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Important questions remain about the sources of transmission of pneumococcus to older adults in the community. This is critical for understanding the potential effects of using pneumococcal conjugate vaccines (PCVs) in children and older adults. For non-institutionalized individuals, we hypothesized that the most likely source of adult-to-adult transmission is within the household. We designed a longitudinal study to sample adults ≥60 years of age living in the same household (New Haven, CT, USA), without younger residents in the household. Saliva samples and social and health questionnaires were obtained every 2 weeks for a period of 10 weeks. DNA extracted from culture-enriched saliva was tested using qPCR for pneumococcus genes piaB, lytA , and serotype. Across two study seasons (November 2020-August 2021, November 2021-September 2022), 121 individuals from 61 households completed all six visits; 62 individuals were enrolled in both seasons. Overall, 52/1088 (4.8%) samples tested positive for pneumococcus, with 27/121 (22.3%) individuals colonized at least once. Several individuals were colonized at multiple time points; two individuals were colonized at 5/6 time points and two at all six. In 5 instances, both household members were carriers in the same season, though not necessarily at the same time. Pneumococcal carriage was substantially higher among individuals who had contact with children (10.0% vs. 1.6%). Contact with young children was the most important factor that influenced pneumococcal acquisition rates. While there were several instances where both adult household members were colonized at the same time or at sequential visits, these individuals typically had contact with children. As such, PCV immunization can directly protect older adults who have contact with children., Competing Interests: A.L.W. has received consulting and/or advisory board fees from Pfizer, Merck, Diasorin, PPS Health, Primary Health, Co-Diagnostics, and Global Diagnostic Systems for work unrelated to this project, and is Principal Investigator on research grants from Pfizer, Merck, and NIH RADx UP to Yale University and from NIH RADx, Balvi.io and Shield T3 to SalivaDirect, Inc.. D.M.W. has received consulting fees from Pfizer, Merck, GSK, and Matrivax for work unrelated to this project and is the Principal Investigator on research grants and contracts with Pfizer and Merck to Yale University. This work has been previously presented in part at ESCMID Global 2024; the 13th International Symposium on Pneumococci and Pneumococcal Diseases (ISPPD-13), Cape Town, South Africa; and IDWeek 2023, Boston, MA, USA., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Molecular methods have improved the sensitivity of the detection of pneumococcal carriage in saliva. However, they typically require sample culture enrichment and nucleic acid extraction prior to performing the detection assay and may limit scalability for extensive surveillance of pneumococcus, particularly in low-resource settings. We evaluated the performance of a DNA-extraction-free method for the detection of pneumococcus in saliva. We developed a streamlined qPCR-based protocol for the detection of pneumococcus, omitting culture enrichment and DNA extraction. Using saliva samples collected from children attending childcare centers (New Haven, CT, USA), we evaluated the detection of pneumococcus using saliva lysates as compared to purified DNA extracted from culture-enriched aliquots of the paired samples using qPCR targeting the pneumococcal piaB gene. Of the 759 saliva samples tested from 92 children [median age 3.65 years; IQR (2.46-4.78)], pneumococcus was detected in 358 (47.2%) saliva lysates prepared using the extraction-free protocol and in 369 (48.6%) DNA extracted from culture-enriched samples. We observed near-perfect agreement between the two protocols (Cohen's kappa: 0.92; 95% CI: 0.90-0.95). Despite a high correlation between C T values generated by the two methods ( r = 0.93, P < 0.0001), the C T values generated from saliva lysates were higher (lower concentration) than those from culture-enriched samples (Δ C T = 6.69, P < 0.00001). The cost of detecting pneumococcus using saliva lysates was at least fivefold lower (US$2.53) compared to the cost of the culture-enriched method (range: US$13.60-US$19.46). For pneumococcal carriage surveillance in children, our findings suggest that a DNA extraction-free approach may offer a cost-effective alternative to the resource-intensive culture-enrichment method.IMPORTANCESurveillance for carriage of pneumococcus is a key component of evaluating the performance of pneumococcal vaccines and informing new vaccination strategies. To improve the scalability of pneumococcal carriage surveillance, we show that molecular detection of pneumococcus in saliva from children can be performed without culture enrichment and DNA extraction. Our findings show that using the extraction-free method can improve surveillance efforts for pneumococcal carriage in children, overcoming the resource-intensive hurdle that comes with the use of molecular methods, particularly in low-resource settings., Competing Interests: D.M.W. has received consulting fees from Pfizer, Merck, and GSK and is a PI on research grants from Pfizer and Merck. A.L.W. has received consulting and/or advisory board fees from Pfizer, Merck, Diasorin, PPS Health, Co-Diagnostics, and Global Diagnostic Systems for work unrelated to this project and is a principal investigator on research grants from Pfizer, Merck, NIH RADx UP, and SalivaDirect, Inc. to Yale University and on research grants from NIH RADx, Balvi.io, and Shield T3 to SalivaDirect, Inc. All other co-authors declare no conflict of interest.

