Nationwide WWSCAN August 2024 Update

Hi everyone,

This newsletter contains updates on the WastewaterSCAN program with wastewater samples analyzed by the lab through August 31, 2024. Below you will find information on the program in terms of participating partners, review of targets being measured, as well as current wastewater categories (low, medium, high) of the infectious disease targets.

Ali & Marlene

WastewaterSCAN Participant Stats:

• 148 plants sampling
• >39 million people covered through sampling efforts
• 40 states

The methods for our assays are in the public domain and links for these are provided at the end of the newsletter.

New WastewaterSCAN Published Body of Work document:

• We are excited to share a document that provides an overview of the program’s peer-reviewed scientific literature, organized by topics and themes. Click here to access the WastewaterSCAN published body of work.

New Peer-Reviewed Scientific Publications:

• Prospective study of Candida auris nucleic acids in wastewater solids in 190 wastewater treatment plants in the United States suggests widespread occurrence
• Detection of Hemagglutinin H5 Influenza A Virus Sequence in Municipal Wastewater Solids at Wastewater Treatment Plants with Increases in Influenza A in Spring, 2024

SARS-CoV-2 N gene RNA median concentration across all sites in August 2024 was 182,288 copies/gram. For reference, the median concentration in July 2024 was 188,660 copies/gram. Below is the population weighted average SARS-CoV-2 N gene concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country over the last 365 days.

Nationally, SARS-CoV-2 is in the "High" category with aggregated measurements showing high concentrations and no significant upward or downward trend over the last 3 weeks.

Below are the population-weighted average SARS-CoV-2 N gene concentrations (normalized by PMMoV) from sites in the Midwest US (top left panel), Northeast US (top right panel), Southern US (bottom left panel) and Western US (bottom right panel) over the last 365 days. Raw data have been omitted from the plots for clarity of presentation but are still available in the chart settings on dashboard.

The Midwest, Northeast and South are in the "High" Wastewater Category due to high concentrations, with the South showing a downward trend in the last 3 weeks and the Midwest and Northeast showing no significant trend over the last 3 weeks. The West is in the "Medium" Wastewater Category due to medium concentrations and a downward trend in the last 3 weeks.

Below are the current SARS-CoV-2 wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level):

• 23 are in the LOW Wastewater Category
• 39 are in the MEDIUM Wastewater Category
• 76 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Sequencing of SARS-CoV-2 from wastewater is available for a select number of individual sites. The sequencing protocol is available on protocol.io. The plots below show the relative proportions of different variants inferred from sequencing the entire genome of SARS-CoV-2. Results are based on sequencing of 2 samples per week, combined to provide a weekly value.

Sequencing data is also available in aggregate across all sites in WWSCAN with sequencing data. That plot is shown below and also suggests that KP.3 (shown in light blue) is the most abundant variant across all sites (30.9%) followed by KP.2 (shown in slate blue) (29%) and JN.1.1.16 (shown in pink) (12.6%).

Influenza A (IAV) RNA median concentration across all sites August 2024 was below the limit of detection. For reference, the median concentration in July 2024 was below the limit of detection as well. Below is the population weighted average IAV concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country over the last 365 days.

Nationally, Influenza A is in the "Low" Wastewater Category with <50% of sites showing onset of IAV in wastewater.

Below are the current Influenza A wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level when in seasonal onset):

• 134 are in the LOW Wastewater Category
• 3 are in the MEDIUM Wastewater Category
• 1 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

H1 Marker in Influenza A (IAV) Since we started monitoring for the H1 marker in influenza A, 5 WastewaterSCAN sites have seen positive detections. You can find a heat map of positive detections here.

H3 Marker in Influenza A (IAV) Since we started monitoring for the H3 marker in influenza A, 5 WastewaterSCAN sites have seen positive detections. You can find a heat map of positive detections here.

H5 Marker in Influenza A (IAV) Since we started monitoring for the H5 marker in influenza A, 27 WastewaterSCAN sites have seen positive detections.

Influenza B (IBV) RNA was detected in ~1.7% of samples in August 2024. IBV RNA median concentration across all sites in August 2024 was below the limit of detection. Below is the population-weighted average IBV RNA concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country over the last 365 days.

Nationally, Influenza B is in the "Low" Wastewater Category with <50% of sites showing onset of IBV in wastewater.

Below are the current Influenza B wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level when in seasonal onset):

• 138 are in the LOW Wastewater Category
• 0 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Respiratory syncytial virus (RSV) RSV RNA median concentration across all sites in August 2024 was below the limit of detection. Below are the RSV RNA concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country over the last 365 days shown as the population-weighted average.

Nationally, RSV is in the "Low" Wastewater Category, with <50% of sites showing onset of RSV in wastewater.

Below are the current RSV wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level when in seasonal onset):

• 138 are in the LOW Wastewater Category
• 0 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

HMPV (human metapneumovirus) RNA median concentrations across all sites in August 2024 was below the level of detection. HMPV RNA was detected in ~6% of samples in August 2024. Below is the population weighted average HMPV RNA concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country over the last 365 days.

