Environmental Science & Technology Letters

BackgroundContinuous, non-invasive viral surveillance is essential to monitor emerging pathogens and guide public health responses. Most environmental surveillance studies use targeted qPCR approaches, and comparisons between wastewater and indoor air surveillance remain limited. We aimed to compare the utility of emergency department indoor air and urban wastewater for tracking circulating viruses and resolving genomic information. MethodsWe conducted a matched-pair study comparing 19 weekly indoor air samples from the central ventilation exhaust shaft of an emergency department and 19 24-hour composite municipal wastewater samples in Leuven, Belgium, from December 2024 to April 2025. Both sample sets were processed using probe-based hybrid-capture viral metagenomics targeting over 3000 viral species, using influenza A as a clinically relevant test case. FindingsWastewater captured higher overall viral diversity (233 versus 106 species) and more complete genomes compared to indoor air, showing a relatively stable composition, mainly of enteric and animal-associated viruses. Indoor air demonstrated lower overall diversity but was enriched for respiratory viruses, including influenza A, coronaviruses, metapneumovirus, and respiratory syncytial virus, and more frequently achieved high genome coverage for these pathogens. Although both sample types permitted influenza A subtype characterization, influenza A genomes from wastewater were often less well covered. When coverage thresholds were met, indoor air supported targeted antiviral resistance-site screening for influenza A and RSV-A. InterpretationWastewater and indoor air generate distinct but complementary viromes. Wastewater acts as a diverse, population-level monitor for One-Health applications, whereas indoor air serves as a targeted, human-centric sentinel system facilitating further genomic characterization for respiratory viruses. FundingMustafa Karatas is supported by a Research Foundation Flanders (FWO) fundamental research scholarship (number: 11P7I24N). C.G., L.C., E.H., S.G. and E.A. acknowledge support from the DURABLE project. The DURABLE project has been co-funded by the European Union, under the EU4Health Programme (EU4H), project no. 101102733. Research in context Evidence before this studyWe searched PubMed for studies published between Jan 2000 and March 2024 using the terms "wastewater surveillance", "metagenomics", "indoor air", and "viral metagenomics". Previous studies have shown that wastewater surveillance can detect population-level viral circulation, and more recent work has explored indoor air sampling as a method for monitoring respiratory virus transmission. However, environmental metagenomic studies have largely examined these two sample types separately. Furthermore, most studies relied on untargeted sequencing approaches, which often yield fragmented genomes in these environments. To date, no study has systematically compared indoor air and wastewater using a comprehensive hybrid-capture viral metagenomics approach for virus surveillance. Added value of this studyWe conducted a matched comparison of indoor air from a hospital emergency department and municipal wastewater collected during the same weeks in Leuven, Belgium. We analyzed both sample types using an identical hybrid-capture viral metagenomics workflow targeting more than 3000 viral species. This design enabled a direct evaluation of how the two environmental surveillance lenses differ in viral diversity, genomic recovery, and epidemiological relevance. Wastewater captured broader viral diversity and a stable background dominated by enteric and animal-associated viruses, whereas indoor air captured more respiratory viruses and more frequently yielded high genome completeness for these pathogens. When genome coverage thresholds were met, indoor air data enabled influenza subtype identification and screening for antiviral resistance markers. Implications of all the available evidenceOur findings support a layered environmental surveillance strategy in which different environmental samples provide complementary information. Wastewater offers a stable, population-level view of viral circulation and captures broad viral diversity, including human and animal-associated viruses. Indoor air sampling in human-dominated settings provides a more direct signal of respiratory virus circulation and can yield genomes suitable for subtype and mutation-level characterization. Combining these approaches could strengthen metagenomic surveillance frameworks by improving the interpretation of environmental viral signals, supporting early detection of emerging pathogens, and helping distinguish human virus circulation from environmental or animal-derived detections.

