Unbiased metagenomic detection of RNA viruses for rapid identification of viral pathogens in clinical samples (preprint)

Unbiased long read sequencing approaches for clinical metagenomic sample analysis holds enormous potential for pathogen detection, including improved detection of unknown, novel or emerging viruses. However, the rapid rate of development in nanopore sequencing and library preparation methods complicates the process of selecting a standardized method for unbiased RNA virus detection. Here, we evaluate multiple sequencing approaches to identify a workflow with sufficient sensitivity, limits of detection, and throughput for potential utilization in a clinical laboratory setting. Four separate library preparation methods for the Oxford Nanopore Technologies MinION sequencer are compared, including direct RNA, direct cDNA, rapid cDNA, and double stranded cDNA. We also establish that depletion of host RNA is not required and can be deleterious for viral RNA detection in some instances when using samples in viral transport media (VTM) or plasma. Using unbiased whole genome amplification following reverse transcription, we achieve limits of detection on the order of 1.95E03 GE/mL of Venezuelan Equine Encephalitis Virus (VEEV) spiked in human plasma. We also report initial detection of 5.43E06 GE/mL of coronavirus 229E spiked into VTM samples containing human background RNA which are expected to decrease significantly during upcoming testing. These metrics were achieved within a 6-plex multiplex reaction, illustrating the potential to increase throughput and decrease costs for relevant sample analysis. Data analysis was performed using EPI2ME Labs framework and open access tools that are readily accessible to most clinical laboratories. Taken together, this work describes an optimized method for unbiased nanopore sequencing and analysis of RNA viruses present in two common clinical matrices.

Whole-genome demography of COVID-19 virus during its pandemic period and on “panvalent” vaccine design (preprint)

With over 16 million submitted genomic sequences, the SARS-CoV-2 (SC2) virus, the cause of the most recent worldwide pandemic COVID-19 pandemic, has become the most sequenced genome of all known viruses (1), revealing, for example, a vast number of expanding viral lineages.  Since the pandemic phase appears to be over, we performed a retrospective re-examination of the demographic grouping pattern and their genomic characteristics during the entire pandemic period up to the peak of the last pandemic wave. For our study, we extracted from the NCBI (2) only unique viral sequences and converted each sequence data to a relationalvector, indicating the presence/absence of each variational event compared to a “reference” sequence. Our study revealed several genomic features that are unexpected or different from those of previous studies (3 – 9).  For example, approximately 44,000 variants with unique sequences emerged during the pandemic period; they group into only four major demographic groups and each has a set of mostly unique highly-conserved variant-genotypes (HCVGs); and the set from the first group was passed down to the remaining three groups, suggesting that HCVGs in the next group may be predictable from the HCVGs of the current group(s). Such a concept may be potentially important in designing “panvalent” vaccines against the current and future waves of viral infections.

Identification and analysis of co-disease genes between COVID-19 and osteoarthritis (preprint)

Background: The molecular mechanisms of corona virus disease 2019 (COVID-19) and osteoarthritis (OA) are unclear, and there is an urgent need to identify new biomarkers and explore their potential molecular mechanisms in COVID-19 and OA. Methods: The GSE57218, GSE157103 training sets and the GSE82107, GSE171110 validation sets were acquired via gene expression omnibus (GEO) database. First, differentially expressed genes (DEGs) between disease and normal samples in the GSE57218 and GSE157103 training sets were respectively sifted out by differential expression analysis. The modules with the highest correlation with OA and normal, COVID-19 and non COVID-19 were gained by weighted gene co-expression network analysis (WGCNA), individually. Then, OA-DEGs were intersected with the module genes that had significant correlation with OA, and COVID-19-DEGs were intersected with the module genes which were dramatically correlated with COVID-19 to yield OA-intersected genes and COVID-19 intersected genes, respectively. The OA-intersected genes and COVID-19 intersected genes were intersected to yield candidate genes, and they were analyzed for function enrichment analysis. Next, the seven algorithms (Closeness, MCC, Degree, MNC, Radality, Stress and EPC) were performed on candidate genes to sift out biomarkers. Finally, we constructed the competing endogenous RNA (ceRNA), transcription factor (TF)/miRNA-mRNA and drug-target regulatory networks. Results:There were 1135 OA-DEGs and 4336 COVID-19-DEGs between disease and normal samples in the GSE57218 and GSE157103 training sets, respectively. The pink, blue and brown modules had significant correlations with OA in the GSE57218 training set, while in the GSE157103 training set, the pink and brown modules were notably correlated with COVID-19. We finally yield 715 OA-intersected genes and 2282 COVID-19-intersected genes. After intersecting the above two intersected genes, we gained 106 candidate genes, and they were involved in ADP metabolic process, nucleoside diphosphate phosphorylation, etc.. The 7 biomarkers, namely AK1, APP, ENO1, TPI1, HSP90B1, HSPB1 and ESR1, were acquired based on seven algorithms. Finally, we successfully constructed the ceRNA, TF/miRNA-mRNA and drug-target networks. Conclusion: Through bioinformatic methods, we explored the biomarkers (AK1, APP, ENO1, TPI1, HSP90B1, HSPB1 and ESR1) of COVID-19 combined OA, providing new ideas for studies related to molecular mechanisms and treatment of comorbidity.

