SwabExpress: An End-to-End Protocol for Extraction-Free COVID-19 Testing.

BACKGROUND: The urgent need for massively scaled clinical testing for SARS-CoV-2, along with global shortages of critical reagents and supplies, has necessitated development of streamlined laboratory testing protocols. Conventional nucleic acid testing for SARS-CoV-2 involves collection of a clinical specimen with a nasopharyngeal swab in transport medium, nucleic acid extraction, and quantitative reverse-transcription PCR (RT-qPCR). As testing has scaled across the world, the global supply chain has buckled, rendering testing reagents and materials scarce. To address shortages, we developed SwabExpress, an end-to-end protocol developed to employ mass produced anterior nares swabs and bypass the requirement for transport media and nucleic acid extraction. METHODS: We evaluated anterior nares swabs, transported dry and eluted in low-TE buffer as a direct-to-RT-qPCR alternative to extraction-dependent viral transport media. We validated our protocol of using heat treatment for viral inactivation and added a proteinase K digestion step to reduce amplification interference. We tested this protocol across archived and prospectively collected swab specimens to fine-tune test performance. RESULTS: After optimization, SwabExpress has a low limit of detection at 2-4 molecules/µL, 100% sensitivity, and 99.4% specificity when compared side by side with a traditional RT-qPCR protocol employing extraction. On real-world specimens, SwabExpress outperforms an automated extraction system while simultaneously reducing cost and hands-on time. CONCLUSION: SwabExpress is a simplified workflow that facilitates scaled testing for COVID-19 without sacrificing test performance. It may serve as a template for the simplification of PCR-based clinical laboratory tests, particularly in times of critical shortages during pandemics.

Trends in Risk Factors and Symptoms Associated With SARS-CoV-2 and Rhinovirus Test Positivity in King County, Washington, June 2020 to July 2022.

Importance: Few US studies have reexamined risk factors for SARS-CoV-2 positivity in the context of widespread vaccination and new variants or considered risk factors for cocirculating endemic viruses, such as rhinovirus. Objectives: To evaluate how risk factors and symptoms associated with SARS-CoV-2 test positivity changed over the course of the pandemic and to compare these with the risk factors associated with rhinovirus test positivity. Design, Setting, and Participants: This case-control study used a test-negative design with multivariable logistic regression to assess associations between SARS-CoV-2 and rhinovirus test positivity and self-reported demographic and symptom variables over a 25-month period. The study was conducted among symptomatic individuals of all ages enrolled in a cross-sectional community surveillance study in King County, Washington, from June 2020 to July 2022. Exposures: Self-reported data for 15 demographic and health behavior variables and 16 symptoms. Main Outcomes and Measures: Reverse transcription-polymerase chain reaction-confirmed SARS-CoV-2 or rhinovirus infection. Results: Analyses included data from 23 498 individuals. The median (IQR) age of participants was 34.33 (22.42-45.08) years, 13 878 (59.06%) were female, 4018 (17.10%) identified as Asian, 654 (2.78%) identified as Black, and 2193 (9.33%) identified as Hispanic. Close contact with an individual with SARS-CoV-2 (adjusted odds ratio [aOR], 3.89; 95% CI, 3.34-4.57) and loss of smell or taste (aOR, 3.49; 95% CI, 2.77-4.41) were the variables most associated with SARS-CoV-2 test positivity, but both attenuated during the Omicron period. Contact with a vaccinated individual with SARS-CoV-2 (aOR, 2.03; 95% CI, 1.56-2.79) was associated with lower odds of testing positive than contact with an unvaccinated individual with SARS-CoV-2 (aOR, 4.04; 95% CI, 2.39-7.23). Sore throat was associated with Omicron infection (aOR, 2.27; 95% CI, 1.68-3.20) but not Delta infection. Vaccine effectiveness for participants fully vaccinated with a booster dose was 93% (95% CI, 73%-100%) for Delta, but not significant for Omicron. Variables associated with rhinovirus test positivity included being younger than 12 years (aOR, 3.92; 95% CI, 3.42-4.51) and experiencing a runny or stuffy nose (aOR, 4.58; 95% CI, 4.07-5.21). Black race, residing in south King County, and households with 5 or more people were significantly associated with both SARS-CoV-2 and rhinovirus test positivity. Conclusions and Relevance: In this case-control study of 23 498 symptomatic individuals, estimated risk factors and symptoms associated with SARS-CoV-2 infection changed over time. There was a shift in reported symptoms between the Delta and Omicron variants as well as reductions in the protection provided by vaccines. Racial and sociodemographic disparities persisted in the third year of SARS-CoV-2 circulation and were also present in rhinovirus infection. Trends in testing behavior and availability may influence these results.

Genomic surveillance of SARS-CoV-2 Omicron variants on a university campus.