Wastewater monitoring can provide insights into respiratory disease occurrence in communities that contribute to the wastewater system. Using daily measurements of RNA of influenza A (IAV), respiratory syncytial virus (RSV), and human metapneumovirus (HMPV), as well as SARS-CoV-2 in wastewater solids from eight publicly owned treatment works in the Greater San Francisco Bay Area of California between July 2022 until early May 2023, we identify a "tripledemic" when concentrations of IAV, RSV, and SARS-CoV-2 peaked at approximately the same time. HMPV was also widely circulating. We designed novel hydrolysis probe RT-PCR assays for different IAV subtype makers to discern that the dominant circulating IAV subtype was H3N2. We show that wastewater data can be used to identify onset and offset of wastewater disease occurrence events that can provide insight into disease epidemiology and timely, localized information to inform hospital staffing and clinical decision making to respond to circulating viruses. Whereas RSV and IAV wastewater events were mostly regionally coherent, HMPV events displayed localized occurrence patterns.

BackgroundRespiratory disease is a major cause of morbidity and mortality; however, current surveillance for circulating respiratory viruses is passive and biased. Seasonal circulation of respiratory viruses changed dramatically during the COVID-19 pandemic. More active methods for understanding respiratory disease dynamics are needed to better inform public health response and to guide clinical decision making. Wastewater-based epidemiology has been used to understand COVID-19, influenza A, and RSV infection rates at a community level, but has not been used to investigate other respiratory viruses. MethodsWe measured concentrations of influenza A and B, RSV A and B, human parainfluenza (1-4), rhinovirus, seasonal human coronaviruses, and human metapneumovirus RNA in wastewater solids three times per week for 17 months spanning the COVID-19 pandemic at a wastewater treatment plant in California, USA. Novel probe-based assays were developed and validated for non-influenza viral targets. We compared viral concentrations to positivity rates for viral infections from clinical specimens submitted to sentinel laboratories. FindingsWe detected RNA from all target viruses in wastewater solids. Human rhinovirus and seasonal coronaviruses were found at highest concentrations. Concentrations of viruses correlated significantly and positively with positivity rates of associated viral diseases from sentinel laboratories. Measurements from wastewater indicated limited circulation of RSV A and influenza B, and human coronavirus OC43 dominated the seasonal human coronavirus infections while human parainfluenza 1 and 4A dominated among parainfluenza infections. InterpretationWastewater-based epidemiology can be used to obtain information on circulation of respiratory viruses at a community level without the need to test many individuals because a single sample of wastewater represents the entire contributing community. Results from wastewater can be available within 24 hours of sample collection, allowing real time information to inform public health response, clinical decision making, and individual behavior modifications.

Wastewater surveillance of respiratory pathogens can provide timely estimates of viral activity and disease trends in a population. Indoor air surveillance could be used similarly with some advantages but remains largely unvalidated at the community-scale. Here, an indoor air surveillance program was employed as part of public health environmental surveillance in Chicago, Illinois, USA. Ten air samplers were placed in healthcare and congregate living settings across the city. Weekly air samples were evaluated for influenza A, influenza B, respiratory syncytial virus, and SARS-CoV-2 over two respiratory virus seasons. Citywide, aggregated air sample positivity and viral load were closely correlated with local clinical case and wastewater surveillance data across all respiratory viruses. Virus trends in air data often preceded clinical and wastewater, although this varied across pathogens and respiratory virus seasons. Further, whole-genome sequencing of SARS-CoV-2 showed close correlation of variant proportions across all datasets. At the building-scale, air samples obtained from a single sampling device provided efficient respiratory virus surveillance, with well-correlated estimates of respiratory pathogens. These data demonstrate that air surveillance can provide accurate estimates of respiratory virus infections and variants at a building or community-scale, serving as an alternative or complementary tool for public health environmental surveillance.

Avian influenza (serotype H5N1) is a highly pathogenic virus that emerged in domestic waterfowl in 1996. Over the past decade, zoonotic transmission to mammals, including humans, has been reported. Although human to human transmission is rare, infection has been fatal in nearly half of patients who have contracted the virus in past outbreaks. The increasing presence of the virus in domesticated animals raises substantial concerns that viral adaptation to immunologically naive humans may result in the next flu pandemic. Wastewater-based epidemiology (WBE) to track viruses was historically used to track polio and has recently been implemented for SARS-CoV2 monitoring during the COVID-19 pandemic. Here, using an agnostic, hybrid-capture sequencing approach, we report the detection of H5N1 in wastewater in nine Texas cities, with a total catchment area population in the millions, over a two-month period from March 4th to April 25th, 2024. Sequencing reads uniquely aligning to H5N1 covered all eight genome segments, with best alignments to clade 2.3.4.4b. Notably, 19 of 23 monitored sites had at least one detection event, and the H5N1 serotype became dominant over seasonal influenza over time. A variant analysis suggests avian or bovine origin but other potential sources, especially humans, could not be excluded. We report the value of wastewater sequencing to track avian influenza.

