Host transcriptional profiling identifies B cell associated genes to be upregulated in individuals with asymptomatic COVID-19 and latent tuberculosis

Host transcriptional profiling identifies B cell associated genes to be upregulated in individuals with asymptomatic COVID-19 and latent tuberculosis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Short Report Host transcriptional profiling identifies B cell associated genes to be upregulated in individuals with asymptomatic COVID-19 and latent tuberculosis Fizza Fatima, Javaria Ashraf, Kiran Masood, Maliha Yameen, Rabia Hussain, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8500902/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background SARS-CoV-2 infections resulted in a global pandemic with variable rates of COVID-19 morbidity worldwide. In high tuberculosis (TB) endemic regions it was observed that COVID-19 associated mortality was lower. Infection with Mycobacterium tuberculosis (MTB) has been shown to impact COVID-19 infections but the mechanisms involved are as yet unclear. Here we investigated the effect of latent MTB infection (LTBI) by studying host blood transcriptional profiles in individuals with Asymptomatic COVID-19. Methods We studied participants who had asymptomatic COVID-19 compared with those who had LTBI as well COVID (LTBI-COVID). Participants were aged 35.5 (+-9.2) years and comprised 27% females. Whole blood transcriptional profiles were investigated using RNA microarray analysis of individuals. Results Comparison of the LTBI-COVID and COVID groups revealed 58 differentially expressed genes (DEGs) of which 44 were upregulated and 13 were downregulated. Cluster profiler analysis showed upregulation of innate immunity associated genes (CLEC12A, TRIM15, FCRL3) upregulated in LTBI-COVID. A GO enrich analysis showed upregulation of B cell activation associated with (IL-7, SHLD1, TFRC, and FCRL3). There was a downregulation of RNF5 associated with the STING pathway. Conclusions B cell activation would lead to increased humoral immunity and enhanced uptake of pathogens by macrophages. CLEC12A and RNF5 are associated with dampening of excessive inflammation which reduces risks of COVID-19. These data support the protective role of latent TB infection associated with reduced severity of COVID-19. Latent TB SARS-CoV-2 B cell innate immunity inflammation Figures Figure 1 Introduction The COVID-19 pandemic resulted in 700 million cases and nearly 7 million deaths worldwide between 2019 and 2024 ( 1 ). The severity of SARS-CoV-2 infections ranged from asymptomatic and mild infections, moderate to severe and critical disease as well as post-acute sequelae of infections (PACS) ( 2 ). There were differences observed in global mortality from COVID-19 particularly evident in the early pandemic period 2020–2021 prior to the introduction of vaccinations (S Fig. 1 ). COVID-19 associated death rates in this early pandemic period were; approximately 1.2 million from USA, 200,000 from the UK and 31,000 from Pakistan, respectively. Of note, Pakistan with a high tuberculosis (TB) burden was amongst the South Asian countries that suffered relatively less from COVID-19 ( 3 ). Pakistan delivers ongoing BCG vaccination through the Extended Program of Immunization (EPI) however it ranks 5th amongst high TB burden countries with an incidence of 341/100,000 annually. Both innate and adaptive immunity are required for protection against SARS-CoV-2. Innate immune activation is essential for viral clearance and type I interferon driven responses protect against severe disease ( 4 ). Humoral immunity and effect of antibodies through their virus neutralizing activity as well as T cell activation is required for protection against SARS-CoV-2 and COVID-19 ( 5 ). Factors associated with protection against COVID-19 have been younger age, cross-reactive serological protection from human coronaviruses as well as BCG vaccinations ( 6 – 9 ). Mycobacterium tuberculosis (MTB) infection remains the world’s leading infectious causes of death with over 1 million deaths each year ( 10 ). Most infected people remain asymptomatic with 5–10% thought to develop latent tuberculosis (LTBI) ( 11 ). LTBI has been associated with less severe SARS-CoV-2 infections ( 12 ). Here we investigated host blood transcriptional profiles in those with asymptomatic COVID-19 to investigate the immune mechanisms associated with LTBI that lead to less severe disease. Our study participants were unvaccinated individuals recruited in the early pandemic period between March and October 2020, when SARS-CoV-2 S, L and G clade strains were in circulation ( 13 ). Our results show an upregulation of B cell pathway and CLEC12A associated genes in the LTBI-COVID group. This suggests that LTBI may afford protection by upregulating humoral immunity and dampening of excessive inflammation in those with latent TB. Methods Study Description This study received approval from the Ethics Review Committee of the Aga Khan University (AKU) with the project number RRG 236. All methods were performed in accordance with the relevant guidelines and regulations. All the participants were adults (> 18 years of age) and written informed consent was obtained from each participant. COVID-19 cases were classified according to their disease severity fined by the WHO ordinal score ( 14 ) and those with scores 1–2 as Asymptomatic cases. These individuals were either completely asymptomatic or had a short (1–2 days) history of fever, sore throat or myalgia which resolved without medical treatment. Respiratory swabs were tested for SARS-CoV-2 using the COBAS SARS-CoV-2 assay (COBAS 6800 Roche platform) at the AKU Hospital Clinical laboratories. All study participants had PCR-confirmed COVID-19 and data collected included information regarding clinical history, co-morbid conditions. None had received COVID-19 vaccinations. A single blood sample was taken from study participants within 2–3 days of a positive PCR test. All study participants underwent screening for LTBI using the QuantiFERON-TB Gold (QFT) Interferon-Gamma Release Assay (IGRA) which employs TB1 and TB2 antigens based on ESAT6 and CFP10 peptides. IGRA results were interpreted as per the manufacturer’s instructions (Cellestus, USA). RNA microarray data RNA was extracted from whole blood using the Qiagen RNA Blood Mini Kit (Qiagen, GmbH, Germany). One hundred nanogram of RNA was used for the preparation of cRNA. The samples were hybridized to the Clariom S Assay human microarray(Affymetrix, USA), containing probes for 21,488 genes. Functional enrichment analysis CEL files were analysed using the TAC Transcriptome Analysis Software Suite (TACS version 2) using the Summarization Method: Gene Level - SST-RMA Pos vs Neg AUC Threshold: 0.7 against Genome Version: hg38 (Homo sapiens). DEGs from groups LTBI-COVID and COVID were analyzed. Cellular pathway analysis of significant differentially expressed genes (DEGs) up- or down-regulated (log FC (fold change) 2; p value < 0.05) were identified by TACS. Significantly modified pathways were sub-grouped. Analyses were performed using R (RStudio) with the ClusterProfiler and gProfiler packages. Functional analysis of the DEGs was carried out by employing the enricher function within the Cluster Profiler package. GO enrichment for biological pathways was applied onto the data resulting in a dotplot along with a corresponding heatmap. Additional exploration of gene-level expression was carried out using the gProfiler package which enabled the formation of a gene-membership heatmap illustrating the genes contributing to each pathway. Data Availability Raw array output (CEL) and processed (CHP) files were submitted to Gene Expression Omnibus (GEO) NCBI, where accession numbers were assigned and are listed in the supplementary table. All data generated or analysed during this study are included with this published article (S Table 1). Data for Quality Metrics and sample signals of probes are available in S Tables 2 and 3. Results Description of study subjects Asymptomatic COVID-19 patients were recruited between March and October 2020. Study participants comprised SARS-CoV-2 positive unvaccinated individuals recruited in our previous study conducted in the early pandemic period before COVID-19 ( 4 ). The age range of participants was 19–49 years (Mean SD 35.5, ± 9.2 years). There were 6 females and 16 males. IGRA testing identified two of 22 male participants to have LTBI. Investigating the host blood transcriptomes in COVID-19 cases with and without LTBI We compared host blood transcriptome profiles of study participants in LTBI-COVID and COVID groups. Unhierarchical Principal Component Analysis (PCA) of transcriptional signals showed some separation between different samples but no difference between the samples with LTBI-COVID and cases of COVID (Fig. 1 A). Fifty-eight differentially expressed genes (DEGs) were observed with 44 upregulated (75.8%) and 13 downregulated genes (Fig. 1 B). A volcano plot generated to visualization of DEGs depicted that CLEC12A (inhibitory C-type lectin-like receptor on innate immune cells) showed the greatest upregulation followed by FCRL3 (immunoregulatory receptor that modulates innate B cell signaling). A subset of genes demonstrated notable downregulation, including ARMCX6 (part of ARMC subfamily; active in the mitochondria), ATG13 (a protein involved in the induction of type I interferon), TMEM126A (role in TLR4-mediated immune response), STX10 (part of SNARE complex), and FAU (regulates T cell function and survival). GO enrich analysis of biological pathways analysis showed that upregulation of genes was associated with B cell activation (IL-7, SHLD1, TFRC, and FCRL3) LTBI-COVID (Fig. 1 C). The accompanying heatmap identifies; IL-7 (essential for lymphocyte development), SHLD1 (crucial for antibody class switching) and FCRL3 (immunoregulatory receptor that impedes the activation of human memory T lymphocytes). Downregulation of RNF5 associated with the STING pathway was also observed in LTBI-COVID. Cluster profiler analysis showed upregulation of predominantly innate immunity genes (CLEC12A, TRIM15, FCRL3) associated with LTBI-COVID. Its associated heatmap shows three functional pathway categories where CLEC12A showed the highest level of upregulation (Fig. 1 D). These were enzyme binding and organelle membrane associated pathways. Additional DEGs were associated with B and T lymphocyte differentiation and maturation, regulation or signaling (OSTM1, ART1, LMBR1L, FCRL3 and MGAT2 were evident. Also, innate immune response genes; GCC2 (NK cell-mediated immunity), APAF1 (regulation of apoptosis and inflammatory responses) and ATP1B1 (antiviral innate immune response). A further description of DEGs involved in B and T cell or innate immunity are provided in S Table 2. This identifies upregulation of genes involved in T cell receptor function. Additionally, we there was upregulation of immune response genes related to vesicular trafficking, endocytosis, golgi apparatus and cellular signaling including, the STING (stimulator of interferon genes) pathway. Discussion Our study provides important insights into how MTB infection may affect the host immune response to SARS-CoV-2. Through host blood transcriptional profiling we observed upregulation of genes associated with B memory cells in those with asymptomatic COVID-19 as well as LTBI. Importantly, upregulation of CLEC12A associated with downmodulation of inflammatory responses was observed in the LTBI-COVID-19 group. These data fit with our recent work showing reduced rates of LTBI amongst those with mild COVID-19, suggesting a protective effect of MTB ( 12 ). Earlier, we have observed higher type I interferon responses in those with asymptomatic COVID-19 ( 4 ). Here, by focusing studying the same asymptomatic COVID-19 cohort we observed that in those with LTBI, there was the upregulation of genes associated with B cell activation. Specifically, IL-7 plays an important role in B cell development ( 15 ). FCRL3 modulates lymphocyte activity. These data are important as humoral immunity plays an important role in protection against SARS-CoV-2 infection ( 5 ). In Pakistan, we observed high rising titers of seropositivity to SARS-CoV-2 in unvaccinated individuals in the prepandemic period ( 16 ). Further, serosurveys in the population revealed increasing antibody titers associated with SARS-CoV-2 in asymptomatic individuals ( 17 ). Our study fits with reports by Rajamanickam et al that LTBI-positive individuals exhibit elevated levels of humoral, cytokine, and systemic immune responses ( 18 ). Our work is also supported by studies showing raised cytokine responses to Spike in IGRA positive COVID-19 