Evaluation of a novel university-based testing platform to increase access to SARS-CoV-2 testing during the COVID-19 pandemic in a cohort study.

OBJECTIVE: We aimed to evaluate the feasibility and utility of an unsupervised testing mechanism, in which participants pick up a swab kit, self-test (unsupervised) and return the kit to an on-campus drop box, as compared with supervised self-testing at staffed locations. DESIGN: University SARS-CoV-2 testing cohort. SETTING: Husky Coronavirus Testing provided voluntary SARS-CoV-2 testing at a university in Seattle, USA. OUTCOME MEASURES: We computed descriptive statistics to describe the characteristics of the study sample. Adjusted logistic regression implemented via generalised estimating equations was used to estimate the odds of a self-swab being conducted through unsupervised versus supervised testing mechanisms by participant characteristics, including year of study enrolment, pre-Omicron versus post-Omicron time period, age, sex, race, ethnicity, affiliation and symptom status. RESULTS: From September 2021 to July 2022, we received 92 499 supervised and 26 800 unsupervised self-swabs. Among swabs received by the laboratory, the overall error rate for supervised versus unsupervised swabs was 0.3% vs 4%, although this declined to 2% for unsupervised swabs by the spring of the academic year. Results were returned for 92 407 supervised (5% positive) and 25 836 unsupervised (4%) swabs from 26 359 participants. The majority were students (79%), 61% were female and most identified as white (49%) or Asian (34%). The use of unsupervised testing increased during the Omicron wave when testing demand was high and stayed constant in spring 2022 even when testing demand fell. We estimated the odds of using unsupervised versus supervised testing to be significantly greater among those <25 years of age (p<0.001), for Hispanic versus non-Hispanic individuals (OR 1.2, 95% CI 1.0 to 1.3, p=0.01) and lower among individuals symptomatic versus asymptomatic or presymptomatic (0.9, 95% CI 0.8 to 0.9, p<0.001). CONCLUSIONS: Unsupervised swab collection permitted increased testing when demand was high, allowed for access to a broader proportion of the university community and was not associated with a substantial increase in testing errors.

Impacts of human mobility on the citywide transmission dynamics of 18 respiratory viruses in pre- and post-COVID-19 pandemic years.

Many studies have used mobile device location data to model SARS-CoV-2 dynamics, yet relationships between mobility behavior and endemic respiratory pathogens are less understood. We studied the effects of population mobility on the transmission of 17 endemic viruses and SARS-CoV-2 in Seattle over a 4-year period, 2018-2022. Before 2020, visits to schools and daycares, within-city mixing, and visitor inflow preceded or coincided with seasonal outbreaks of endemic viruses. Pathogen circulation dropped substantially after the initiation of COVID-19 stay-at-home orders in March 2020. During this period, mobility was a positive, leading indicator of transmission of all endemic viruses and lagging and negatively correlated with SARS-CoV-2 activity. Mobility was briefly predictive of SARS-CoV-2 transmission when restrictions relaxed but associations weakened in subsequent waves. The rebound of endemic viruses was heterogeneously timed but exhibited stronger, longer-lasting relationships with mobility than SARS-CoV-2. Overall, mobility is most predictive of respiratory virus transmission during periods of dramatic behavioral change and at the beginning of epidemic waves.

Comparative diagnostic utility of SARS-CoV-2 rapid antigen and molecular testing in a community setting.

BACKGROUND: SARS-CoV-2 antigen-detection rapid diagnostic tests (Ag-RDTs) have become widely utilized but longitudinal characterization of their community-based performance remains incompletely understood. METHODS: This prospective longitudinal study at a large public university in Seattle, WA utilized remote enrollment, online surveys, and self-collected nasal swab specimens to evaluate Ag-RDT performance against real-time reverse transcription polymerase chain reaction (rRT-PCR) in the context of SARS-CoV-2 Omicron. Ag-RDT sensitivity and specificity within 1 day of rRT-PCR were evaluated by symptom status throughout the illness episode and Orf1b cycle threshold (Ct). RESULTS: From February to December 2022, 5,757 participants reported 17,572 Ag-RDT results and completed 12,674 rRT-PCR tests, of which 995 (7.9%) were rRT-PCR-positive. Overall sensitivity and specificity were 53.0% (95% CI: 49.6-56.4%) and 98.8% (98.5-99.0%), respectively. Sensitivity was comparatively higher for Ag-RDTs used 1 day after rRT-PCR (69.0%), 4 to 7 days post-symptom onset (70.1%), and Orf1b Ct ≤20 (82.7%). Serial Ag-RDT sensitivity increased with repeat testing ≥2 (68.5%) and ≥4 (75.8%) days after an initial Ag-RDT-negative result. CONCLUSION: Ag-RDT performance varied by clinical characteristics and temporal testing patterns. Our findings support recommendations for serial testing following an initial Ag-RDT-negative result, especially among recently symptomatic persons or those at high-risk for SARS-CoV-2 infection.

