Pandemic and post-pandemic dynamics of respiratory viruses in a Spanish middle-size city using a long-term wastewater surveillance

Wastewater-based surveillance has emerged over the past decade as one of the most informative complements to clinical testing for tracking viral pathogens at the population level. The fundamental premise rests on the fact that individuals infected with respiratory viruses shed viral genetic material not only through respiratory secretions but also, to varying degrees, through the gastrointestinal tract, which means that fragments of viral genomes routinely find their way into sewage systems. Because wastewater sampling aggregates material from entire sewersheds, a single composite sample can effectively represent the infection status of tens of thousands of people, capturing symptomatic cases, asymptomatic infections, and individuals who never seek medical care. This aggregation property became especially valuable during the COVID-19 pandemic, when clinical testing capacity was strained and testing policies changed repeatedly, making case counts unreliable indicators of true transmission dynamics. The Spanish study of a middle-size city contributes to a growing body of literature demonstrating that wastewater signals can anticipate or corroborate clinical trends for multiple respiratory pathogens simultaneously.

One of the distinguishing features of this research is its long-term scope, spanning both the acute pandemic phase and the post-pandemic transition period. Most wastewater surveillance studies published to date have focused on relatively short windows, often limited to pandemic waves of SARS-CoV-2, which restricts the ability to draw conclusions about how viral circulation behaves under more ordinary epidemiological conditions. By continuing collection through the period when public health interventions were lifted and society returned to pre-pandemic patterns of contact, the researchers were able to observe the re-establishment of seasonal respiratory virus dynamics that had been dramatically suppressed during 2020 and much of 2021. This before-and-after contrast is scientifically precious because it documents, in a single location with consistent methodology, how the near-total interruption of transmission for viruses such as influenza and respiratory syncytial virus was followed by unusual out-of-season resurgences and subsequently by a gradual return to typical winter seasonality.

The concept of a middle-size city is relevant to the broader applicability of wastewater surveillance. Much of the foundational work in this field has been conducted in large metropolitan areas, where sewersheds serve millions of people and dilution effects are substantial but signal magnitude is high. Smaller cities present a different set of conditions: the contributing population is smaller, which can make signals more sensitive to localized outbreaks but also more variable, and the sewer network characteristics, industrial discharges, and demographic composition differ from those of megacities. Demonstrating that a standardized analytical pipeline can produce interpretable, reproducible data in a middle-size urban setting strengthens the case for deploying such systems across heterogeneous municipalities, which is precisely what many national and regional surveillance programs in Europe now aim to do under frameworks supported by the European Commission and coordinated through initiatives involving public health institutes across member states.

Methodologically, long-term wastewater studies of respiratory viruses must contend with several persistent analytical challenges. Viral RNA in sewage degrades over time depending on temperature, pH, and the presence of nucleases, so normalization strategies are needed to distinguish true changes in viral shedding from artifacts introduced by variable wastewater flow, rainfall dilution, or sample processing efficiency. Common approaches include normalizing to fecal indicators such as human adenovirus or pepper mild mottle virus, or to physicochemical parameters like ammonium concentration and flow volume. Recovery controls, typically spiked surrogate viruses, allow laboratories to estimate extraction efficiency for each sample. The choice of concentration method, whether electronegative membrane filtration, ultrafiltration, or polyethylene glycol precipitation, influences sensitivity for different viruses. Studies that maintain the same protocol over years, as this one did, gain an important advantage: temporal comparisons become more reliable because methodological noise is held constant, allowing genuine epidemiological trends to stand out more clearly.

The multipathogen panel typical of such studies generally includes SARS-CoV-2, influenza A and B viruses, respiratory syncytial virus, and often additional targets such as human metapneumovirus, parainfluenza viruses, seasonal coronaviruses, rhinoviruses, and adenoviruses. Quantitative reverse transcription PCR remains the workhorse detection technology because it provides absolute or relative quantification with well-characterized performance. Multiplexing several assays in a single reaction conserves sample volume and reduces cost, which matters when hundreds of samples are processed over multi-year campaigns. The resulting time series can be analyzed for peak timing, peak height, epidemic onset, and the lead time between wastewater signal and clinical indicators such as hospital admissions or sentinel physician reports. Across many studies, wastewater signals for influenza and RSV have tended to lead or coincide with clinical peaks by roughly one to two weeks, a window that can be operationally meaningful for hospital preparedness, staffing decisions, and the timing of public health communications.

