Integrated time-series transcriptomic and metabolomic analyses reveal different inflammatory and adaptive immune responses contributing to host resistance to PRRSV.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a highly contagious disease that affects the global pig industry. To understand mechanisms of susceptibility/resistance to PRRSV, this study profiled the time-serial white blood cells transcriptomic and serum metabolomic responses to PRRSV in piglets from a crossbred population of PRRSV-resistant Tongcheng pigs and PRRSV-susceptible Large White pigs. Gene set enrichment analysis (GSEA) illustrated that PRRSV infection up-regulated the expression levels of marker genes of dendritic cells, monocytes and neutrophils and inflammatory response, but down-regulated T cells, B cells and NK cells markers. CIBERSORT analysis confirmed the higher T cells proportion in resistant pigs during PRRSV infection. Resistant pigs showed a significantly higher level of T cell activation and lower expression levels of monocyte surface signatures post infection than susceptible pigs, corresponding to more severe suppression of T cell immunity and inflammatory response in susceptible pigs. Differentially expressed genes between resistant/susceptible pigs during the course of infection were significantly enriched in oxidative stress, innate immunity and humoral immunity, cell cycle, biotic stimulated cellular response, wounding response and behavior related pathways. Fourteen of these genes were distributed in 5 different QTL regions associated with PRRSV-related traits. Chemokine CXCL10 levels post PRRSV infection were differentially expressed between resistant pigs and susceptible pigs and can be a promising marker for susceptibility/resistance to PRRSV. Furthermore, the metabolomics dataset indicated differences in amino acid pathways and lipid metabolism between pre-infection/post-infection and resistant/susceptible pigs. The majority of metabolites levels were also down-regulated after PRRSV infection and were significantly positively correlated to the expression levels of marker genes in adaptive immune response. The integration of transcriptome and metabolome revealed concerted molecular events triggered by the infection, notably involving inflammatory response, adaptive immunity and G protein-coupled receptor downstream signaling. This study has increased our knowledge of the immune response differences induced by PRRSV infection and susceptibility differences at the transcriptomic and metabolomic levels, providing the basis for the PRRSV resistance mechanism and effective PRRS control.

Human leukocyte antigen class II gene diversity tunes antibody repertoires to common pathogens.

Allelic diversity of human leukocyte antigen (HLA) class II genes may help maintain humoral immunity against infectious diseases. In this study, we investigated germline genetic variation in classical HLA class II genes and employed a systematic, unbiased approach to explore the relative contribution of this genetic variation in the antibody repertoire to various common pathogens. We leveraged a well-defined cohort of 800 adults representing the general Arab population in which genetic material is shared because of the high frequency of consanguineous unions. By applying a high-throughput method for large-scale antibody profiling to this well-defined cohort, we were able to dissect the overall effect of zygosity for classical HLA class II genes, as well as the effects associated with specific HLA class II alleles, haplotypes and genotypes, on the antimicrobial antibody repertoire breadth and antibody specificity with unprecedented resolution. Our population genetic studies revealed that zygosity of the classical HLA class II genes is a strong predictor of antibody responses to common human pathogens, suggesting that classical HLA class II gene heterozygosity confers a selective advantage. Moreover, we demonstrated that multiple HLA class II alleles can have additive effects on the antibody repertoire to common pathogens. We also identified associations of HLA-DRB1 genotypes with specific antigens. Our findings suggest that HLA class II gene polymorphisms confer specific humoral immunity against common pathogens, which may have contributed to the genetic diversity of HLA class II loci during hominine evolution.

Global patterns of antigen receptor repertoire disruption across adaptive immune compartments in COVID-19.

