Retrospective monkeypox virus (MPXV) surveillance among male users of I Want The Kit in Maryland, United States.

Retrospective surveillance leveraging male rectal swab sample remnants from I Want The Kit from July 2021 through October 2023, identified one symptomatic and one asymptomatic mpox case at the peak of transmission in 2022. Although sporadic cases continue to be reported in Maryland, additional asymptomatic cases were not identified in this leveraged surveillance.

Application of machine learning models to identify serological predictors of COVID-19 severity and outcomes.

Critically ill people with COVID-19 have greater antibody titers than those with mild to moderate illness, but their association with recovery or death from COVID-19 has not been characterized. In 178 COVID-19 patients, 73 non-hospitalized and 105 hospitalized patients, mucosal swabs and plasma samples were collected at hospital enrollment and up to 3 months post-enrollment (MPE) to measure virus RNA, cytokines/chemokines, binding antibodies, ACE2 binding inhibition, and Fc effector antibody responses against SARS-CoV-2. The association of demographic variables and >20 serological antibody measures with intubation or death due to COVID-19 was determined using machine learning algorithms. Predictive models revealed that IgG binding and ACE2 binding inhibition responses at 1 MPE were positively and C1q complement activity at enrollment was negatively associated with an increased probability of intubation or death from COVID-19 within 3 MPE. Serological antibody measures were more predictive than demographic variables of intubation or death among COVID-19 patients.

Respiratory viruses in rural Zambia during the second year of the COVID-19 pandemic.

Objectives: Limited data on respiratory infections are available from sub-Saharan Africa during the COVID-19 pandemic. The objective of this study was to evaluate the burden of respiratory viruses in rural Zambia from 2019-2021. Methods: Surveillance was initiated at Macha Hospital in Zambia in December 2018. Each week, patients with respiratory symptoms were enrolled from the outpatient clinic. Nasopharyngeal samples were collected and tested for respiratory pathogens. The prevalence of respiratory symptoms and viruses in 2021 was compared to results from 2019 and 2020. Results: After seeing few cases of influenza virus and respiratory syncytial virus in 2020, a return to prepandemic levels was observed in 2021. Rhinovirus/enterovirus, parainfluenza virus 1-4, and adenovirus circulated from 2019 to 2021, while human metapneumovirus and human coronaviruses (HKU1, 229E, OC43, and NL63 subtypes) were observed sporadically. SARS-CoV-2 was observed consistently in 2021 after being first identified in December 2020. The proportion of participants with co-infections in 2021 (11.6%) was significantly higher than in 2019 (6.9%) or 2020 (7.7%). Conclusion: Declines in influenza virus and respiratory syncytial virus were reversed once public health measures were lifted. Respiratory viruses contributed to a significant burden of respiratory infections in 2021. This study provides important information about respiratory viruses in this changing context and underrepresented region.

Diversity of Anal HPV and Non-HPV Sexually Transmitted Infections and Concordance with Genital Infections in HIV-Infected and HIV-Uninfected Women in the Tapajós Region, Amazon, Brazil.

The aim of this study was to classify the diversity of anal HPV and non-HPV sexually transmitted infections (STIs) and compare the concordance between anal and genital infections in HIV-infected and uninfected women living in the Tapajós region, Amazon, Brazil. A cross-sectional study was performed with 112 HIV-uninfected and 41 HIV-infected nonindigenous women. Anal and cervical scrapings were collected and analyzed for HPV, Chlamydia trachomatis (CT), Neisseria gonorrheae (NG), Trichomonas vaginalis (TV), Mycoplasma genitalium (MG), and Human alphaherpesvirus 2 (HSV-2). The Kappa test evaluated the concordance between anal and genital infections. The overall prevalence of anal HPV infection was 31.3% in HIV-uninfected and 97.6% in HIV-infected women. The most frequent anal high-risk HPV (hrHPV) types were HPV18 and HPV16 in HIV-uninfected women and HPV51, HPV59, HPV31, and HPV58 in HIV-infected women. Anal HPV75 Betapapillomavirus was also identified. Anal non-HPV STIs were identified in 13.0% of all participants. The concordance analysis was fair for CT, MG, and HSV-2, almost perfect agreement for NG, moderate for HPV, and variable for the most frequent anal hrHPV types. Thus, a high prevalence of anal HPV infection with moderate and fair concordance between anal and genital HPV and non-HPV STIs was observed in our study.