Most pneumococcal disease occurs among infants and older adults and is thought to be driven by the transmission of Streptococcus pneumoniae from young children to these vulnerable age groups. However, pneumococcal disease outbreaks also affect non-elderly adults living or working in congregate, close-contact settings. Little is known about pneumococcal carriage in such populations. From July to November 2020, we collected saliva from low-income adult farmworkers in Monterey County, California, and tested for pneumococcal carriage following culture enrichment via quantitative PCR assays targeting the pneumococcal lytA and piaB genes. Participants were considered to carry pneumococci if lytA and piaB cycle threshold values were both below 40. Among 1,283 participants enrolled in our study, 117 (9.1%) carried pneumococci. Carriers tended more often than non-carriers to be exposed to children aged <5 years [odds ratio (OR) = 1.45 (0.95-2.20)] and overcrowding [OR = 1.48 (0.96-2.30) and 2.84 (1.20-6.73), respectively, for participants in households with >2-4 and >4 persons per bedroom vs ≤2 persons per bedroom]. Household overcrowding remained associated with increased risk of carriage among participants not exposed to children aged <5 years [OR = 2.05 (1.18-3.59) for participants living in households with >2 vs ≤2 persons per bedroom]. Exposure to children aged <5 years and overcrowding were each associated with increased pneumococcal density among carriers [ piaB c T difference of 2.04 (0.36-3.73) and 2.44 (0.80-4.11), respectively]. While exposure to young children was a predictor of pneumococcal carriage, associations of overcrowding with increased prevalence and density of carriage in households without young children suggest that transmission also occurs among adults in close-contact settings.IMPORTANCEAlthough infants and older adults are the groups most commonly affected by pneumococcal disease, outbreaks are known to occur among healthy, working-age populations exposed to overcrowding, including miners, shipyard workers, military recruits, and prisoners. Carriage of Streptococcus pneumoniae is the precursor to pneumococcal disease, and its relation to overcrowding in adult populations is poorly understood. We used molecular methods to characterize pneumococcal carriage in culture-enriched saliva samples from low-income adult farmworkers in Monterey County, CA. While exposure to children in the household was an important risk factor for pneumococcal carriage, living in an overcrowded household without young children was an independent predictor of carriage as well. Moreover, participants exposed to children or overcrowding carried pneumococci at higher density than those without such exposures, suggesting recent transmission. Our findings suggest that, in addition to transmission from young children, pneumococcal transmission may occur independently among adults in overcrowded settings., Competing Interests: J.A.L. discloses receipt of grant funding from Pfizer, Inc. and Merck, Sharp & Dohme and consulting honoraria from Pfizer, Inc.; Merck, Sharp & Dohme; and VaxCyte, Inc. A.L.W. discloses receipt of grant funding from Pfizer, Inc.; Merck, Sharp & Dohme; SalivaDirect, Inc.; Valvi.io; and Shield T3 and consulting honoraria from Pfizer, Inc.; Merck, Sharp & Dohme; Diasorin; PPS Health; Co-Diagnostics; and Global Diagnostic Systems. All other authors declare no competing interests.

Public perception regarding diagnostic sample types as well as personal experiences can influence willingness to test. As such, public preferences for specific sample type(s) should be used to inform diagnostic and surveillance testing programs to improve public health response efforts. To understand where preferences lie, we conducted an international survey regarding the sample types used for SARS-CoV-2 tests. A Qualtrics survey regarding SARS-CoV-2 testing preferences was distributed via social media and email. The survey collected preferences regarding sample methods and key demographic data. Python was used to analyze survey responses. From March 30th to June 15th, 2022, 2,094 responses were collected from 125 countries. Participants were 55% female and predominantly aged 25-34 years (27%). Education and employment were skewed: 51% had graduate degrees, 26% had bachelor's degrees, 27% were scientists/researchers, and 29% were healthcare workers. By rank sum analysis, the most preferred sample type globally was the oral swab, followed by saliva, with parents/guardians preferring saliva-based testing for children. Respondents indicated a higher degree of trust in PCR testing (84%) vs. rapid antigen testing (36%). Preferences for self- or healthcare worker-collected sampling varied across regions. This international survey identified a preference for oral swabs and saliva when testing for SARS-CoV-2. Notably, respondents indicated that if they could be assured that all sample types performed equally, then saliva was preferred. Overall, survey responses reflected the region-specific testing experiences during the COVID-19. Public preferences should be considered when designing future response efforts to increase utilization, with oral sample types (either swabs or saliva) providing a practical option for large-scale, accessible diagnostic testing., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: ALW has received consulting and/or advisory board fees from Pfizer, Merck, Diasorin, PPS Health, Co-Diagnostics, and Global Diagnostic Systems for work unrelated to this project, and is Principal Investigator on research grants from Pfizer, Merck, NIH RADx UP and SalivaDirect, Inc. to Yale University and from NIH RADx, Balvi.io and Shield T3 to SalivaDirect, Inc. All other co-authors declare no potential conflict of interest., (Copyright: © 2024 Salzano et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Streptococcus pneumoniae is a leading cause of morbidity and mortality in children and older adults. Yet knowledge on the development of pneumococcal protein-specific antibody responses throughout life is limited. To investigate this, we measured serum IgG levels to 55 pneumococcal proteins in 11-month old infants (n=73), 24-month old children (n=101), parents (n=99), adults without children <6 years of age (n= 99) and older adults aged >60 years (n=100). Our findings revealed low IgG levels in infancy, with distinct development patterns peaking in adults. A decrease in levels was observed for 27 antigens towards older age. Adults and older adults had increased IgG levels during pneumococcal carriage and at increased exposure risk to S. pneumoniae. Carriage was a stronger predictor than exposure or age for antibody responses. These findings highlight the dynamic nature of naturally acquired humoral immunity to pneumococcal proteins throughout life, offering insights for age-targeted interventions., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Emergency departments are high-risk settings for severe acute respiratory coronavirus virus 2 (SARS-CoV-2) surface contamination. Environmental surface samples were obtained in rooms with patients suspected of having COVID-19 who did or did not undergo aerosol-generating procedures (AGPs). SARS-CoV-2 RNA surface contamination was most frequent in rooms occupied by coronavirus disease 2019 (COVID-19) patients who received no AGPs.

Despite its prominence in early scientific records, the usefulness of saliva as a respiratory specimen has been de-emphasised over the past century. However, due to its low cost and reliance on specific supply chains and the non-invasive nature of its collection, its benefits over swab-based specimens are again becoming increasingly recognised. These benefits were highlighted over the course of the COVID-19 pandemic, where saliva emerged as a more practical, clinically non-inferior sample type for the detection of SARS-CoV-2 and saw numerous saliva-based diagnostic tests approved for clinical use. Looking forward, as saliva uniquely contains both respiratory secretions and immunological components, it has potentially wide applications, ranging from clinical diagnostics to post-vaccine disease burden and immunity surveillance. This Personal View seeks to summarise the existing evidence for the use of saliva in detecting respiratory pathogens, beyond SARS-CoV-2, as well as detailing methodological factors that can influence sample quality and thus, clinical utility., Competing Interests: Declaration of interests SP has received consulting or travel fees from Pfizer, Inventprise, Global Diagnostic Systems, and Vaxcyte, and is the Principal Investigator on a grant from Merck to Yale University, for work not related to this Personal View. ALW has received consulting or advisory board fees from Pfizer, RADx, Diasorin, PPS Health, Co-Diagnostics, Filtration Group, and Global Diagnostics Systems for work not related to this Personal View. ALW is a Principal Investigator for research grants funded by Pfizer, Merck, Flambeau Diagnostics, Tempus Labs, and The Rockefeller Foundation to Yale University. All other authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY-NC-ND 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