Remote surveillance and detection of SARS-CoV-2 transmission among household members in King County, Washington.

BACKGROUND: Early during the COVID-19 pandemic, it was important to better understand transmission dynamics of SARS-CoV-2, the virus that causes COVID-19. Household contacts of infected individuals are particularly at risk for infection, but delays in contact tracing, delays in testing contacts, and isolation and quarantine posed challenges to accurately capturing secondary household cases. METHODS: In this study, 346 households in the Seattle region were provided with respiratory specimen collection kits and remotely monitored using web-based surveys for respiratory illness symptoms weekly between October 1, 2020, and June 20, 2021. Symptomatic participants collected respiratory specimens at symptom onset and mailed specimens to the central laboratory in Seattle. Specimens were tested for SARS-CoV-2 using RT-PCR with whole genome sequencing attempted when positive. SARS-CoV-2-infected individuals were notified, and their household contacts submitted specimens every 2 days for 14 days. RESULTS: In total, 1371 participants collected 2029 specimens that were tested; 16 individuals (1.2%) within 6 households tested positive for SARS-CoV-2 during the study period. Full genome sequences were generated from 11 individuals within 4 households. Very little genetic variation was found among SARS-CoV-2 viruses sequenced from different individuals in the same household, supporting transmission within the household. CONCLUSIONS: This study indicates web-based surveillance of respiratory symptoms, combined with rapid and longitudinal specimen collection and remote contact tracing, provides a viable strategy to monitor households and detect household transmission of SARS-CoV-2. TRIAL REGISTRATION IDENTIFIER: NCT04141930, Date of registration 28/10/2019.

Utilizing a university testing program to estimate relative effectiveness of monovalent COVID-19 mRNA booster vaccine versus two-dose primary series against symptomatic SARS-CoV-2 infection.

Vaccine effectiveness (VE) studies utilizing the test-negative design are typically conducted in clinical settings, rather than community populations, leading to bias in VE estimates against mild disease and limited information on VE in healthy young adults. In a community-based university population, we utilized data from a large SARS-CoV-2 testing program to estimate relative VE of COVID-19 mRNA vaccine primary series and monovalent booster dose versus primary series only against symptomatic SARS-CoV-2 infection from September 2021 to July 2022. We used the test-negative design and logistic regression implemented via generalized estimating equations adjusted for age, calendar time, prior SARS-CoV-2 infection, and testing frequency (proxy for test-seeking behavior) to estimate relative VE. Analyses included 2,218 test-positive cases (59 % received monovalent booster dose) and 9,615 test-negative controls (62 %) from 9,066 individuals, with median age of 21 years, mostly students (71 %), White (56 %) or Asian (28 %), and with few comorbidities (3 %). More cases (23 %) than controls (6 %) had COVID-19-like illness. Estimated adjusted relative VE of primary series and monovalent booster dose versus primary series only against symptomatic SARS-CoV-2 infection was 40 % (95 % CI: 33-47 %) during the overall analysis period and 46 % (39-52 %) during the period of Omicron circulation. Relative VE was greater for those without versus those with prior SARS-CoV-2 infection (41 %, 34-48 % versus 33 %, 9 %-52 %, P < 0.001). Relative VE was also greater in the six months after receiving a booster dose (41 %, 33-47 %) compared to more than six months (27 %, 8-42 %), but this difference was not statistically significant (P = 0.06). In this relatively young and healthy adult population, an mRNA monovalent booster dose provided increased protection against symptomatic SARS-CoV-2 infection, overall and with the Omicron variant. University testing programs may be utilized for estimating VE in healthy young adults, a population that is not well-represented by routine VE studies.

Risk of Subsequent Respiratory Virus Detection After Primary Virus Detection in a Community Household Study-King County, Washington, 2019-2021.