Point-of-Care Amenable Detection of Mycoplasma genitalium and Its Antibiotic Resistance Mutations.

Mycoplasma genitalium (MG) is an emerging sexually transmitted bacterium. Due to its fastidious and slow-growing nature, MG is difficult to detect through culture-based diagnostics. Like Neisseria gonorrheae, another bacterial pathogen linked to sexually transmitted infections (STIs), MG has developed resistance to macrolide and fluoroquinolone antibiotics used to treat STIs. The ability to detect MG and identify genomic mutations associated with antibiotic resistance simultaneously can enable antibiotic stewardship and mitigate the spread of antibiotic-resistant MG. Toward this end, we first developed a multiplexed probe-based PCR-melt assay that detects MG and the presence of macrolide resistance mutations in the 23S rRNA gene and fluoroquinolone resistance mutations in the parC gene. Each target was identified via its unique combination of fluorescence label and melting temperature. This approach allowed differentiation between the different types of mutations at the genes of interest. Following initial assay optimization, the assay was integrated into a droplet magnetofluidic cartridge used in a portable platform to integrate automated sample extraction, PCR amplification, and detection. Lastly, we demonstrated that the integrated assay and droplet magnetofluidic platform could detect MG and antibiotic resistance-associated mutations in clinical isolates spiked into urine samples in 40 min.

Limitations of Molecular and Antigen Test Performance for SARS-CoV-2 in Symptomatic and Asymptomatic COVID-19 Contacts.

COVID-19 has brought unprecedented attention to the crucial role of diagnostics in pandemic control. We compared severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) test performance by sample type and modality in close contacts of SARS-CoV-2 cases. Close contacts of SARS-CoV-2-positive individuals were enrolled after informed consent. Clinician-collected nasopharyngeal (NP) swabs in viral transport media (VTM) were tested with a routine clinical reference nucleic acid test (NAT) and PerkinElmer real-time reverse transcription-PCR (RT-PCR) assay; positive samples were tested for infectivity using a VeroE6TMPRSS2 cell culture model. Self-collected passive drool was also tested using the PerkinElmer RT-PCR assay. For the first 4 months of study, midturbinate swabs were tested using the BD Veritor rapid antigen test. Between 17 November 2020 and 1 October 2021, 235 close contacts of SARS-CoV-2 cases were recruited, including 95 with symptoms (82% symptomatic for ≤5 days) and 140 asymptomatic individuals. Reference NATs were positive for 53 (22.6%) participants; 24/50 (48%) were culture positive. PerkinElmer testing of NP and saliva samples identified an additional 28 (11.9%) SARS-CoV-2 cases who tested negative by reference NAT. Antigen tests performed for 99 close contacts showed 83% positive percent agreement (PPA) with reference NAT among early symptomatic persons, but 18% PPA in others; antigen tests in 8 of 11 (72.7%) culture-positive participants were positive. Contacts of SARS-CoV-2 cases may be falsely negative early after contact, but more sensitive platforms may identify these cases. Repeat or serial SARS-CoV-2 testing with both antigen and molecular assays may be warranted for individuals with high pretest probability for infection.

Evaluation of Four Point of Care (POC) Antigen Assays for the Detection of the SARS-CoV-2 Variant Omicron.