Measles incidence has increased in recent years as vaccination rates have dropped globally. However, there are challenges in surveillance of measles; measles presents similarly to other diseases and can be misdiagnosed. The lag between infectivity and symptom onset also poses a challenge for surveillance, as measles is highly infectious and significant transmission can occur before case identification. Wastewater monitoring of measles RNA could help to fill gaps in clinical surveillance. In this study, we developed a novel assay to detect wild-type measles virus in wastewater; through both in silico and in vivo tests, we demonstrated assay specificity and sensitivity. We conducted both retrospective and prospective monitoring in a sewershed adjacent to ongoing outbreak areas in the region in the United States from December 2024 - May 2025. In total, 11 of 105 (10.5%) of samples were positive for measles with a median concentration of 6,900 gene copies per dry gram of wastewater solids. Overall, we demonstrate that measles is detectable in wastewater during an ongoing outbreak and that wastewater monitoring of measles can result in early warning over clinical surveillance.

Wastewater represents a composite biological sample from the entire contributing population. People infected with monkeypox excrete monkeypox virus DNA via skin lesions, saliva, feces and urine and these can enter the wastewater via toilets, sinks, and shower drains. To test whether monkeypox can be detected and monitored in wastewater during a period when publicly reported monkey cases in the region were increasing, we deployed digital PCR assays that target genomic DNA from the monkeypox virus in our routine, ongoing wastewater surveillance program in the Greater Bay Area of California, USA. We measured monkeypox virus DNA daily in settled solids samples from nine wastewater plants over the period of approximately 4 weeks. During that period, we detected monkeypox virus DNA in wastewater solids at nearly all the wastewater plants we routinely sample. Frequency of occurrence and concentrations were highest at plants serving San Francisco County. To confirm the presence of monkeypox DNA, we used two assays that target distinct sequences on the monkeypox genome on a subset of samples and results from both assays were in close agreement strongly suggesting true positives in the wastewater. Additionally, we show that concentrations of monkeypox DNA is 103 times higher in the solid fraction compared to the liquid fraction of wastewater on a mass-equivalent basis.

Early detection of outbreaks and emerging pathogens is critical for public health and global biosecurity. Airports, as major international travel hubs with dense, enclosed populations, are high-risk settings for disease transmission and potential pathogen introduction. The U.S. Centers for Disease Control and Prevention, in collaboration with Ginkgo Biosecurity and the University of Wisconsin-Madison, implemented air monitoring for pathogen surveillance in congregate areas at four U.S. international airports. From October 2023 to August 2024, SARS-CoV-2 was detected by PCR in 98.3% of air samples and influenza A in 17.2%. These results correlated with positivity trends from other sample modalities, including aviation wastewater, traveler nasal swabs, and national clinical surveillance data. Targeted amplicon sequencing of SARS-CoV-2 from air samples correlated with contemporaneous lineages in wastewater collected and sequenced from the same airports. Metagenomic enrichment sequencing detected 30 viral species and recovered high-quality genomes for SARS-CoV-2, influenza, bocavirus, and seasonal coronaviruses. Together, these findings demonstrate that air sampling is a complementary surveillance modality to aviation wastewater for early pathogen detection at ports of entry.

