Comparison of gene targets and sampling regimes for SARS-CoV-2 quantification for wastewater epidemiology in UK prisons.

Prisons are high-risk settings for infectious disease transmission, due to their enclosed and semi-enclosed environments. The proximity between prisoners and staff, and the diversity of prisons reduces the effectiveness of non-pharmaceutical interventions, such as social distancing. Therefore, alternative health monitoring methods, such as wastewater-based epidemiology (WBE), are needed to track pathogens, including SARS-CoV-2. This pilot study assessed WBE to quantify SARS-CoV-2 prevalence in prison wastewater to determine its utility within a health protection system for residents. The study analysed 266 samples from six prisons in England over a 12-week period for nucleoprotein 1 (N1 gene) and envelope protein (E gene) using quantitative reverse transcriptase-polymerase chain reaction. Both gene assays successfully detected SARS-CoV-2 fragments in wastewater samples, with both genes significantly correlating with COVID-19 case numbers across the prisons (p < 0.01). However, in 25% of the SARS-positive samples, only one gene target was detected, suggesting that both genes be used to reduce false-negative results. No significant differences were observed between 14- and 2-h composite samples, although 2-h samples showed greater signal variance. Population normalisation did not improve correlations between the N1 and E genes and COVID-19 case data. Overall, WBE shows considerable promise for health protection in prison settings.

Comparison of collection methods for Phlebotomus argentipes sand flies to use in a molecular xenomonitoring system for the surveillance of visceral leishmaniasis.

BACKGROUND: The kala-azar elimination programme has resulted in a significant reduction in visceral leishmaniasis (VL) cases across the Indian Subcontinent. To detect any resurgence of transmission, a sensitive cost-effective surveillance system is required. Molecular xenomonitoring (MX), detection of pathogen DNA/RNA in vectors, provides a proxy of human infection in the lymphatic filariasis elimination programme. To determine whether MX can be used for VL surveillance in a low transmission setting, large numbers of the sand fly vector Phlebotomus argentipes are required. This study will determine the best method for capturing P. argentipes females for MX. METHODOLOGY/PRINCIPAL FINDINGS: The field study was performed in two programmatic and two non-programmatic villages in Bihar, India. A total of 48 households (12/village) were recruited. Centers for Disease Control and Prevention light traps (CDC-LTs) were compared with Improved Prokopack (PKP) and mechanical vacuum aspirators (MVA) using standardised methods. Four 12x12 Latin squares, 576 collections, were attempted (12/house, 144/village,192/method). Molecular analyses of collections were conducted to confirm identification of P. argentipes and to detect human and Leishmania DNA. Operational factors, such as time burden, acceptance to householders and RNA preservation, were also considered. A total of 562 collections (97.7%) were completed with 6,809 sand flies captured. Females comprised 49.0% of captures, of which 1,934 (57.9%) were identified as P. argentipes. CDC-LTs collected 4.04 times more P. argentipes females than MVA and 3.62 times more than PKP (p<0.0001 for each). Of 21,735 mosquitoes in the same collections, no significant differences between collection methods were observed. CDC-LTs took less time to install and collect than to perform aspirations and their greater yield compensated for increased sorting time. No significant differences in Leishmania RNA detection and quantitation between methods were observed in experimentally infected sand flies maintained in conditions simulating field conditions. CDC-LTs were favoured by householders. CONCLUSIONS/SIGNIFICANCE: CDC-LTs are the most useful collection tool of those tested for MX surveillance since they collected higher numbers of P. argentipes females without compromising mosquito captures or the preservation of RNA. However, capture rates are still low.

Research needs for optimising wastewater-based epidemiology monitoring for public health protection.

Wastewater-based epidemiology (WBE) is an unobtrusive method used to observe patterns in illicit drug use, poliovirus, and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The pandemic and need for surveillance measures have led to the rapid acceleration of WBE research and development globally. With the infrastructure available to monitor SARS-CoV-2 from wastewater in 58 countries globally, there is potential to expand targets and applications for public health protection, such as other viral pathogens, antimicrobial resistance (AMR), pharmaceutical consumption, or exposure to chemical pollutants. Some applications have been explored in academic research but are not used to inform public health decision-making. We reflect on the current knowledge of WBE for these applications and identify barriers and opportunities for expanding beyond SARS-CoV-2. This paper critically reviews the applications of WBE for public health and identifies the important research gaps for WBE to be a useful tool in public health. It considers possible uses for pathogenic viruses, AMR, and chemicals. It summarises the current evidence on the following: (1) the presence of markers in stool and urine; (2) environmental factors influencing persistence of markers in wastewater; (3) methods for sample collection and storage; (4) prospective methods for detection and quantification; (5) reducing uncertainties; and (6) further considerations for public health use.