to its public health interest.

Adapted methods for monitoring influenza virus and respiratory syncytial virus in sludge and wastewater.

Wastewater-based surveillance constitutes a valuable methodology for the continuous monitoring of viral circulation, with the capacity to function as an early warning system. It holds particular significance in scenarios where respiratory viruses exhibit overlapping clinical presentations, as occurs with SARS-CoV-2, influenza virus (IV), and respiratory syncytial virus (RSV), and allows seasonal virus outbreaks to be distinguished from COVID-19 peaks. Furthermore, sewage sludge, given it harbors concentrated human waste from a large population, serves as a substantial reservoir for pathogen detection. To effectively integrate wastewater-based epidemiology into infectious disease surveillance, the detection methods employed in wastewater samples must be adapted to the distinct characteristics of sludge matrices. In this study, we adapted and applied protocols for the detection of IV and RSV in sewage sludge, comparing their performance with the results obtained in wastewater. To assess the efficiency of these protocols, sludge and wastewater samples were spiked with IV and RSV RNA, either free or incorporated in lentiviral particles. Samples were concentrated using the aluminum hydroxide adsorption-precipitation method before viral RNA extraction. Absolute virus quantification was carried out by RT-qPCR, including an internal control to monitor potential inhibitory factors. Recovery efficiencies for both free IV and RSV RNA were 60 % in sludge, and 75 % and 71 % respectively in wastewater, whereas the values for IV and RSV RNA in lentiviral particles were 16 % and 10 % in sludge and 21 % and 17 % in wastewater respectively. Additionally, the protocol enabled the quantification of naturally occurring IV and RSV in wastewater and sludge samples collected from two wastewater treatment plants during the winter months, thus affirming the efficacy of the employed methodologies.

Wastewater surveillance for antibiotics and resistance genes in a river catchment: Spatiotemporal variations and the main drivers.

Wastewater-based epidemiology (WBE) is used for mining information about public health such as antibiotics resistance. This study investigated the distribution profiles of six types of antibiotic resistance genes (ARGs) in wastewater and rivers in Wuhu City, China. The levels of ARGs found in the Qingyijiang River were significantly higher than other rivers, and were comparable to effluent levels. Among the ARGs, sulfonamides ARGs and intI1 were the predominant in both wastewaters and rivers. Additionally, the concentrations of ARGs were higher on weekends as opposed to weekdays. Their distribution patterns remained consistent inter-week and inter-season using linear regression analysis (p < 0.001). Interestingly, the occurrence levels of ARGs in wastewaters during spring were significantly higher than in autumn, although insignificant in rivers. The apparent removal rate of ARGs in domestic wastewater sources ranged from 61.52-99.29%, except for qepA (-1.91% to 81.09%), whereas the removal rates in mixed domestic and industrial wastewaters showed a marked decrease (-92.94% to 76.67%). A correlation network analysis revealed that azithromycin and erythromycin were key antibiotics, while blaNDM-1, tetM, tetB, and ermB were identified as key ARGs. Sulfonamide and fluoroquinolone antibiotics, and tetracycline and macrolide ARGs were the primary contributors. Linear mixed models demonstrated that socio-economic variables positively impacted the occurrence levels of ARGs, whereas wastewater flow and river runoff were the negative drivers for their concentrations in wastewaters and surface waters, respectively. Overall, this WBE study contributes to the understanding of spatiotemporal profiles and main drivers of the occurrence of ARGs in wastewater and receiving water.

Estimation of anti-diabetes drug metformin in Turkiye using wastewater-based epidemiology.

Metformin is the most commonly used drug for the treatment of type 2 diabetes (T2D), which is dramatically increasing due to factors such as increasing obesity, physical inactivity, and aging of the population. Metformin analysis was carried out in composite wastewater samples seasonally collected from wastewater treatment plants in 10 cities in 2019 and 2020 30 cities in 2021 in Turkiye. Metformin was measured in all wastewater samples, with an average concentration of 97.81 µg/l in 2019, 75.19 µg/l in 2020, and 69.13 µg/l in 2021. This study was utilized to predict metformin usage in different sociodemographic regions in Turkiye using a wastewater-based epidemiology (WBE) approach. As a result of the analysis, the average metformin consumption in Turkiye was estimated to be 22.2 ± 9.6 [1.9-63.8] g/d/1,000 persons (mean ± SD [range]). Furthermore, these estimates were compared with data for time, sociodemographic characteristics, and patient numbers. Assessing the correlation with estimates and the socioeconomic classes of the cities in question revealed that cities with high-income levels had the lowest metformin use rate. Finally, the study provides supporting data aiding the development of public health strategies for decreasing the overall load of T2D across Turkiye.

Shotgun proteomics to characterize wastewater proteins.

Classically, the characterization of wastewater components has been restricted to the measurement of indirect parameters (chemical and biological oxygen demand, total nitrogen) and small molecules of interest in epidemiology or for environmental control. Despite the fact that metaproteomics has provided important knowledge about the microbial communities in these waters, practically nothing is known about other non-microbial proteins transported in the wastewater. The method described here has allowed us to perform a large-scale characterization of the wastewater proteome. Wastewater protein profiles have shown to be very different in different collection sites probably reflecting their human population and industrial activities. We believe that wastewater proteomics is opening the doors to the discovery of new environmental and health biomarkers and the development of new, more effective monitoring devices for issues like monitorization of population health, pest control, or control of industry discharges. The method developed is relatively simple and combines procedures for the separation of the soluble and particulate fractions of wastewater and their concentration, and conventional shotgun proteomics using high-resolution mass spectrometry for protein identification. •Unprecedented method for wastewater proteome characterization.•Proteins as new potential biomarkers for sewage chemical-information mining, wastewater epidemiology and environmental monitoring.•Wastewater protein profiles reflect human and industrial activities.