BACKGROUND: The epidemiology of respiratory viral infections is complex. How infection with one respiratory virus affects risk of subsequent infection with the same or another respiratory virus is not well described. METHODS: From October 2019 to June 2021, enrolled households completed active surveillance for acute respiratory illness (ARI), and participants with ARI self-collected nasal swab specimens; after April 2020, participants with ARI or laboratory-confirmed severe acute respiratory syndrome coronavirus 2 and their household members self-collected nasal swab specimens. Specimens were tested using multiplex reverse-transcription polymerase chain reaction for respiratory viruses. A Cox regression model with a time-dependent covariate examined risk of subsequent detections following a specific primary viral detection. RESULTS: Rhinovirus was the most frequently detected pathogen in study specimens (406 [9.5%]). Among 51 participants with multiple viral detections, rhinovirus to seasonal coronavirus (8 [14.8%]) was the most common viral detection pairing. Relative to no primary detection, there was a 1.03-2.06-fold increase in risk of subsequent virus detection in the 90 days after primary detection; risk varied by primary virus: human parainfluenza virus, rhinovirus, and respiratory syncytial virus were statistically significant. CONCLUSIONS: Primary virus detection was associated with higher risk of subsequent virus detection within the first 90 days after primary detection.

Tiny swabs: nasal swabs integrated into tube caps facilitate large-scale self-collected SARS-CoV-2 testing.

The COVID-19 pandemic spurred the development of innovative solutions for specimen collection and molecular detection for large-scale community testing. Among these developments is the RHINOstic nasal swab, a plastic anterior nares swab built into the cap of a standard matrix tube that facilitates automated processing of up to 96 specimens at a time. In a study of unsupervised self-collection utilizing these swabs, we demonstrate comparable analytic performance and shipping stability compared to traditional anterior nares swabs, as well as significant improvements in laboratory processing efficiency. The use of these swabs may allow laboratories to accommodate large numbers of sample collections during periods of high testing demand. Automation-friendly nasal swabs are an important tool for high-throughput processing of samples that may be adopted in response to future respiratory viral pandemics.

Incidence of SARS-CoV-2 infection and associated risk factors among staff and residents at homeless shelters in King County, Washington: an active surveillance study.

Homeless shelter residents and staff may be at higher risk of SARS-CoV-2 infection. However, SARS-CoV-2 infection estimates in this population have been reliant on cross-sectional or outbreak investigation data. We conducted routine surveillance and outbreak testing in 23 homeless shelters in King County, Washington, to estimate the occurrence of laboratory-confirmed SARS-CoV-2 infection and risk factors during 1 January 2020-31 May 2021. Symptom surveys and nasal swabs were collected for SARS-CoV-2 testing by RT-PCR for residents aged ≥3 months and staff. We collected 12,915 specimens from 2,930 unique participants. We identified 4.74 (95% CI 4.00-5.58) SARS-CoV-2 infections per 100 individuals (residents: 4.96, 95% CI 4.12-5.91; staff: 3.86, 95% CI 2.43-5.79). Most infections were asymptomatic at the time of detection (74%) and detected during routine surveillance (73%). Outbreak testing yielded higher test positivity than routine surveillance (2.7% versus 0.9%). Among those infected, residents were less likely to report symptoms than staff. Participants who were vaccinated against seasonal influenza and were current smokers had lower odds of having an infection detected. Active surveillance that includes SARS-CoV-2 testing of all persons is essential in ascertaining the true burden of SARS-CoV-2 infections among residents and staff of congregate settings.

Streptococcus pneumoniae nasal carriage patterns with and without common respiratory virus detections in households in Seattle, WA, USA before and during the COVID-19 pandemic.

