Extreme differences in SARS-CoV-2 viral loads among respiratory specimen types during presumed pre-infectious and infectious periods

Creators: Winnett, Alexander Viloria; Akana, Reid; Shelby, Natasha; Davich, Hannah; Caldera, Saharai; Yamada, Taikun; Reyna, John Raymond B.; Romano, Anna E.; Carter, Alyssa M.; Kim, Mi Kyung; Thomson, Matt; Tognazzini, Colten; Feaster, Matthew; Goh, Ying-Ying; Chew, Yap Ching; Ismagilov, Rustem F.

Abstract

SARS-CoV-2 viral load measurements from a single specimen type are used to establish diagnostic strategies, interpret clinical-trial results for vaccines and therapeutics, model viral transmission, and understand virus-host interactions. However, measurements from a single specimen type are implicitly assumed to be representative of other specimen types. We quantified viral-load timecourses from individuals who began daily self-sampling of saliva, anterior nares (nasal), and oropharyngeal (throat) swabs before or at the incidence of infection with the Omicron variant. Viral loads in different specimen types from the same person at the same timepoint exhibited extreme differences, up to 109 copies/mL. These differences were not due to variation in sample self-collection, which was consistent. For most individuals, longitudinal viral-load timecourses in different specimen types did not correlate. Throat-swab and saliva viral loads began to rise up to 7 days earlier than nasal-swab viral loads in most individuals, leading to very low clinical sensitivity of nasal swabs during the first days of infection. Individuals frequently exhibited presumably infectious viral loads in one specimen type while viral loads were low or undetectable in other specimen types. Therefore, defining an individual as infectious based on assessment of a single specimen type underestimates the infectious period, and overestimates the ability of that specimen type to detect infectious individuals. For diagnostic COVID-19 testing, these three single specimen types have low clinical sensitivity, whereas a combined throat-nasal swab, and assays with high analytical sensitivity, were inferred to have significantly better clinical sensitivity to detect presumed pre-infectious and infectious individuals.