Cell culture differentiation and proliferation conditions influence the in vitro regeneration of the human airway epithelium (preprint)

The human airway mucociliary epithelium can be recapitulated in vitro using primary cells cultured in an Air-Liquid Interface (ALI), a reliable surrogate to perform pathophysiological studies. As tremendous variations exist between media used for ALI-cultured human airway epithelial cells, our study aimed to evaluate the impact of several media (BEGM, PneumaCult, "Half&Half" and "Clancy") on cell type distribution using single-cell RNA sequencing and imaging. Our work revealed the impact of these media on cell composition, gene expression profile, cell signaling or epithelial morphology. We found higher proportions of multiciliated cells in PneumaCult-ALI and Half&Half, stronger EGF signaling from basal cells in BEGM-ALI, differential expression of the SARS-CoV-2 entry factor ACE2, and distinct secretome transcripts depending on media used. We also established that proliferation in PneumaCult-Ex Plus favored secretory cell fate, showing the key influence of proliferation media on late differentiation epithelial characteristics. Altogether, our data offer a comprehensive repertoire for evaluating the effects of culture conditions on airway epithelial differentiation and will help to choose the most relevant medium according to the processes to be investigated such as cilia, mucus biology or viral infection. We detail useful parameters that should be explored to document airway epithelial cell fate and morphology.

Vaccine-Induced Viral Reactivation and Autism Spectrum Disorder: A Review, Hypothesis and Implications. (preprint)

Understanding the origins of autism spectrum disorder is imperative given its increasing prevalence and significant global impact. This review examines existing research on the role of viral infections in the etiology of autism and scrutinizes emerging data on post-vaccination viral reactivation. It introduces a novel hypothesis, that vaccines may inadvertently reactivate latent viral infections, triggering the onset or exacerbation of autism. Through a comprehensive literature review across multiple databases and search engines, this review analyzed studies, case reports, and observational research focusing on viral infections, post-vaccination viral reactivation, and autism. This search yielded 12 studies implicating viral infections and viral encephalitis as potential causal factors in the development of autism spectrum disorder. Moreover, 10 studies were identified suggesting an association of viral reactivation following vaccination. This connection raises important questions about the potential role of vaccines in the onset of autism. The findings advocate for continued vigilance in vaccine safety research, particularly concerning neurodevelopmental disorders.

Clinical outcomes of coronavirus disease 2019 in people living with human immunodeficiency virus in South Korea: A nationwide population-based cohort study (preprint)

Background: We aimed to compare the epidemiological and clinical characteristics of coronavirus disease 2019 (COVID-19) in people living with human immunodeficiency virus (HIV) (PLWH) with those in the general population. Methods: This nationwide descriptive epidemiological study was conducted in South Korea between January 2020 and February 2022. The National Health Insurance claims data covering the whole nation were collected through the Health Insurance Review and Assessment Service. Results: Among 3,653,808 individuals who were diagnosed with COVID-19, 1,311 (0.04%) were PLWH. All the PLWH received antiretroviral therapy, and 26.47% had more than one underlying disease other than HIV infection. The overall in-hospital mortality rates of PLWH and the general population were 0.76% and 0.25%, respectively (P=0.002). According to the Cox proportional hazard model, no significant difference was observed in the in-hospital mortality rate [hazard ratio (HR): 1.80, 95% confidence interval (CI): 0.70–4.67] between PLWH and the general population. However, progression to severe or critical COVID-19 was more common in PLWH (HR: 2.70, 95% CI: 1.37–5.33). In PLWH diagnosed with COVID-19, a multivariable Cox regression analysis found old age (≥60 years old) (HR: 6.9; 95% CI 2.57–18.56) and diabetes mellitus (HR: 5.13; 95% CI: 2.02–13.00) as the independent risk factors for severe or critical COVID-19. Conclusions: PLWH had a significantly higher risk of severe or critical COVID-19 than that of the general population. Our findings suggest the need for applying differentiated strategies to decrease the impact of COVID-19 on PLWH.