For children, the gold standard for the detection of pneumococcal carriage is conventional culture of a nasopharyngeal swab. Saliva, however, has a history as one of the most sensitive methods for surveillance of pneumococcal colonization and has recently been shown to improve carriage detection in older age groups. Here, we compared the sensitivity of paired nasopharyngeal and saliva samples from PCV7-vaccinated 24-month-old children for pneumococcal carriage detection using conventional and molecular detection methods. Nasopharyngeal and saliva samples were collected from 288 24-month-old children during the autumn/winter, 2012/2013. All samples were first processed by conventional diagnostic culture. Next, DNA extracted from all plate growth was tested by qPCR for the presence of the pneumococcal genes piaB and lytA and a subset of serotypes. By culture, 161/288 (60 %) nasopharyngeal swabs tested positive for pneumococcus, but detection was not possible from saliva due to abundant polymicrobial growth on culture plates. By qPCR, 155/288 (54 %) culture-enriched saliva samples and 187/288 (65 %) nasopharyngeal swabs tested positive. Altogether, 219/288 (76 %) infants tested positive for pneumococcus, with qPCR-based carriage detection of culture-enriched nasopharyngeal swabs detecting significantly more carriers compared to either conventional culture ( P <0.001) or qPCR detection of saliva ( P =0.002). However, 32/219 (15 %) carriers were only positive in saliva, contributing significantly to the overall number of carriers detected ( P =0.002). While testing nasopharyngeal swabs by qPCR proved most sensitive for pneumococcal detection in infants, saliva sampling could be considered as complementary to provide additional information on carriage and serotypes that may not be detected in the nasopharynx and may be particularly useful in longitudinal studies, requiring repeated sampling of study participants.

Early in the pandemic, a simple, open-source, RNA extraction-free RT-qPCR protocol for SARS-CoV-2 detection in saliva was developed and made widely available. This simplified approach (SalivaDirect) requires only sample treatment with proteinase K prior to PCR testing. However, feedback from clinical laboratories highlighted a need for a flexible workflow that can be seamlessly integrated into their current health and safety requirements for the receiving and handling of potentially infectious samples. To address these varying needs, we explored additional pre-PCR workflows. We built upon the original SalivaDirect workflow to include an initial incubation step (95 °C for 30 min, 95 °C for 5 min or 65 °C for 15 min) with or without addition of proteinase K. The limit of detection for the workflows tested did not significantly differ from that of the original SalivaDirect workflow. When tested on de-identified saliva samples from confirmed COVID-19 individuals, these workflows also produced comparable virus detection and assay sensitivities, as determined by RT-qPCR analysis. Exclusion of proteinase K did not negatively affect the sensitivity of the assay. The addition of multiple heat pretreatment options to the SalivaDirect protocol increases the accessibility of this cost-effective SARS-CoV-2 test as it gives diagnostic laboratories the flexibility to implement the workflow which best suits their safety protocols., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Anne L. Wyllie reports financial support was provided by Tempus Labs Inc. Anne L. Wyllie reports financial support was provided by Fast Grant., (Copyright © 2023. Published by Elsevier Ltd.)

Reported rates of invasive pneumococcal disease were markedly lower than normal during the 2020/2021 winter in the Northern Hemisphere, the first year after the start of the COVID-19 pandemic. However, little is known about rates of carriage of pneumococcus among adults during this period. Between October 2020-August 2021, couples in the Greater New Haven Area, USA, were enrolled if both individuals were aged 60 years and above and did not have any individuals under the age of 60 years living in the household. Saliva samples and questionnaires regarding social activities and contacts and medical history were obtained every 2 weeks for a period of 10 weeks. Following culture-enrichment, extracted DNA was tested using qPCR for pneumococcus-specific sequences piaB and lytA . Individuals were considered positive for pneumococcal carriage when Ct values for piaB were ≤40. Results. We collected 567 saliva samples from 95 individuals (47 household pairs and 1 singleton). Of those, 7.1% of samples tested positive for pneumococcus, representing 22/95 (23.2%) individuals and 16/48 (33.3%) households. Study participants attended few social events during this period. However, many participants continued to have regular contact with children. Individuals who had regular contact with preschool and school-aged children (i.e., 2 to 9 year olds) had a higher prevalence of carriage (15.9% versus 5.4%). Despite COVID-19-related disruptions, a large proportion of older adults continued to carry pneumococcus. Prevalence was particularly high among those who had contact with school-aged children, but carriage was not limited to this group. IMPORTANCE Carriage of Streptococcus pneumoniae (pneumococcus) in the upper respiratory tract is considered a prerequisite to invasive pneumococcal disease. During the first year of the COVID-19 pandemic, markedly lower rates of invasive pneumococcal disease were reported worldwide. Despite this, by testing saliva samples with PCR, we found that older adults continued to carry pneumococcus at pre-pandemic levels. Importantly, this study was conducted during a period when transmission mitigation measures related to the COVID-19 pandemic were in place. However, our observations are in line with reports from Israel and Belgium where carriage was also found to persist in children. In line with this, we observed that carriage prevalence was particularly high among the older adults in our study who maintained contact with school-aged children., Competing Interests: The authors declare a conflict of interest. A.L.W. has received consulting and/or advisory board fees from Pfizer, RADx, Diasorin, PPS Health, Co-Diagnostics, Filtration Group, and Global Diagnostic Systems for work unrelated to this project, and is Principal Investigator on research grants with Pfizer, Merck, Flambeau Diagnostics, Tempus Labs, and The Rockefeller Foundation to Yale University. D.M.W. has received consulting fees from Pfizer, Merck, GSK, Affinivax, and Matrivax for work unrelated to this project and is Principal Investigator on research grants and contracts with Pfizer and Merck to Yale University. This work has been previously presented in part at IDweek 2021 (virtually); the 15th European Meeting on the Molecular Biology of the Pneumococcus, Liverpool, United Kingdom; and the 12th International Symposium on Pneumococci and Pneumococcal Diseases (ISPPD-12), Toronto, Canada. Adriano Arguedas (A.A.) is a Pfizer employee and may have stock options at Pfizer.