Additional Information

The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY 4.0 International license. Paper in collection COVID-19 SARS-CoV-2 preprints from medRxiv and bioRxiv. This work was funded in part by a grant from the Ronald and Maxine Linde Center for New Initiatives at the California Institute of Technology (to RFI), a grant from the Jacobs Institute for Molecular Engineering for Medicine at the California Institute of Technology (to RFI), and a DGSOM Geffen Fellowship at the University of California, Los Angeles (to AVW). We sincerely thank the study participants for making this work possible. We thank Lauriane Quenee, Grace Fisher-Adams, Junie Hildebrandt, Megan Hayashi, RuthAnne Bevier, Chantal D'Apuzzo, Ralph Adolphs, Victor Rivera, Steve Chapman, Gary Waters, Leonard Edwards, Gaylene Ursua, Cynthia Ramos, and Shannon Yamashita for their assistance and advice on study implementation and/or administration. We thank Jessica Leong, Ojas Pradhan, Si Hyung Jin, Emily Savela, Bridget Yang, Ekta Patel, Hsiuchen Chen , Paresh Samantaray, Zeynep Turan, Cindy Kim, Trinity Lee, Vanessa Mechan, Katherine Stiefel, Rosie Zedan, Rahulijeet Chadha, Minkyo Lee, and Jenny Ji for volunteering their time to help with this study. We thank Prabhu Gounder, Tony Chang, Jennifer Howes, and Nari Shin for their support with recruitment. Finally, we thank all the case investigators and contact tracers at the Pasadena Public Health Department and Caltech Student Wellness Services for their efforts in study recruitment and their work in the pandemic response. AUTHOR CONTRIBUTIONS (listed alphabetically by last name): Reid Akana (RA): Collaborated with AVW in creating digital participant symptom surveys; assisted with data quality control/curation with NS, HD, SC; created current laboratory information management system (LIMS) for specimen logging and tracking. Creation of iOS application for sample logging/tracking. Configured an SQL database for data storage. Created an Apache server and websites to view study data. Configured FTPS server to catalog PCR data. Wrote a Python package to access study data. Trained study coordinators on SQL. Troubleshooting and QC of LIMS. Made Fig 3(C-D) and SI Figs S3, S4, S5, Table S2, S3, S4, S5, S6. Wrote and edited the manuscript with AVW and NS. Alyssa M. Carter (AMC): Assisted with the inventory and archiving of >6,000 samples at Caltech; coordinated shipment of samples to Caltech with AER and JRBR; assisted with procurement of antigen tests; assisted with organizing volunteers and making participant kits; assisted AER in developing and implementing QC for participant kits. Provided feedback and edited the manuscript. Yap Ching Chew (YCC): Primary liaison with Caltech team. Prepared and provided Zymo SafeCollect kits and related materials to Caltech team. Supervised the extraction, PCR, and QC teams at Pangea Laboratory. Sent PCR results daily to Caltech team. Arranged for Pangea team to perform viral-variant sequencing on selected samples; reported results and provided sequencing files. Saharai Caldera (SC): Study coordinator; recruited, enrolled and maintained study participants with NS and HD; study-data quality control, curation and archiving with RA, NS, HD and MKK; supplies acquisition with AER, NS, HD and MKK. Hannah Davich (HD): Lead study coordinator; co-wrote participant informational sheets with NS; developed recruitment strategies and did outreach with NS; participant kit creation and co-coordinated kit-making by volunteers with AER; recruited, enrolled and maintained study participants with NS and SC; managed the study-coordinator inventory; study-data quality control, curation and archiving with RA, NS, SC and MKK; supplies acquisition with AER, NS, SC and MKK. Matthew Feaster (MF): Co-investigator; collaborated with AVW, MMC, NS, YG, RFI on study design and recruitment strategies; provided guidance and expertise on SARS-CoV-2 epidemiology and local trends. Ying-Ying Goh (Y-YG): Co-investigator; collaborated with AVW, MMC, NS, MF, RFI on study design and recruitment strategies; provided guidance and expertise on SARS-CoV-2 epidemiology and local trends. Rustem F. Ismagilov (RFI): Principal investigator; collaborated with AVW, MMC, NS, MF, YYG on study design and recruitment strategies; provided leadership, technical guidance, oversight of all analyses, and was responsible for obtaining the primary funding for the study. Mi Kyung Kim (MKK): Study coordinator (part-time); maintaining participants with NS, HD, and SC; study-data quality control, curation and archiving with RA, NS, SC and HD; supplies acquisition with AER, NS, SC and HD; collected contact info for local university/college student health centers for recruitment outreach; assembled Table S1 with NS. John Raymond B. Reyna (JRBR): Organized sample labeling and short-term storage of all samples at Pangea Laboratories. Arranged shipment of all samples to Caltech team. Assisted with processing of the specimens. Anna E. Romano (AER): Co-coordinated kit-making by volunteers with HD; implemented QC process for kit-making; participated in kit-making; managed logistics for the inventory and archiving of >6,000 samples at Caltech; supplies acquisition with HD, NS, SC and MKK; assisted with securing funding; compiled Table S3; organized and performed QC on sequencing data. Provided feedback and edited the manuscript. Natasha Shelby (NS): Study administrator; collaborated with AVW, RFI, YG, MF on initial study design and recruitment strategies; co-wrote IRB protocol and informed consent with AVW; co-wrote enrollment questionnaire and post-study questionnaire with AVW; initiated the collaboration with Zymo and served as primary liaison throughout study; reviewed pilot sampling data and amended instructional sheets/graphics for specimen collections in collaboration with Zymo; co-wrote participant informational sheets with HD; hired, trained, and supervised the study-coordinator team; developed recruitment strategies and did outreach with HD; recruited, enrolled and maintained study participants with HD and SC; co-developed participant keep/drop criteria with AVW; performed the daily upload, review, and QC of PCR data received from Zymo; made the daily keep/drop decisions based on viral-load trajectories in each household; made all phone calls to alert presumptive positives of their status and provide resources; study-data quality control, curation and archiving with RA, HD, SC and MKK; organized archiving of all participant data and antigen-test photographs; supplies acquisition with AER, HD, SC and MKK; assisted with securing funding; managed the overall study budget; assembled Fig 1 with AVW; assembled Table S1 with MKK; made Fig 2 with AVW; managed citations and reference library; verified the underlying data with AVW and RA; co-wrote and edited the manuscript with AVW and RA. Matt Thomson (MT): Assisted with statistical approach and analyses. Colten Tognazzini (CT): Coordinated the recruitment efforts at PPHD with case investigators and contact tracers; provided guidance and expertise on SARS-CoV-2 epidemiology and local trends. Alexander Viloria Winnett (AVW): Collaborated with NS, RFI, YG, MF on initial study design and recruitment strategies; co-wrote IRB protocol and informed consent with NS; co-wrote enrollment questionnaire and post-study questionnaire with NS; co-developed participant keep/drop criteria with NS; funding acquisition; designed and coordinated LOD validation experiments; selected and prepared specimen for viral-variant sequencing with NS, YC, and AER; assisted with the inventory and archiving of >6,000 specimen at Caltech with AER and AMC; minor role supporting outreach by HD and NS; minor role supporting kit-making by AER, HD and AMC; verified the underlying data with NS and RA; assembled Fig 1 with NS; made Fig 2 with NS; performed analysis and prepared Figs 4-7, Table S2, Fig S1, S2, S6, S7, S8, S9. Co-wrote and edited the manuscript with NS and RA. Taikun Yamada (TY): Performed the RT-qPCR COVID-19 testing at Pangea Laboratory. DATA AVAILABILITY. The data underlying the results presented in the study can be accessed at CaltechDATA: https://data.caltech.edu/records/20223. COMPETING INTERESTS STATEMENT. RFI is a co-founder, consultant, and a director and has stock ownership of Talis Biomedical Corp.

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