Exploring the therapeutic potential of defective interfering particles in reducing the replication of SARS-CoV-2 (preprint)

SARS-CoV-2 still presents a global threat to human health due to the continued emergence of new strains and waning immunity amongst vaccinated populations. Therefore, it is still relevant to investigate potential therapeutics, such as therapeutic interfering particles (TIPs). Mathematical and computational modelling are valuable tools to study viral infection dynamics for predictive analysis. Here, we expand on the previous work by Grebennikov et al. (2021) on SARS-CoV-2 intra-cellular replication dynamics to include defective interfering particles (DIPs) as potential therapeutic agents. We formulate a deterministic model that describes the replication of wild-type (WT) SARS-CoV-2 virus in the presence of DIPs. Sensitivity analysis of parameters to several model outputs is employed to inform us on those parameters to be carefully calibrated from experimental data. We then study the effects of co-infection on WT replication and how DIP dose perturbs the release of WT viral particles. Furthermore, we provide a stochastic formulation of the model that is compared to the deterministic one. These models could be further developed into population-level models or used to guide the development and dose of TIPs.

Resolution of SARS-CoV-2 infection in human lung tissues is driven by extravascular CD163+ monocytes (preprint)

The lung-resident immune mechanisms driving resolution of SARS-CoV-2 infection in humans remain elusive. Using mice co-engrafted with a genetically matched human immune system and fetal lung xenograft (fLX), we mapped the immunological events defining resolution of SARS-CoV-2 infection in human lung tissues. Viral infection is rapidly cleared from fLX following a peak of viral replication. Acute replication results in the emergence of cell subsets enriched in viral RNA, including extravascular inflammatory monocytes (iMO) and macrophage-like T-cells, which dissipate upon infection resolution. iMO display robust antiviral responses, are transcriptomically unique among myeloid lineages, and their emergence associates with the recruitment of circulating CD4+ monocytes. Consistently, mice depleted for human CD4+ cells but not CD3+ T-cells failed to robustly clear infectious viruses and displayed signatures of chronic infection. Our findings uncover the transient differentiation of extravascular iMO from CD4+ monocytes as a major hallmark of SARS-CoV-2 infection resolution and open avenues for unravelling viral and host adaptations defining persistently active SARS-CoV-2 infection.

The relationship between omega-3 intake and the severity and symptoms of Covid-19 (preprint)

Background and Aims: Omega 3 polyunsaturated fatty acids (n3 PUFAs) may exert beneficial effects on the immune system of patients with viral infections. This paper aimed to examine the effect of n3 PUFA on severity and symptoms in critically ill patients with COVID 19Methods In a cross-sectional study, 250 COVID-19 patients between the ages of 18 and 65 were enrolled. To gauge dietary omega-3 consumption, a validated 168-item online food frequency questionnaire (FFQ) was used. The severity of COVID-19 was determined using the COVID-19 Treatment Guidelines, and symptoms were assessed using a common questionnaire. Crude and modified analyses were carried out. (Model 1: age, sex, and energy intake; Model 2: Model 1 + physical activity, supplements, corticosteroids, and antiviral drugs; Model 3: Model 2 + body mass index).Results The mean age of participants was 44.06 ± 11.84 years, and 46% of them had severe COVID-19. Patients at the highest tertile of dietary omega-3 intake had lower serum levels of inflammatory biomarkers, including CRP (10.25 ± 13.12 vs. 25.20 ± 24.89 mg/L, p < 0.001) and ESR (14.55 ± 15.02 vs. 29.13 ± 2.26 mm/hr, p < 0.001), than those at the lowest tertile. After controlling for potential confounders, we observed that a higher dietary omega-3 intake was associated with a lower odds of severe COVID-19 (OR: 0.42; 95% CI: 0.22–0.79).Conclusion We found that higher intake of dietary omega-3 was inversely associated with COVID-19 severity and symptoms.

