Morning SARS-CoV-2 testing yields better detection of infection due to higher viral loads in saliva and nasal swabs upon waking

BackgroundThe analytical sensitivities of SARS-CoV-2 diagnostic tests span 6 orders of magnitude. Optimizing sample-collection methods to achieve the most reliable detection for a given sensitivity would increase the effectiveness of testing and minimize COVID-19 outbreaks. MethodsFrom September 2020 to April 2021 we performed a household-transmission study in which participants self-collected samples every morning and evening throughout acute SARS-CoV-2 infection. Seventy mildly symptomatic participants collected saliva and, of those, 29 also collected nasal-swab samples. Viral load was quantified in 1194 saliva and 661 nasal-swab samples using a high-analytical-sensitivity RT-qPCR assay (LOD, 1,000 SARS-CoV-2 RNA copies/mL). FindingsViral loads in both saliva and nasal-swab samples were significantly higher in morning-collected samples than evening-collected samples after symptom onset. We used these quantitative measurements to infer which diagnostic tests would have detected infection (based on sample type and test analytical sensitivity). We find that morning collection would have resulted in significantly improved detection and that this advantage would be most pronounced for tests with low to moderate analytical sensitivity, which would likely have missed infections if sampling in the evening. InterpretationCollecting samples for COVID-19 testing in the morning offers a simple and low-cost improvement to clinical diagnostic sensitivity of low- to moderate-analytical-sensitivity tests. The phenomenon of higher viral loads in the morning may also have implications related to when transmission is more likely to occur. FundingBill & Melinda Gates Foundation, Ronald and Maxine Linde Center for New Initiatives (Caltech), Jacobs Institute for Molecular Engineering for Medicine (Caltech) RESEARCH IN CONTEXTO_ST_ABSEvidence before this studyC_ST_ABSReliable COVID-19 diagnostic testing is critical to reducing transmission of SARS-CoV-2 and reducing cases of severe or fatal disease, particularly in areas with limited vaccine access or uptake. Saliva and anterior-nares nasal swabs are common sample types; however, different diagnostic tests using these sample types have a range of analytical sensitivities spanning 6 orders of magnitude, with limits of detection (LODs) between 102 and 108 genomic copy equivalents of SARS-CoV-2 RNA (copies) per mL of sample. Due to limitations in clinical laboratory capacity, many low-resource settings rely on COVID-19 tests that fall on the moderate (LODs of 104 to 105 copies/mL) to lower (LODs of 105 to 108 copies/mL) end of this spectrum of analytical sensitivity. Alterations in sample collection methods, including time of sample collection, may improve the performance of these diagnostics. Added value of this studyThis study quantifies viral loads from saliva and nasal-swab samples that were longitudinally self-collected by symptomatic patients in the morning immediately after waking and in the evening just prior to sleeping throughout the course of acute SARS-CoV-2 infection. The study cohort was composed of mildly or moderately symptomatic individuals (outpatients). This analysis demonstrates significantly higher viral loads in samples collected in the morning, relative to those collected in the evening. When using moderate to lower analytical sensitivity test methods, these loads are inferred to result in significantly better detection of infected individuals in the morning. Implications of available evidenceThese findings suggest that samples collected in the morning immediately after waking will better detect SARS-CoV-2 infection in symptomatic individuals tested by moderate to lower analytical sensitivity COVID-19 diagnostic tests (LODs at or above 104 viral copies per mL of sample), such as many rapid antigen tests currently available.