Building-level wastewater surveillance localizes interseasonal influenza variation

Abstract

Influenza virus poses a recurring threat to public health and infects many populations in annual waves of generally unpredictable magnitude and timing. We aimed to detect the arrival and estimate the case magnitude of seasonal influenza A in urban New York City college dormitory buildings. Our wastewater-based surveillance (WBS) program measured viral RNA in the sewage outflow of three dormitories at Barnard College in 2021 and 2022. Wastewater test positivity strongly correlated with New York County clinical cases (Kendall's τ = 0.58). Positive wastewater samples are also associated with campus clinical cases. The 2022 data stand in stark contrast to the 2021 results by revealing the more frequent and earlier presence of influenza A. The increase in positive tests is significant (P < 0.01). It is further noteworthy that positive samples were not evenly distributed among buildings. Surveillance additionally identified the influenza A H3 subtype but did not detect any influenza B. We also systematically analyzed our viral purification protocol to identify in which fraction influenza can be found. While virus can be found in solid fractions, a substantial quantity remains in the final liquid fraction. Our work focuses on individual buildings rather than larger sewersheds because buildings may localize interseasonal influenza variation to specific subpopulations. Our results highlight the potential value of building-level WBS in measuring influenza incidence to help guide public health intervention.IMPORTANCESeasonal influenza remains a major public health burden. We monitored influenza A in dormitory wastewater of a New York City college in 2021 and 2022. Longitudinal samples acquired over consecutive years allowed measurement of individual buildings between seasons. We uncovered building-level changes in the magnitude and timing of test positivity concordant with clinical cases. Surveillance also localized the heterogeneity of influenza variation during the large 2022 seasonal surge. The ability to detect such changes could be leveraged as part of a public health response.

Keywords: New York City; influenza A virus; students; wastewater; wastewater-based epidemiological monitoring.

Fig 1

Wastewater-based surveillance of influenza A in dormitory building wastewater compared to local clinical case reports. Results for PCR testing of building wastewater from three dormitories during the (A) 2021 and (B) 2022 influenza seasons. Each box indicates a week consisting of two wastewater sampling dates. Positive and negative wastewater tests are depicted as reddish purple and bluish green squares, respectively. Invalid wastewater tests or the absence of sampling is depicted as a white square. A 3 denotes a positive H3 subtype PCR test. An X denotes a clinical confirmed case of a building resident. The height of the blue bars indicates the number of clinical confirmed cases reported in New York County for CDC weeks 40–52.

Fig 2

Quantitative assessment of the association between influenza A campus wastewater-based surveillance and local clinical case reports. Correlation between the number of clinical confirmed cases reported in New York County and (A) weekly wastewater test positivity or (B) C_t values of undiluted RNA. Data from 2021 and 2022 were combined. The correlation coefficient is shown as Kendall’s τ with a P value of <0.05.

Fig 3

Wastewater-based surveillance of influenza B in dormitory building wastewater compared to clinical case reports. Results for PCR testing of building wastewater from three dormitories during the 2022 influenza season. Each box indicates a week consisting of two wastewater sampling dates. Positive and negative wastewater tests are depicted as reddish purple and bluish green squares, respectively. Invalid wastewater tests or the absence of sampling is depicted as a white square. The height of the blue bars indicates the number of clinical confirmed cases reported in New York County for CDC weeks 40–52.

Fig 4

Partitioning and stability of influenza A in wastewater. (A) Schematic of accessible preparation of influenza A from wastewater for RNA analysis. Underlined labels indicate fractions subjected to RNA purification. (B) Quantitation of influenza A RNA purification from different fractions. C_t values of undiluted RNA were obtained in technical duplicate for each fraction. ND indicates not detected. Dot colors indicate n = 5 independent biological replicates. (C) Quantitation of influenza A RNA purification from the same sample over time. C_t values of undiluted RNA were obtained in technical duplicate for each time point. ND indicates not detected. A positive signal in the last 5 of the 45 PCR cycles is assigned a value of 40. Dot colors indicate n = 4 independent biological replicates.