skip to main content

Explore Research Products in the NSF-PAR

Title: Bacterial metatranscriptomes in wastewater can differentiate virally infected human populations
Monitoring wastewater samples at building-level resolution screens large populations for SARS-CoV-2, prioritizing testing and isolation efforts. Here we perform untargeted metatranscriptomics on virally-enriched wastewater samples from 10 locations on the UC San Diego campus, demonstrating that resulting bacterial taxonomic and functional profiles discriminate SARS-CoV-2 status even without direct detection of viral transcripts. Our proof-of-principle reveals emergent threats through changes in the human microbiome, suggesting new approaches for untargeted wastewater-based epidemiology.  more » « less
Award ID(s):
2038509
NSF-PAR ID:
10335834
Author(s) / Creator(s):
; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; more » ; ; « less
Date Published:
Journal Name:
bioRxiv
ISSN:
2692-8205
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. ; ; ; ; ; ; ; ; ; ; et al ( , mBio)
    Swanson, Michele S. (Ed.)
    ABSTRACT Wastewater surveillance (WS), when coupled with advanced molecular techniques, offers near real-time monitoring of community-wide transmission of SARS-CoV-2 and allows assessing and mitigating COVID-19 outbreaks, by evaluating the total microbial assemblage in a community. Composite wastewater samples (24 h) were collected weekly from a manhole between December 2020 and November 2021 in Maryland, USA. RT-qPCR results showed concentrations of SARS-CoV-2 RNA recovered from wastewater samples reflected incidence of COVID-19 cases. When a drastic increase in COVID-19 was detected in February 2021, samples were selected for microbiome analysis (DNA metagenomics, RNA metatranscriptomics, and targeted SARS-CoV-2 sequencing). Targeted SARS-CoV-2 sequencing allowed for detection of important genetic mutations, such as spike: K417N, D614G, P681H, T716I, S982A, and D1118H, commonly associated with increased cell entry and reinfection. Microbiome analysis (DNA and RNA) provided important insight with respect to human health-related factors, including detection of pathogens and their virulence/antibiotic resistance genes. Specific microbial species comprising the wastewater microbiome correlated with incidence of SARS-CoV-2 RNA, suggesting potential association with SARS-CoV-2 infection. Climatic conditions, namely, temperature, were related to incidence of COVID-19 and detection of SARS-CoV-2 in wastewater, having been monitored as part of an environmental risk score assessment carried out in this study. In summary, the wastewater microbiome provides useful public health information, and hence, a valuable tool to proactively detect and characterize pathogenic agents circulating in a community. In effect, metagenomics of wastewater can serve as an early warning system for communicable diseases, by providing a larger source of information for health departments and public officials. IMPORTANCE Traditionally, testing for COVID-19 is done by detecting SARS-CoV-2 in samples collected from nasal swabs and/or saliva. However, SARS-CoV-2 can also be detected in feces of infected individuals. Therefore, wastewater samples can be used to test all individuals of a community contributing to the sewage collection system, i.e., the infrastructure, such as gravity pipes, manholes, tanks, lift stations, control structures, and force mains, that collects used water from residential and commercial sources and conveys the flow to a wastewater treatment plant. Here, we profile community wastewater collected from a manhole, detect presence of SARS-CoV-2, identify genetic mutations of SARS-CoV-2, and perform COVID-19 risk score assessment of the study area. Using metagenomics analysis, we also detect other microorganisms (bacteria, fungi, protists, and viruses) present in the samples. Results show that by analyzing all microorganisms present in wastewater, pathogens circulating in a community can provide an early warning for contagious diseases. 
    more » « less
  2. ; ; ; ; ; ; ; ; ; ; et al ( , Journal of Travel Medicine)
    null (Ed.)