cases ( 19 ). Trained immunity plays a role in responses of a range of cell types from B and T to macrophages and Natural killer cells ( 25 , 26 ). Recall responses to BCG were found present in adults from the prepandemic period whilst their plasma had cross-reactivity to SARS-CoV-2 RBD protein, identifying the recognition of pathogen motifs by BCG-induced antibodies ( 27 ). BCG has been shown to control mortality in children due to respiratory tract infections ( 28 ) and appropriate innate B cell gene activation may be one such mechanism.B cells primed with cross reactive antigens have been shown to elicit stronger anti-viral responses against LCMV infection ( 29 ). Recently a subset of long-term memory IgM switched IgG + B cells has been identified to play an important role in in antiviral immunity with the characteristic of long-term epigenetic changes ( 30 ). Hence, it is likely that B cell expansion to SARS-CoV-2 antigens is facilitated in a BCG vaccination population such as ours. COVID-19 outcomes have been associated with the severity of the initial inflammatory response to infection, with acute outcomes associated with a heightened inflammasome response. We found CLEC12A/B to be regulated. These C-type lectin- receptors play an important role in the phagocytosis of mycobacteria and in facilitating their trafficking to the phagolysosomal pathway ( 20 , 21 ). This is supported by the pathway analysis which identifies genes linked with organelle and membrane binding to be differentially regulated in the COVID-19 sub-groups. Importantly, CLEC12A contains an inhibitory motif that binds to mycobacterial mycolic acids, and can sense DAMPs which lead to the suppression of immune responses ( 22 ). Interestingly, we found raised TRIM15, an interferon stimulated genes (ISG) but downregulation of RNF5 which is an activator of the STING pathway that is linked to inflammasome responses ( 23 ). Hence, this data suggests that mycobacteria may dampen host immune responses reducing excess inflammatory response to SARS-CoV-2, associated with unfavourable outcomes ( 24 ). Our study had limitations that was a relatively small study and we did not have any laboratory data such as hematological parameters for the study cohort. However, we know that the study participants all had asymptomatic COVID-19 with no lasting symptoms. We only had two individuals in the LTBI-COVID-19 group. This may be due to the lower frequency of LTBI in those with mild COVID-19 observed earlier ( 31 ). We had mostly males in this group as this testing was conducted in the first cohort of healthcare workers who had returned to on campus work after the first lockdown in Pakistan. Importantly, the individuals were relatively young, all aged less than 50 years old and therefore there should be no confounding effects of an older age group present here. Overall, our data fits with upregulation of a protective response in those with MTB infection in the context of SARS-CoV-2 infections ( 32 ). BCG vaccination. These may explain the reduced disease severity from COVID-19 observed in unvaccinated individuals in the early pandemic period in high TB burden countries such as Pakistan. Declarations Ethics approval and consent to participate Ethics approval was granted by the Ethics Review Committee of the Aga Khan University. Informed consent was taken from all study participants. Consent for publication Consent for publication is given by all authors. Availability of data and materials All data after available with the submitted manuscript and are publicly available. Competing interests The authors declare that they have no conflicts of interest. Funding The study was supported by a COVID-19 Rapid Research Grant (RRG-236) and GCF-913 grand awarded by the Higher Education Commission, Pakistan. Fizza Fatima was supported by the Gulamali Hirji endowed Professorship fund. Authors' contributions ZH, designed the study; KIM, MY: implementation and execution; FA, MY, JA: data analysis; ZH, FA, RH: drafted the initial manuscript; KIM, MY, JA: manuscript revision with critical input. All authors reviewed and approved the manuscript. Acknowledgements We thank the Department of Pathology and Laboratory Medicine, Clinical Laboratories, Aga Khan University, Pakistan for facilitating the recruitment of COVID-19 patients for the study. References JHU. COVID-19 Data Repository by the Center for Systems Science and Engineering (CSSE) at Johns Hopkins University: John Hopkins University; 2021 [Available from: https://github.com/CSSEGISandData/COVID-19 . Rubio-Rivas M, Mora-Luján JM, Formiga F, Arévalo-Cañas C, Lebrón Ramos JM, Villalba García MV, et al. WHO ordinal scale and inflammation risk categories in COVID-19. Comparative study of the severity scales. 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Supplementary Files FigS1.docx STable12v2.docx STable2.xlsx STable3.xlsx STable4.docx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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07:46:02","extension":"xml","order_by":11,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":70127,"visible":true,"origin":"","legend":"","description":"","filename":"fea3f64e1da044d2b8e4f04bcaab37c51structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/9e2f33ffc4a7d67bfa65c37f.xml"},{"id":100010505,"identity":"022f9fed-d531-4835-a660-ff1ced68b7b7","added_by":"auto","created_at":"2026-01-12 06:06:39","extension":"html","order_by":12,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":80685,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/c3eb29d9c0f3522e936a1b22.html"},{"id":100010487,"identity":"3de67199-cdcd-41fb-b258-0f50eefad156","added_by":"auto","created_at":"2026-01-12 06:06:38","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":114356,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHost transcriptional profiles of COVID-19 cases with and without LTBI.\u003c/strong\u003e Twenty-two individuals with Asymptomatic COVID-19 were studied and transcriptional profiles of LTBI-COVID as compared with COVID cases were compared.\u003c/p\u003e\n\u003cp\u003eA. Show Principal Component Analysis (PCA) of the dataset with each individual represented by a circle (red, LTBI-COVID and red, COVID).