The sequencing of human virus genomes from wastewater samples is an efficient method for tracking viral transmission and evolution at the community level. However, this requires the recovery of viral nucleic acids of high quality. We developed a reusable tangential-flow filtration system to concentrate and purify viruses from wastewater for whole-genome sequencing. A pilot study was conducted with 94 wastewater samples from four local sewersheds, from which viral nucleic acids were extracted, and the whole genome of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was sequenced using the ARTIC V4.0 primers. Our method yielded a high probability (0.9) of recovering complete or near-complete SARS-CoV-2 genomes (>90% coverage at 10x depth) from wastewater when the COVID-19 incidence rate exceeded 33 cases per 100 000 people. The relative abundances of sequenced SARS-CoV-2 variants followed the trends observed from patient-derived samples. We also identified SARS-CoV-2 lineages in wastewater that were underrepresented or not present in the clinical whole-genome sequencing data. The developed tangential-flow filtration system can be easily adopted for the sequencing of other viruses in wastewater, particularly those at low concentrations. SYNOPSISThe tangential-flow filtration method extracts viral nucleic acids of high enough quality from wastewater for robust and successful whole-genome sequencing. GRAPHIC FOR TABLE OF CONTENTS (TOC) O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/22279692v2_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@566377org.highwire.dtl.DTLVardef@19c3ba7org.highwire.dtl.DTLVardef@106c70org.highwire.dtl.DTLVardef@3f3f8f_HPS_FORMAT_FIGEXP M_FIG C_FIG

BackgroundWidespread use of at-home COVID-19 tests hampers determination of community COVID-19 incidence. Using nationwide data available through the US National Wastewater Surveillance System, we examined the performance of two wastewater metrics in predicting high case and hospitalizations rates both before and after widespread use of at-home tests. MethodsWe performed area under the receiver operating characteristic (ROC) curve analysis (AUC) for two wastewater metrics--viral concentration relative to the peak of January 2022 ("wastewater percentile") and 15-day percent change in SARS-CoV-2 ("percent change"). Dichotomized reported cases ([&ge;] 200 or <200 cases per 100,000) and new hospitalizations ([&ge;] 10 or <10 per 100,000) were our dependent variables, stratified by calendar quarter. Using logistic regression, we assessed the performance of combining wastewater metrics. ResultsAmong 268 counties across 22 states, wastewater percentile detected high reported case and hospitalizations rates in the first quarter of 2022 (AUC 0.95 and 0.86 respectively) whereas the percent change did not (AUC 0.54 and 0.49 respectively). A wastewater percentile of 51% maximized sensitivity (0.93) and specificity (0.82) for detecting high case rates. A model inclusive of both metrics performed no better than using wastewater percentile alone. The predictive capability of wastewater percentile declined over time (AUC 0.84 and 0.72 for cases for second and third quarters of 2022). ConclusionNationwide, county wastewater levels above 51% relative to the historic peak predicted high COVID rates and hospitalization in the first quarter of 2022, but performed less well in subsequent quarters. Decline over time in predictive performance of this metric likely reflects underreporting of cases, reduced testing, and possibly lower virulence of infection due to vaccines and treatments.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), an elusive and highly pathogenic agent, has resulted in the ongoing COVID-19 pandemic affecting numerous populations worldwide. New studies investigating the tenacity of SARS-CoV-2 have highlighted its ability to persist on a myriad of surfaces for several days, including gowns and shoes. As a result, there is a global need for sterilization of a variety of potentially-contaminated items, ranging from clothing to personal protective equipment like face coverings. To this end, we have designed and constructed a cost-effective, scalable, and sustainable sterilization system that uses ozone gas to inactivate viral particles. We sought to determine the efficacy of the system in the sterilization of viral particles as well as its ability to sterilize N95 respirators for reuse. N95 respirators inoculated with P22 bacteriophage and sterilized in the ozone system showed a 6-log10 reduction in viral load when treated at 25 ppm for 150 minutes. Further, N95 respirators treated with five 150-minute cycles at 35 ppm for a total concentration-time product (CT) of 26,250 ppm min in the ozone system showed comparable filtration efficiency to untreated N95 respirators in a 50 to 200 nmr particulate challenge filtration test. Interestingly, the surgical N95 respirators tested showed complete inactivation of fluid resistance and degradation of the elasticity of polyisoprene straps after five cycles in the sterilization system. Taken together, these data suggest that while our ozone system may negatively affect certain protective aspects of surgical N95 respirators, it does effectively sterilize viral particles and can be utilized for a multitude of other use cases, including sterilizing polypropylene face coverings after potential SARS-CoV-2 contamination. In addition to providing long-term environmental benefits, deployment of this system during the ongoing pandemic reduces the risk of COVID-19 community transmission while conserving monetary resources otherwise spent on the continuous purchase of disposable face coverings.