While the sensitivity of detection of pneumococcal carriage can be improved by testing respiratory tract samples with quantitative PCR (qPCR), concerns have been raised regarding the specificity of this approach. We therefore investigated the reliability of the widely used lytA qPCR assay when applied to saliva samples from older adults in relation to a more specific qPCR assay ( piaB ). During the autumn/winter seasons of 2018/2019 and 2019/2020, saliva was collected at multiple time points from 103 healthy adults aged 21 to 39 ( n = 34) and >64 ( n = 69) years ( n = 344 total samples). Following culture enrichment, extracted DNA was tested using qPCR for piaB and lytA . By sequencing the variable region of rpsB (S2 typing), we identified the species of bacteria isolated from samples testing lytA -positive only. While 30 of 344 (8.7%) saliva samples (16.5% individuals) tested qPCR-positive for both piaB and lytA , 52 (15.1%) samples tested lytA -positive only. No samples tested piaB -positive only. Through extensive reculture attempts of the lytA -positive samples collected in 2018/2019, we isolated 23 strains (in 8 samples from 5 individuals) that were also qPCR-positive for only lytA . Sequencing determined that Streptococcus mitis and Streptococcus infantis were predominantly responsible for this lytA -positive qPCR signal. We identified a comparatively large proportion of samples generating positive signals with the widely used lytA qPCR and identified nonpneumococcal Streptococcus species responsible for this signal. This highlights the importance of testing for the presence of multiple gene targets in tandem for reliable and specific detection of pneumococcus in polymicrobial respiratory tract samples. IMPORTANCE Testing saliva samples with quantitative PCR (qPCR) improves the sensitivity of detection of pneumococcal carriage. The qPCR assay targeting lytA , the gene encoding the major pneumococcal autolysin, has become widely accepted for the identification of pneumococcus and is even considered the "gold standard" by many. However, when applying this approach to investigate the prevalence of pneumococcal carriage in adults in New Haven, CT, USA, we identified nonpneumococcal Streptococcus spp. that generate positive signals in this widely used assay. By testing also for piaB (encoding the iron acquisition ABC transporter lipoprotein, PiaB), our findings demonstrate the importance of testing for the presence of multiple gene targets in tandem for reliable molecular detection of pneumococcus in respiratory tract samples; targeting only lytA may lead to an overestimation of true carriage rates., Competing Interests: The authors declare a conflict of interest. A.L.W has received consulting and/or advisory board fees from Pfizer, Diasorin, PPS Health, Co-Diagnostics, and Global Diagnostic Systems for work unrelated to this project, and is Principal Investigator on research grants from Pfizer, Merck, and Flambeau Diagnostics to Yale University. D.M.W has received consulting fees from Pfizer, Merck, GSK, Affinivax, and Matrivax for work unrelated to this project and is Principal Investigator on research grants and contracts with Pfizer and Merck to Yale University. All other co-authors declare no potential conflict of interest.

The key to limiting SARS-CoV-2 spread is to identify virus-infected individuals (both symptomatic and asymptomatic) and isolate them from the general population. Hence, routine weekly testing for SARS-CoV-2 in all asymptomatic (capturing both infected and non-infected) individuals is considered critical in situations where a large number of individuals co-congregate such as schools, prisons, aged care facilities and industrial workplaces. Such testing is hampered by operational issues such as cost, test availability, access to healthcare workers and throughput. We developed the SalivaDirect RT-qPCR assay to increase access to SARS-CoV-2 testing via a low-cost, streamlined protocol using self-collected saliva. To expand the single sample testing protocol, we explored multiple extraction-free pooled saliva testing workflows prior to testing with the SalivaDirect RT-qPCR assay. A pool size of five, with or without heat inactivation at 65 °C for 15 min prior to testing resulted in a positive agreement of 98% and 89%, respectively, and an increased Ct value shift of 1.37 and 1.99 as compared to individual testing of the positive clinical saliva specimens. Applying this shift in Ct value to 316 individual, sequentially collected, SARS-CoV-2 positive saliva specimen results reported from six clinical laboratories using the original SalivaDirect assay, 100% of the samples would have been detected (Ct value < 45) had they been tested in the 1:5 pool strategy. The availability of multiple pooled testing workflows for laboratories can increase test turnaround time, permitting results in a more actionable time frame while minimizing testing costs and changes to laboratory operational flow., (© 2023. The Author(s).)

Background: Streptococcus pneumoniae interacts with numerous viral respiratory pathogens in the upper airway. It is unclear whether similar interactions occur with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)., Methods: We collected saliva specimens from working-age adults undergoing SARS-CoV-2 molecular testing at outpatient clinics and via mobile community-outreach testing between July and November 2020 in Monterey County, California. After bacterial culture enrichment, we tested for pneumococci by means of quantitative polymerase chain reaction targeting the lytA and piaB genes, and we measured associations with SARS-CoV-2 infection using conditional logistic regression., Results: Analyses included 1278 participants, with 564 enrolled in clinics and 714 enrolled through outreach-based testing. The prevalence of pneumococcal carriage was 9.2% (117 of 1278) among all participants (11.2% [63 of 564] in clinic-based testing and 7.6% [54 of 714] in outreach-based testing). The prevalence of SARS-CoV-2 infection was 27.4% (32 of 117) among pneumococcal carriers and 9.6% (112 of 1161) among noncarriers (adjusted odds ratio [aOR], 2.73 [95% confidence interval (CI): 1.58-4.69). Associations between SARS-CoV-2 infection and pneumococcal carriage were enhanced in the clinic-based sample (aOR, 4.01 [95% CI: 2.08-7.75]) and among symptomatic participants (3.38 [1.35-8.40]), compared with findings within the outreach-based sample and among asymptomatic participants. The adjusted odds of SARS-CoV-2 coinfection increased 1.24-fold (95% CI: 1.00-1.55-fold) for each 1-unit decrease in piaB quantitative polymerase chain reaction cycle threshold value among pneumococcal carriers. Finally, pneumococcal carriage modified the association of SARS-CoV-2 infection with recent exposure to a suspected coronavirus disease 2019 case (aOR, 7.64 [95% CI: 1.91-30.7] and 3.29 [1.94-5.59]) among pneumococcal carriers and noncarriers, respectively)., Conclusions: Associations of pneumococcal carriage detection and density with SARS-CoV-2 suggest a synergistic relationship in the upper airway. Longitudinal studies are needed to determine interaction mechanisms between pneumococci and SARS-CoV-2., Competing Interests: Potential conflicts of interest. J. A. L. discloses receipt of grant funding and consulting honoraria from Pfizer and from Merck, Sharp & Dohme and consulting honoraria from VaxCyte. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2022. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The separation of pneumococcal serotypes from a complex polymicrobial mixture may be required for different applications. For instance, a minority strain could be present at a low frequency in a clinical sample, making it difficult to identify and isolate by traditional culture-based methods. We therefore developed an assay to separate mixed pneumococcal samples using serotype-specific antiserum and a magnetic bead-based separation method. Using qPCR and colony counting methods, we first show that serotypes (12F, 23F, 3, 14, 19A, and 15A) present at ∼0.1% of a dual serotype mixture can be enriched to between 10% and 90% of the final sample. We demonstrate two applications for this method: extraction of known pneumococcal serotypes from saliva samples and efficient purification of capsule switch variants from experimental transformation experiments. This method may have further laboratory or clinical applications when the selection of specific serotypes is required., Competing Interests: D.M.W. has received consulting fees from Pfizer, Merck, GSK, Affinivax, and Matrivax and is PI on research grants from Pfizer and Merck to Yale. A.L.W. has received consulting fees from Pfizer and is PI on research grants from Pfizer to Yale., (© 2023 The Authors.)