Whereas pathogen-specific T and B cells are a primary focus of interest during infectious disease, we have used COVID-19 to ask whether their emergence comes at a cost of broader B cell and T cell repertoire disruption. We applied a genomic DNA-based approach to concurrently study the immunoglobulin-heavy (IGH) and T cell receptor (TCR) ß and δ chain loci of 95 individuals. Our approach detected anticipated repertoire focusing for the IGH repertoire, including expansions of clusters of related sequences temporally aligned with SARS-CoV-2-specific seroconversion, and enrichment of some shared SARS-CoV-2-associated sequences. No significant age-related or disease severity-related deficiencies were noted for the IGH repertoire. By contrast, whereas focusing occurred at the TCRß and TCRδ loci, including some TCRß sequence-sharing, disruptive repertoire narrowing was almost entirely limited to many patients aged older than 50 y. By temporarily reducing T cell diversity and by risking expansions of nonbeneficial T cells, these traits may constitute an age-related risk factor for COVID-19, including a vulnerability to new variants for which T cells may provide key protection.

Upper airway gene expression shows a more robust adaptive immune response to SARS-CoV-2 in children.

Unlike other respiratory viruses, SARS-CoV-2 disproportionately causes severe disease in older adults whereas disease burden in children is lower. To investigate whether differences in the upper airway immune response may contribute to this disparity, we compare nasopharyngeal gene expression in 83 children (<19-years-old; 38 with SARS-CoV-2, 11 with other respiratory viruses, 34 with no virus) and 154 older adults (>40-years-old; 45 with SARS-CoV-2, 28 with other respiratory viruses, 81 with no virus). Expression of interferon-stimulated genes is robustly activated in both children and adults with SARS-CoV-2 infection compared to the respective non-viral groups, with only subtle distinctions. Children, however, demonstrate markedly greater upregulation of pathways related to B cell and T cell activation and proinflammatory cytokine signaling, including response to TNF and production of IFNγ, IL-2 and IL-4. Cell type deconvolution confirms greater recruitment of B cells, and to a lesser degree macrophages, to the upper airway of children. Only children exhibit a decrease in proportions of ciliated cells, among the primary targets of SARS-CoV-2, upon infection. These findings demonstrate that children elicit a more robust innate and especially adaptive immune response to SARS-CoV-2 in the upper airway that likely contributes to their protection from severe disease in the lower airway.

The Impact of IFNλ4 on the Adaptive Immune Response to SARS-CoV-2 Infection.

Genetic polymorphisms at the IFNL4 loci are known to influence the clinical outcome of several different infectious diseases. Best described is the association between the IFNL4 genotype and hepatitis C virus clearance. However, an influence of the IFNL4 genotype on the adaptive immune system was suggested by several studies but never investigated in humans. In this cross-sectional study, we have genotyped 201 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-positive participants for 3 IFNL4 polymorphisms (rs368234815, rs12979860, and rs117648444) and stratified them according to the IFNλ4 activity. Based on this stratification, we investigated the association between the IFNL4 genotype and the antibody as well as the CD8+ T cell response in the acute phase of the SARS-CoV-2 infection. We observed no differences in the genotype distribution compared with a Danish reference cohort or the 1,000 Genome Project, and we were not able to link the IFNL4 genotype to changes in either the antibody or CD8+ T cell responses of these patients.

T-Cell Repertoire Characteristics of Asymptomatic and Re-Detectable Positive COVID-19 Patients.

The prevention of the COVID-19 pandemic is highly complicated by the prevalence of asymptomatic and recurrent infection. Many previous immunological studies have focused on symptomatic and convalescent patients, while the immune responses in asymptomatic patients and re-detectable positive cases remain unclear. Here we comprehensively analyzed the peripheral T-cell receptor (TCR) repertoire of 54 COVID-19 patients in different courses, including asymptomatic, symptomatic, convalescent, and re-detectable positive cases. We identified a set of V-J gene combinations characterizing the upward immune responses through asymptomatic and symptomatic courses. Furthermore, some of these V-J combinations could be awakened in the re-detectable positive cases, which may help predict the risk of recurrent infection. Therefore, TCR repertoire examination has the potential to strengthen the clinical surveillance and the immunotherapy development for COVID-19.

