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The impact of adjuvants on malaria vaccine-induced antibody repertoire is poorly understood. Here, we characterize the impact of two adjuvants, Alhydrogel® and AS01, on antibody clonotype diversity, binding and function, post malaria vaccination. We expressed 132 recombinant anti-Pfs230D1 human monoclonal antibodies (mAbs) from participants immunized with malaria transmission-blocking vaccine Pfs230D1, formulated with either Alhydrogel® or AS01. Anti-Pfs230D1 mAbs generated by Alhydrogel® formulation showed higher binding frequency to Pfs230D1 compared to AS01 formulation, although the frequency of functional mAbs was similar between adjuvant groups. Overall, the AS01 formulation induced anti-Pfs230D1 functional antibodies from a broader array of germline sequences versus the Alhydrogel® formulation. All mAbs using IGHV1-69 gene from the Alhydrogel® cohort bound to recombinant Pfs230D1, but did not block parasite transmission to mosquitoes, similar to the IGHV1-69 mAbs isolated from the AS01 cohort. These findings may help inform vaccine design and adjuvant selection for immunization with Plasmodium antigens.
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Adaptive Immune Receptor Repertoire sequencing (AIRR-seq) is a valuable experimental tool to study the immune state in health and following immune challenges such as infectious diseases, (auto)immune diseases, and cancer. Several tools have been developed to reconstruct B cell and T cell receptor sequences from AIRR-seq data and infer B and T cell clonal relationships. However, currently available tools offer limited parallelization across samples, scalability or portability to high-performance computing infrastructures. To address this need, we developed nf-core/airrflow, an end-to-end bulk and single-cell AIRR-seq processing workflow which integrates the Immcantation Framework following BCR and TCR sequencing data analysis best practices. The Immcantation Framework is a comprehensive toolset, which allows the processing of bulk and single-cell AIRR-seq data from raw read processing to clonal inference. nf-core/airrflow is written in Nextflow and is part of the nf-core project, which collects community contributed and curated Nextflow workflows for a wide variety of analysis tasks. We assessed the performance of nf-core/airrflow on simulated sequencing data with sequencing errors and show example results with real datasets. To demonstrate the applicability of nf-core/airrflow to the high-throughput processing of large AIRR-seq datasets, we validated and extended previously reported findings of convergent antibody responses to SARS-CoV-2 by analyzing 97 COVID-19 infected individuals and 99 healthy controls, including a mixture of bulk and single-cell sequencing datasets. Using this dataset, we extended the convergence findings to 20 additional subjects, highlighting the applicability of nf-core/airrflow to validate findings in small in-house cohorts with reanalysis of large publicly available AIRR datasets.
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COVID-19 , Biologia Computacional , Receptores de Antígenos de Linfócitos T , SARS-CoV-2 , Fluxo de Trabalho , Humanos , COVID-19/imunologia , COVID-19/virologia , COVID-19/genética , SARS-CoV-2/imunologia , SARS-CoV-2/genética , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Biologia Computacional/métodos , Receptores de Antígenos de Linfócitos B/genética , Receptores de Antígenos de Linfócitos B/imunologia , Software , Análise de Célula Única/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Imunidade Adaptativa/genética , Linfócitos B/imunologia , Linfócitos T/imunologiaRESUMO
The emergence of highly contagious and immune-evasive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants has required reformulation of coronavirus disease 2019 (COVID-19) vaccines to target those new variants specifically. While previous infections and booster vaccinations can enhance variant neutralization, it is unclear whether the monovalent version, administered using either mRNA or protein-based vaccine platforms, can elicit de novo B-cell responses specific for Omicron XBB.1.5 variants. Here, we dissected the genetic antibody repertoire of 603 individual plasmablasts derived from five individuals who received a monovalent XBB.1.5 vaccination either with mRNA (Moderna or Pfizer/BioNtech) or adjuvanted protein (Novavax). From these sequences, we expressed 100 human monoclonal antibodies and determined binding, affinity and protective potential against several SARS-CoV-2 variants, including JN.1. We then select two vaccine-induced XBB.1.5 mAbs, M2 and M39. M2 mAb was a de novo, antibody, i.e., specific for XBB.1.5 but not ancestral SARS-CoV-2. M39 bound and neutralized both XBB.1.5 and JN.1 strains. Our high-resolution cryo-electron microscopy (EM) structures of M2 and M39 in complex with the XBB.1.5 spike glycoprotein defined the epitopes engaged and revealed the molecular determinants for the mAbs' specificity. These data show, at the molecular level, that monovalent, variant-specific vaccines can elicit functional antibodies, and shed light on potential functional and genetic differences of mAbs induced by vaccinations with different vaccine platforms.\.