High-level exposure to indoor air pollutants (IAPs), including volatile organic compounds (VOCs), has substantially contributed to the burden of disease in China over the past two decades. However, the source contributions to the indoor VOC-related health burden remain unknown. This study utilized a novel approach based on positive matrix factorization (PMF) of indoor multipollutant data to estimate the source-specific residential VOC concentrations and associated burden of disease. Indoor concentrations of 39 VOCs were collected repeatedly in different seasons from 2016 to 2017 in 249 residences across nine cities in China. In 2017, the disability-adjusted life years (DALYs) attributable to residential VOC exposure across nine provinces in China reached 134.2 (95% UI: 65.7 - 225.0) per 100,000, resulting in financial costs of 28.1 (13.8 - 47.1) billion CNY. Contributions to indoor VOC concentrations from six indoor sources and three outdoor sources were derived by PMF. The top three sources, i.e., wood building materials and furniture, outdoor vehicle exhaust, and cooking and indoor combustion, accounted for 42.7%, 25.9%, and 11.0% of the VOC-attributable DALYs, which suggests prioritizing controlling these sources in China. This approach can be extended to other IAPs and provide fundamental data for future cost-benefit analysis of source control interventions. TOC Art O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/25333590v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@1a69a9corg.highwire.dtl.DTLVardef@f07ec4org.highwire.dtl.DTLVardef@1129103org.highwire.dtl.DTLVardef@1ee68d1_HPS_FORMAT_FIGEXP M_FIG C_FIG SynopsisThis novel method leverages multi-seasonal and multi-room residential VOC measurements to identify emission sources, quantify source-specific exposure concentrations, and estimate source-specific health burden, thus prioritizing the sources needing control.

Respiratory Syncytial Virus (RSV) is responsible for a substantial health burden worldwide, particularly among children and older adults. In 2023, novel immunoprophylactic interventions for RSV were approved, underscoring the need to monitor circulating RSV lineages and detect mutations that could compromise intervention effectiveness. Here, we implemented wastewater-based genomic RSV surveillance by integrating digital PCR and amplicon-based sequencing within Switzerland's national wastewater monitoring program. We tracked RSV subtypes and individual mutations across the 2024-2025 peak season in six Swiss cities. RSV-A and RSV-B co-circulated nationwide, and both exhibited similar epidemiological dynamics estimated from their subtype-specific effective reproduction numbers. No previously reported F protein mutations relevant to prophylaxis efficacy were identified. Genetic diversity analysis of wastewater-derived sequences reflected patterns previously reported in clinical data, with higher diversity in RSV-A than RSV-B and greater variability in the G compared to the F gene. These findings demonstrate the potential of wastewater-based RSV surveillance for monitoring RSV dynamics and diversity and establish a national baseline for RSV evolution during the first season following vaccine implementation in Switzerland.

The outbreak of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has dramatically transformed policies and practices surrounding public health. One such shift is the expanded emphasis on environmental surveillance for pathogens. Environmental surveillance methods have primarily relied upon wastewater and indoor surface testing, and despite substantial evidence that SARS-CoV-2 commonly travels through space in aerosols, there has been limited indoor air surveillance. This study investigated the effectiveness of integrated surveillance including an active air sampler, surface swabs and passive settling plates to detect SARS-CoV-2 in hospital rooms with COVID-19 patients and compared detection efficacy among sampling methods. The AerosolSense active air sampler was found to detect SARS-CoV-2 in 53.8% of all samples collected compared to 12.1% detection by passive air sampling and 14.8% detection by surface swabs. Approximately 69% of sampled rooms (22/32) returned a positive environmental sample of any type. Among positive rooms, ~32% had only active air samples that returned positive, while ~27% and ~9% had only one or more surface swabs or passive settling plates that returned a positive respectively, and ~32% had more than one sample type that returned a positive result. This study demonstrates the potential for the AerosolSense to detect SARS-CoV-2 RNA in real-world healthcare environments and suggests that integrated sampling that includes active air sampling is an important addition to environmental pathogen surveillance in support of public health.