Nationally, HMPV is in the "LOW" Wastewater Category with <50% of sites showing onset of HMPV in wastewater.

Below are the current HMPV wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level):

• 138 are in the LOW Wastewater Category
• 0 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Enterovirus D68 (EV-D68) RNA was detected in ~60% of samples collected in August 2024 across WWSCAN sites and the median concentration across all sites was 3,509 copies/gram (for reference median concentration in July 2024 was below the limit of detection).

Nationally, EV-D68 is in the "Medium" Wastewater Category with >50% of positive detections in samples from the last 3 weeks.

The chart below plants in Texas as rows, and the days as columns (labeled at the bottom axis). White indicates no samples, blue is non-detect for EV-D68 RNA and red is detect for EV-D68 RNA.

Below are the current EV-D68 wastewater categories for all WWSCAN sites (recall the category is determined by the frequency of detection):

• 48 are in the LOW Wastewater Category
• 90 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Norovirus GII (HuNoV GII) RNA was detected in 100% of samples analyzed in August 2024. The median concentrations across all sites in August 2024 is 4,426,500 copies/gram (for reference median concentration in July 2024 was 7,252,88 copies/gram). Below are the HuNoV GII RNA concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country over the last 365 days. The population-weighted average is shown.

Nationally, Norovirus is in the "Medium" Wastewater Category with aggregated measurements showing medium concentrations and no significant upward or downward trend over the last 3 weeks.

Below are the current Norovirus wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level):

• 86 are in the LOW Wastewater Category
• 34 are in the MEDIUM Wastewater Category
• 18 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Rotavirus RNA median concentrations across all sites in August 2024 is 8,205 copies/gram. For reference, the median concentration in July 2024 was 21,367 copies/gram. Below are the Rotavirus RNA concentrations (normalized by PMMoV) from all participating WWSCAN sites across the country. The population-weighted average is shown.

Rotavirus is in the "Low" Wastewater Category nationally with aggregated measurements showing low concentrations with a downward trend in the last 3 weeks.

Below are the current Rotavirus wastewater categories for all WWSCAN sites (recall the category is determined by both the trend and the level):

• 105 are in the LOW Wastewater Category
• 27 are in the MEDIUM Wastewater Category
• 6 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Mpox clade II DNA - We have only detected mpox DNA in 9 samples in the month of August 2024. Nationally, Mpox clade II is in the "Low" category due to limited detections.

The chart below plants in HHS region 4 as rows, and days as columns (labeled at the bottom axis). White indicates no samples, blue is non-detect for mpox DNA and red is detect for mpox DNA.

Below are the Mpox clade II wastewater categories for all WastewaterSCAN sites (recall the category is determined by the frequency of detection):

• 138 are in the LOW Wastewater Category
• 0 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Candida auris (C.auris) DNA - We have detected C. auris DNA in ~4% of samples from all WWSCAN sites in August 2024. Nationally, C. auris is in the "Low" category due to limited detections.

The chart below shows plants in HHS region 5 as rows, and days as columns (labeled at the bottom axis). White indicates no samples, blue is non-detect for C. auris DNA and red is detect for C. auris DNA.

Below are the C. auris wastewater categories for all WastewaterSCAN sites (recall the category is determined by the frequency of detection):

• 137 are in the LOW Wastewater Category
• 1 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Hepatitis A (Hep A) RNA - We have detected Hep A RNA in ~13% of samples from all WWSCAN sites in August 2024. Nationally, HepA is in "Low" category due to limited detections.

The chart below shows each WWSCAN plant in HHS Region 1 as rows, and days as columns (labeled at the bottom axis). White indicates no samples, blue is a non-detect for Hep A RNA and red is a detection for Hep A RNA.

Below are the current HepA wastewater categories for all WWSCAN sites (the category is determined by the frequency of detection):

• 123 are in the LOW Wastewater Category
• 15 are in the MEDIUM Wastewater Category
• 0 are in the HIGH Wastewater Category
• 10 do not have enough data to calculate

Protocols (Pre-analytical methods and SARS-CoV-2 analytical methods including controls):

• High Throughput RNA Extraction and PCR Inhibitor Removal of Settled Solids for Wastewater Surveillance of SARS-CoV-2 RNA
• High Throughput pre-analytical processing of wastewater settled solids for SARS-CoV-2 RNA analyses
• Quantification of SARS-CoV-2 variant mutations (HV69-70, E484K/N501Y, del156-157/R158G, del143-145, LPPA24S, S:477-505, and ORF1a Del 141-143) in settled solids using digital RT-PCR
• High Throughput SARS-COV-2, PMMoV, and BCoV quantification in settled solids using digital RT-PCR

Pre-prints and peer-reviewed publications provided WWSCAN methods: 

• SARS-CoV-2 
• Influenza, Human metapneumovirus, Respiratory syncytial virus, Human parainfluenza
• Mpox clade II
• Norovirus GII
• Adenovirus group F, Rotavirus
• Candida auris
• Hepatitis A
• Enterovirus D68

If you have questions about the website or anything else, please email: wwscan_stanford_emory@lists.stanford.edu