Multiple early factors anticipate post-acute COVID-19 sequelae.

Post-acute sequelae of COVID-19 (PASC) represent an emerging global crisis. However, quantifiable risk factors for PASC and their biological associations are poorly resolved. We executed a deep multi-omic, longitudinal investigation of 309 COVID-19 patients from initial diagnosis to convalescence (2-3 months later), integrated with clinical data and patient-reported symptoms. We resolved four PASC-anticipating risk factors at the time of initial COVID-19 diagnosis: type 2 diabetes, SARS-CoV-2 RNAemia, Epstein-Barr virus viremia, and specific auto-antibodies. In patients with gastrointestinal PASC, SARS-CoV-2-specific and CMV-specific CD8+ T cells exhibited unique dynamics during recovery from COVID-19. Analysis of symptom-associated immunological signatures revealed coordinated immunity polarization into four endotypes, exhibiting divergent acute severity and PASC. We find that immunological associations between PASC factors diminish over time, leading to distinct convalescent immune states. Detectability of most PASC factors at COVID-19 diagnosis emphasizes the importance of early disease measurements for understanding emergent chronic conditions and suggests PASC treatment strategies.

Single-cell multi-omics reveals dyssynchrony of the innate and adaptive immune system in progressive COVID-19.

Dysregulated immune responses against the SARS-CoV-2 virus are instrumental in severe COVID-19. However, the immune signatures associated with immunopathology are poorly understood. Here we use multi-omics single-cell analysis to probe the dynamic immune responses in hospitalized patients with stable or progressive course of COVID-19, explore V(D)J repertoires, and assess the cellular effects of tocilizumab. Coordinated profiling of gene expression and cell lineage protein markers shows that S100Ahi/HLA-DRlo classical monocytes and activated LAG-3hi T cells are hallmarks of progressive disease and highlights the abnormal MHC-II/LAG-3 interaction on myeloid and T cells, respectively. We also find skewed T cell receptor repertories in expanded effector CD8+ clones, unmutated IGHG+ B cell clones, and mutated B cell clones with stable somatic hypermutation frequency over time. In conclusion, our in-depth immune profiling reveals dyssynchrony of the innate and adaptive immune interaction in progressive COVID-19.

Time-Resolved Gene Expression Analysis Monitors the Regulation of Inflammatory Mediators and Attenuation of Adaptive Immune Response by Vitamin D.

Peripheral blood mononuclear cells (PBMCs) belong to the innate and adaptive immune system and are highly sensitive and responsive to changes in their systemic environment. In this study, we focused on the time course of transcriptional changes in freshly isolated human PBMCs 4, 8, 24 and 48 h after onset of stimulation with the active vitamin D metabolite 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3). Taking all four time points together, 662 target genes were identified and segregated either by time of differential gene expression into 179 primary and 483 secondary targets or by driver of expression change into 293 direct and 369 indirect targets. The latter classification revealed that more than 50% of target genes were primarily driven by the cells' response to ex vivo exposure than by the nuclear hormone and largely explained its down-regulatory effect. Functional analysis indicated vitamin D's role in the suppression of the inflammatory and adaptive immune response by down-regulating ten major histocompatibility complex class II genes, five alarmins of the S100 calcium binding protein A family and by affecting six chemokines of the C-X-C motif ligand family. Taken together, studying time-resolved responses allows to better contextualize the effects of vitamin D on the immune system.