Ensuring SARS-CoV-2 diagnostics that can reliably detect emerging variants has been an ongoing challenge. Due to the rapid spread of the Omicron variant, point-of-care (POC) antigen tests have become more widely used. This study aimed at (i) comparing the analytical sensitivity (LOD) of 4 POC antigen assays, BD Veritor, Abbott BinaxNow, Orasure InteliSwab and Quidel QuickVue, for the Omicron versus the Delta variant and (ii) verifying the reproducible detection of Omicron by the 4 antigen assays. The LOD for all four assays were evaluated using Omicron and Delta virus stocks quantified for infectivity and genome copies. The four assays detected all replicates of Omicron and Delta dilutions at 104 and 105 TCID50/mL, respectively. We quantified both viral stocks using droplet digital PCR (ddPCR), which revealed that the Omicron stock had equivalent copies of the N gene to Delta at a one log lower infectious virus. The Abbott BinaxNow and Orasure InteliSwab had the highest analytical sensitivity for Omicron while the Orasure InteliSwab and the Quidel QuickVue had the highest analytical sensitivity for Delta. When 14 SARS-CoV-2 real-time PCR positive nasal/nasopharyngeal swab samples (12 Omicron and 2 Delta, mean Ct = 19.1), were tested by the four assays, only the QuickVue detected all samples. Antigen test positivity correlated with recovery of infectious virus on cell culture in 9 out of 13 tested specimens from symptomatic, asymptomatic, unvaccinated, and vaccinated individuals. Although our study confirms the reduced analytical sensitivity of antigen testing compared to molecular methods, the Omicron variant was detectable by the four evaluated rapid antigen tests. IMPORTANCE In the manuscript, we report an evaluation of the capability of 4 point of care (POC) antigen assays, the BD Veritor, Abbott BinaxNow, Orasure InteliSwab and Quidel QuickVue to detect the Omicron variant of SARS-CoV-2, and we compared their analytical sensitivity for Omicron versus Delta. In this analysis we found that all four assays detected Omicron and Delta at 104 and 105 TCID50/mL, respectively. We further quantified the viral stocks used by droplet digital (ddPCR) and found that the Omicron stock had equivalent copies of the N gene to Delta at a one log lower infectious virus titer and that an increased RNA to infectious virus ratio may be contributing to discrepancies in limit of detection in Omicron compared to Delta. We evaluated 14 SARS-CoV-2 real-time PCR positive nasal/nasopharyngeal swab samples (12 Omicron and 2 Delta), with an average cycle threshold value of 19.1, and only the QuickVue showed 100% agreement.

Respiratory viruses in rural Zambia before and during the COVID-19 pandemic.

OBJECTIVES: With the emergence of the COVID-19 pandemic, restrictions were implemented globally to control the virus. Data on respiratory pathogens in sub-Saharan Africa during the COVID-19 pandemic are scarce. This analysis was conducted to evaluate patterns of respiratory pathogens in rural Zambia before and during the first year of the pandemic. METHODS: Surveillance was established in December 2018 at Macha Hospital in southern Zambia. Patients with respiratory symptoms in the outpatient and inpatient clinics were recruited. Nasopharyngeal samples were collected and tested for respiratory pathogens. The prevalence of respiratory symptoms and pathogens was evaluated and compared in the first (December 10, 2018-December 9, 2019) and second (December 10, 2019-November 30, 2020) years of surveillance. RESULTS: Outpatient visits and admissions for respiratory illness significantly decreased from the first to second year, especially among children. SARS-CoV-2 was not detected from any participants in Year 2. Among outpatients and inpatients with respiratory symptoms, the prevalence of respiratory syncytial virus and influenza viruses decreased from the first to second year. In contrast, the prevalence of rhinovirus/enterovirus, metapneumovirus and parainfluenza virus increased. CONCLUSIONS: The epidemiology of respiratory viruses in rural Zambia changed during the first year of the COVID-19 pandemic, suggesting that public health interventions may have had an impact on the introduction and circulation of respiratory pathogens in this area.

Improving the specificity of nucleic acid detection with endonuclease-actuated degradation.

Nucleic acid detection is essential for numerous biomedical applications, but often requires complex protocols and/or suffers false-positive readouts. Here, we describe SENTINEL, an approach that combines isothermal amplification with a sequence-specific degradation method to detect nucleic acids with high sensitivity and sequence-specificity. Target single-stranded RNA or double-stranded DNA molecules are amplified by loop-mediated isothermal amplification (LAMP) and subsequently degraded by the combined action of lambda exonuclease and a sequence-specific DNA endonuclease (e.g., Cas9). By combining the sensitivity of LAMP with the precision of DNA endonucleases, the protocol achieves attomolar limits of detection while differentiating between sequences that differ by only one or two base pairs. The protocol requires less than an hour to complete using a 65 °C heat block and fluorometer, and detects SARS-CoV-2 virus particles in human saliva and nasopharyngeal swabs with high sensitivity.

Retrospective Analysis of Ugandan Men with Urethritis Reveals Mycoplasma genitalium and Associated Macrolide Resistance.

