Evaluation and deployment of isotype-specific salivary antibody assays for detecting previous SARS-CoV-2 infection in children and adults
Authors: Thomas Amy C. , Oliver Elizabeth , Baum Holly E. , Gupta Kapil , Shelley Kathryn L. , Long Anna E. , Jones Hayley E. , Smith Joyce , Hitchings Benjamin , di Bartolo Natalie , Vasileiou Kate , Rabi Fruzsina , Alamir Hanin , Eghleilib Malak , Francis Ore , Oliver Jennifer , Morales-Aza Begonia , Obst Ulrike , Shattock Debbie , Barr Rachael , Collingwood Lucy , Duale Kaltun , Grace Niall , Livera Guillaume Gonnage , Bishop Lindsay , Downing Harriet , Rodrigues Fernanda , Timpson Nicholas , Relton Caroline L. , Toye Ashley , Woolfson Derek N. , Berger Imre , Goenka Anu , Davidson Andrew D. , Gillespie Kathleen M. , Williams Alistair J. K. , Bailey Mick , Brooks-Pollock Ellen , Finn Adam , Halliday Alice ,   the CoMMinS Study Team, Alamir Hanin , Baum Holly E. , Goenka Anu , Halliday Alice , Hitchings Ben , Oliver Elizabeth , Shattock Debbie , Smith Joyce , Thomas Amy C. , Adegbite David , Antico Rupert , Atkins Jamie , Baxter Edward , Bishop Lindsay , Boon Adam , Bridgeman Emma , Collingwood Lucy , Derrick Catherine , Fleming Leah , Garcia Ricardo Garcia , Liveria Guillaume Gonnage , Grace Niall , Grimwood Lucy , Kinney Jane , Myrtou Rafaella , O’Rouke Alice , Oliver Jenny , Payne Chloe , Pennie Rhian , Powell Millie , Garcia Laura Ratero , Storer-Martin Aoife , Summerhill John , Taylor Amy , Taylor Zoe , Thompson Helen , Thomson-Hill Samantha , Underwood Louis , Valentine Gabriella , Vergnano Stefania , Way Amelia , White Maddie , Williams Arthur , Allen David , Anderson Josh , Ardeshir Mariella , Booth Michael , Butler Charles , Chaulagain Monika , Darling Alex , Dayrell-Armes Nicholas , Duale Kaltun , Eghleilib Malak , Farren Chloe , Freestone Danny , Harkness Jason , Healy William , Flanagan Milo Jeenes , Khalique Maria , King Nadine , Koi Anna , Lyall Maia , Morales-Aza Begonia , Pozo Maria , Pereira Ainhoa Rodriguez , Rosa Jessica , Setter Louise , Thomas Liam , Thomas Dylan , Vowles Jonathan 
Published: March 2023

Abstract
Background
Saliva is easily obtainable non-invasively and potentially suitable for detecting both current and previous SARS-CoV-2 infection, but there is limited evidence on the utility of salivary antibody testing for community surveillance.

Methods
We established 6 ELISAs detecting IgA and IgG antibodies to whole SARS-CoV-2 spike protein, to its receptor binding domain region and to nucleocapsid protein in saliva. We evaluated diagnostic performance, and using paired saliva and serum samples, correlated mucosal and systemic antibody responses. The best-performing assays were field-tested in 20 household outbreaks.

Results
We demonstrate in test accuracy (N = 320), spike IgG (ROC AUC: 95.0%, 92.8–97.3%) and spike IgA (ROC AUC: 89.9%, 86.5–93.2%) assays to discriminate best between pre-pandemic and post COVID-19 saliva samples. Specificity was 100% in younger age groups (0–19 years) for spike IgA and IgG. However, sensitivity was low for the best-performing assay (spike IgG: 50.6%, 39.8–61.4%). Using machine learning, diagnostic performance was improved when a combination of tests was used. As expected, salivary IgA was poorly correlated with serum, indicating an oral mucosal response whereas salivary IgG responses were predictive of those in serum. When deployed to household outbreaks, antibody responses were heterogeneous but remained a reliable indicator of recent infection. Intriguingly, unvaccinated children without confirmed infection showed evidence of exposure almost exclusively through specific IgA responses.

Conclusions
Through robust standardisation, evaluation and field-testing, this work provides a platform for further studies investigating SARS-CoV-2 transmission and mucosal immunity with the potential for expanding salivo-surveillance to other respiratory infections in hard-to-reach settings.