Single-cell profiling of T and B cell repertoires following SARS-CoV-2 mRNA vaccine.

mRNA vaccines for SARS-CoV-2 have shown exceptional clinical efficacy, providing robust protection against severe disease. However, our understanding of transcriptional and repertoire changes following full vaccination remains incomplete. We used scRNA-Seq and functional assays to compare humoral and cellular responses to 2 doses of mRNA vaccine with responses observed in convalescent individuals with asymptomatic disease. Our analyses revealed enrichment of spike-specific B cells, activated CD4+ T cells, and robust antigen-specific polyfunctional CD4+ T cell responses following vaccination. On the other hand, although clonally expanded CD8+ T cells were observed following both vaccination and natural infection, CD8+ T cell responses were relatively weak and variable. In addition, TCR gene usage was variable, reflecting the diversity of repertoires and MHC polymorphism in the human population. Natural infection induced expansion of CD8+ T cell clones that occupy distinct clusters compared to those induced by vaccination and likely recognize a broader set of viral antigens of viral epitopes presented by the virus not seen in the mRNA vaccine. Our study highlights a coordinated adaptive immune response in which early CD4+ T cell responses facilitate the development of the B cell response and substantial expansion of effector CD8+ T cells, together capable of contributing to future recall responses.

Single-Cell Immunogenomic Approach Identified SARS-CoV-2 Protective Immune Signatures in Asymptomatic Direct Contacts of COVID-19 Cases.

The response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is largely impacted by the level of virus exposure and status of the host immunity. The nature of protection shown by direct asymptomatic contacts of coronavirus disease 2019 (COVID-19)-positive patients is quite intriguing. In this study, we have characterized the antibody titer, SARS-CoV-2 surrogate virus neutralization, cytokine levels, single-cell T-cell receptor (TCR), and B-cell receptor (BCR) profiling in asymptomatic direct contacts, infected cases, and controls. We observed significant increase in antibodies with neutralizing amplitude in asymptomatic contacts along with cytokines such as Eotaxin, granulocyte-colony stimulating factor (G-CSF), interleukin 7 (IL-7), migration inhibitory factor (MIF), and macrophage inflammatory protein-1α (MIP-1α). Upon single-cell RNA (scRNA) sequencing, we explored the dynamics of the adaptive immune response in few representative asymptomatic close contacts and COVID-19-infected patients. We reported direct asymptomatic contacts to have decreased CD4+ naive T cells with concomitant increase in CD4+ memory and CD8+ Temra cells along with expanded clonotypes compared to infected patients. Noticeable proportions of class switched memory B cells were also observed in them. Overall, these findings gave an insight into the nature of protection in asymptomatic contacts.

Severe Adaptive Immune Suppression May Be Why Patients With Severe COVID-19 Cannot Be Discharged From the ICU Even After Negative Viral Tests.

During the COVID-19 pandemic, a phenomenon emerged in which some patients with severe disease were critically ill and could not be discharged from the ICU even though they exhibited negative viral tests. To explore the underlying mechanism, we collected blood samples from these patients and analyzed the gene expression profiles of peripheral immune cells. We found that all enrolled patients, regardless of changes in genes related to different symptoms and inflammatory responses, showed universally and severely decreased expression of adaptive immunity-related genes, especially those related to T/B cell arms and HLA molecules, and that these patients exhibited long-term secondary infections. In addition, no significant change was found in the expression of classic immunosuppression molecules including PD-1, PD-L1, and CTLA-4, suggesting that the adaptive immune suppression may not be due to the change of these genes. According to the published literatures and our data, this adaptive immunosuppression is likely to be caused by the "dysregulated host response" to severe infection, similar to the immunosuppression that exists in other severely infected patients with sepsis.

Normal T and B Cell Responses Against SARS-CoV-2 in a Family With a Non-Functional Vitamin D Receptor: A Case Report.

The coronavirus disease 2019 (COVID-19) pandemic has severely impacted daily life all over the world. Any measures to slow down the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and to decrease disease severity are highly requested. Recent studies have reported inverse correlations between plasma levels of vitamin D and susceptibility to SARS-CoV-2 infection and COVID-19 severity. Therefore, it has been proposed to supplement the general population with vitamin D to reduce the impact of COVID-19. However, by studying the course of COVID-19 and the immune response against SARS-CoV-2 in a family with a mutated, non-functional vitamin D receptor, we here demonstrate that vitamin D signaling was dispensable for mounting an efficient adaptive immune response against SARS-CoV-2 in this family. Although these observations might not directly be transferred to the general population, they question a central role of vitamin D in the generation of adaptive immunity against SARS-CoV-2.