While considerable attention was placed on SARS-CoV-2 testing and surveillance programs in the K-12 setting, younger age groups in childcare centers were largely overlooked. Childcare facilities are vital to communities, allowing parents/guardians to remain at work and providing safe environments for both children and staff. Therefore, early in the COVID-19 pandemic (October 2020), we established a PCR-based COVID-19 surveillance program in childcare facilities, testing children and staff with the goal of collecting actionable public health data and aiding communities in the progressive resumption of standard operations and ways of life. In this study we describe the development of a weekly saliva testing program and provide early results from our experience implementing this in childcare centers. We enrolled children (aged 6 months to 7 years) and staff at seven childcare facilities and trained participants in saliva collection using video chat technology. Weekly surveys were sent out to assess exposures, symptoms, and vaccination status changes. Participants submitted weekly saliva samples at school. Samples were transported to a partnering clinical laboratory or RT-PCR testing using SalivaDirect and results were uploaded to each participant's online patient portal within 24 h. SARS-CoV-2 screening and routine testing programs have focused less on the childcare population, resulting in knowledge gaps in this critical age group, especially as many are still ineligible for vaccination. SalivaDirect testing for SARS-CoV-2 provides a feasible method of asymptomatic screening and symptomatic testing for children and childcare center staff. Given the relative aversion to nasal swabs in younger age groups, an at-home saliva collection method provides an attractive alternative, especially as a routine surveillance tool. Results can be shared rapidly electronically through participants' private medical chart portals, and video chat technology allows for discussion and instruction between investigators and participants. This study fosters a cooperative partnership with participating childcare centers, parents/guardians, and staff with the goal of mitigating COVID-19 transmission in childcare centers. Age-related challenges in saliva collection can be overcome by working with parents/guardians to conceptualize new collection strategies and by offering parents/guardians continued virtual guidance and support., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Rayack, Askari, Zirinsky, Lapidus, Sheikha, Peno, Kazemi, Yolda-Carr, Liu, Grubaugh, Ko, Wyllie, Spatz, Oliveira and Bei.)

Nasopharyngeal swabs are considered the gold-standard sample type for the detection of Streptococcus pneumoniae carriage, but recent studies have demonstrated the utility of saliva in improving the detection of carriage in adults. Saliva is generally collected in its raw, unsupplemented state, unlike nasopharyngeal swabs, which are collected into stabilizing transport media. Few data exist regarding the stability of pneumococci in unsupplemented saliva during transport and laboratory storage. We therefore evaluated the effect of storage conditions on the detection of pneumococci in saliva samples using strains representing eight pneumococcal serotypes. The bacteria were spiked into raw saliva from asymptomatic individuals, and we assessed sample viability after storage at 4°C, room temperature, and 30°C for up to 72 h; at 40°C for 24 h; and following three freeze-thaw cycles. We observed little decrease in pneumococcal detection following culture enrichment and quantitative PCR (qPCR) detection of the piaB and lytA genes compared to testing fresh samples, indicating the prolonged viability of pneumococci in neat saliva samples. This sample stability makes saliva a viable sample type for pneumococcal carriage studies conducted in remote or low-resource settings and provides insight into the effect of the storage of saliva samples in the laboratory. IMPORTANCE For pneumococcal carriage studies, saliva is a sample type that can overcome some of the issues typically seen with nasopharyngeal and oropharyngeal swabs. Understanding the limitations of saliva as a sample type is important for maximizing its use. This study sought to better understand how different storage conditions and freeze-thaw cycles affect pneumococcal survival over time. These findings support the use of saliva as an alternative sample type for pneumococcal carriage studies, particularly in remote or low-resource settings with reduced access to health care facilities.

As the coronavirus disease 2019 (COVID-19) continues to disproportionately affect low- and middle-income countries, the need for simple, accessible and frequent diagnostic testing grows. In lower-resource settings, case detection is often limited by a lack of available testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). To address global inequities in testing, alternative sample types could be used to increase access to testing by reducing the associated costs. Saliva is a sensitive, minimally invasive and inexpensive diagnostic sample for SARS-CoV-2 detection that is appropriate for asymptomatic surveillance, symptomatic testing and at-home collection. Saliva testing can lessen two major challenges faced by lower- and middle-income countries: constrained resources and overburdened health workers. Saliva sampling enables convenient self-collection and requires fewer resources than swab-based methods. However, saliva testing for SARS-CoV-2 diagnostics has not been implemented on a large scale in low- and middle-income countries. While numerous studies based in these settings have demonstrated the usefulness of saliva sampling, there has been insufficient attention on optimizing its implementation in practice. We argue that implementation science research is needed to bridge this gap between evidence and practice. Low- and middle-income countries face many barriers as they continue their efforts to provide mass COVID-19 testing in the face of substantial inequities in global access to vaccines. Laboratories should look to replicate successful approaches for sensitive detection of SARS-CoV-2 in saliva, while governments should act to facilitate mass testing by lifting restrictions that limit implementation of saliva-based methods., ((c) 2022 The authors; licensee World Health Organization.)