The rising rates of antimicrobial resistance (AMR) in Mycoplasma genitalium globally and the association of this sexually transmitted infection (STI) with cervicitis, urethritis, and HIV are potentially of great public health concern. Data on the epidemiology of M. genitalium in men in sub-Saharan Africa are limited. We sought to determine the prevalence of M. genitalium and macrolide resistance in men with urethritis in Kampala, Uganda. Self-collected penile-meatal swabs and/or urine samples from men with symptomatic urethritis (n = 250) were retrospectively analyzed for the presence of M. genitalium and macrolide resistance markers with the Aptima M. genitalium and ResistancePlus M. genitalium assays. Additionally, demographic and STI coinfection data were used to investigate associations with M. genitalium infection. M. genitalium was detected in 12.8% (32/250) of individuals; 40.6% (n = 13) had M. genitalium monoinfection. Mutations associated with macrolide resistance were detected in 10.7% (3/28) of participants. Coinfection with Neisseria gonorrhoeae was common (41.0%), but M. genitalium was more prevalent in participants without N. gonorrhoeae coinfection (P = 0.001). M. genitalium is common in Ugandan men with urethritis both as a monoinfection and as a coinfection with other curable STIs. Macrolide resistance was present and warrants further research on treatment outcomes and the association between untreated M. genitalium and subsequent morbidity. IMPORTANCE Mycoplasma genitalium is a common sexually transmitted infection associated with urethritis in men. Little is known about M. genitalium infection in men with urethritis in Uganda. We report that 12% of participants in this study were positive for M. genitalium and that resistance to azithromycin, a macrolide antibiotic, is present. Furthermore, we show that either self-collected penile-meatal swabs or urine can be used for detection of M. genitalium.

Viral co-infections are associated with increased rates of hospitalization in those with influenza.

BACKGROUND: Influenza causes significant morbidity and mortality in the United States. Among patients infected with influenza, the presence of bacterial co-infection is associated with worse clinical outcomes; less is known regarding the clinical importance of viral co-infections. The objective of this study was to determine rates of viral co-infections in emergency department (ED) patients with confirmed influenza and association of co-infection with disease severity. METHODS: Secondary analysis of a biorepository and clinical database from a parent study where rapid influenza testing was implemented in four U.S. academic EDs, during the 2014-2015 influenza season. Patients were systematically tested for influenza virus using a validated clinical decision guideline. Demographic and clinical data were extracted from medical records; nasopharyngeal specimens from influenza-positive patients were tested for viral co-infections (ePlex, Genmark Diagnostics). Patterns of viral co-infections were evaluated using chi-square analysis. The association of viral co-infection with hospital admission was assessed using univariate and multivariate regression. RESULTS: The overall influenza A/B positivity rate was 18.1% (1071/5919). Of the 999 samples with ePlex results, the prevalence of viral co-infections was 7.9% (79/999). The most common viral co-infection was rhinovirus/enterovirus (RhV/EV), at 3.9% (39/999). The odds of hospital admission (OR 2.33, 95% CI: 1.01-5.34) increased significantly for those with viral co-infections (other than RhV/EV) versus those with influenza A infection only. CONCLUSION: Presence of viral co-infection (other than RhV/EV) in ED influenza A/B positive patients was independently associated with increased risk of hospital admission. Further research is needed to determine clinical utility of ED multiplex testing.

Limitations of molecular and antigen test performance for SARS-CoV-2 in symptomatic and asymptomatic COVID-19 contacts.

OBJECTIVES: COVID-19 has brought unprecedented attention to the crucial role of diagnostics in pandemic control. We compared SARS-CoV-2 test performance by sample type and modality in close contacts of SARS-CoV-2 cases. METHODS: Close contacts of SARS-CoV-2 positive individuals were enrolled after informed consent. Clinician-collected nasopharyngeal (NP) swabs in viral transport media (VTM) were tested with a nucleic acid test (NAT). NP VTM and self-collected passive drool were tested using the PerkinElmer real-time reverse transcription PCR (RT-PCR) assay. For the first 4 months of study, mid-turbinate swabs were tested using the BD Veritor rapid antigen test. NAT positive NP samples were tested for infectivity using a VeroE6TMPRSS2 cell culture model. RESULTS: Between November 17, 2020, and October 1, 2021, 235 close contacts of SARS-CoV-2 cases were recruited, including 95 with symptoms (82% symptomatic for < 5 days) and 140 asymptomatic individuals. NP swab reference tests were positive for 53 (22.6%) participants; 24/50 (48%) were culture positive. PerkinElmer testing of NP and saliva samples identified an additional 28 (11.9%) SARS-CoV-2 cases who tested negative by clinical NAT. Antigen tests performed for 99 close contacts showed 83% positive percent agreement (PPA) with reference NAT among early symptomatic persons, but 18% PPA in others; antigen tests in 8 of 11 (72.7%) culture-positive participants were positive. CONCLUSIONS: Contacts of SARS-CoV-2 cases may be falsely negative early after contact, which more sensitive platforms may identify. Repeat or serial SARS-CoV-2 testing with both antigen and molecular assays may be warranted for individuals with high pretest probability for infection.