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The differentiation of naive and memory B cells into antibody-secreting cells (ASCs) is a key feature of adaptive immunity. The requirement for phosphoinositide 3-kinase-delta (PI3Kδ) to support B cell biology has been investigated intensively; however, specific functions of the related phosphoinositide 3-kinase-gamma (PI3Kγ) complex in B lineage cells have not. In the present study, we report that PI3Kγ promotes robust antibody responses induced by T cell-dependent antigens. The inborn error of immunity caused by human deficiency in PI3Kγ results in broad humoral defects, prompting our investigation of roles for this kinase in antibody responses. Using mouse immunization models, we found that PI3Kγ functions cell intrinsically within activated B cells in a kinase activity-dependent manner to transduce signals required for the transcriptional program supporting differentiation of ASCs. Furthermore, ASC fate choice coincides with upregulation of PIK3CG expression and is impaired in the context of PI3Kγ disruption in naive B cells on in vitro CD40-/cytokine-driven activation, in memory B cells on toll-like receptor activation, or in human tonsillar organoids. Taken together, our study uncovers a fundamental role for PI3Kγ in supporting humoral immunity by integrating signals instructing commitment to the ASC fate.
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Formação de Anticorpos , Linfócitos B , Diferenciação Celular , Classe Ib de Fosfatidilinositol 3-Quinase , Animais , Classe Ib de Fosfatidilinositol 3-Quinase/metabolismo , Classe Ib de Fosfatidilinositol 3-Quinase/imunologia , Camundongos , Diferenciação Celular/imunologia , Humanos , Linfócitos B/imunologia , Linfócitos B/metabolismo , Formação de Anticorpos/imunologia , Camundongos Knockout , Células Produtoras de Anticorpos/imunologia , Ativação Linfocitária/imunologia , Camundongos Endogâmicos C57BL , Transdução de Sinais/imunologia , Células B de Memória/imunologia , Células B de Memória/metabolismoRESUMO
Immunological health has been challenging to characterize but could be defined as the absence of immune pathology. While shared features of some immune diseases and the concept of immunologic resilience based on age-independent adaptation to antigenic stimulation have been developed, general metrics of immune health and its utility for assessing clinically healthy individuals remain ill defined. Here we integrated transcriptomics, serum protein, peripheral immune cell frequency and clinical data from 228 patients with 22 monogenic conditions impacting key immunological pathways together with 42 age- and sex-matched healthy controls. Despite the high penetrance of monogenic lesions, differences between individuals in diverse immune parameters tended to dominate over those attributable to disease conditions or medication use. Unsupervised or supervised machine learning independently identified a score that distinguished healthy participants from patients with monogenic diseases, thus suggesting a quantitative immune health metric (IHM). In ten independent datasets, the IHM discriminated healthy from polygenic autoimmune and inflammatory disease states, marked aging in clinically healthy individuals, tracked disease activities and treatment responses in both immunological and nonimmunological diseases, and predicted age-dependent antibody responses to immunizations with different vaccines. This discriminatory power goes beyond that of the classical inflammatory biomarkers C-reactive protein and interleukin-6. Thus, deviations from health in diverse conditions, including aging, have shared systemic immune consequences, and we provide a web platform for calculating the IHM for other datasets, which could empower precision medicine.
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Germinal centers (GC) are microanatomical lymphoid structures where affinity-matured memory B cells and long-lived bone marrow plasma cells are primarily generated. It is unclear how the maturation of B cells within the GC impacts the breadth and durability of B cell responses to influenza vaccination in humans. We used fine needle aspiration of draining lymph nodes to longitudinally track antigen-specific GC B cell responses to seasonal influenza vaccination. Antigen-specific GC B cells persisted for at least 13 wk after vaccination in two out of seven individuals. Monoclonal antibodies (mAbs) derived from persisting GC B cell clones exhibit enhanced binding affinity and breadth to influenza hemagglutinin (HA) antigens compared with related GC clonotypes isolated earlier in the response. Structural studies of early and late GC-derived mAbs from one clonal lineage in complex with H1 and H5 HAs revealed an altered binding footprint. Our study shows that inducing sustained GC reactions after influenza vaccination in humans supports the maturation of responding B cells.