Novel variants continue to emerge in the SARS-CoV-2 pandemic. University testing programs may provide timely epidemiologic and genomic surveillance data to inform public health responses. We conducted testing from September 2021 to February 2022 in a university population under vaccination and indoor mask mandates. A total of 3,048 of 24,393 individuals tested positive for SARS-CoV-2 by RT-PCR; whole genome sequencing identified 209 Delta and 1,730 Omicron genomes of the 1,939 total sequenced. Compared to Delta, Omicron had a shorter median serial interval between genetically identical, symptomatic infections within households (2 versus 6 days, P = 0.021). Omicron also demonstrated a greater peak reproductive number (2.4 versus 1.8), and a 1.07 (95% confidence interval: 0.58, 1.57; P < 0.0001) higher mean cycle threshold value. Despite near universal vaccination and stringent mitigation measures, Omicron rapidly displaced the Delta variant to become the predominant viral strain and led to a surge in cases in a university population.

Associations Between Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Variants and Risk of Coronavirus Disease 2019 (COVID-19) Hospitalization Among Confirmed Cases in Washington State: A Retrospective Cohort Study.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic is dominated by variant viruses; the resulting impact on disease severity remains unclear. Using a retrospective cohort study, we assessed the hospitalization risk following infection with 7 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants. METHODS: Our study includes individuals with positive SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) in the Washington Disease Reporting System with available viral genome data, from 1 December 2020 to 14 January 2022. The analysis was restricted to cases with specimens collected through sentinel surveillance. Using a Cox proportional hazards model with mixed effects, we estimated hazard ratios (HR) for hospitalization risk following infection with a variant, adjusting for age, sex, calendar week, and vaccination. RESULTS: In total, 58 848 cases were sequenced through sentinel surveillance, of which 1705 (2.9%) were hospitalized due to COVID-19. Higher hospitalization risk was found for infections with Gamma (HR 3.20, 95% confidence interval [CI] 2.40-4.26), Beta (HR 2.85, 95% CI 1.56-5.23), Delta (HR 2.28 95% CI 1.56-3.34), or Alpha (HR 1.64, 95% CI 1.29-2.07) compared to infections with ancestral lineages; Omicron (HR 0.92, 95% CI .56-1.52) showed no significant difference in risk. Following Alpha, Gamma, or Delta infection, unvaccinated patients show higher hospitalization risk, while vaccinated patients show no significant difference in risk, both compared to unvaccinated, ancestral lineage cases. Hospitalization risk following Omicron infection is lower with vaccination. CONCLUSIONS: Infection with Alpha, Gamma, or Delta results in a higher hospitalization risk, with vaccination attenuating that risk. Our findings support hospital preparedness, vaccination, and genomic surveillance.

SARS-CoV-2 Epidemiology on a Public University Campus in Washington State.

BACKGROUND: We aimed to evaluate a testing program to facilitate control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission at a large university and measure spread in the university community using viral genome sequencing. METHODS: Our prospective longitudinal study used remote contactless enrollment, daily mobile symptom and exposure tracking, and self-swab sample collection. Individuals were tested if the participant was exposed to a known SARS-CoV-2-infected person, developed new symptoms, or reported high-risk behavior (such as attending an indoor gathering without masking or social distancing), if a member of a group experiencing an outbreak, or at enrollment. Study participants included students, staff, and faculty at an urban public university during the Autumn quarter of 2020. RESULTS: We enrolled 16 476 individuals, performed 29 783 SARS-CoV-2 tests, and detected 236 infections. Seventy-five percent of positive cases reported at least 1 of the following: symptoms (60.8%), exposure (34.7%), or high-risk behaviors (21.5%). Greek community affiliation was the strongest risk factor for testing positive, and molecular epidemiology results suggest that specific large gatherings were responsible for several outbreaks. CONCLUSIONS: A testing program focused on individuals with symptoms and unvaccinated persons who participate in large campus gatherings may be effective as part of a comprehensive university-wide mitigation strategy to control the spread of SARS-CoV-2.

Comparable specimen collection from both ends of at-home mid-turbinate swabs.

Unsupervised upper respiratory specimen collection is a key factor in the ability to massively scale SARS-CoV-2 testing. But there is concern that unsupervised specimen collection may produce inferior samples. Across two studies that included unsupervised at-home mid-turbinate specimen collection, ∼1% of participants used the wrong end of the swab. We found that molecular detection of respiratory pathogens and a human biomarker were comparable between specimens collected from the handle of the swab and those collected correctly. Older participants were more likely to use the swab backwards. Our results suggest that errors made during home-collection of nasal specimens do not preclude molecular detection of pathogens and specialized swabs may be an unnecessary luxury during a pandemic.

Incidence of Medically Attended Acute Respiratory Illnesses Due to Respiratory Viruses Across the Life Course During the 2018/19 Influenza Season.