    Abstract Background Wastewater-based epidemiology (WBE) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be an important source of information for coronavirus disease 2019 (COVID-19) management during and after the pandemic. Currently, governments and transportation industries around the world are developing strategies to minimize SARS-CoV-2 transmission associated with resuming activity. This study investigated the possible use of SARS-CoV-2 RNA wastewater surveillance from airline and cruise ship sanitation systems and its potential use as a COVID-19 public health management tool. Methods Aircraft and cruise ship wastewater samples (n = 21) were tested for SARS-CoV-2 using two virus concentration methods, adsorption–extraction by electronegative membrane (n = 13) and ultrafiltration by Amicon (n = 8), and five assays using reverse-transcription quantitative polymerase chain reaction (RT-qPCR) and RT-droplet digital PCR (RT-ddPCR). Representative qPCR amplicons from positive samples were sequenced to confirm assay specificity. Results SARS-CoV-2 RNA was detected in samples from both aircraft and cruise ship wastewater; however concentrations were near the assay limit of detection. The analysis of multiple replicate samples and use of multiple RT-qPCR and/or RT-ddPCR assays increased detection sensitivity and minimized false-negative results. Representative qPCR amplicons were confirmed for the correct PCR product by sequencing. However, differences in sensitivity were observed among molecular assays and concentration methods. Conclusions The study indicates that surveillance of wastewater from large transport vessels with their own sanitation systems has potential as a complementary data source to prioritize clinical testing and contact tracing among disembarking passengers. Importantly, sampling methods and molecular assays must be further optimized to maximize detection sensitivity. The potential for false negatives by both wastewater testing and clinical swab testing suggests that the two strategies could be employed together to maximize the probability of detecting SARS-CoV-2 infections amongst passengers. 
    more » « less
  3. ; ; ; ; ; ; ; ( , Science of The Total Environment)
    Early detection of the COVID-19 virus, SARS-CoV-2, is key to mitigating the spread of new outbreaks. Data from individual testing is increasingly difficult to obtain as people conduct non-reported home tests, defer tests due to logistics or attitudes, or ignore testing altogether. Wastewater based epidemiology is an alternative method for surveilling a community while maintaining individual anonymity; however, a problem is that SARS-CoV-2 markers in wastewater vary throughout the day. Collecting grab samples at a single time may miss marker presence, while autosampling throughout a day is technically challenging and expensive. This study investigates a passive sampling method that would be expected to accumulate greater amounts of viral material from sewers over a period of time. Tampons were tested as passive swab sampling devices from which viral markers could be eluted with a Tween-20 surfactant wash. Six sewersheds in Detroit were sampled 16–22 times by paired swab (4 h immersion before retrieval) and grab methods over a five-month period and enumerated for N1 and N2 SARS-CoV-2 markers using ddPCR. Swabs detected SARS-CoV-2 markers significantly more frequently (P < 0.001) than grab samples, averaging two to three-fold more copies of SARS-CoV-2 markers than their paired grab samples (p < 0.0001) in the assayed volume (10 mL) of wastewater or swab eluate. No significant difference was observed in the recovery of a spiked-in control (Phi6), indicating that the improved sensitivity is not due to improvements in nucleic acid recovery or reduction of PCR inhibition. The outcomes of swab-based sampling varied significantly between sites, with swab samples providing the greatest improvements in counts for smaller sewersheds that otherwise tend to have greater variation in grab sample counts. Swab-sampling with tampons provides significant advantages in detection of SARS-CoV-2 wastewater markers and are expected to provide earlier detection of new outbreaks than grab samples, with consequent public health benefits. 