Background: Respiratory viruses might influence Streptococcus pneumoniae nasal carriage and subsequent disease risk. We estimated the association between common respiratory viruses and semiquantitative S. pneumoniae nasal carriage density in a household setting before and during the COVID-19 pandemic. Methods: From November 2019-June 2021, we enrolled participants in a remote household surveillance study of respiratory pathogens. Participants submitted weekly reports of acute respiratory illness (ARI) symptoms. Mid-turbinate or anterior nasal swabs were self-collected at enrollment, when ARI occurred, and, in the second year of the study only, from household contacts after SARS-CoV-2 was detected in a household member. Specimens were tested using multiplex reverse-transcription PCR for respiratory pathogens, including S. pneumoniae, rhinovirus, adenovirus, common human coronavirus, influenza A/B virus, respiratory syncytial virus (RSV) A/B, human metapneumovirus, enterovirus, and human parainfluenza virus. We estimated differences in semiquantitative S. pneumoniae nasal carriage density, estimated by the inverse of S. pneumoniae relative cycle threshold (Crt) values, with and without viral detection for any virus and for specific respiratory viruses using linear generalized estimating equations of S. pneumoniae Crt values on virus detection adjusted for age and swab type and accounting for clustering of swabs within households. Results: We collected 346 swabs from 239 individuals in 151 households that tested positive for S. pneumoniae (n = 157 with and 189 without ≥1 viruses co-detected). Difficulty breathing, cough, and runny nose were more commonly reported among individuals with specimens with viral co-detection compared to without (15%, 80% and 93% vs. 8%, 57%, and 51%, respectively) and ear pain and headache were less commonly reported (3% and 26% vs. 16% and 41%, respectively). For specific viruses among all ages, semiquantitative S. pneumoniae nasal carriage density was greater with viral co-detection for enterovirus, RSV A/B, adenovirus, rhinovirus, and common human coronavirus (P < 0.01 for each). When stratified by age, semiquantitative S. pneumoniae nasal carriage density was significantly greater with viral co-detection among children aged <5 (P = 0.002) and 5-17 years (P = 0.005), but not among adults aged 18-64 years (P = 0.29). Conclusion: Detection of common respiratory viruses was associated with greater concurrent S. pneumoniae semiquantitative nasal carriage density in a household setting among children, but not adults.

CASCADIA: a prospective community-based study protocol for assessing SARS-CoV-2 vaccine effectiveness in children and adults using a remote nasal swab collection and web-based survey design.

INTRODUCTION: Although SARS-CoV-2 vaccines were first approved under Emergency Use Authorization by the Food and Drug Administration in late 2020 for adults, authorisation for young children 6 months to <5 years of age did not occur until 2022. These authorisations were based on clinical trials, understanding real-world vaccine effectiveness (VE) in the setting of emerging variants is critical. The primary goal of this study is to evaluate SARS-CoV-2 VE against infection among children aged >6 months and adults aged <50 years. METHODS: CASCADIA is a 4-year community-based prospective study of SARS-CoV-2 VE among 3500 adults and paediatric populations aged 6 months to 49 years in Oregon and Washington, USA. At enrolment and regular intervals, participants complete a sociodemographic questionnaire. Individuals provide a blood sample at enrolment and annually thereafter, with optional blood draws every 6 months and after infection and vaccination. Participants complete weekly self-collection of anterior nasal swabs and symptom questionnaires. Swabs are tested for SARS-CoV-2 and other respiratory pathogens by reverse transcription-PCR, with results of selected pathogens returned to participants; nasal swabs with SARS-CoV-2 detected will undergo whole genome sequencing. Participants who test positive for SARS-CoV-2 undergo serial swab collection every 3 days for 21 days. Serum samples are tested for SARS-CoV-2 antibody by binding and neutralisation assays. ANALYSIS: The primary outcome is SARS-CoV-2 infection. Cox regression models will be used to estimate the incidence rate ratio associated with SARS-CoV-2 vaccination among the paediatric and adult population, controlling for demographic factors and other potential confounders. ETHICS AND DISSEMINATION: All study materials including the protocol, consent forms, data collection instruments, participant communication and recruitment materials, were approved by the Kaiser Permanente Interregional Institutional Review Board, the IRB of record for the study. Results will be disseminated through peer-reviewed publications, presentations, participant newsletters and appropriate general news media.

Results from a test-and-treat study for influenza among residents of homeless shelters in King County, WA: A stepped-wedge cluster-randomized trial.