BACKGROUND: Although multiple respiratory viruses circulate in humans, few studies have compared the incidence of different viruses across the life course. We estimated the incidence of outpatient illness due to 12 different viruses during November 2018 through April 2019 in a fully enumerated population. METHODS: We conducted active surveillance for ambulatory care visits for acute respiratory illness (ARI) among members of Kaiser Permanente Washington (KPWA). Enrolled patients provided respiratory swab specimens which were tested for 12 respiratory viruses using reverse transcription polymerase chain reaction (RT-PCR). We estimated the cumulative incidence of infection due to each virus overall and by age group. RESULTS: The KPWA population under surveillance included 202 562 individuals, of whom 2767 (1.4%) were enrolled in the study. Influenza A(H3N2) was the most commonly detected virus, with an overall incidence of 21 medically attended illnesses per 1000 population; the next most common viruses were influenza A(H1N1) (18 per 1000), coronaviruses (13 per 1000), respiratory syncytial virus (RSV, 13 per 1000), and rhinovirus (9 per 1000). RSV was the most common cause of medically attended ARI among children aged 1-4 years; coronaviruses were the most common among adults aged ≥65 years. CONCLUSIONS: Consistent with other studies focused on single viruses, we found that influenza and RSV were major causes of acute respiratory illness in persons of all ages. In comparison, coronaviruses and rhinovirus were also important pathogens. Prior to the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), coronaviruses were the second-most common cause of medically attended ARI during the 2018/19 influenza season.

Characteristics of COVID-19 in Homeless Shelters : A Community-Based Surveillance Study.

BACKGROUND: Homeless shelters are a high-risk setting for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission because of crowding and shared hygiene facilities. OBJECTIVE: To investigate SARS-CoV-2 case counts across several adult and family homeless shelters in a major metropolitan area. DESIGN: Cross-sectional, community-based surveillance study. (ClinicalTrials.gov: NCT04141917). SETTING: 14 homeless shelters in King County, Washington. PARTICIPANTS: A total of 1434 study encounters were done in shelter residents and staff, regardless of symptoms. INTERVENTION: 2 strategies were used for SARS-CoV-2 testing: routine surveillance and contact tracing ("surge testing") events. MEASUREMENTS: The primary outcome measure was test positivity rate of SARS-CoV-2 infection at shelters, determined by dividing the number of positive cases by the total number of participant encounters, regardless of symptoms. Sociodemographic, clinical, and virologic variables were assessed as correlates of viral positivity. RESULTS: Among 1434 encounters, 29 (2% [95% CI, 1.4% to 2.9%]) cases of SARS-CoV-2 infection were detected across 5 shelters. Most (n = 21 [72.4%]) were detected during surge testing events rather than routine surveillance, and most (n = 21 [72.4% {CI, 52.8% to 87.3%}]) were asymptomatic at the time of sample collection. Persons who were positive for SARS-CoV-2 were more frequently aged 60 years or older than those without SARS-CoV-2 (44.8% vs. 15.9%). Eighty-six percent of persons with positive test results slept in a communal space rather than in a private or shared room. LIMITATION: Selection bias due to voluntary participation and a relatively small case count. CONCLUSION: Active surveillance and surge testing were used to detect multiple cases of asymptomatic and symptomatic SARS-CoV-2 infection in homeless shelters. The findings suggest an unmet need for routine viral testing outside of clinical settings for homeless populations. PRIMARY FUNDING SOURCE: Gates Ventures.

Viral genomes reveal patterns of the SARS-CoV-2 outbreak in Washington State.

The rapid spread of SARS-CoV-2 has gravely impacted societies around the world. Outbreaks in different parts of the globe are shaped by repeated introductions of new lineages and subsequent local transmission of those lineages. Here, we sequenced 3940 SARS-CoV-2 viral genomes from Washington State to characterize how the spread of SARS-CoV-2 in Washington State (USA) was shaped by differences in timing of mitigation strategies across counties, as well as by repeated introductions of viral lineages into the state. Additionally, we show that the increase in frequency of a potentially more transmissible viral variant (614G) over time can potentially be explained by regional mobility differences and multiple introductions of 614G, but not the other variant (614D) into the state. At an individual level, we see evidence of higher viral loads in patients infected with the 614G variant. However, using clinical records data, we do not find any evidence that the 614G variant impacts clinical severity or patient outcomes. Overall, this suggests that at least to date, the behavior of individuals has been more important in shaping the course of the pandemic than changes in the virus.

Cryptic transmission of SARS-CoV-2 in Washington state.

After its emergence in Wuhan, China, in late November or early December 2019, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus rapidly spread globally. Genome sequencing of SARS-CoV-2 allows the reconstruction of its transmission history, although this is contingent on sampling. We analyzed 453 SARS-CoV-2 genomes collected between 20 February and 15 March 2020 from infected patients in Washington state in the United States. We find that most SARS-CoV-2 infections sampled during this time derive from a single introduction in late January or early February 2020, which subsequently spread locally before active community surveillance was implemented.