Filopodial Mechanotransduction is regulated by Angiotensin-Converting Enzyme 2 (ACE2) and by SARS-CoV-2 spike protein (preprint)

Filopodia are dynamic, actin-rich cellular protrusions, increasingly linked to cellular mechanotransduction. However, how dynamic filopodia translate external mechanical cues remains poorly understood. Recent studies show that the SARS-CoV-2 spike (S) protein binds the ACE2 receptor on airway multicilia and that cilia are required for viral infection(1) and sufficient to induce filopodial extension and viral binding. To test if spike protein is sufficient to induce filopodial expansion, we employed live-cell single-particle tracking with quantum dots targeting ACE2, to reveal a robust filopodia extension and virus binding mechanism requiring the enzymatic activity of ACE2. Using time-lapse imaging, we reveal that spike protein binding to filopodia is associated with intracellular actin remodeling, alterations in bulk cell stiffness, and an elevation in intracellular calcium levels linked to actin-rearrangement, filopodia initiation, and persistence. We propose the activation of ACE2 creates an active signaling and mechanosensory environment within adherent cells and airway epithelial cells that allows the remodeling of actin in filopodia to trap virus and potentially organize viral exit from cells.

Targeting ARF4-mediated intracellular transport as broad-spectrum antivirals (preprint)

Host factors that regulate cellular vesicular trafficking also contribute to progeny virions’ destination, thus representing as potential antiviral drug targets. Here we demonstrate that genetic deletion of ARF4, a regulator in vesicle transport, repressed multiple pathogenic RNA viral infections including Zika virus (ZIKV), influenza A virus (IAV), SARS-CoV-2 and Vesicular Stomatitis virus (VSV). ARF4 activation was stimulated upon viral infection, and viral production was rescued when reconstituted with the activated ARF4, but not the inactivated mutants. Mechanically, ARF4 deletion obstructed viral normal translocation into Golgi complex, but led to mis-sorting for lysosomal degradation, consequently caused the blockage of final release. More importantly, ARF4 targeting peptides achieved significant therapeutic efficacy against ZIKV and IAV challenge in mice by blocking ARF4 activation. Hence, we clarify the critical role of ARF4 during viral infection, providing a broad-spectrum antiviral target and the basis for further pharmaceutical development.

A retrospective clinical study in Graves disease with COVID-19 infection in China (preprint)

Background Corona Virus Disease 2019 (COVID-19) is the most prevalent global pandemic in recent times. Graves disease (GD), an autoimmune thyroid disease, is a clinical syndrome caused by excessive thyroid hormones. Our study is to understand the current epidemiological situation of COVID-19 infection in GD patients, and to analyze whether COVID-19 will affect the thyroid function, thyroid autoantibody and metabolism of GD patients.Methods 109 GD patients were followed by Shanghai General Hospital Thyroid Disease Center (TDC) from November 2022 to June 2023. There were three groups defined, i.e., pre, one-month after and three months after infection with COVID-19. SPSS was used to analyze the recruited data.Results 109 GD patients are infected with COVID-19 (72.48%), uncontrolled GD patients with high FT3 had a higher COVID-19 infection rate (79.31%). As for thyroid function in 35 GD patients with antithyroid drug (ATD) maintenance stage, there were significant differences in FT3, FT4, TT3 and TT4 before and after being infected with COVID-19. What’s more, there’s a significant difference between GD patients in one month and three months after COVID-19 infection of high TSAb group (p = 0.048) but no significant difference between pre and one month. What’s more, there were significant differences in TT3, TT4 of GD patients after infected COVID-19 in non. And Phosphorus (P), 25-hydroxyvitamin D (25-OH-D3), Procollagen type 1 N-terminal propeptide (P1NP) in GD patients were be affected by COVID-19 infection.Conclusion GD patients with uncontrolled thyroid function group are susceptible to COVID-19. COVID-19 may affect the thyroid function of GD in TT3, TT4, TSAb high level group infection. COVID-19 vaccine is conducive to the stability of GD patients' condition. And COVID-19 may affect the bone metabolism in GD patients before and after COVID-19 infection. But there is no effect on glucose metabolism or lipid metabolism.