BACKGROUND: Persons experiencing homelessness face increased risk of influenza as overcrowding in congregate shelters can facilitate influenza virus spread. Data regarding on-site influenza testing and antiviral treatment within homeless shelters remain limited. METHODS: We conducted a cluster-randomized stepped-wedge trial of point-of-care molecular influenza testing coupled with antiviral treatment with baloxavir or oseltamivir in residents of 14 homeless shelters in Seattle, WA, USA. Residents ≥3 months with cough or ≥2 acute respiratory illness (ARI) symptoms and onset <7 days were eligible. In control periods, mid-nasal swabs were tested for influenza by reverse transcription polymerase chain reaction (RT-PCR). The intervention period included on-site rapid molecular influenza testing and antiviral treatment for influenza-positives if symptom onset was <48 h. The primary endpoint was monthly influenza virus infections in the control versus intervention periods. Influenza whole genome sequencing was performed to assess transmission and antiviral resistance. RESULTS: During 11/15/2019-4/30/2020 and 11/2/2020-4/30/2021, 1283 ARI encounters from 668 participants were observed. Influenza virus was detected in 51 (4%) specimens using RT-PCR (A = 14; B = 37); 21 influenza virus infections were detected from 269 (8%) intervention-eligible encounters by rapid molecular testing and received antiviral treatment. Thirty-seven percent of ARI-participant encounters reported symptom onset < 48 h. The intervention had no effect on influenza virus transmission (adjusted relative risk 1.73, 95% confidence interval [CI] 0.50-6.00). Of 23 influenza genomes, 86% of A(H1N1)pdm09 and 81% of B/Victoria sequences were closely related. CONCLUSION: Our findings suggest feasibility of influenza test-and-treat strategies in shelters. Additional studies would help discern an intervention effect during periods of increased influenza activity.

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.

Human Parainfluenza Virus in Homeless Shelters before and during the COVID-19 Pandemic, Washington, USA.

To determine the epidemiology of human parainfluenza virus in homeless shelters during the COVID-19 pandemic, we analyzed data and sequences from respiratory specimens collected in 23 shelters in Washington, USA, during 2019-2021. Two clusters in children were genetically similar by shelter of origin. Shelter-specific interventions are needed to reduce these infections.

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.

The clinical and genomic epidemiology of seasonal human coronaviruses in congregate homeless shelter settings: A repeated cross-sectional study.

Background: The circulation of respiratory viruses poses a significant health risk among those residing in congregate settings. Data are limited on seasonal human coronavirus (HCoV) infections in homeless shelter settings. Methods: We analysed data from a clinical trial and SARS-CoV-2 surveillance study at 23 homeless shelter sites in King County, Washington between October 2019-May 2021. Eligible participants were shelter residents aged ≥3 months with acute respiratory illness. We collected enrolment data and nasal samples for respiratory virus testing using multiplex RT-PCR platform including HCoV. Beginning April 1, 2020, eligibility expanded to shelter residents and staff regardless of symptoms. HCoV species was determined by RT-PCR with species-specific primers, OpenArray assay or genomic sequencing for samples with an OpenArray relative cycle threshold <22. Findings: Of the 14,464 samples from 3281 participants between October 2019-May 2021, 107 were positive for HCoV from 90 participants (median age 40 years, range: 0·9-81 years, 38% female). HCoV-HKU1 was the most common species identified before and after community-wide mitigation. No HCoV-positive samples were identified between May 2020-December 2020. Adults aged ≥50 years had the highest detection of HCoV (11%) among virus-positive samples among all age-groups. Species and sequence data showed diversity between and within HCoV species over the study period. Interpretation: HCoV infections occurred in all congregate homeless shelter site age-groups with the greatest proportion among those aged ≥50 years. Species and sequencing data highlight the complexity of HCoV epidemiology within and between shelters sites. Funding: Gates Ventures, Centers for Disease Control and Prevention, National Institute of Health.

Multiplex Target-Redundant RT-LAMP for Robust Detection of SARS-CoV-2 Using Fluorescent Universal Displacement Probes.