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Linfócitos B , Centro Germinativo , Vacinas contra Influenza , Vacinação , Centro Germinativo/imunologia , Humanos , Vacinas contra Influenza/imunologia , Linfócitos B/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Influenza Humana/imunologia , Influenza Humana/prevenção & controle , Anticorpos Antivirais/imunologia , Anticorpos Monoclonais/imunologia , Adulto , Feminino , Masculino , Pessoa de Meia-IdadeRESUMO
Motivation: With the increased reliance on multi-omics data for bulk and single cell analyses, the availability of robust approaches to perform unsupervised analysis for clustering, visualization, and feature selection is imperative. Joint dimensionality reduction methods can be applied to multi-omics datasets to derive a global sample embedding analogous to single-omic techniques such as Principal Components Analysis (PCA). Multiple co-inertia analysis (MCIA) is a method for joint dimensionality reduction that maximizes the covariance between block- and global-level embeddings. Current implementations for MCIA are not optimized for large datasets such such as those arising from single cell studies, and lack capabilities with respect to embedding new data. Results: We introduce nipalsMCIA, an MCIA implementation that solves the objective function using an extension to Non-linear Iterative Partial Least Squares (NIPALS), and shows significant speed-up over earlier implementations that rely on eigendecompositions for single cell multi-omics data. It also removes the dependence on an eigendecomposition for calculating the variance explained, and allows users to perform out-of-sample embedding for new data. nipalsMCIA provides users with a variety of pre-processing and parameter options, as well as ease of functionality for down-stream analysis of single-omic and global-embedding factors. Availability: nipalsMCIA is available as a BioConductor package at https://bioconductor.org/packages/release/bioc/html/nipalsMCIA.html, and includes detailed documentation and application vignettes. Supplementary Materials are available online.
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BACKGROUNDPatients hospitalized for COVID-19 exhibit diverse clinical outcomes, with outcomes for some individuals diverging over time even though their initial disease severity appears similar to that of other patients. A systematic evaluation of molecular and cellular profiles over the full disease course can link immune programs and their coordination with progression heterogeneity.METHODSWe performed deep immunophenotyping and conducted longitudinal multiomics modeling, integrating 10 assays for 1,152 Immunophenotyping Assessment in a COVID-19 Cohort (IMPACC) study participants and identifying several immune cascades that were significant drivers of differential clinical outcomes.RESULTSIncreasing disease severity was driven by a temporal pattern that began with the early upregulation of immunosuppressive metabolites and then elevated levels of inflammatory cytokines, signatures of coagulation, formation of neutrophil extracellular traps, and T cell functional dysregulation. A second immune cascade, predictive of 28-day mortality among critically ill patients, was characterized by reduced total plasma Igs and B cells and dysregulated IFN responsiveness. We demonstrated that the balance disruption between IFN-stimulated genes and IFN inhibitors is a crucial biomarker of COVID-19 mortality, potentially contributing to failure of viral clearance in patients with fatal illness.CONCLUSIONOur longitudinal multiomics profiling study revealed temporal coordination across diverse omics that potentially explain the disease progression, providing insights that can inform the targeted development of therapies for patients hospitalized with COVID-19, especially those who are critically ill.TRIAL REGISTRATIONClinicalTrials.gov NCT04378777.FUNDINGNIH (5R01AI135803-03, 5U19AI118608-04, 5U19AI128910-04, 4U19AI090023-11, 4U19AI118610-06, R01AI145835-01A1S1, 5U19AI062629-17, 5U19AI057229-17, 5U19AI125357-05, 5U19AI128913-03, 3U19AI077439-13, 5U54AI142766-03, 5R01AI104870-07, 3U19AI089992-09, 3U19AI128913-03, and 5T32DA018926-18); NIAID, NIH (3U19AI1289130, U19AI128913-04S1, and R01AI122220); and National Science Foundation (DMS2310836).