Background: Infections with respiratory viruses [e.g. influenza and respiratory syncytial virus (RSV)] can increase the risk of severe pneumococcal infections. Likewise, pneumococcal coinfection is associated with poorer outcomes in viral respiratory infection. However, there are limited data describing the frequency of pneumococcus and SARS-CoV-2 coinfection and the role of coinfection in influencing COVID-19 severity. We, therefore, investigated the detection of pneumococcus in COVID-19 inpatients during the early pandemic period., Methods: The study included patients aged 18 years and older, admitted to the Yale-New Haven Hospital who were symptomatic for respiratory infection and tested positive for SARS-CoV-2 during March-August 2020. Patients were tested for pneumococcus through culture-enrichment of saliva followed by RT-qPCR (to identify carriage) and serotype-specific urine antigen detection (UAD) assays (to identify presumed lower respiratory tract pneumococcal disease)., Results: Among 148 subjects, the median age was 65 years; 54.7% were male; 50.7% had an ICU stay; 64.9% received antibiotics; and 14.9% died while admitted. Pneumococcal carriage was detected in 3/96 (3.1%) individuals tested by saliva RT-qPCR. Additionally, pneumococcus was detected in 14/127 (11.0%) individuals tested by UAD, and more commonly in severe than moderate COVID-19 [OR: 2.20; 95% CI: (0.72, 7.48)]; however, the numbers were small with a high degree of uncertainty. None of the UAD-positive individuals died., Conclusions: Pneumococcal lower respiratory tract infection (LRTI), as detected by positive UAD, occurred in patients hospitalized with COVID-19. Moreover, pneumococcal LRTI was more common in those with more serious COVID-19 outcomes. Future studies should assess how pneumococcus and SARS-CoV-2 interact to influence COVID-19 severity in hospitalized patients., (© The Author(s) 2022. Published by Oxford University Press on behalf of FEMS.)

The characteristics of pneumococcal carriage vary between infants and adults. Host immune factors have been shown to contribute to these age-specific differences, but the role of pathogen sequence variation is currently less well-known. Identification of age-associated pathogen genetic factors could leadto improved vaccine formulations. We therefore performed genome sequencing in a large carriage cohort of children and adults and combined this with data from an existing age-stratified carriage study. We compiled a dictionary of pathogen genetic variation, including serotype, strain, sequence elements, single-nucleotide polymorphisms (SNPs), and clusters of orthologous genes (COGs) for each cohort - all of which were used in a genome-wide association with host age. Age-dependent colonization showed weak evidence of being heritable in the first cohort ( h 2 = 0.10, 95% CI 0.00-0.69) and stronger evidence in the second cohort ( h 2 = 0.56, 95% CI 0.23-0.87). We found that serotypes and genetic background (strain) explained a proportion of the heritability in the first cohort ( h 2 serotype = 0.07, 95% CI 0.04-0.14 and h 2 GPSC = 0.06, 95% CI 0.03-0.13) and the second cohort ( h 2 serotype = 0.11, 95% CI 0.05-0.21 and h 2 GPSC = 0.20, 95% CI 0.12-0.31). In a meta-analysis of these cohorts, we found one candidate association (p=1.2 × 10 -9 ) upstream of an accessory Sec-dependent serine-rich glycoprotein adhesin. Overall, while we did find a small effect of pathogen genome variation on pneumococcal carriage between child and adult hosts, this was variable between populations and does not appear to be caused by strong effects of individual genes. This supports proposals for adaptive future vaccination strategies that are primarily targeted at dominant circulating serotypes and tailored to the composition of the pathogen populations., Competing Interests: PK, BF, HB, NR, AW, ES, KT, AW, PT, Av, MB, SB, Dv, JL No competing interests declared, (© 2022, Kremer et al.)

Introduction: Symptomatic testing and asymptomatic screening for SARS-CoV-2 continue to be essential tools for mitigating virus transmission. Though COVID-19 diagnostics initially defaulted to oropharyngeal or nasopharyngeal sampling, the worldwide urgency to expand testing efforts spurred innovative approaches and increased diversity of detection methods. Strengthening innovation and facilitating widespread testing remains critical for global health, especially as additional variants emerge and other mitigation strategies are recalibrated., Areas Covered: A growing body of evidence reflects the need to expand testing efforts and further investigate the efficiency, sensitivity, and acceptability of saliva samples for SARS-CoV-2 detection. Countries have made pandemic response decisions based on resources, costs, procedures, and regional acceptability - the adoption and integration of saliva-based testing among them. Saliva has demonstrated high sensitivity and specificity while being less invasive relative to nasopharyngeal swabs, securing saliva's position as a more acceptable sample type., Expert Opinion: Despite the accessibility and utility of saliva sampling, global implementation remains low compared to swab-based approaches. In some cases, countries have validated saliva-based methods but face challenges with testing implementation or expansion. Here, we review the localities that have demonstrated success with saliva-based SARS-CoV-2 testing approaches and can serve as models for transforming concepts into globally-implemented best practices.

Background: There is an urgent need to expand testing for SARS-CoV-2 and other respiratory pathogens as the global community struggles to control the COVID-19 pandemic. Current diagnostic methods can be affected by supply chain bottlenecks and require the assistance of medical professionals, impeding the implementation of large-scale testing. Self-collection of saliva may solve these problems, as it can be completed without specialized training and uses generic materials., Methods: We observed 30 individuals who self-collected saliva using four different collection devices and analyzed their feedback. Two of these devices, a funnel and bulb pipette, were used to evaluate at-home saliva collection by 60 individuals. SARS-CoV-2-spiked saliva samples were subjected to temperature cycles designed to simulate the conditions the samples might be exposed to during the summer and winter seasons and sensitivity of detection was evaluated., Results: All devices enabled the safe, unsupervised self-collection of saliva. The quantity and quality of the samples received were acceptable for SARS-CoV-2 diagnostic testing, as determined by human RNase P detection. There was no significant difference in SARS-CoV-2 nucleocapsid gene (N1) detection between the freshly spiked samples and those incubated with the summer and winter profiles., Conclusion: We demonstrate inexpensive, generic, buffer free collection devices suitable for unsupervised and home saliva self-collection., (© 2022. The Author(s).)