Preclinical Validation of a Novel Injection-Molded Swab for the Molecular Assay Detection of SARS-CoV-2.

During the COVID-19 public health emergency, many actions have been undertaken to help ensure that patients and health care providers have timely and continued access to high-quality medical devices to respond effectively. The development and validation of new testing supplies and equipment, including collection swabs, has helped to expand the availability and capability for various diagnostic, therapeutic, and protective medical devices in high demand during the COVID-19 emergency. Here, we report the initial validation of a new injection-molded anterior nasal swab, ClearTip™, that was experimentally validated in a laboratory setting as well as in independent clinical studies in comparison to gold standard flocked swabs. We have also developed an in vitro anterior nasal tissue model which offers a novel, efficient, and clinically relevant validation tool to replicate the clinical swabbing workflow with high fidelity, while being accessible, safe, reproducible, and time- and cost-effective. ClearTip™ displayed greater inactivated virus release in the benchtop model, confirmed by its greater ability to report positive samples in a small clinical study in comparison to flocked swabs. We also quantified the detection of biological materials, as a proxy for viral material, in multi-center pre-clinical and clinical studies which showed a statistically significant difference in one study and a reduction in performance in comparison to flocked swabs. Taken together, these results emphasize the compelling benefits of non-absorbent injection-molded anterior nasal swabs for COVID-19 detection, comparable to standard flocked swabs. Injection-molded swabs, as ClearTip™, could have the potential to support future swab shortages, due to its manufacturing advantages, while offering benefits in comparison to highly absorbent swabs in terms of comfort, limited volume collection, and potential multiple usage.

Impact of coinfection status and comorbidity on disease severity in adult emergency department patients with influenza B.

BACKGROUND: Influenza B accounts for approximately one fourth of the seasonal influenza burden. However, research on the importance of influenza B has received less attention compared to influenza A. We sought to describe the association of both coinfections and comorbidities with disease severity among adults presenting to emergency departments (ED) with influenza B. METHODS: Nasopharyngeal samples from patients found to be influenza B positive in four US and three Taiwanese ED over four consecutive influenza seasons (2014-2018) were tested for coinfections with the ePlex RP RUO panel. Multivariable logistic regressions were fitted to model adjusted odds ratios (aOR) for two severity outcomes separately: hospitalization and pneumonia diagnosis. Adjusting for demographic factors, underlying health conditions, and the National Early Warning Score (NEWS), we estimated the association of upper respiratory coinfections and comorbidity with disease severity (including hospitalization or pneumonia). RESULTS: Amongst all influenza B positive individuals (n = 446), presence of another upper respiratory pathogen was associated with an increased likelihood of hospitalization (aOR = 2.99 [95% confidence interval (95% CI): 1.14-7.85, p = 0.026]) and pneumonia (aOR = 2.27 [95% CI: 1.25-4.09, p = 0.007]). Chronic lung diseases (CLD) were the strongest predictor for hospitalization (aOR = 3.43 [95% CI: 2.98-3.95, p < 0.001]), but not for pneumonia (aOR = 1.73 [95% CI: 0.80-3.78, p = 0.166]). CONCLUSION: Amongst ED patients infected with influenza B, the presence of other upper respiratory pathogens was independently associated with both hospitalization and pneumonia; presence of CLD was also associated with hospitalization. These findings may be informative for ED clinician's in managing patients infected with influenza B.

Nosocomial Respiratory Infections in a Rural Zambian Hospital.