\u003c/p\u003e\n\u003cp\u003eB. The \u0026nbsp;\u0026nbsp;\u0026nbsp;\u0026nbsp;\u0026nbsp;volcano plot represents the differentially expressed genes (DEGs) between groups with 44 upregulated (red dots) and 13 downregulated (green dots) \u0026nbsp;\u0026nbsp;\u0026nbsp;\u0026nbsp;\u0026nbsp;genes. Details of DEGs are seen in the inset plot.\u003c/p\u003e\n\u003cp\u003eC. Dotplot of enriched gene ontology (GO) pathway analysis applied on DEGs. Upper panel; Gene ratio is displayed on the X-axis while the pathway details are mentioned on the Y-axis. The color and size of the dot correspond to the p-adjusted value and the number of genes involved. The figure shows only one active biological pathway. Lower panel: Heatmap of biological pathways generated to depict the gene expression data. Pathway details are stated on the Y-axis while the genes involved are displayed on the X-axis. The color scale on the right indicates the FC for each gene.\u003c/p\u003e\n\u003cp\u003eD. The heatmap DEGs as analysed through GO pathways. Pathway details are mentioned on the Y-axis while the genes contributing to them are mentioned on the X-axis.\u003c/p\u003e","description":"","filename":"Fig1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/bca81f171feaf3a90d5d6c33.jpg"},{"id":107868302,"identity":"1fed2f7f-1d0b-4583-a022-487a217f1ae7","added_by":"auto","created_at":"2026-04-27 07:09:54","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":296194,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/71ef78c4-e668-4978-9ccc-8d7b35687437.pdf"},{"id":100361204,"identity":"89c4c1c4-cb7d-4ba1-b435-48ef3664d387","added_by":"auto","created_at":"2026-01-16 07:44:39","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":226355,"visible":true,"origin":"","legend":"","description":"","filename":"FigS1.docx","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/98daa51211a18051f7d22c6b.docx"},{"id":100010489,"identity":"1fb7b0bf-dbc8-4cf7-96c5-294b59bdb87c","added_by":"auto","created_at":"2026-01-12 06:06:38","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":18274,"visible":true,"origin":"","legend":"","description":"","filename":"STable12v2.docx","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/6cde05a2362c0955aaceb1a4.docx"},{"id":100361216,"identity":"062cdc4b-5ccf-44a8-8fc0-d141149dda79","added_by":"auto","created_at":"2026-01-16 07:44:40","extension":"xlsx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":30673,"visible":true,"origin":"","legend":"","description":"","filename":"STable2.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/816f8d1737f1c4785988d47f.xlsx"},{"id":100361511,"identity":"3a2cf043-644e-4cb7-8118-7da4908b2d24","added_by":"auto","created_at":"2026-01-16 07:45:14","extension":"xlsx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":21428,"visible":true,"origin":"","legend":"","description":"","filename":"STable3.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/ac6871b7c5f5c5df46ec8f26.xlsx"},{"id":100010494,"identity":"1aa01035-c7bc-4c6e-a00a-88229a40b5ac","added_by":"auto","created_at":"2026-01-12 06:06:38","extension":"docx","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":19982,"visible":true,"origin":"","legend":"","description":"","filename":"STable4.docx","url":"https://assets-eu.researchsquare.com/files/rs-8500902/v1/6b01deeeb1c6c3124d68b5cb.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Host transcriptional profiling identifies B cell associated genes to be upregulated in individuals with asymptomatic COVID-19 and latent tuberculosis","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe COVID-19 pandemic resulted in 700\u0026nbsp;million cases and nearly 7\u0026nbsp;million deaths worldwide between 2019 and 2024 (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). The severity of SARS-CoV-2 infections ranged from asymptomatic and mild infections, moderate to severe and critical disease as well as post-acute sequelae of infections (PACS) (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e). There were differences observed in global mortality from COVID-19 particularly evident in the early pandemic period 2020\u0026ndash;2021 prior to the introduction of vaccinations (S Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). COVID-19 associated death rates in this early pandemic period were; approximately 1.2\u0026nbsp;million from USA, 200,000 from the UK and 31,000 from Pakistan, respectively. Of note, Pakistan with a high tuberculosis (TB) burden was amongst the South Asian countries that suffered relatively less from COVID-19 (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). Pakistan delivers ongoing BCG vaccination through the Extended Program of Immunization (EPI) however it ranks 5th amongst high TB burden countries with an incidence of 341/100,000 annually.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eBoth innate and adaptive immunity are required for protection against SARS-CoV-2. Innate immune activation is essential for viral clearance and type I interferon driven responses protect against severe disease (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). Humoral immunity and effect of antibodies through their virus neutralizing activity as well as T cell activation is required for protection against SARS-CoV-2 and COVID-19 (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). Factors associated with protection against COVID-19 have been younger age, cross-reactive serological protection from human coronaviruses as well as BCG vaccinations (\u003cspan additionalcitationids=\"CR7 CR8\" citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cem\u003eMycobacterium tuberculosis\u003c/em\u003e (MTB) infection remains the world\u0026rsquo;s leading infectious causes of death with over 1\u0026nbsp;million deaths each year (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e). Most infected people remain asymptomatic with 5\u0026ndash;10% thought to develop latent tuberculosis (LTBI) (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eLTBI has been associated with less severe SARS-CoV-2 infections (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). Here we investigated host blood transcriptional profiles in those with asymptomatic COVID-19 to investigate the immune mechanisms associated with LTBI that lead to less severe disease. Our study participants were unvaccinated individuals recruited in the early pandemic period between March and October 2020, when SARS-CoV-2 S, L and G clade strains were in circulation (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e). Our results show an upregulation of B cell pathway and CLEC12A associated genes in the LTBI-COVID group. This suggests that LTBI may afford protection by upregulating humoral immunity and dampening of excessive inflammation in those with latent TB.