    more » « less
  4. ; ; ; ; ; ; ; ( , Environmental Science: Water Research & Technology)
    Background: recent applications of wastewater-based epidemiology (WBE) have demonstrated its ability to track the spread and dynamics of COVID-19 at the community level. Despite the growing body of research, quantitative synthesis of SARS-CoV-2 RNA levels in wastewater generated from studies across space and time using diverse methods has not been performed. Objective: the objective of this study is to examine the correlations between SARS-CoV-2 RNA levels in wastewater and epidemiological indicators across studies, stratified by key covariates in study methodologies. In addition, we examined the association of proportions of positive detections in wastewater samples and methodological covariates. Methods: we systematically searched the Web of Science for studies published by February 16th, 2021, performed a reproducible screening, and employed mixed-effects models to estimate the levels of SARS-CoV-2 viral RNA quantities in wastewater samples and their correlations to the case prevalence, the sampling mode (grab or composite sampling), and the wastewater fraction analyzed ( i.e. , solids, solid–supernatant mixtures, or supernatants/filtrates). Results: a hundred and one studies were found; twenty studies (671 biosamples and 1751 observations) were retained following a reproducible screening. The mean positivity across all studies was 0.68 (95%-CI, [0.52; 0.85]). The mean viral RNA abundance was 5244 marker copies per mL (95%-CI, [0; 16 432]). The Pearson correlation coefficients between the viral RNA levels and case prevalence were 0.28 (95%-CI, [0.01; 0.51]) for daily new cases or 0.29 (95%-CI, [−0.15; 0.73]) for cumulative cases. The fraction analyzed accounted for 12.4% of the variability in the percentage of positive detections, followed by the case prevalence (9.3% by daily new cases and 5.9% by cumulative cases) and sampling mode (0.6%). Among observations with positive detections, the fraction analyzed accounted for 56.0% of the variability in viral RNA levels, followed by the sampling mode (6.9%) and case prevalence (0.9% by daily new cases and 0.8% by cumulative cases). While the sampling mode and fraction analyzed both significantly correlated with the SARS-CoV-2 viral RNA levels, the magnitude of the increase in positive detection associated with the fraction analyzed was larger. The mixed-effects model treating studies as random effects and case prevalence as fixed effects accounted for over 90% of the variability in SARS-CoV-2 positive detections and viral RNA levels. Interpretations: positive pooled means and confidence intervals in the Pearson correlation coefficients between the SARS-CoV-2 viral RNA levels and case prevalence indicators provide quantitative evidence that reinforces the value of wastewater-based monitoring of COVID-19. Large heterogeneities among studies in proportions of positive detections, viral RNA levels, and Pearson correlation coefficients suggest a strong demand for methods to generate data accounting for cross-study heterogeneities and more detailed metadata reporting. Large variance was explained by the fraction analyzed, suggesting sample pre-processing and fractionation as a direction that needs to be prioritized in method standardization. Mixed-effects models accounting for study level variations provide a new perspective to synthesize data from multiple studies. 
    more » « less
  5. ; ; ; ; ; ; ; ; ; ; et al ( , Environmental Science: Water Research & Technology)
    Wastewater-based epidemiology has gained attention throughout the world for detection of SARS-CoV-2 RNA in wastewater to supplement clinical testing. Raw wastewater consists of small particles, or solids, suspended in liquid. Methods have been developed to measure SARS-CoV-2 RNA in the liquid and the solid fraction of wastewater, with some studies reporting higher concentrations in the solid fraction. To investigate this relationship further, six laboratories collaborated to conduct a study across five publicly owned treatment works (POTWs) where both primary settled solids obtained from primary clarifiers and raw wastewater influent samples were collected and quantified for SARS-CoV-2 RNA. Settled solids and influent samples were processed by participating laboratories using their respective methods and retrospectively paired based on date of collection. SARS-CoV-2 RNA concentrations, on a mass equivalent basis, were higher in settled solids than in influent by approximately three orders of magnitude. Concentrations in matched settled solids and influent were positively and significantly correlated at all five POTWs. RNA concentrations in both settled solids and influent were correlated to COVID-19 incidence rates in the sewersheds and thus representative of disease occurrence; the settled solids methods appeared to produce a comparable relationship between SARS-CoV-2 RNA concentration measurements and incidence rates across all POTWs. Settled solids and influent methods showed comparable sensitivity, N gene detection frequency, and calculated empirical incidence rate lower limits. Analysis of settled solids for SARS-CoV-2 RNA has the advantage of using less sample volume to achieve similar sensitivity to influent methods. 
    more » « less
  • Citation Formats
  • MLA
  • APA
  • Chicago
  • BibTeX
  • Save / Share this Record
  • Send to Email