The burden of nosocomial respiratory infections in rural southern Africa is poorly understood. We established a surveillance program at a rural Zambian hospital to detect influenza-like illness (ILI) and respiratory infections among hospitalized patients and a cohort of healthcare workers (HCWs). Nasopharyngeal specimens from symptomatic patients and HCWs underwent broadly multiplexed molecular testing to detect viruses and atypical bacteria. During 1 year of surveillance, 15 patients (1.7% of admissions) developed ILI more than 48 hours after admission. Among 44 HCWs, 19 (43%) experienced at least one ILI episode, with a total of 31 ILI episodes detected. Respiratory viruses were detected in 45% of patient and 55% of HCW specimens. The cumulative incidence of influenza infection among HCWs over 1 year was 9%. Overall, respiratory viruses were commonly found among patients and HCWs in a rural Zambian hospital with limited infection control infrastructure.

Delayed Rise of Oral Fluid Antibodies, Elevated BMI, and Absence of Early Fever Correlate With Longer Time to SARS-CoV-2 RNA Clearance in a Longitudinally Sampled Cohort of COVID-19 Outpatients.

BACKGROUND: Sustained molecular detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in the upper respiratory tract (URT) in mild to moderate coronavirus disease 2019 (COVID-19) is common. We sought to identify host and immune determinants of prolonged SARS-CoV-2 RNA detection. METHODS: Ninety-five symptomatic outpatients self-collected midturbinate nasal, oropharyngeal (OP), and gingival crevicular fluid (oral fluid) samples at home and in a research clinic a median of 6 times over 1-3 months. Samples were tested for viral RNA, virus culture, and SARS-CoV-2 and other human coronavirus antibodies, and associations were estimated using Cox proportional hazards models. RESULTS: Viral RNA clearance, as measured by SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR), in 507 URT samples occurred a median (interquartile range) 33.5 (17-63.5) days post-symptom onset. Sixteen nasal-OP samples collected 2-11 days post-symptom onset were virus culture positive out of 183 RT-PCR-positive samples tested. All participants but 1 with positive virus culture were negative for concomitant oral fluid anti-SARS-CoV-2 antibodies. The mean time to first antibody detection in oral fluid was 8-13 days post-symptom onset. A longer time to first detection of oral fluid anti-SARS-CoV-2 S antibodies (adjusted hazard ratio [aHR], 0.96; 95% CI, 0.92-0.99; P = .020) and body mass index (BMI) ≥25 kg/m2 (aHR, 0.37; 95% CI, 0.18-0.78; P = .009) were independently associated with a longer time to SARS-CoV-2 viral RNA clearance. Fever as 1 of first 3 COVID-19 symptoms correlated with shorter time to viral RNA clearance (aHR, 2.06; 95% CI, 1.02-4.18; P = .044). CONCLUSIONS: We demonstrate that delayed rise of oral fluid SARS-CoV-2-specific antibodies, elevated BMI, and absence of early fever are independently associated with delayed URT viral RNA clearance.

Identification of H3N2 NA and PB1-F2 genetic variants and their association with disease symptoms during the 2014-15 influenza season.

The 2014-15 influenza season saw the emergence of an H3N2 antigenic drift variant that formed the 3C.2a HA clade. Whole viral genomes were sequenced from nasopharyngeal swabs of ninety-four patients with confirmed influenza A virus infection and primary human nasal epithelial cell cultures used to efficiently isolate H3N2 viruses. The isolates were classified by HA clade and the presence of a new set of co-selected mutations in NA (a glycosylation site, NAg+) and PB1-F2 (H75P). The NA and PB1-F2 mutations were present in a subset of clade 3C.2a viruses (NAg+F2P), which dominated during the subsequent influenza seasons. In human nasal epithelial cell cultures, a virus with the novel NAg+F2P genotype replicated less well compared with a virus with the parental genotype. Retrospective analyses of clinical data showed that NAg+F2P genotype viruses were associated with increased cough and shortness of breath in infected patients.

Delayed rise of oral fluid antibodies, elevated BMI, and absence of early fever correlate with longer time to SARS-CoV-2 RNA clearance in an longitudinally sampled cohort of COVID-19 outpatients.