Background: The underlying immunologic deficiencies enabling severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reinfection are currently unknown. We describe deep longitudinal immune profiling of a transplant recipient hospitalized twice for coronavirus disease 2019 (COVID-19)., Methods: A 66-year-old male renal transplant recipient was hospitalized with COVID-19 March 2020 then readmitted to the hospital with COVID-19 233 days after initial diagnosis. Virologic and immunologic investigations were performed on samples from the primary and secondary infections., Results: Whole viral genome sequencing and phylogenetic analysis revealed that viruses causing both infections were caused by distinct genetic lineages without evidence of immune escape mutations. Longitudinal comparison of cellular and humoral responses during primary SARS-CoV-2 infection revealed that this patient responded to the primary infection with low neutralization titer anti-SARS-CoV-2 antibodies that were likely present at the time of reinfection., Conclusions: The development of neutralizing antibodies and humoral memory responses in this patient failed to confer protection against reinfection, suggesting that they were below a neutralizing titer threshold or that additional factors may be required for efficient prevention of SARS-CoV-2 reinfection. Development of poorly neutralizing antibodies may have been due to profound and relatively specific reduction in naive CD4 T-cell pools. Seropositivity alone may not be a perfect correlate of protection in immunocompromised patients., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Dysregulated immune responses against the SARS-CoV-2 virus are instrumental in severe COVID-19. However, the immune signatures associated with immunopathology are poorly understood. Here we use multi-omics single-cell analysis to probe the dynamic immune responses in hospitalized patients with stable or progressive course of COVID-19, explore V(D)J repertoires, and assess the cellular effects of tocilizumab. Coordinated profiling of gene expression and cell lineage protein markers shows that S100A hi /HLA-DR lo classical monocytes and activated LAG-3 hi T cells are hallmarks of progressive disease and highlights the abnormal MHC-II/LAG-3 interaction on myeloid and T cells, respectively. We also find skewed T cell receptor repertories in expanded effector CD8 + clones, unmutated IGHG + B cell clones, and mutated B cell clones with stable somatic hypermutation frequency over time. In conclusion, our in-depth immune profiling reveals dyssynchrony of the innate and adaptive immune interaction in progressive COVID-19., (© 2022. The Author(s).)

The COVID-19 pandemic has highlighted the need and benefits for all communities to be permitted timely access to on-demand screening for infectious respiratory diseases. This can be achieved with simplified testing approaches and affordable access to core resources. While RT-qPCR-based tests remain the gold standard for SARS-CoV-2 detection due to their high sensitivity, implementation of testing requires high upfront costs to obtain the necessary instrumentation. This is particularly restrictive in low-resource settings. The Ubiquitome Liberty16 system was developed as an inexpensive, portable, battery-operated single-channel RT-qPCR device with an associated iPhone app to simplify assay set-up and data reporting. When coupled with the SalivaDirect protocol for testing saliva samples for SARS-CoV-2, the Liberty16 device yielded a limit of detection (LOD) of 12 SARS-CoV-2 RNA copies/µL, comparable to the upper end of the LOD range for the standard SalivaDirect protocol when performed on larger RT-qPCR instruments. While further optimization may deliver even greater sensitivity and assay speed, findings from this study indicate that small portable devices such as the Liberty16 can deliver reliable results and provide the opportunity to further increase access to gold standard SARS-CoV-2 testing., Competing Interests: ST was an employee of Ubiquitome Limited at the time of this study and PP is the CEO of Ubiquitome Limited. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yolda-Carr, Thammavongsa, Vega, Turner, Pickering and Wyllie.)

Genomic sequencing is crucial to understanding the epidemiology and evolution of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Often, genomic studies rely on remnant diagnostic material, typically nasopharyngeal (NP) swabs, as input into whole-genome SARS-CoV-2 next-generation sequencing pipelines. Saliva has proven to be a safe and stable specimen for the detection of SARS-CoV-2 RNA via traditional diagnostic assays; however, saliva is not commonly used for SARS-CoV-2 sequencing. Using the ARTIC Network amplicon-generation approach with sequencing on the Oxford Nanopore MinION, we demonstrate that sequencing SARS-CoV-2 from saliva produces genomes comparable to those from NP swabs, and that RNA extraction is necessary to generate complete genomes from saliva. In this study, we show that saliva is a useful specimen type for genomic studies of SARS-CoV-2., (© The Author(s) 2022. Published by Oxford University Press.)

Pooled testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection is instrumental for increasing test capacity while decreasing test cost. Pooled testing programs permit sustainable, long-term surveillance measures, which are essential for the early detection of virus resurgence in communities or the emergence of variants of concern. While numerous pooled approaches have been proposed to increase test capacity, uptake by laboratories has been limited. On 9 December 2020, we invited 362 U.S. laboratories that inquired about the Yale School of Public Health SalivaDirect test to participate in a survey to evaluate testing constraints and pooling strategies for SARS-CoV-2 testing. The survey was distributed using Qualtrics, and three reminders were sent. The survey closed on 21 January 2021. Of 93 responses received (25.7% response rate), 90 were from Clinical Laboratory Improvement Amendments (CLIA)-certified laboratories conducting SARS-CoV-2 testing. The remaining three were excluded from the analyses. Responses indicated that the major barriers to the uptake of pooled testing in the United States may not simply be the number of tests a laboratory can process per day, but rather the lack of clear protocols and adequate resources; laboratories are working with fixed physical and human capital constraints. Importantly, laboratories across the country are heterogeneous in infrastructure and workflow. The need for SARS-CoV-2 testing will remain for years to come. Testing programs can be maintained through pooled PCR testing strategies, and while statisticians, operations researchers, and others with expertise in sampling design have important value to add, laboratories require support on how to transition from traditional diagnostic testing to pooled surveillance. IMPORTANCE While numerous pooled SARS-CoV-2 testing approaches have been described in an effort to increase testing capacity and decrease test prices, uptake by laboratories has been limited. Responses to our survey of United States-based laboratories highlight the importance of consulting end-users-those that solutions are being designed for-so challenges can be addressed in a manner tailored to meet the specific needs out in the field. It may be surprising to those designing pooled testing strategies to learn that laboratories view pooling as more time-consuming than testing samples individually, and therefore that it is thought to create delays in test reporting.

As the second year of the COVID-19 pandemic begins, it remains clear that a massive increase in the ability to test for SARS-CoV-2 infections in a myriad of settings is critical to controlling the pandemic and to preparing for future outbreaks. The current gold standard for molecular diagnostics is the polymerase chain reaction (PCR), but the extraordinary and unmet demand for testing in a variety of environments means that both complementary and supplementary testing solutions are still needed. This review highlights the role that loop-mediated isothermal amplification (LAMP) has had in filling this global testing need, providing a faster and easier means of testing, and what it can do for future applications, pathogens, and the preparation for future outbreaks. This review describes the current state of the art for research of LAMP-based SARS-CoV-2 testing, as well as its implications for other pathogens and testing. The authors represent the global LAMP (gLAMP) Consortium, an international research collective, which has regularly met to share their experiences on LAMP deployment and best practices; sections are devoted to all aspects of LAMP testing, including preanalytic sample processing, target amplification, and amplicon detection, then the hardware and software required for deployment are discussed, and finally, a summary of the current regulatory landscape is provided. Included as well are a series of first-person accounts of LAMP method development and deployment. The final discussion section provides the reader with a distillation of the most validated testing methods and their paths to implementation. This review also aims to provide practical information and insight for a range of audiences: for a research audience, to help accelerate research through sharing of best practices; for an implementation audience, to help get testing up and running quickly; and for a public health, clinical, and policy audience, to help convey the breadth of the effect that LAMP methods have to offer., (© 2021 ABRF.)