The pandemic-to-post-pandemic transition also offers a natural experiment in viral interference and immune landscape dynamics. During the period of intense SARS-CoV-2 circulation and non-pharmaceutical interventions, the near-disappearance of influenza and RSV created a substantial immunity debt, particularly among children born during those years who had never encountered RSV. When restrictions eased, many countries in the Northern Hemisphere, including Spain, experienced an out-of-season RSV wave in the summer of 2021 and an unusually early and intense influenza and RSV season in late 2022. A wastewater time series that spans these events provides an independent record of how quickly viral circulation rebounded and how the relative timing of different pathogens shifted, information that is difficult to reconstruct from clinical data alone because testing practices for non-COVID respiratory viruses were themselves disrupted during the pandemic.

Another dimension of long-term wastewater data is its potential to capture the emergence and replacement of SARS-CoV-2 variants. Variant-specific assays or sequencing of wastewater samples can reveal the rise of Alpha, Delta, Omicron, and subsequent lineages weeks before genomic surveillance of clinical samples detects the same shifts, simply because wastewater aggregates infections across the whole community without the sampling biases introduced by who gets tested. Even when the primary focus of a study is quantitative viral load rather than lineage tracking, the overall SARS-CoV-2 signal reflects the cumulative effect of variant-driven changes in transmissibility, immune evasion, and shedding kinetics. The post-pandemic period, characterized by the evolution of Omicron sublineages and the transition of COVID-19 toward an endemic, wave-like pattern, is particularly interesting in this respect, as wastewater data can help define whether SARS-CoV-2 settles into winter seasonality similar to influenza or retains a distinct periodicity.

From a public health operations standpoint, the value of a multi-year dataset lies in establishing baselines. A single season of data cannot tell decision-makers whether a given viral load measurement represents a normal winter peak or an anomalous surge. After several years of consistent monitoring, thresholds can be defined empirically, for example as multiples of the median off-season concentration, and these thresholds can trigger predefined responses such as enhanced clinical testing, hospital surge planning, or targeted vaccination campaigns. The European Union’s recommendation in 2023 that member states extend wastewater surveillance beyond SARS-CoV-2 to include other pathogens of concern reflects exactly this logic: sustained, standardized monitoring is what converts raw measurements into actionable intelligence. Studies conducted in individual cities, with fully documented protocols and openly reported concentrations, provide the calibration points that such larger programs depend upon.

It is also worth noting the complementary relationship between wastewater surveillance and clinical sentinel systems. Clinical data provide information that wastewater cannot: which individuals are infected, their age distribution, vaccination status, symptom severity, and the identification of specific strains through patient sampling. Wastewater data, conversely, provide population-level coverage without dependence on healthcare-seeking behavior or testing policy, and they are available even when clinical laboratories scale back routine respiratory panels during off-seasons. Integrating the two streams, for instance by correlating wastewater concentrations with hospitalization rates or by using wastewater to trigger more intensive clinical sampling, generally yields better situational awareness than either source alone. The Spanish middle-size city dataset, by covering both pandemic and post-pandemic phases, illustrates how this integration can be evaluated across very different epidemiological regimes, from emergency-driven mass testing to routine seasonal monitoring.

Finally, the scientific community’s interest in studies of this kind reflects a broader shift in how infectious disease surveillance is conceptualized. Rather than reacting to outbreaks after they become clinically visible, public health authorities increasingly seek leading indicators drawn from environmental monitoring, genomic sequencing, and digital data sources. Wastewater surveillance occupies a central place in this vision because it is relatively inexpensive per capita, technologically accessible to regional laboratories, and demonstrably effective across a growing list of pathogens, including not only respiratory viruses but also enteroviruses, hepatitis A, mpox, and antimicrobial resistance genes. Long-term, single-site studies with consistent methodology, such as the one conducted in this Spanish city, serve as the empirical backbone for this transition, demonstrating that the signals are stable, interpretable, and reproducible over years rather than weeks, and that the infrastructure built during the COVID-19 emergency can be repurposed into durable, routine surveillance capacity capable of informing responses to future epidemic threats.

Subject of Research: Pandemic and post-pandemic dynamics of respiratory viruses in a Spanish middle-size city using a long-term wastewater surveillance

Article Title: Pandemic and post-pandemic dynamics of respiratory viruses in a Spanish middle-size city using a long-term wastewater surveillance

Article References: Casado-Martín, L., Hernández, M., Pérez-Alonso, D., Yeramian, N., Alves-Elois, M., Dorighello-Cadamuro, R., Fongaro, G., Eiros, J. M., & Rodríguez-Lázaro, D. (2026). Pandemic and post-pandemic dynamics of respiratory viruses in a Spanish middle-size city using a long-term wastewater surveillance. npj Viruses. https://doi.org/10.1038/s44298-026-00232-2

Image Credits: AI Generated

DOI: 10.1038/s44298-026-00232-2

Keywords: Pandemic, post-pandemic, dynamics, respiratory, viruses, Spanish, middle-size, city, long-term, wastewater, surveillance, scientific research