BACKGROUND: Sustained molecular detection of SARS-CoV-2 RNA in the upper respiratory tract (URT) in mild to moderate COVID-19 is common. We sought to identify host and immune determinants of prolonged SARS-CoV-2 RNA detection. METHODS: Ninety-five outpatients self-collected mid-turbinate nasal, oropharyngeal (OP), and gingival crevicular fluid (oral fluid) samples at home and in a research clinic a median of 6 times over 1-3 months. Samples were tested for viral RNA, virus culture, and SARS-CoV-2 and other human coronavirus antibodies, and associations were estimated using Cox proportional hazards models. RESULTS: Viral RNA clearance, as measured by SARS-CoV-2 RT-PCR, in 507 URT samples occurred a median (IQR) 33.5 (17-63.5) days post-symptom onset. Sixteen nasal-OP samples collected 2-11 days post-symptom onset were virus culture positive out of 183 RT-PCR positive samples tested. All participants but one with positive virus culture were negative for concomitant oral fluid anti-SARS-CoV-2 antibodies. The mean time to first antibody detection in oral fluid was 8-13 days post-symptom onset. A longer time to first detection of oral fluid anti-SARS-CoV-2 S antibodies (aHR 0.96, 95% CI 0.92-0.99, p=0.020) and BMI ≥ 25kg/m 2 (aHR 0.37, 95% CI 0.18-0.78, p=0.009) were independently associated with a longer time to SARS-CoV-2 viral RNA clearance. Fever as one of first three COVID-19 symptoms correlated with shorter time to viral RNA clearance (aHR 2.06, 95% CI 1.02-4.18, p=0.044). CONCLUSIONS: We demonstrate that delayed rise of oral fluid SARS-CoV-2-specific antibodies, elevated BMI, and absence of early fever are independently associated with delayed URT viral RNA clearance.

Identification of pathogens from the upper respiratory tract of adult emergency department patients at high risk for influenza complications in a pre-Sars-CoV-2 environment.

The emergence of SARS-CoV-2 and subsequent COVID-19 pandemic highlights the morbidity and potential disease severity caused by respiratory viruses. To elucidate pathogen prevalence, etiology of coinfections and URIs from symptomatic adult Emergency department patients in a pre-SARS-CoV-2 environment, we evaluated specimens from four geographically diverse Emergency departments in the United States from 2013-2014 utilizing ePlex RP RUO cartridges (Genmark Diagnostics). The overall positivity was 30.1% (241/799), with 6.6% (16/241) coinfections. Noninfluenza pathogens from most to least common were rhinovirus/enterovirus, coronavirus, human metapneumovirus and RSV, respectively. Broad differences in disease prevalence and pathogen distributions were observed across geographic regions; the site with the highest detection rate (for both mono and coinfections) demonstrated the greatest pathogen diversity. A variety of respiratory pathogens and geographic variations in disease prevalence and copathogen type were observed. Further research is required to evaluate the clinical relevance of these findings, especially considering the SARS-CoV-2 pandemic and related questions regarding SARS-CoV-2 disease severity and the presence of co-infections.

Self-Collected Oral Fluid Saliva Is Insensitive Compared With Nasal-Oropharyngeal Swabs in the Detection of Severe Acute Respiratory Syndrome Coronavirus 2 in Outpatients.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic control will require widespread access to accurate diagnostics. Salivary sampling circumvents swab supply chain bottlenecks, is amenable to self-collection, and is less likely to create an aerosol during collection compared with the nasopharyngeal swab. METHODS: We compared real-time reverse-transcription polymerase chain reaction Abbott m2000 results from matched salivary oral fluid (gingival crevicular fluid collected in an Oracol device) and nasal-oropharyngeal (OP) self-collected specimens in viral transport media from a nonhospitalized, ambulatory cohort of coronavirus disease 2019 (COVID-19) patients at multiple time points. These 2 sentences should be at the beginning of the results. RESULTS: There were 171 matched specimen pairs. Compared with nasal-OP swabs, 41.6% of the oral fluid samples were positive. Adding spit to the oral fluid percent collection device increased the percent positive agreement from 37.2% (16 of 43) to 44.6% (29 of 65). The positive percent agreement was highest in the first 5 days after symptoms and decreased thereafter. All of the infectious nasal-OP samples (culture positive on VeroE6 TMPRSS2 cells) had a matched SARS-CoV-2 positive oral fluid sample. CONCLUSIONS: In this study of nonhospitalized SARS-CoV-2-infected persons, we demonstrate lower diagnostic sensitivity of self-collected oral fluid compared with nasal-OP specimens, a difference that was especially prominent more than 5 days from symptom onset. These data do not justify the routine use of oral fluid collection for diagnosis of SARS-CoV-2 despite the greater ease of collection. It also underscores the importance of considering the method of saliva specimen collection and the time from symptom onset especially in outpatient populations.