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COVID-19 , Índice de Gravidade de Doença , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , COVID-19/imunologia , COVID-19/mortalidade , COVID-19/sangue , Citocinas/sangue , Citocinas/imunologia , Estudos Longitudinais , MultiômicaRESUMO
Antibodies play a crucial role in adaptive immune responses by determining B cell specificity to antigens and focusing immune function on target pathogens. Accurate prediction of antibody-antigen specificity directly from antibody sequencing data would be a great aid in understanding immune responses, guiding vaccine design, and developing antibody-based therapeutics. In this study, we present a method of supervised fine-tuning for antibody language models, which improves on previous results in binding specificity prediction to SARS-CoV-2 spike protein and influenza hemagglutinin. We perform supervised fine-tuning on four pre-trained antibody language models to predict specificity to these antigens and demonstrate that fine-tuned language model classifiers exhibit enhanced predictive accuracy compared to classifiers trained on pre-trained model embeddings. The change of model attention activations after supervised fine-tuning suggested that this performance was driven by an increased model focus on the complementarity determining regions (CDRs). Application of the supervised fine-tuned models to BCR repertoire data demonstrated that these models could recognize the specific responses elicited by influenza and SARS-CoV-2 vaccination. Overall, our study highlights the benefits of supervised fine-tuning on pre-trained antibody language models as a mechanism to improve antigen specificity prediction.
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Age is a major risk factor for severe coronavirus disease 2019 (COVID-19), yet the mechanisms behind this relationship have remained incompletely understood. To address this, we evaluated the impact of aging on host immune response in the blood and the upper airway, as well as the nasal microbiome in a prospective, multicenter cohort of 1031 vaccine-naïve patients hospitalized for COVID-19 between 18 and 96 years old. We performed mass cytometry, serum protein profiling, anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody assays, and blood and nasal transcriptomics. We found that older age correlated with increased SARS-CoV-2 viral abundance upon hospital admission, delayed viral clearance, and increased type I interferon gene expression in both the blood and upper airway. We also observed age-dependent up-regulation of innate immune signaling pathways and down-regulation of adaptive immune signaling pathways. Older adults had lower naïve T and B cell populations and higher monocyte populations. Over time, older adults demonstrated a sustained induction of pro-inflammatory genes and serum chemokines compared with younger individuals, suggesting an age-dependent impairment in inflammation resolution. Transcriptional and protein biomarkers of disease severity differed with age, with the oldest adults exhibiting greater expression of pro-inflammatory genes and proteins in severe disease. Together, our study finds that aging is associated with impaired viral clearance, dysregulated immune signaling, and persistent and potentially pathologic activation of pro-inflammatory genes and proteins.
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COVID-19 , Humanos , Idoso , Adolescente , Adulto Jovem , Adulto , Pessoa de Meia-Idade , Idoso de 80 Anos ou mais , SARS-CoV-2 , Estudos Prospectivos , Multiômica , QuimiocinasRESUMO
MOTIVATION: Predictive biological signatures provide utility as biomarkers for disease diagnosis and prognosis, as well as prediction of responses to vaccination or therapy. These signatures are identified from high-throughput profiling assays through a combination of dimensionality reduction and machine learning techniques. The genes, proteins, metabolites, and other biological analytes that compose signatures also generate hypotheses on the underlying mechanisms driving biological responses, thus improving biological understanding. Dimensionality reduction is a critical step in signature discovery to address the large number of analytes in omics datasets, especially for multi-omics profiling studies with tens of thousands of measurements. Latent factor models, which can account for the structural heterogeneity across diverse assays, effectively integrate multi-omics data and reduce dimensionality to a small number of factors that capture correlations and associations among measurements. These factors provide biologically interpretable features for predictive modeling. However, multi-omics integration and predictive modeling are generally performed independently in sequential steps, leading to suboptimal factor construction. Combining these steps can yield better multi-omics signatures that are more predictive while still being biologically meaningful. RESULTS: We developed a supervised variational Bayesian factor model that extracts multi-omics signatures from high-throughput profiling datasets that can span multiple data types. Signature-based multiPle-omics intEgration via lAtent factoRs (SPEAR) adaptively determines factor rank, emphasis on factor structure, data relevance and feature sparsity. The method improves the reconstruction of underlying factors in synthetic examples and prediction accuracy of coronavirus disease 2019 severity and breast cancer tumor subtypes. AVAILABILITY AND IMPLEMENTATION: SPEAR is a publicly available R-package hosted at https://bitbucket.org/kleinstein/SPEAR.