Inhibition of SARS-CoV-2 infection by Porphyromonas gingivalis and the oral microbiome (preprint)

The COVID-19 pandemic persists despite the availability of vaccines, and it is therefore crucial to develop new therapeutic and preventive approaches. In this study, we investigated the potential role of the oral microbiome in SARS-CoV-2 infection. Using an in vitro SARS-CoV-2 pseudovirus infection assay, we found a potent inhibitory effect exerted by Porphyromonas gingivalis on SARS-CoV-2 infection mediated by known P. gingivalis compounds such as phosphoglycerol dihydroceramide (PGDHC) and gingipains as well as by unknown bacterial factors. We found that the gingipain-mediated inhibition of infection is likely due to cytotoxicity, while PGDHC inhibited virus infection by an unknown mechanism. Unidentified factors present in P. gingivalis supernatant inhibited SARS-CoV-2 likely via the fusion step of the virus life cycle. We addressed the role of other oral bacteria and found certain periodontal pathogens capable of inhibiting SARS-CoV-2 pseudovirus infection by inducing cytotoxicity on target cells. In the human oral cavity, we observed the modulatory activity of oral microbial communities varied among individuals in that some saliva-based cultures were capable of inhibiting while others were enhancing infection. These findings contribute to our understanding of the complex relationship between the oral microbiome and viral infections, offering potential avenues for innovative therapeutic strategies in combating COVID-19.

Secondary structure of the SARS-CoV-2 genome is predictive of nucleotide substitution frequency (preprint)

Accurate estimation of the effects of mutations on SARS-CoV-2 viral fitness can inform public-health responses such as vaccine development and predicting the impact of a new variant; it can also illuminate biological mechanisms including those underlying the emergence of variants of concern. Recently, Lan et al reported a high-quality model of SARS-CoV-2 secondary structure and its underlying dimethyl sulfate (DMS) reactivity data. I investigated whether secondary structure can explain some variability in the frequency of observing different nucleotide substitutions across millions of patient sequences in the SARS-CoV-2 phylogenetic tree. Nucleotide basepairing was compared to the estimated mutational fitness of substitutions, a measurement of the difference between observed and expected substitution frequency that is correlated with other estimates of viral fitness. This comparison revealed that secondary structure is often predictive of substitution frequency, with significant decreases in substitution frequencies at basepaired positions. Focusing on the mutational fitness of C[->]T, the most common type of substitution, I describe C[->]T substitutions at basepaired positions that characterize major SARS-CoV-2 variants; such mutations may have a greater impact on fitness than appreciated when considering substitution frequency alone.

Comparative Clinical Assessment and Risk Stratification of COVID-19 and Influenza Infections in Adults and Children: A Comprehensive Systematic Review and Meta-Analysis (preprint)

Background and objective Coronavirus disease 2019 (COVID-19) is a viral disease that rapidly spread over the world, prompting to it to be declared a global pandemic. Since the illness exhibits similar symptoms as influenza, it can be challenging to tell the two diseases apart, especially during the influenza season. Therefore, it was necessary to carry out a comparative study to assess the clinical risks and outcomes of COVID-19 and influenza.Methods The search for relevant articles was carried out through the database search method and a manual search which involved going through the reference lists of articles related to the topic for additional studies. The Quality appraisal was carried out using the Newcastle Ottawa tool, while data analysis was done using the Review Manager Software (RevMan 5.4.1).Results The meta-analysis results show that COVID-19 patients had similar lengths of hospital stay (SMD: -0.25; 95% CI: -0.60 to 0.11; p = 0.17). However, COVID-19 patients had significantly higher mortality rates (RR: 0.28; 95% CI: 0.21 to 0.37; p < 0.0001), in-hospital complications (RR: 0.57; 95% CI: 0.50 to 0.65; p < 0.00001), intensive care unit (ICU) admissions (OR: 0.48; 95% CI: 0.37 to 0.61; p < 0.00001), length of ICU stay (SMD: -0.45; 95% CI: -0.83 to 0.06; p = 0.02), and mechanical ventilation use (OR: 0.36; 95% CI: 0.28 to 0.46; p < 0.00001).Conclusion The findings suggest that COVID-19 is more severe than influenza. Therefore, “flu-like” symptoms should not be dismissed without a clear diagnosis, especially during the winter seasons when influenza is more common.