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy Description\u003c/h2\u003e \u003cp\u003e This study received approval from the Ethics Review Committee of the Aga Khan University (AKU) with the project number RRG 236. All methods were performed in accordance with the relevant guidelines and regulations. All the participants were adults (\u0026gt;\u0026thinsp;18 years of age) and written informed consent was obtained from each participant.\u003c/p\u003e \u003cp\u003eCOVID-19 cases were classified according to their disease severity fined by the WHO ordinal score (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e) and those with scores 1\u0026ndash;2 as Asymptomatic cases. These individuals were either completely asymptomatic or had a short (1\u0026ndash;2 days) history of fever, sore throat or myalgia which resolved without medical treatment.\u003c/p\u003e \u003cp\u003eRespiratory swabs were tested for SARS-CoV-2 using the COBAS SARS-CoV-2 assay (COBAS 6800 Roche platform) at the AKU Hospital Clinical laboratories. All study participants had PCR-confirmed COVID-19 and data collected included information regarding clinical history, co-morbid conditions. None had received COVID-19 vaccinations.\u003c/p\u003e \u003cp\u003eA single blood sample was taken from study participants within 2\u0026ndash;3 days of a positive PCR test. All study participants underwent screening for LTBI using the QuantiFERON-TB Gold (QFT) Interferon-Gamma Release Assay (IGRA) which employs TB1 and TB2 antigens based on ESAT6 and CFP10 peptides. IGRA results were interpreted as per the manufacturer\u0026rsquo;s instructions (Cellestus, USA).\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eRNA microarray data\u003c/h3\u003e\n\u003cp\u003eRNA was extracted from whole blood using the Qiagen RNA Blood Mini Kit (Qiagen, GmbH, Germany). One hundred nanogram of RNA was used for the preparation of cRNA. The samples were hybridized to the Clariom S Assay human microarray(Affymetrix, USA), containing probes for 21,488 genes.\u003c/p\u003e\n\u003ch3\u003eFunctional enrichment analysis\u003c/h3\u003e\n\u003cp\u003eCEL files were analysed using the TAC Transcriptome Analysis Software Suite (TACS version 2) using the Summarization Method: Gene Level - SST-RMA Pos vs Neg AUC Threshold: 0.7 against Genome Version: hg38 (Homo sapiens). DEGs from groups LTBI-COVID and COVID were analyzed. Cellular pathway analysis of significant differentially expressed genes (DEGs) up- or down-regulated (log FC (fold change) \u0026lt; -2 or \u0026gt;\u0026thinsp;2; p value\u0026thinsp;\u0026lt;\u0026thinsp;0.05) were identified by TACS. Significantly modified pathways were sub-grouped. Analyses were performed using R (RStudio) with the ClusterProfiler and gProfiler packages. Functional analysis of the DEGs was carried out by employing the enricher function within the Cluster Profiler package. GO enrichment for biological pathways was applied onto the data resulting in a dotplot along with a corresponding heatmap. Additional exploration of gene-level expression was carried out using the gProfiler package which enabled the formation of a gene-membership heatmap illustrating the genes contributing to each pathway.\u003c/p\u003e\n\u003ch3\u003eData Availability\u003c/h3\u003e\n\u003cp\u003eRaw array output (CEL) and processed (CHP) files were submitted to Gene Expression Omnibus (GEO) NCBI, where accession numbers were assigned and are listed in the supplementary table. All data generated or analysed during this study are included with this published article (S Table\u0026nbsp;1). Data for Quality Metrics and sample signals of probes are available in S Tables\u0026nbsp;2 and 3.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eDescription of study subjects\u003c/h2\u003e \u003cp\u003eAsymptomatic COVID-19 patients were recruited between March and October 2020. Study participants comprised SARS-CoV-2 positive unvaccinated individuals recruited in our previous study conducted in the early pandemic period before COVID-19 (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). The age range of participants was 19\u0026ndash;49 years (Mean SD 35.5, \u0026plusmn; 9.2 years). There were 6 females and 16 males. IGRA testing identified two of 22 male participants to have LTBI.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eInvestigating the host blood transcriptomes in COVID-19 cases with and without LTBI\u003c/h3\u003e\n\u003cp\u003eWe compared host blood transcriptome profiles of study participants in LTBI-COVID and COVID groups. Unhierarchical Principal Component Analysis (PCA) of transcriptional signals showed some separation between different samples but no difference between the samples with LTBI-COVID and cases of COVID (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA). Fifty-eight differentially expressed genes (DEGs) were observed with 44 upregulated (75.8%) and 13 downregulated genes (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB). A volcano plot generated to visualization of DEGs depicted that CLEC12A (inhibitory C-type lectin-like receptor on innate immune cells) showed the greatest upregulation followed by FCRL3 (immunoregulatory receptor that modulates innate B cell signaling). A subset of genes demonstrated notable downregulation, including ARMCX6 (part of ARMC subfamily; active in the mitochondria), ATG13 (a protein involved in the induction of type I interferon), TMEM126A (role in TLR4-mediated immune response), STX10 (part of SNARE complex), and FAU (regulates T cell function and survival).\u003c/p\u003e \u003cp\u003eGO enrich analysis of biological pathways analysis showed that upregulation of genes was associated with B cell activation (IL-7, SHLD1, TFRC, and FCRL3) LTBI-COVID (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC). The accompanying heatmap identifies; IL-7 (essential for lymphocyte development), SHLD1 (crucial for antibody class switching) and FCRL3 (immunoregulatory receptor that impedes the activation of human memory T lymphocytes). Downregulation of RNF5 associated with the STING pathway was also observed in LTBI-COVID.