BackgroundWastewater represents a composite biological sample from the entire contributing population. People infected with monkeypox virus (MPXV)1 may excrete viral DNA into wastewater via multiple ways such as in feces, urine, skin lesions, and/or saliva. We describe results from rapid establishment of wastewater surveillance in selected regions in California within a month of the first reported case of monkeypox in the United States. MethodsPCR assays targeting genomic DNA from MPXV were deployed in an ongoing wastewater surveillance program in California. MPXV DNA concentrations were measured daily in settled solids samples from nine wastewater plants. Results over a four-week period were validated across different MPXV assays, compared using influent and solids samples, and correlated using non-parametric methods (Kendalls tau) with the number of monkeypox cases reported from each sewershed. ResultsMPXV DNA was detected at all nine sites between June 19 and August 1, 2022; 5 of 9 sites detected MPXV DNA prior to or within a day of the first case identified in the source sewershed. At the four sites with >10 positive detections, we observed a positive, statistically significant correlation (p <0.001) between MPXV DNA in wastewater solids and incidence rate of reported cases. ConclusionsOur findings suggest wastewater can be used to effectively detect the introduction of MPXV and monitor its circulation in the community to inform public health and clinical response. This flexible wastewater surveillance infrastructure may be rapidly leveraged to monitor other pathogens of public health importance that are shed into wastewater.

Tracking the emergence of new SARS-CoV-2 variants is important for a comprehensive understanding of the pandemics progression. However, it remains challenging due to the low variant prevalence in the early stage of an outbreak. Here, we present an integrated framework that combines three key components: early variant detection in wastewater, validation through clinical genome sequencing, and transmissibility assessment using mathematical modeling. Using the SARS-CoV-2 Omicron variant as a proof of concept, we developed a novel nested allele-specific RT-qPCR assay (NAS-PCR) for wastewater surveillance. Our framework detected Omicron in Greater Boston wastewater samples starting from September 2021, over two months before the first U.S. clinical case. We validated these findings by analyzing GISAID clinical sequence data, which revealed 172 previously unreported Omicron genomes predating its official identification in South Africa. To assess transmissibility, we developed a Susceptible-Infected-Viral load model using quantified wastewater concentrations, which estimated Omicrons basic reproduction number (R0) between 2.36 and 3.09, showing robust consistency across varying population sizes, data points, and viral shedding rates. This integrated approach unifies molecular diagnostics, wastewater epidemiology, and mathematical modeling for comprehensive variant surveillance. Our framework provides a systematic solution for early warning and risk assessment of emerging variants, which can strengthen public health preparedness for future viral threats.

Wastewater, which contains everything from pathogens to pollutants, is a geospatially-and temporally-linked microbial fingerprint of a given population. As a result, it can be leveraged for monitoring multiple dimensions of public health across locales and time. Here, we integrate targeted and bulk RNA sequencing (n=1,419 samples) to track the viral, bacterial, and functional content over geospatially distinct areas within Miami Dade County from 2020-2022. First, we used targeted amplicon sequencing (n=966) to track diverse SARS-CoV-2 variants across space and time, and we found a tight correspondence with clinical caseloads from University students (N = 1,503) and Miami-Dade County hospital patients (N = 3,939 patients), as well as an 8-day earlier detection of the Delta variant in wastewater vs. in patients. Additionally, in 453 metatranscriptomic samples, we demonstrate that different wastewater sampling locations have clinically and public-health-relevant microbiota that vary as a function of the size of the human population they represent. Through assembly, alignment-based, and phylogenetic approaches, we also detect multiple clinically important viruses (e.g., norovirus) and describe geospatial and temporal variation in microbial functional genes that indicate the presence of pollutants. Moreover, we found distinct profiles of antimicrobial resistance (AMR) genes and virulence factors across campus buildings, dorms, and hospitals, with hospital wastewater containing a significant increase in AMR abundance. Overall, this effort lays the groundwork for systematic characterization of wastewater to improve public health decision making and a broad platform to detect emerging pathogens.