The increasing prevalence of variant lineages during the COVID-19 pandemic has the potential to disrupt molecular diagnostics due to mismatches between primers and variant templates. Point-of-care molecular diagnostics, which often lack the complete functionality of their high-throughput laboratory counterparts, are particularly susceptible to this type of disruption, which can result in false-negative results. To address this challenge, we have developed a robust Loop Mediated Isothermal Amplification assay with single tube multiplexed multitarget redundancy and an internal amplification control. A convenient and cost-effective target-specific fluorescence detection system allows amplifications to be grouped by signal using adaptable probes for pooled reporting of SARS-CoV-2 target amplifications or differentiation of the Internal Amplification Control. Over the course of the pandemic, primer coverage of viral lineages by the three redundant sub-assays has varied from assay to assay as they have diverged from the Wuhan-Hu-1 isolate sequence, but aggregate coverage has remained high for all variant sequences analyzed, with a minimum of 97.4% (Variant of Interest: Eta). In three instances (Delta, Gamma, Eta), a high-frequency mismatch with one of the three sub-assays was observed, but overall coverage remained high due to multitarget redundancy. When challenged with extracted human samples the multiplex assay showed 87% or better sensitivity (of 30 positive samples), with 100% sensitivity for samples containing greater than 30 copies of viral RNA per reaction (of 21 positive samples), and 100% specificity (of 60 negative samples). These results are further evidence that conventional laboratory methodologies can be leveraged at the point of care for robust performance and diagnostic stability over time. IMPORTANCE The COVID-19 pandemic has had tremendous impact, and the ability to perform molecular diagnostics in resource limited settings has emerged as a key resource for mitigating spread of the disease. One challenge in COVID-19 diagnosis, as well as other viruses, is ongoing mutation that can allow viruses to evade detection by diagnostic tests. We developed a test that detects multiple parts of the virus genome in a single test to reduce the chance of missing a virus due to mutation, and it is designed to be simpler and faster than typical laboratory tests while maintaining high sensitivity. This capability is enabled by a novel fluorescent probe technology that works with a simple constant temperature reaction condition.

The Clinical and Genomic Epidemiology of Rhinovirus in Homeless Shelters-King County, Washington.

BACKGROUND: Rhinovirus (RV) is a common cause of respiratory illness in all people, including those experiencing homelessness. RV epidemiology in homeless shelters is unknown. METHODS: We analyzed data from a cross-sectional homeless shelter study in King County, Washington, October 2019-May 2021. Shelter residents or guardians aged ≥3 months reporting acute respiratory illness completed questionnaires and submitted nasal swabs. After 1 April 2020, enrollment expanded to residents and staff regardless of symptoms. Samples were tested by multiplex RT-PCR for respiratory viruses. A subset of RV-positive samples was sequenced. RESULTS: There were 1066 RV-positive samples with RV present every month of the study period. RV was the most common virus before and during the coronavirus disease 2019 (COVID-19) pandemic (43% and 77% of virus-positive samples, respectively). Participants from family shelters had the highest prevalence of RV. Among 131 sequenced samples, 33 RV serotypes were identified with each serotype detected for ≤4 months. CONCLUSIONS: RV infections persisted through community mitigation measures and were most prevalent in shelters housing families. Sequencing showed a diversity of circulating RV serotypes, each detected over short periods of time. Community-based surveillance in congregate settings is important to characterize respiratory viral infections during and after the COVID-19 pandemic. CLINICAL TRIALS REGISTRATION: NCT04141917.

Simultaneous monitoring of HIV viral load and screening of SARS-CoV-2 employing a low-cost RT-qPCR test workflow.

The COVID-19 pandemic interrupted routine care for individuals living with HIV, putting them at risk of virologic failure and HIV-associated illness. Often this population is at high risk for exposure to SARS-CoV-2 infection, and once infected, for severe disease. Therefore, close monitoring of HIV plasma viral load (VL) and screening for SARS-CoV-2 infection are needed. We developed a non-proprietary method to isolate RNA from plasma, nasal secretions (NS), or both. The extracted RNA is then submitted to RT-qPCR to estimate the VL and classify HIV/SARS-CoV-2 status (i.e., HIV virologic failure or suppressed; SARS-CoV-2 as positive, presumptive positive, negative, or indeterminate). In contrived samples, the in-house RNA extraction workflow achieved a detection limit of 200-copies per mL for HIV RNA in plasma and 100-copies per mL for SARS-CoV-2 RNA in NS. Similar detection limits were observed for HIV and SARS-CoV-2 in pooled plasma/NS contrived samples. When comparing in-house with standard extraction methods, we found high agreement (>0.91) between input and measured RNA copies for HIV LTR in contrived plasma; SARS-CoV-2 N1/N2 in contrived NS; and LTR, N1, and N2 in pooled plasma/NS samples. We further evaluated this workflow on 133 clinical specimens: 40 plasma specimens (30 HIV-positive), 67 NS specimens (31 SARS-CoV-2-positive), and 26 combined plasma/NS specimens (26 HIV-positive with 10 SARS-CoV-2-positive), and compared the results obtained using the in-house RNA extraction to those using a commercial kit (standard extraction method). The in-house extraction and standard extraction of clinical specimens were positively correlated: plasma HIV VL (R2 of 0.81) and NS SARS-CoV-2 VL (R2 of 0.95 and 0.99 for N1 and N2 genes, respectively); and pooled plasma/NS HIV VL (R2 of 0.71) and SARS-CoV-2 VL (R2 of 1 both for N1 and N2 genes). Our low-cost molecular test workflow ($1.85 per pooled sample extraction) for HIV RNA and SARS-CoV-2 RNA could serve as an alternative to current standard assays ($12 per pooled sample extraction) for laboratories in low-resource settings.