\u003c/p\u003e \u003cp\u003eCluster profiler analysis showed upregulation of predominantly innate immunity genes (CLEC12A, TRIM15, FCRL3) associated with LTBI-COVID. Its associated heatmap shows three functional pathway categories where CLEC12A showed the highest level of upregulation (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD). These were enzyme binding and organelle membrane associated pathways. Additional DEGs were associated with B and T lymphocyte differentiation and maturation, regulation or signaling (OSTM1, ART1, LMBR1L, FCRL3 and MGAT2 were evident. Also, innate immune response genes; GCC2 (NK cell-mediated immunity), APAF1 (regulation of apoptosis and inflammatory responses) and ATP1B1 (antiviral innate immune response).\u003c/p\u003e \u003cp\u003eA further description of DEGs involved in B and T cell or innate immunity are provided in S Table\u0026nbsp;2. This identifies upregulation of genes involved in T cell receptor function. Additionally, we there was upregulation of immune response genes related to vesicular trafficking, endocytosis, golgi apparatus and cellular signaling including, the STING (stimulator of interferon genes) pathway.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eOur study provides important insights into how MTB infection may affect the host immune response to SARS-CoV-2. Through host blood transcriptional profiling we observed upregulation of genes associated with B memory cells in those with asymptomatic COVID-19 as well as LTBI. Importantly, upregulation of CLEC12A associated with downmodulation of inflammatory responses was observed in the LTBI-COVID-19 group. These data fit with our recent work showing reduced rates of LTBI amongst those with mild COVID-19, suggesting a protective effect of MTB (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eEarlier, we have observed higher type I interferon responses in those with asymptomatic COVID-19 (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). Here, by focusing studying the same asymptomatic COVID-19 cohort we observed that in those with LTBI, there was the upregulation of genes associated with B cell activation. Specifically, IL-7 plays an important role in B cell development (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). FCRL3 modulates lymphocyte activity. These data are important as humoral immunity plays an important role in protection against SARS-CoV-2 infection (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). In Pakistan, we observed high rising titers of seropositivity to SARS-CoV-2 in unvaccinated individuals in the prepandemic period (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). Further, serosurveys in the population revealed increasing antibody titers associated with SARS-CoV-2 in asymptomatic individuals (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e). Our study fits with reports by Rajamanickam et al that LTBI-positive individuals exhibit elevated levels of humoral, cytokine, and systemic immune responses (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e). Our work is also supported by studies showing raised cytokine responses to Spike in IGRA positive COVID-19 cases (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTrained immunity plays a role in responses of a range of cell types from B and T to macrophages and Natural killer cells (\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e). Recall responses to BCG were found present in adults from the prepandemic period whilst their plasma had cross-reactivity to SARS-CoV-2 RBD protein, identifying the recognition of pathogen motifs by BCG-induced antibodies (\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eBCG has been shown to control mortality in children due to respiratory tract infections (\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e) and appropriate innate B cell gene activation may be one such mechanism.B cells primed with cross reactive antigens have been shown to elicit stronger anti-viral responses against LCMV infection (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e). Recently a subset of long-term memory IgM switched IgG\u0026thinsp;+\u0026thinsp;B cells has been identified to play an important role in in antiviral immunity with the characteristic of long-term epigenetic changes (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e). Hence, it is likely that B cell expansion to SARS-CoV-2 antigens is facilitated in a BCG vaccination population such as ours.\u003c/p\u003e \u003cp\u003eCOVID-19 outcomes have been associated with the severity of the initial inflammatory response to infection, with acute outcomes associated with a heightened inflammasome response. We found CLEC12A/B to be regulated. These C-type lectin- receptors play an important role in the phagocytosis of mycobacteria and in facilitating their trafficking to the phagolysosomal pathway (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e). This is supported by the pathway analysis which identifies genes linked with organelle and membrane binding to be differentially regulated in the COVID-19 sub-groups.\u003c/p\u003e \u003cp\u003eImportantly, CLEC12A contains an inhibitory motif that binds to mycobacterial mycolic acids, and can sense DAMPs which lead to the suppression of immune responses (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e). Interestingly, we found raised TRIM15, an interferon stimulated genes (ISG) but downregulation of RNF5 which is an activator of the STING pathway that is linked to inflammasome responses (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e). Hence, this data suggests that mycobacteria may dampen host immune responses reducing excess inflammatory response to SARS-CoV-2, associated with unfavourable outcomes (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOur study had limitations that was a relatively small study and we did not have any laboratory data such as hematological parameters for the study cohort. However, we know that the study participants all had asymptomatic COVID-19 with no lasting symptoms. We only had two individuals in the LTBI-COVID-19 group. This may be due to the lower frequency of LTBI in those with mild COVID-19 observed earlier (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e). We had mostly males in this group as this testing was conducted in the first cohort of healthcare workers who had returned to on campus work after the first lockdown in Pakistan. Importantly, the individuals were relatively young, all aged less than 50 years old and therefore there should be no confounding effects of an older age group present here.