The Genetic Interface of Immunity and COVID-19: Insights from a Two-Sample Mendelian Randomization Approach (preprint)

Introduction Coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has emerged as a global public health emergency since late 2019. Immune cells are crucial for host defense against viral infection and disease progression. However, the specific immune cell characteristics that influence susceptibility to COVID-19 remain unclear. This study aimed to investigate the causal relationship between immune cell signatures and COVID-19 using MR analysis.Materials and Methods This study utilized publicly available genetic datasets from the COVID-19 Host Genetics Initiative and the Blueprint Consortium and applied a two-sample Mendelian randomization approach to examine the association between 731 immune cell signatures and the risk of COVID-19. We included four types of immune signatures: median fluorescence intensity (MFI), relative cell count (RC), absolute cell count (AC), and morphological parameter (MP) data. We used the inverse-variance weighted (IVW) method as the primary analysis and performed several sensitivity analyses to test the robustness of the results.Results Our analysis revealed 30 distinct immune cell characteristics that were directly associated with the risk of COVID-19, including CD4 + regulatory T cells (CD4 + Treg cells), CCR2 + CD14- CD16 + monocytes, CD86 + plasmacytoid DC AC, CCR2 + plasmacytoid DC (CCR2 + pDC), CCR2 + CD62L + plasmacytoid DC (CCR2 + CD62L + pDC), and CD80 + CD62L + plasmacytoid DC (CD80 + pDC). However, among these findings, only the expression of CCR2 on CD14-CD16 + monocytes had a significant impact (P = 0.0249, OR = 1.0427, 95% CI=[1.0053, 1.0814]) on immune cell attributes in the context of COVID-19. Sensitivity analyses confirmed the validity of the IVW results and ruled out the possibility of horizontal pleiotropy.Conclusion Through two-sample Mendelian randomization analysis, we demonstrated a significant causal relationship between specific immune cell characteristics and the risk of COVID-19. These findings provide important genetic evidence for the development of future vaccines and treatment strategies.

Viral Infections, a Trigger and Risk Factor of Alzheimer’s Disease? (preprint)

Alzheimer’s Disease (AD), a progressive and debilitating condition, is reported to be the most common type of dementia, with at least 55 million people believed to be currently affected. Many causation hypotheses of AD exist, yet the intriguing link between viral infection and its possible contribution to the known etiology of AD has become an attractive focal point of research for the field and a challenging study task. In this review, we will explore the historical perspective and milestones that led the field to investigate the viral connection to AD. Specifically, several viruses such as Herpes Simplex Virus 1 (HSV-1), Zika virus (ZIKV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), along with several others mentioned, include the various viruses presently considered within the field. We delve into the strong evidence implicating these viruses in the development of AD. We will also extend beyond these mere associations by carefully analyzing the potential mechanisms by which viruses may contribute to AD pathology. This includes but is not limited to direct neuronal infections, dysregulation of immune responses, and the impact on protein processing. Controversies and challenges of the viral-AD relationship emerge as we tease out these potential mechanisms considered. Looking forward, we emphasize the future directions the field should take to tackle the remaining unanswered questions and the glaring research gaps that persist. Overall, this review aims to provide a comprehensive survey of the past, present, and future of the potential link between viral infections and their association with AD development.

SARS-CoV-2 infection elucidates unique features of pregnancy-specific immunity (preprint)