Immune Signature Linked to COVID-19 Severity: A SARS-Score for Personalized Medicine.

SARS-CoV-2 infection leads to a highly variable clinical evolution, ranging from asymptomatic to severe disease with acute respiratory distress syndrome, requiring intensive care units (ICU) admission. The optimal management of hospitalized patients has become a worldwide concern and identification of immune biomarkers predictive of the clinical outcome for hospitalized patients remains a major challenge. Immunophenotyping and transcriptomic analysis of hospitalized COVID-19 patients at admission allow identifying the two categories of patients. Inflammation, high neutrophil activation, dysfunctional monocytic response and a strongly impaired adaptive immune response was observed in patients who will experience the more severe form of the disease. This observation was validated in an independent cohort of patients. Using in silico analysis on drug signature database, we identify differential therapeutics that specifically correspond to each group of patients. From this signature, we propose a score-the SARS-Score-composed of easily quantifiable biomarkers, to classify hospitalized patients upon arrival to adapt treatment according to their immune profile.

The role of PD-L1 in the immune dysfunction that mediates hypoxia-induced multiple organ injury.

Hypoxia is a pathological condition common to many diseases, although multiple organ injuries induced by hypoxia are often overlooked. There is increasing evidence to suggest that the hypoxic environment may activate innate immune cells and suppress adaptive immunity, further stimulating inflammation and inhibiting immunosurveillance. We found that dysfunctional immune regulation may aggravate hypoxia-induced tissue damage and contribute to secondary injury. Among the diverse mechanisms of hypoxia-induced immune dysfunction identified to date, the role of programmed death-ligand 1 (PD-L1) has recently attracted much attention. Besides leading to tumour immune evasion, PD-L1 has also been found to participate in the progression of the immune dysfunction which mediates hypoxia-induced multiple organ injury. In this review, we aimed to summarise the role of immune dysfunction in hypoxia-induced multiple organ injury, the effects of hypoxia on the cellular expression of PD-L1, and the effects of upregulated PD-L1 expression on immune regulation. Furthermore, we summarise the latest information pertaining to the involvement, diagnostic value, and therapeutic potential of immunosuppression induced by PD-L1 in various types of hypoxia-related diseases, including cancers, ischemic stroke, acute kidney injury, and obstructive sleep apnoea. Video Abstract.

Innate and Adaptive Immune Genes Associated with MERS-CoV Infection in Dromedaries.

The recent SARS-CoV-2 pandemic has refocused attention to the betacoronaviruses, only eight years after the emergence of another zoonotic betacoronavirus, the Middle East respiratory syndrome coronavirus (MERS-CoV). While the wild source of SARS-CoV-2 may be disputed, for MERS-CoV, dromedaries are considered as source of zoonotic human infections. Testing 100 immune-response genes in 121 dromedaries from United Arab Emirates (UAE) for potential association with present MERS-CoV infection, we identified candidate genes with important functions in the adaptive, MHC-class I (HLA-A-24-like) and II (HLA-DPB1-like), and innate immune response (PTPN4, MAGOHB), and in cilia coating the respiratory tract (DNAH7). Some of these genes previously have been associated with viral replication in SARS-CoV-1/-2 in humans, others have an important role in the movement of bronchial cilia. These results suggest similar host genetic pathways associated with these betacoronaviruses, although further work is required to better understand the MERS-CoV disease dynamics in both dromedaries and humans.

Immune transcriptomes of highly exposed SARS-CoV-2 asymptomatic seropositive versus seronegative individuals from the Ischgl community.