Mobile monitoring campaigns to estimate long-term air pollution levels are becoming increasingly common. Still, many campaigns have not conducted temporally-balanced sampling, and few have looked at the implications of such study designs for epidemiologic exposure assessment. We carried out a simulation study of fixed-site air quality monitors to better understand how different mobile monitoring designs involving short-term stationary measurements at fixed locations impact the resulting exposure surfaces. We used Monte Carlo resampling to simulate three archetypal monitoring designs using oxides of nitrogen (NOx) monitoring data from 69 regulatory sites in California: a year-around Balanced Design that sampled during all seasons of the year, days of the week, and all or various hours of the day; a temporally reduced Rush Hours Design; and a temporally reduced Business Hours Design. We evaluated the performance of each designs land use regression prediction model. The Balanced Design consistently yielded the most accurate annual averages; while the reduced Rush Hours and Business Hours Designs generally produced more biased results. A temporally-balanced sampling design is crucial for mobile monitoring campaigns aiming to assess accurate long-term exposure in epidemiologic cohorts. SynopsisAir pollution mobile monitoring campaigns rarely conduct temporally balanced sampling. We show that this results in biased annual average exposure estimates. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/21255641v2_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@126c8ddorg.highwire.dtl.DTLVardef@14d52e5org.highwire.dtl.DTLVardef@17d390dorg.highwire.dtl.DTLVardef@2cc3d1_HPS_FORMAT_FIGEXP M_FIG C_FIG

BackgroundInfluenza A virus (IAV) is a major cause of morbidity and mortality globally, causing seasonal influenza in humans and infecting birds and some mammals. In 2024, IAV H5N1 highly pathogenic avian influenza (HPAI) in the United States moved into cattle. While the outbreak is currently of low risk to the public, there is an urgent need to monitor the disease and prevent spread. MethodsWe conducted a nationwide study evaluating the relationship between H5 hemagglutinin gene RNA concentrations in wastewater and reported outbreaks of IAV H5N1 in animals and humans. We utilized an H5-specific droplet digital RT-PCR test to quantify H5 RNA in wastewater in 40 states across the United States, and 1) examined the temporal association between outbreaks and wastewater detections and 2) utilized linear mixed models (LMM) to determine the relationship between measurements in wastewater and outbreak-related factors in the local area. ResultsWe find that there is a significant temporal association between wastewater H5 detections and the incidence of outbreaks in poultry and wild birds, but not in cattle or with human infections. However, outbreaks tended to occur at the same time across populations - wild bird detections were also associated with H5N1 in herds, poultry, and humans. Utilizing a LMM, we find that for individual sites, there is a relationship between H5 measurements in wastewater and both poultry outbreaks and the presence of dairy industry locally, but that there was either no relationship or a negative relationship with H5 measurements and either combined systems that accept storm water or those with detection of H5 in wild birds. ConclusionsThe study highlights how wastewater monitoring can supplement traditional surveillance, providing vital data that reflects public health threats. The findings underscore the potential of scaled wastewater surveillance as a proactive tool in monitoring and managing future outbreaks.

Far-UVC (222 nm UV-C light) is a promising tool to mitigate aerosol transmission of pathogens indoors. However, recent studies have raised concerns related to ozone (O3) production and secondary chemistry. In this study, we measured indoor O3 and ultra-fine particle (UFP, 17.5-289 nm) concentrations with and without 222 nm far-UVC (average fluence rate 1.7-1.8 {micro}W/cm2) in a hotel quarantine facility in Baltimore (MD, USA). We obtained nearby outdoor O3 concentrations from the Environmental Protection Agency (EPA) website. In a sealed empty guest room, the average O3 concentrations were 3 ppb (UV off, 0.1-0.5 ACH), 16 ppb (UV on, 0.1 ACH) and 9 ppb (UV on, 0.5 ACH). In a standard guest room, the average O3 concentrations were 12 ppb (UV off, 1.4 ACH) and 14 ppb (UV on, 1.4 ACH), and correlated with outdoor concentrations ({rho} = 0.65 - 0.74, p = 2*10-12 - 2*10-29). A linear regression model, adjusted for outdoor O3, estimated that use of far-UVC lamps increased the O3 concentration by 5.7 ppb (95% confidence interval (CI) 4.9 - 6.5 ppb) in the standard hotel room. Indoor O3 concentrations increased with far-UVC usage, however, the concentrations remained 6-12 ppb lower, on average, than outdoors and well below EPA ambient limits. We did not find a clear relationship between indoor UFP concentrations and UV usage. Although our study was limited by absence of direct outdoor measurements of local O3 and UFPs, our findings do not support a major impact of far-UVC on UFP concentrations in the real-world environment that we studied.