Highly Sensitive Immunoresistive Sensor for Point-Of-Care Screening for COVID-19.

Current point-of-care (POC) screening of Coronavirus disease 2019 (COVID-19) requires further improvements to achieve highly sensitive, rapid, and inexpensive detection. Here we describe an immunoresistive sensor on a polyethylene terephthalate (PET) film for simple, inexpensive, and highly sensitive COVID-19 screening. The sensor is composed of single-walled carbon nanotubes (SWCNTs) functionalized with monoclonal antibodies that bind to the spike protein of SARS-CoV-2. Silver electrodes are silkscreen-printed on SWCNTs to reduce contact resistance. We determine the SARS-CoV-2 status via the resistance ratio of control- and SARS-CoV-2 sensor electrodes. A combined measurement of two adjacent sensors enhances the sensitivity and specificity of the detection protocol. The lower limit of detection (LLD) of the SWCNT assay is 350 genome equivalents/mL. The developed SWCNT sensor shows 100% sensitivity and 90% specificity in clinical sample testing. Further, our device adds benefits of a small form factor, simple operation, low power requirement, and low assay cost. This highly sensitive film sensor will facilitate rapid COVID-19 screening and expedite the development of POC screening platforms.

Diagnostic Accuracy of an At-Home, Rapid Self-test for Influenza: Prospective Comparative Accuracy Study.

BACKGROUND: Rapid diagnostic tests (RDTs) for influenza used by individuals at home could potentially expand access to testing and reduce the impact of influenza on health systems. Improving access to testing could lead to earlier diagnosis following symptom onset, allowing more rapid interventions for those who test positive, including behavioral changes to minimize spread. However, the accuracy of RDTs for influenza has not been determined in self-testing populations. OBJECTIVE: This study aims to assess the accuracy of an influenza RDT conducted at home by lay users with acute respiratory illness compared with that of a self-collected sample by the same individual mailed to a laboratory for reference testing. METHODS: We conducted a comparative accuracy study of an at-home influenza RDT (Ellume) in a convenience sample of individuals experiencing acute respiratory illness symptoms. Participants were enrolled in February and March 2020 from the Greater Seattle region in Washington, United States. Participants were mailed the influenza RDT and reference sample collection materials, which they completed and returned for quantitative reverse-transcription polymerase chain reaction influenza testing in a central laboratory. We explored the impact of age, influenza type, duration, and severity of symptoms on RDT accuracy and on cycle threshold for influenza virus and ribonuclease P, a marker of human DNA. RESULTS: A total of 605 participants completed all study steps and were included in our analysis, of whom 87 (14.4%) tested positive for influenza by quantitative reverse-transcription polymerase chain reaction (70/87, 80% for influenza A and 17/87, 20% for influenza B). The overall sensitivity and specificity of the RDT compared with the reference test were 61% (95% CI 50%-71%) and 95% (95% CI 93%-97%), respectively. Among individuals with symptom onset ≤72 hours, sensitivity was 63% (95% CI 48%-76%) and specificity was 94% (95% CI 91%-97%), whereas, for those with duration >72 hours, sensitivity and specificity were 58% (95% CI 41%-74%) and 96% (95% CI 93%-98%), respectively. Viral load on reference swabs was negatively correlated with symptom onset, and quantities of the endogenous marker gene ribonuclease P did not differ among reference standard positive and negative groups, age groups, or influenza subtypes. The RDT did not have higher sensitivity or specificity among those who reported more severe illnesses. CONCLUSIONS: The sensitivity and specificity of the self-test were comparable with those of influenza RDTs used in clinical settings. False-negative self-test results were more common when the test was used after 72 hours of symptom onset but were not related to inadequate swab collection or severity of illness. Therefore, the deployment of home tests may provide a valuable tool to support the management of influenza and other respiratory infections.