SARS-CoV-2 infection ranges from asymptomatic to severe with lingering symptomatology in some. This prompted investigation of whether or not asymptomatic disease results in measurable immune activation post-infection. Immune activation following asymptomatic SARS-CoV-2 infection was characterized through a comparative investigation of the immune cell transcriptomes from 43 asymptomatic seropositive and 52 highly exposed seronegative individuals from the same community 4-6 weeks following a superspreading event. Few of the 95 individuals had underlying health issues. One seropositive individual reported Cystic Fibrosis and one individual reported Incontinentia pigmenti. No evidence of immune activation was found in asymptomatic seropositive individuals with the exception of the Cystic Fibrosis patient. There were no statistically significant differences in immune transcriptomes between asymptomatic seropositive and highly exposed seronegative individuals. Four positive controls, mildly symptomatic seropositive individuals whose blood was examined 3 weeks following infection, showed immune activation. Negative controls were four seronegative individuals from neighboring communities without COVID-19. All individuals remained in their usual state of health through a five-month follow-up after sample collection. In summary, whole blood transcriptomes identified individual immune profiles within a community population and showed that asymptomatic infection within a super-spreading event was not associated with enduring immunological activation.

Genetic variation analyses indicate conserved SARS-CoV-2-host interaction and varied genetic adaptation in immune response factors in modern human evolution.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a pandemic as of early 2020. Upon infection, SARS-CoV-2 attaches to its receptor, that is, angiotensin-converting enzyme 2 (ACE2), on the surface of host cells and is then internalized into host cells via enzymatic machineries. This subsequently stimulates immune response factors. Since the host immune response and severity of COVID-19 vary among individuals, genetic risk factors for severe COVID-19 cases have been investigated. Our research group recently conducted a survey of genetic variants among SARS-CoV-2-interacting molecules across populations, noting near absence of difference in allele frequency spectrum between populations in these genes. Recent genome-wide association studies have identified genetic risk factors for severe COVID-19 cases in a segment of chromosome 3 that involves six genes encoding three immune-regulatory chemokine receptors and another three molecules. The risk haplotype seemed to be inherited from Neanderthals, suggesting genetic adaptation against pathogens in modern human evolution. Therefore, SARS-CoV-2 uses highly conserved molecules as its virion interaction, whereas its immune response appears to be genetically biased in individuals to some extent. We herein review the molecular process of SARS-CoV-2 infection as well as our further survey of genetic variants of its related immune effectors. We also discuss aspects of modern human evolution.

Sex differences in COVID-19: candidate pathways, genetics of ACE2, and sex hormones.

Biological sex is increasingly recognized as a critical determinant of health and disease, particularly relevant to the topical COVID-19 pandemic caused by the SARS-CoV-2 coronavirus. Epidemiological data and observational reports from both the original SARS epidemic and the most recent COVID-19 pandemic have a common feature: males are more likely to exhibit enhanced disease severity and mortality than females. Sex differences in cardiovascular disease and COVID-19 share mechanistic foundations, namely, the involvement of both the innate immune system and the canonical renin-angiotensin system (RAS). Immunological differences suggest that females mount a rapid and aggressive innate immune response, and the attenuated antiviral response in males may confer enhanced susceptibility to severe disease. Furthermore, the angiotensin-converting enzyme 2 (ACE2) is involved in disease pathogenesis in cardiovascular disease and COVID-19, either to serve as a protective mechanism by deactivating the RAS or as the receptor for viral entry, respectively. Loss of membrane ACE2 and a corresponding increase in plasma ACE2 are associated with worsened cardiovascular disease outcomes, a mechanism attributed to a disintegrin and metalloproteinase (ADAM17). SARS-CoV-2 infection also leads to ADAM17 activation, a positive feedback cycle that exacerbates ACE2 loss. Therefore, the relationship between cardiovascular disease and COVID-19 is critically dependent on the loss of membrane ACE2 by ADAM17-mediated proteolytic cleavage. This article explores potential mechanisms involved in COVID-19 that may contribute to sex-specific susceptibility focusing on the innate immune system and the RAS, namely, genetics and sex hormones. Finally, we highlight here the added challenges of gender in the COVID-19 pandemic.