Pregnancy is a risk factor for increased severity of SARS-CoV-2 and other respiratory infections. The mechanisms underlying this risk have not been well-established, partly due to a limited understanding of how pregnancy shapes immune responses. To gain insight into the role of pregnancy in modulating immune responses at steady state and upon perturbation, we collected peripheral blood mononuclear cells (PBMC), plasma, and stool from 226 women, including 152 pregnant individuals (n = 96 with SARS-CoV-2 infection and n = 56 healthy controls) and 74 non-pregnant women (n = 55 with SARS-CoV-2 and n = 19 healthy controls). We found that SARS-CoV-2 infection was associated with altered T cell responses in pregnant compared to non-pregnant women. Differences included a lower percentage of memory T cells, a distinct clonal expansion of CD4-expressing CD8+ T cells, and the enhanced expression of T cell exhaustion markers, such as programmed cell death-1 (PD-1) and T cell immunoglobulin and mucin domain-3 (Tim-3), in pregnant women. We identified additional evidence of immune dysfunction in severely and critically ill pregnant women, including a lack of expected elevation in regulatory T cell (Treg) levels, diminished interferon responses, and profound suppression of monocyte function. Consistent with earlier data, we found maternal obesity was also associated with altered immune responses to SARS-CoV-2 infection, including enhanced production of inflammatory cytokines by T cells. Certain gut bacterial species were altered in pregnancy and upon SARS-CoV-2 infection in pregnant individuals compared to non-pregnant women. Shifts in cytokine and chemokine levels were also identified in the sera of pregnant individuals, most notably a robust increase of interleukin-27 (IL-27), a cytokine known to drive T cell exhaustion, in the pregnant uninfected control group compared to all non-pregnant groups. IL-27 levels were also significantly higher in uninfected pregnant controls compared to pregnant SARS-CoV-2-infected individuals. Using two different preclinical mouse models of inflammation-induced fetal demise and respiratory influenza viral infection, we found that enhanced IL-27 protects developing fetuses from maternal inflammation but renders adult female mice vulnerable to viral infection. These combined findings from human and murine studies reveal nuanced pregnancy-associated immune responses, suggesting mechanisms underlying the increased susceptibility of pregnant individuals to viral respiratory infections.

Causal relationship between COVID-19 and the risk of asthma: A Mendelian Randomisation study (preprint)

Background Existing research has focused on new-onset asthma and viral infections, particularly respiratory syncytial virus (RSV). However, studies on whether COVID-19 can induce asthma are limited.Methods We performed bidirectional two-sample Mendelian Randomization (MR) to assess the potential causal relationship between COVID-19 and asthma using genome-wide association study (GWAS) summary data obtained from the COVID-19 Host Genetic Initiative GWAS Meta-analysis Round 5 (release date: 18 January 2021). Several methods (random-effects inverse variance weighted, weighted median, MR-Egger regression, and MR-PRESSO) were used to ensure the robustness of the causal effects. Heterogeneity was measured using Cochran's Q value. Horizontal pleiotropy was evaluated using MR-Egger regression and leave-one-out analyses.Results We observed a significant causal association between COVID-19 hospitalisation and asthma (odds ratio (OR) = 1.042, 95% Confidence Interval (CI) = 1.004–1.081, p = 0.031), indicating a significantly increased risk of COVID-19 hospitalisation associated with asthma. However, no statistically significant causal relationships were observed for COVID-19 susceptibility (OR = 1.023, 95% CI = 0.931–1.124, p = 0.637), COVID-19 severity (OR = 1.006, 95% CI = 0.978–1.035, p = 0.669), and asthma.Conclusions COVID-19 can trigger the onset of asthma. Individuals experiencing prolonged coughing, chest tightness, or difficulty in breathing long after recovery from COVID-19 should remain vigilant about the possibility of developing asthma.

SARS-CoV-2-Specific Immune Responses in Vaccination and Infection during the Pandemic in 2020–2022 (preprint)

To better understand how immunity develops against SARS-CoV-2 during the pandemic period of 2020 to 2022, we analyzed the immune response of a small group of university staff and students who were either infected or vaccinated. We investigated the levels of receptor-binding domain (RBD)-specific and nucleocapsid (N)-specific IgG and IgA antibodies in serum and saliva samples taken early (around 10 days after infection or vaccination) and later (around 1 month later), as well as N-specific T-cell responses. One patient who had been infected in 2020 developed serum RBD and N-specific IgG antibodies, but declined 8 months later, then mRNA vaccination in 2021 produced a higher level of anti-RBD IgG than natural infection. In the vaccination of naïve individuals, vaccines induced anti-RBD IgG, but it declined after 6 months. A third vaccination boosted the IgG level again, albeit to a lower level than after the second. In 2022, when the Omicron variant became dominant, familial transmission occurred even among vaccinated people. In infected individuals, the levels of serum anti-RBD IgG antibodies increased later, while anti-N IgG peaked earlier. The N-specific activated T cells expressing IFN- or CD107a were detected early. In saliva, SARS-CoV-2-specific IgG but not IgA was detected. Interestingly, two individuals showed a temporary peak in RBD- and N-specific IgA antibodies in their saliva on the second day after infection. Thus, SARS-CoV-2 infection triggers typical immune responses in humans against acute viral infections, but mucosal immune responses, which are temporarily induced early during infection, may be more important for protection than current intramuscular vaccines.