Stringent thresholds in SARS-CoV-2 IgG assays lead to under-detection of mild infections

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Abstract

Background

Thresholds for SARS-CoV-2 antibody assays have typically been determined using samples from symptomatic, often hospitalised, patients. In this setting the sensitivity and specificity of the best performing assays can both exceed 98%. However, antibody assay performance following mild infection is less clear.

Methods

We assessed quantitative IgG responses in a cohort of healthcare workers in Oxford, UK, with a high pre-test probability of Covid-19, in particular the 991/11,475(8.6%) who reported loss of smell/taste. We use anosmia/ageusia and other risk factors as probes for Covid-19 infection potentially undiagnosed by immunoassays by investigating their relationship with antibody readings either side of assay thresholds.

Results

The proportion of healthcare workers reporting anosmia/ageusia increased at antibody readings below diagnostic thresholds using an in-house ELISA ( n = 9324) and the Abbott Architect chemiluminescent microparticle immunoassay (CMIA; n = 11,324): 426/906 (47%) reported anosmia/ageusia with a positive ELISA, 59/449 (13.1%) with high-negative and 326/7969 (4.1%) with low-negative readings. Similarly, by CMIA, 518/1093 (47.4%) with a positive result reported anosmia/ageusia, 106/686 (15.5%) with a high-negative and 358/9563 (3.7%) with a low-negative result. Adjusting for the proportion of staff reporting anosmia/ageusia suggests the sensitivity of both assays in mild infection is lower than previously reported: Oxford ELISA 89.8% (95%CI 86.6–92.8%) and Abbott CMIA 79.3% (75.9–82.7%).

Conclusion

Following mild SARS-CoV-2 infection 10–30% of individuals may have negative immunoassay results. While lowered diagnostic thresholds may result in unacceptable specificity, our findings have implications for epidemiological analyses and result interpretation in individuals with a high pre-test probability. Samples from mild PCR-confirmed infections should be included in SARS-CoV-2 immunoassay evaluations.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12879-021-05878-2.

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Most cited references 25

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Robust T cell immunity in convalescent individuals with asymptomatic or mild COVID-19

Takuya Sekine, André Perez-Potti, Olga Rivera-Ballesteros … (2020)

Summary SARS-CoV-2-specific memory T cells will likely prove critical for long-term immune protection against COVID-19. We here systematically mapped the functional and phenotypic landscape of SARS-CoV-2-specific T cell responses in unexposed individuals, exposed family members, and individuals with acute or convalescent COVID-19. Acute phase SARS-CoV-2-specific T cells displayed a highly activated cytotoxic phenotype that correlated with various clinical markers of disease severity, whereas convalescent phase SARS-CoV-2-specific T cells were polyfunctional and displayed a stem-like memory phenotype. Importantly, SARS-CoV-2-specific T cells were detectable in antibody-seronegative exposed family members and convalescent individuals with a history of asymptomatic and mild COVID-19. Our collective dataset shows that SARS-CoV-2 elicits robust, broad and highly functional memory T cell responses, suggesting that natural exposure or infection may prevent recurrent episodes of severe COVID-19.

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Prevalence of SARS-CoV-2 in Spain (ENE-COVID): a nationwide, population-based seroepidemiological study

Marina Pollán, Beatriz Pérez-Gómez, Roberto Pastor-Barriuso … (2020)

Summary Background Spain is one of the European countries most affected by the COVID-19 pandemic. Serological surveys are a valuable tool to assess the extent of the epidemic, given the existence of asymptomatic cases and little access to diagnostic tests. This nationwide population-based study aims to estimate the seroprevalence of SARS-CoV-2 infection in Spain at national and regional level. Methods 35 883 households were selected from municipal rolls using two-stage random sampling stratified by province and municipality size, with all residents invited to participate. From April 27 to May 11, 2020, 61 075 participants (75·1% of all contacted individuals within selected households) answered a questionnaire on history of symptoms compatible with COVID-19 and risk factors, received a point-of-care antibody test, and, if agreed, donated a blood sample for additional testing with a chemiluminescent microparticle immunoassay. Prevalences of IgG antibodies were adjusted using sampling weights and post-stratification to allow for differences in non-response rates based on age group, sex, and census-tract income. Using results for both tests, we calculated a seroprevalence range maximising either specificity (positive for both tests) or sensitivity (positive for either test). Findings Seroprevalence was 5·0% (95% CI 4·7–5·4) by the point-of-care test and 4·6% (4·3–5·0) by immunoassay, with a specificity–sensitivity range of 3·7% (3·3–4·0; both tests positive) to 6·2% (5·8–6·6; either test positive), with no differences by sex and lower seroprevalence in children younger than 10 years ( 10%) and lower in coastal areas (<3%). Seroprevalence among 195 participants with positive PCR more than 14 days before the study visit ranged from 87·6% (81·1–92·1; both tests positive) to 91·8% (86·3–95·3; either test positive). In 7273 individuals with anosmia or at least three symptoms, seroprevalence ranged from 15·3% (13·8–16·8) to 19·3% (17·7–21·0). Around a third of seropositive participants were asymptomatic, ranging from 21·9% (19·1–24·9) to 35·8% (33·1–38·5). Only 19·5% (16·3–23·2) of symptomatic participants who were seropositive by both the point-of-care test and immunoassay reported a previous PCR test. Interpretation The majority of the Spanish population is seronegative to SARS-CoV-2 infection, even in hotspot areas. Most PCR-confirmed cases have detectable antibodies, but a substantial proportion of people with symptoms compatible with COVID-19 did not have a PCR test and at least a third of infections determined by serology were asymptomatic. These results emphasise the need for maintaining public health measures to avoid a new epidemic wave. Funding Spanish Ministry of Health, Institute of Health Carlos III, and Spanish National Health System.