\u003c/p\u003e \u003cp\u003eOverall, our data fits with upregulation of a protective response in those with MTB infection in the context of SARS-CoV-2 infections (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e). BCG vaccination. These may explain the reduced disease severity from COVID-19 observed in unvaccinated individuals in the early pandemic period in high TB burden countries such as Pakistan.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEthics approval was granted by the Ethics Review Committee of the Aga Khan University. Informed consent was taken from all study participants.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConsent for publication is given by all authors.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll data after available with the submitted manuscript and are publicly available.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no conflicts of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study was supported by a COVID-19 Rapid Research Grant (RRG-236) and GCF-913 grand awarded by the Higher Education Commission, Pakistan. Fizza Fatima was supported by the Gulamali Hirji endowed Professorship fund.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026apos; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eZH, designed the study; KIM, MY: implementation and execution; FA, MY, JA: data analysis; ZH, FA, RH: drafted the initial manuscript; KIM, MY, JA: manuscript revision with critical input. All authors reviewed and approved the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank the Department of Pathology and Laboratory Medicine, Clinical Laboratories, Aga Khan University, Pakistan for facilitating the recruitment of COVID-19 patients for the study.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eJHU. COVID-19 Data Repository by the Center for Systems Science and Engineering (CSSE) at Johns Hopkins University: John Hopkins University; 2021 [Available from: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://github.com/CSSEGISandData/COVID-19\u003c/span\u003e\u003cspan address=\"https://github.com/CSSEGISandData/COVID-19\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRubio-Rivas M, Mora-Luj\u0026aacute;n JM, Formiga F, Ar\u0026eacute;valo-Ca\u0026ntilde;as C, Lebr\u0026oacute;n Ramos JM, Villalba Garc\u0026iacute;a MV, et al. WHO ordinal scale and inflammation risk categories in COVID-19. Comparative study of the severity scales. Journal of general internal medicine. 2022;37(8):1980\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBhutta ZA, Siddiqi S, Hafeez A, Islam M, Nundy S, Qadri F, et al. Beyond the numbers: understanding the diversity of covid-19 epidemiology and response in South Asia. BMJ. 2021;373:n1544.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMasood KI, Yameen M, Ashraf J, Shahid S, Mahmood SF, Nasir A, et al. Upregulated type I interferon responses in asymptomatic COVID-19 infection are associated with improved clinical outcome. Sci Rep. 2021;11(1):22958.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSette A, Crotty S. Adaptive immunity to SARS-CoV-2 and COVID-19. Cell. 2021;184(4):861\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHilligan KL, Namasivayam S, Sher A. BCG mediated protection of the lung against experimental SARS-CoV-2 infection. Front Immunol. 2023;14:1232764.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSarfraz Z, Sarfraz A, Pandav K, Singh Makkar S, Hasan Siddiqui S, Patel G, et al. Variances in BCG protection against COVID-19 mortality: A global assessment. J Clin Tuberc Other Mycobact Dis. 2021;24:100249.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSarfraz A, Hasan Siddiqui S, Iqbal J, Ali SA, Hasan Z, Sarfraz Z, et al. COVID-19 age-dependent immunology and clinical outcomes: implications for vaccines. J Dev Orig Health Dis. 2021:1\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMurray SM, Ansari AM, Frater J, Klenerman P, Dunachie S, Barnes E, et al. The impact of pre-existing cross-reactive immunity on SARS-CoV-2 infection and vaccine responses. Nat Rev Immunol. 2022:1\u0026ndash;13.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWHO. Global tuberculosis report 2021. Geneva, Switzerland: World Health Organization; 2021.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWHO. Latent tuberculosis infection: updated and consolidated guidelines for programmatic management Geneva, Switzerland: sWorld Health Organization; 2018.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAbbas U, Masood KI, Iqbal T, Jamil B, Qaiser S, Yameen M, et al. Individuals with latent tuberculosis in a high TB endemic country show mild COVID-19. PLoS One. 2025;20(12):e0339240.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGhanchi NK, Nasir A, Masood KI, Abidi SH, Mahmood SF, Kanji A, et al. Higher entropy observed in SARS-CoV-2 genomes from the first COVID-19 wave in Pakistan. PLoS One. 2021;16(8):e0256451.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOrganization WH. Ordinal Scale for Clinical Improvement. 2020.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCorfe SA, Paige CJ. The many roles of IL-7 in B cell development; mediator of survival, proliferation and differentiation. Semin Immunol. 2012;24(3):198\u0026ndash;208.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHasan M, Moiz B, Qaiser S, Masood KI, Ghous Z, Hussain A, et al. IgG antibodies to SARS-CoV-2 in asymptomatic blood donors at two time points in Karachi. PLoS One. 2022;17(8):e0271259.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eIqbal J, Hasan Z, Habib MA, Malik AA, Muhammad S, Begum K, et al. Evidence of rapid rise in population immunity from SARS-CoV-2 subclinical infections through pre-vaccination serial serosurveys in Pakistan. J Glob Health. 2025;15:04078.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRajamanickam A, Kumar NP, Padmapriyadarsini C, Nancy A, Selvaraj N, Karunanithi K, et al. Latent tuberculosis co-infection is associated with heightened levels of humoral, cytokine and acute phase responses in seropositive SARS-CoV-2 infection. J Infect. 