Spatial and temporal variation in respiratory syncytial virus (RSV) subtype RNA in wastewater, and relation to clinical specimens

Respiratory syncytial virus (RSV) causes a large burden of respiratory illness, globally. It has two subtypes, RSV A and RSV B, but little is known regarding the predominance of these subtypes during different seasons and their impact on morbidity and mortality. Using molecular methods, we quantified RSV A and RSV B RNA in wastewater solids across multiple seasons and metropolitan areas to gain insight into the predominance of RSV subtypes. We determined the predominant subtype for each group using the proportion of RSV A to total RSV (RSV A + RSV B) in each wastewater sample (PA,WW), and conducted a comparative analysis temporally, spatially and against clinical specimens. A median PA,WW of 0.00 in the first season and 0.58 in the second season indicated a temporal shift in the predominant subtype. Spatially, while we observed dominance of the same subtype, PA,WW was higher in some areas (PA,WW = 0.58 to 0.88). The same subtype predominated in wastewater and clinical samples, but clinical samples showed higher levels of RSV A (RSV A positivity in clinical samples = 79%, median PA,WW = 0.58). These results suggest that wastewater, alongside clinical data, holds promise for enhanced subtype surveillance. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was funded by a Stanford Graduate Fellowship and the Sergey Brin Family Foundation. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: IRB of Stanford University gave ethical approval for this work; IRB 62834. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All wastewater data are available online at the Stanford Digital Repository. Clinical data are available upon request to the authors.