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Viral dynamics in mild and severe cases of COVID-19

Yang Liu, Li-Meng Yan, Lagen Wan … (2020)

Coronavirus disease 2019 (COVID-19) is a new pandemic disease. We previously reported that the viral load of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) peaks within the first week of disease onset.1, 2 Findings from Feb, 2020, indicated that the clinical spectrum of this disease can be very heterogeneous. 3 Here, we report the viral RNA shedding patterns observed in patients with mild and severe COVID-19. 76 patients admitted to the First Affiliated Hospital of Nanchang University (Nanchang, China) from Jan 21 to Feb 4, 2020, were included in the study. All patients were confirmed to have COVID-19 at the time of admission by RT-PCR. The viral loads of their nasopharyngeal swab samples were estimated with the DCt method (Ctsample – Ctref). Patients who had any of the following features at the time of, or after, admission were classified as severe cases: (1) respiratory distress (≥30 breaths per min); (2) oxygen saturation at rest ≤93%; (3) ratio of partial pressure of arterial oxygen to fractional concentration of oxygen inspired air ≤300 mm Hg; or (4) severe disease complications (eg, respiratory failure, requirement of mechanical ventilation, septic shock, or non-respiratory organ failure). 46 (61%) individuals were classified as mild cases and 30 (39%) were classified as severe cases. The basic demographic data and initial clinical symptoms of these patients are shown in the appendix. Parameters did not differ significantly between the groups, except that patients in the severe group were significantly older than those in the mild group, as expected. 4 No patient died from the infection. 23 (77%) of 30 severe cases received intensive care unit (ICU) treatment, whereas none of the mild cases required ICU treatment. We noted that the DCt values of severe cases were significantly lower than those of mild cases at the time of admission (appendix). Nasopharyngeal swabs from both the left and right nasal cavities of the same patient were kept in a sample collection tube containing 3 mL of standard viral transport medium. All samples were collected according to WHO guidelines. 5 The mean viral load of severe cases was around 60 times higher than that of mild cases, suggesting that higher viral loads might be associated with severe clinical outcomes. We further stratified these data according to the day of disease onset at the time of sampling. The DCt values of severe cases remained significantly lower for the first 12 days after onset than those of corresponding mild cases (figure A ). We also studied serial samples from 21 mild and ten severe cases (figure B). Mild cases were found to have an early viral clearance, with 90% of these patients repeatedly testing negative on RT-PCR by day 10 post-onset. By contrast, all severe cases still tested positive at or beyond day 10 post-onset. Overall, our data indicate that, similar to SARS in 2002–03, 6 patients with severe COVID-19 tend to have a high viral load and a long virus-shedding period. This finding suggests that the viral load of SARS-CoV-2 might be a useful marker for assessing disease severity and prognosis. Figure Viral dynamics in patients with mild and severe COVID-19 (A) DCT values (Ctsample-Ctref) from patients with mild and severe COVID-19 at different stages of disease onset. Median, quartile 1, and quartile 3 are shown. (B) DCT values of serial samples from patients with mild and severe COVID-19. COVID-19=coronavirus disease 2019. *p<0·005.

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Author and article information

Contributors

David W. Eyre: david.eyre@bdi.ox.ac.uk

Journal

Journal ID (nlm-ta): BMC Infect Dis

Journal ID (iso-abbrev): BMC Infect Dis

Title: BMC Infectious Diseases

Publisher: BioMed Central (London )

ISSN (Electronic): 1471-2334

Publication date (Electronic): 18 February 2021

Publication date PMC-release: 18 February 2021

Publication date Collection: 2021

Volume: 21

Electronic Location Identifier: 187

Affiliations

[1 ]GRID grid.4991.5, ISNI 0000 0004 1936 8948, Big Data Institute, Nuffield Department of Population Health, , University of Oxford, ; Oxford, UK

[2 ]GRID grid.410556.3, ISNI 0000 0001 0440 1440, Oxford University Hospitals NHS Foundation Trust, ; Oxford, UK

[3 ]GRID grid.4991.5, ISNI 0000 0004 1936 8948, NIHR Oxford Biomedical Research Centre, , University of Oxford, ; Oxford, UK

[4 ]GRID grid.4991.5, ISNI 0000 0004 1936 8948, NIHR Health Protection Research Unit in Healthcare Associated Infections and Antimicrobial Resistance at University of Oxford in partnership with Public Health England, ; Oxford, UK

[5 ]GRID grid.8348.7, ISNI 0000 0001 2306 7492, Microbiology Department, , John Radcliffe Hospital, ; Headley Way, Oxford, OX3 9DU UK

[6 ]GRID grid.4991.5, ISNI 0000 0004 1936 8948, Nuffield Department of Medicine, , University of Oxford, ; Oxford, UK

[7 ]GRID grid.4991.5, ISNI 0000 0004 1936 8948, Kennedy Institute of Rheumatology Research, , University of Oxford, ; Oxford, UK

[8 ]GRID grid.4991.5, ISNI 0000 0004 1936 8948, Target Discovery Institute, University of Oxford, ; Oxford, UK

[9 ]GRID grid.412433.3, ISNI 0000 0004 0429 6814, Oxford University Clinical Research Unit, ; Ho Chi Minh City, Vietnam

Article

Publisher ID: 5878

DOI: 10.1186/s12879-021-05878-2

PMC ID: 7889711

PubMed ID: 33602152

SO-VID: 20022ec3-e675-4241-8fe5-9661fc560df4

License:

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