2021;83(3):339\u0026ndash;46.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAbbas U, Masood KI, Iqbal T, Qaiser S, Rottenberg M, Jamil B, et al. Dysregulated IFN-gamma, IL-6 and TNF-alpha after COVID-19 is suggestive of lowered innate immune responses to SARS-CoV-2 and MTB. Tuberculosis (Edinb). 2025;156:102718.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSchlesinger L, Bellinger-Kawahara C, Payne N, Horwitz M. Phagocytosis of \u003cem\u003eMycobacterium tuberculosis\u003c/em\u003e is mediated by human monocyte complement receptors and complement component C3. J Immunol. 1990;144:2771\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKang B, Schlesinger L. Characterization of mannose receptor-dependent phagocytosis mediated by \u003cem\u003eMycobacterium tuberculosis\u003c/em\u003e lipoarabinomannan. Infect Immun. 1998;66:2769\u0026ndash;77.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNishimura N, Tomiyasu N, Torigoe S, Mizuno S, Fukano H, Ishikawa E, et al. Mycobacterial mycolic acids trigger inhibitory receptor Clec12A to suppress host immune responses. Tuberculosis (Edinb). 2023;138:102294.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhong B, Zhang L, Lei C, Li Y, Mao AP, Yang Y, et al. The ubiquitin ligase RNF5 regulates antiviral responses by mediating degradation of the adaptor protein MITA. Immunity. 2009;30(3):397\u0026ndash;407.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRodrigues TS, Zamboni DS. Inflammasome activation by SARS-CoV-2 and its participation in COVID-19 exacerbation. Curr Opin Immunol. 2023;84:102387.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSherwood ER, Burelbach KR, McBride MA, Stothers CL, Owen AM, Hernandez A, et al. Innate Immune Memory and the Host Response to Infection. J Immunol. 2022;208(4):785\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKleinnijenhuis J, Quintin J, Preijers F, Joosten LA, Jacobs C, Xavier RJ, et al. BCG-induced trained immunity in NK cells: Role for non-specific protection to infection. Clin Immunol. 2014;155(2):213\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eIqbal NT, Ahmed K, Sattar T, Aziz F, Hussain R. BCG activation of trained immunity is associated with induction of cross reactive COVID-19 antibodies in a BCG vaccinated population. PLoS One. 2024;19(5):e0302722.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCampelo TA, Oliveira NS, Souza PFN, de-Oliveira DFG, Frota CC, Antas PRZ. Unleashing the power of the BCG vaccine in modulating viral immunity through heterologous protection: A scoping review. Hum Vaccin Immunother. 2025;21(1):2521190.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePratumchai I, Zak J, Huang Z, Min B, Oldstone MBA, Teijaro JR. B cell-derived IL-27 promotes control of persistent LCMV infection. Proc Natl Acad Sci U S A. 2022;119(3).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhu X, Hong S, Bu J, Liu Y, Liu C, Li R, et al. Antiviral memory B cells exhibit enhanced innate immune response facilitated by epigenetic memory. Sci Adv. 2024;10(13):eadk0858.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAbbas U, Masood KI, Iqbal T, Jamil B, Yameen M, Qaiser S, et al. Immune Reactivity to Mycobacterium Tuberculosis Antigens is Positively Associated with Protection Against Infection with COVID-19. SSRN preprint. 2024.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBooysen P, Wilkinson KA, Sheerin D, Waters R, Coussens AK, Wilkinson RJ. Immune interaction between SARS-CoV-2 and Mycobacterium tuberculosis. Front Immunol. 2023;14:1254206.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Latent TB, SARS-CoV-2, B cell, innate immunity, inflammation","lastPublishedDoi":"10.21203/rs.3.rs-8500902/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8500902/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eSARS-CoV-2 infections resulted in a global pandemic with variable rates of COVID-19 morbidity worldwide. In high tuberculosis (TB) endemic regions it was observed that COVID-19 associated mortality was lower. Infection with \u003cem\u003eMycobacterium tuberculosis\u003c/em\u003e (MTB) has been shown to impact COVID-19 infections but the mechanisms involved are as yet unclear. Here we investigated the effect of latent MTB infection (LTBI) by studying host blood transcriptional profiles in individuals with Asymptomatic COVID-19.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eWe studied participants who had asymptomatic COVID-19 compared with those who had LTBI as well COVID (LTBI-COVID). Participants were aged 35.5 (+-9.2) years and comprised 27% females. Whole blood transcriptional profiles were investigated using RNA microarray analysis of individuals.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eComparison of the LTBI-COVID and COVID groups revealed 58 differentially expressed genes (DEGs) of which 44 were upregulated and 13 were downregulated. Cluster profiler analysis showed upregulation of innate immunity associated genes (CLEC12A, TRIM15, FCRL3) upregulated in LTBI-COVID. A GO enrich analysis showed upregulation of B cell activation associated with (IL-7, SHLD1, TFRC, and FCRL3). There was a downregulation of RNF5 associated with the STING pathway.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e \u003cp\u003eB cell activation would lead to increased humoral immunity and enhanced uptake of pathogens by macrophages. CLEC12A and RNF5 are associated with dampening of excessive inflammation which reduces risks of COVID-19. These data support the protective role of latent TB infection associated with reduced severity of COVID-19.\u003c/p\u003e","manuscriptTitle":"Host transcriptional profiling identifies B cell associated genes to be upregulated in individuals with asymptomatic COVID-19 and latent tuberculosis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-12 06:06:34","doi":"10.21203/rs.3.rs-8500902/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"1e426f2f-6571-40f1-9340-1ea9ee61fd50","owner":[],"postedDate":"January 12th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-04-21T21:23:52+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-12 06:06:34","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8500902","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8500902","identity":"rs-8500902","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}