Background: The negative impact of continued school closures during the height of the COVID-19 pandemic warrants the establishment of cost-effective strategies for surveillance and screening to safely reopen and monitor for potential in-school transmission. Here, we present a novel approach to increase the availability of repetitive and routine COVID-19 testing that may ultimately reduce the overall viral burden in the community., Methods: We implemented a testing program using the SalivaClear࣪ pooled surveillance method that included students, faculty and staff from K-12 schools (student age range 5-18 years) and universities (student age range >18 years) across the country (Mirimus Clinical Labs, Brooklyn, NY). The data analysis was performed using descriptive statistics, kappa agreement, and outlier detection analysis., Findings: From August 27, 2020 until January 13, 2021, 253,406 saliva specimens were self-collected from students, faculty and staff from 93 K-12 schools and 18 universities. Pool sizes of up to 24 samples were tested over a 20-week period. Pooled testing did not significantly alter the sensitivity of the molecular assay in terms of both qualitative (100% detection rate on both pooled and individual samples) and quantitative (comparable cycle threshold (Ct) values between pooled and individual samples) measures. The detection of SARS-CoV-2 in saliva was comparable to the nasopharyngeal swab. Pooling samples substantially reduced the costs associated with PCR testing and allowed schools to rapidly assess transmission and adjust prevention protocols as necessary. In one instance, in-school transmission of the virus was determined within the main office and led to review and revision of heating, ventilating and air-conditioning systems., Interpretation: By establishing low-cost, weekly testing of students and faculty, pooled saliva analysis for the presence of SARS-CoV-2 enabled schools to determine whether transmission had occurred, make data-driven decisions, and adjust safety protocols. We provide strong evidence that pooled testing may be a fundamental component to the reopening of schools by minimizing the risk of in-school transmission among students and faculty., Funding: Skoll Foundation generously provided funding to Mobilizing Foundation and Mirimus for these studies., Competing Interests: None., (© 2021 The Author(s).)

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Recent studies have provided insights into innate and adaptive immune dynamics in coronavirus disease 2019 (COVID-19). However, the exact features of antibody responses that govern COVID-19 disease outcomes remain unclear. In this study, we analyzed humoral immune responses in 229 patients with asymptomatic, mild, moderate and severe COVID-19 over time to probe the nature of antibody responses in disease severity and mortality. We observed a correlation between anti-spike (S) immunoglobulin G (IgG) levels, length of hospitalization and clinical parameters associated with worse clinical progression. Although high anti-S IgG levels correlated with worse disease severity, such correlation was time dependent. Deceased patients did not have higher overall humoral response than discharged patients. However, they mounted a robust, yet delayed, response, measured by anti-S, anti-receptor-binding domain IgG and neutralizing antibody (NAb) levels compared to survivors. Delayed seroconversion kinetics correlated with impaired viral control in deceased patients. Finally, although sera from 85% of patients displayed some neutralization capacity during their disease course, NAb generation before 14 d of disease onset emerged as a key factor for recovery. These data indicate that COVID-19 mortality does not correlate with the cross-sectional antiviral antibody levels per se but, rather, with the delayed kinetics of NAb production., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)

COVID-19 manifests with a wide spectrum of clinical phenotypes that are characterized by exaggerated and misdirected host immune responses 1-6 . Although pathological innate immune activation is well-documented in severe disease 1 , the effect of autoantibodies on disease progression is less well-defined. Here we use a high-throughput autoantibody discovery technique known as rapid extracellular antigen profiling 7 to screen a cohort of 194 individuals infected with SARS-CoV-2, comprising 172 patients with COVID-19 and 22 healthcare workers with mild disease or asymptomatic infection, for autoantibodies against 2,770 extracellular and secreted proteins (members of the exoproteome). We found that patients with COVID-19 exhibit marked increases in autoantibody reactivities as compared to uninfected individuals, and show a high prevalence of autoantibodies against immunomodulatory proteins (including cytokines, chemokines, complement components and cell-surface proteins). We established that these autoantibodies perturb immune function and impair virological control by inhibiting immunoreceptor signalling and by altering peripheral immune cell composition, and found that mouse surrogates of these autoantibodies increase disease severity in a mouse model of SARS-CoV-2 infection. Our analysis of autoantibodies against tissue-associated antigens revealed associations with specific clinical characteristics. Our findings suggest a pathological role for exoproteome-directed autoantibodies in COVID-19, with diverse effects on immune functionality and associations with clinical outcomes.

Background: The importance of specific serotypes causing invasive pneumococcal disease (IPD) differs by age. Data on pneumococcal carriage in different age groups, along with data on serotype-specific invasiveness, could help explain these age-related patterns and their implications for vaccination., Methods: Using pneumococcal carriage and disease data from Israel, we evaluated the association between serotype-specific IPD in adults and serotype-specific carriage prevalence among children in different age categories, while adjusting for serotype-specific invasiveness. We estimated carriage prevalence using different age groupings that were selected a priori. The Deviance Information Criterion was used to determine which age groupings of carriage data best fit the adult IPD data. Serotype-specific disease patterns were further evaluated by stratifying IPD data by comorbidity status., Results: The relative frequency of serotypes causing IPD differed between adults and children, and also differed between older and younger adults and between adults with and without comorbidities. Serotypes overrepresented as causes of IPD in adults were more commonly carried in older children compared with younger children. In line with this, the serotype-specific frequency of carriage in older children, rather than infants, best correlated with serotype-specific IPD in adults., Conclusions: These analyses demonstrate that the serotype patterns in carriage in older children, rather than infants, are best correlated with disease patterns in adults. This might suggest these older children are more influential for disease patterns in adults. These insights could help in optimizing vaccination strategies to reduce disease burden across all ages., (© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)