To sample or not to sample: A governance-focused decision tree for wastewater service providers considering participation in wastewater-based epidemiology (WBE) in support of public health programs.

Wastewater-based epidemiology (WBE) provides value to public health monitoring and protection. Participation of public and private wastewater system operators in WBE efforts is critical to public health surveillance program success and sustainability. However, given the number of WBE solicitations wastewater service providers receive, the limitation of service provider resources, the concerns around privacy, ethics, and equity, and the fatigue associated with responding to COVID-19, operators are becoming more hesitant to participate in WBE efforts. While various ethical concerns and sustainability challenges associated with WBE have been documented, no efforts to date have investigated what factors should systematically influence the decision to provide samples to a WBE effort. Therefore, this study develops a decision-making tool for WBE teams to proactively monitor, manage, and avoid wastewater system operators' operational risks and potential liabilities. Ultimately, using this tool allows WBE program partners in academia, government, and industry to better understand wastewater system operators' needs and challenges surrounding data quality and use, public health ethics, and daily wastewater infrastructure operation.

Evaluation of wastewater-based epidemiology of COVID-19 approaches in Singapore's 'closed-system' scenario: A long-term country-wide assessment.

With the COVID-19 pandemic the use of WBE to track diseases spread has rapidly evolved into a widely applied strategy worldwide. However, many of the current studies lack the necessary systematic approach and supporting quality of epidemiological data to fully evaluate the effectiveness and usefulness of such methods. Use of WBE in a very low disease prevalence setting and for long-term monitoring has yet to be validated and it is critical for its intended use as an early warning system. In this study we seek to evaluate the sensitivity of WBE approaches under low prevalence of disease and ability to provide early warning. Two monitoring scenarios were used: (i) city wide monitoring (population 5,700,000) and (ii) community/localized monitoring (population 24,000 to 240,000). Prediction of active cases by WBE using multiple linear regression shows that a multiplexed qPCR approach with three gene targets has a significant advantage over single-gene monitoring approaches, with R2 = 0.832 (RMSE 0.053) for an analysis using N, ORF1ab and S genes (R2 = 0.677 to 0.793 for single gene strategies). A predicted disease prevalence of 0.001% (1 in 100,000) for a city-wide monitoring was estimated by the multiplexed RT-qPCR approach and was corroborated by epidemiological data evidence in three 'waves'. Localized monitoring setting shows an estimated detectable disease prevalence of ∼0.002% (1 in 56,000) and is supported by the geospatial distribution of active cases and local population dynamics data. Data analysis also shows that this approach has a limitation in sensitivity, or hit rate, of 62.5 % and an associated high miss rate (false negative rate) of 37.5 % when compared to available epidemiological data. Nevertheless, our study shows that, with enough sampling resolution, WBE at a community level can achieve high precision and accuracies for case detection (96 % and 95 %, respectively) with low false omission rate (4.5 %) even at low disease prevalence levels.

Interlaboratory comparison using inactivated SARS-CoV-2 variants as a feasible tool for quality control in COVID-19 wastewater monitoring.

Wastewater-based SARS-CoV-2 epidemiology (WBE) has proven as an excellent tool to monitor pandemic dynamics supporting individual testing strategies. WBE can also be used as an early warning system for monitoring the emergence of novel pathogens or viral variants. However, for a timely transmission of results, sophisticated sample logistics and analytics performed in decentralized laboratories close to the sampling sites are required. Since multiple decentralized laboratories commonly use custom in-house workflows for sample purification and PCR-analysis, comparative quality control of the analytical procedures is essential to report reliable and comparable results. In this study, we performed an interlaboratory comparison at laboratories specialized for PCR and high-throughput-sequencing (HTS)-based WBE analysis. Frozen reserve samples from low COVID-19 incidence periods were spiked with different inactivated authentic SARS-CoV-2 variants in graduated concentrations and ratios. Samples were sent to the participating laboratories for analysis using laboratory specific methods and the reported viral genome copy numbers and the detection of viral variants were compared with the expected values. All PCR-laboratories reported SARS-CoV-2 genome copy equivalents (GCE) for all spiked samples with a mean intra- and inter-laboratory variability of 19 % and 104 %, respectively, largely reproducing the spike-in scheme. PCR-based genotyping was, in dependence of the underlying PCR-assay performance, able to predict the relative amount of variant specific substitutions even in samples with low spike-in amount. The identification of variants by HTS, however, required >100 copies/ml wastewater and had limited predictive value when analyzing at a genome coverage below 60 %. This interlaboratory test demonstrates that despite highly heterogeneous isolation and analysis procedures, overall SARS-CoV-2 GCE and mutations were determined accurately. Hence, decentralized SARS-CoV-2 wastewater monitoring is feasible to generate comparable analysis results. However, since not all assays detected the correct variant, prior evaluation of PCR and sequencing workflows as well as sustained quality control such as interlaboratory comparisons are mandatory for correct variant detection.