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Teorema de Bayes , Biologia Computacional , Multiômica , Humanos , Biologia Computacional/métodos , Genômica/métodos , Aprendizado de Máquina SupervisionadoRESUMO
Abs are vital to human immune responses and are composed of genetically variable H and L chains. These structures are initially expressed as BCRs. BCR diversity is shaped through somatic hypermutation and selection during immune responses. This evolutionary process produces B cell clones, cells that descend from a common ancestor but differ by mutations. Phylogenetic trees inferred from BCR sequences can reconstruct the history of mutations within a clone. Until recently, BCR sequencing technologies separated H and L chains, but advancements in single-cell sequencing now pair H and L chains from individual cells. However, it is unclear how these separate genes should be combined to infer B cell phylogenies. In this study, we investigated strategies for using paired H and L chain sequences to build phylogenetic trees. We found that incorporating L chains significantly improved tree accuracy and reproducibility across all methods tested. This improvement was greater than the difference between tree-building methods and persisted even when mixing bulk and single-cell sequencing data. However, we also found that many phylogenetic methods estimated significantly biased branch lengths when some L chains were missing, such as when mixing single-cell and bulk BCR data. This bias was eliminated using maximum likelihood methods with separate branch lengths for H and L chain gene partitions. Thus, we recommend using maximum likelihood methods with separate H and L chain partitions, especially when mixing data types. We implemented these methods in the R package Dowser: https://dowser.readthedocs.io.
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Linfócitos B , Filogenia , Receptores de Antígenos de Linfócitos B , Humanos , Receptores de Antígenos de Linfócitos B/genética , Receptores de Antígenos de Linfócitos B/imunologia , Linfócitos B/imunologia , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Leves de Imunoglobulina/genética , Cadeias Leves de Imunoglobulina/imunologia , Análise de Célula Única/métodos , MutaçãoRESUMO
Systems vaccinology studies have identified factors affecting individual vaccine responses, but comparing these findings is challenging due to varying study designs. To address this lack of reproducibility, we established a community resource for comparing Bordetella pertussis booster responses and to host annual contests for predicting patients' vaccination outcomes. We report here on our experiences with the "dry-run" prediction contest. We found that, among 20+ models adopted from the literature, the most successful model predicting vaccination outcome was based on age alone. This confirms our concerns about the reproducibility of conclusions between different vaccinology studies. Further, we found that, for newly trained models, handling of baseline information on the target variables was crucial. Overall, multiple co-inertia analysis gave the best results of the tested modeling approaches. Our goal is to engage community in these prediction challenges by making data and models available and opening a public contest in August 2024.
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Multiômica , Vacinas , Humanos , Vacinologia/métodos , Reprodutibilidade dos Testes , Simulação por ComputadorRESUMO
Food allergy is caused by allergen-specific immunoglobulin E (IgE) antibodies, but little is known about the B cell memory of persistent IgE responses. Here, we describe, in human pediatric peanut allergy, a population of CD23+IgG1+ memory B cells arising in type 2 immune responses that contain high-affinity peanut-specific clones and generate IgE-producing cells upon activation. The frequency of CD23+IgG1+ memory B cells correlated with circulating concentrations of IgE in children with peanut allergy. A corresponding population of "type 2-marked" IgG1+ memory B cells was identified in single-cell RNA sequencing experiments. These cells differentially expressed interleukin-4 (IL-4)- and IL-13-regulated genes, such as FCER2/CD23+, IL4R, and germline IGHE, and carried highly mutated B cell receptors (BCRs). In children with high concentrations of serum peanut-specific IgE, high-affinity B cells that bind the main peanut allergen Ara h 2 mapped to the population of "type 2-marked" IgG1+ memory B cells and included clones with convergent BCRs across different individuals. Our findings indicate that CD23+IgG1+ memory B cells transcribing germline IGHE are a unique memory population containing precursors of high-affinity pathogenic IgE-producing cells that are likely to be involved in the long-term persistence of peanut allergy.
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Hipersensibilidade Alimentar , Hipersensibilidade a Amendoim , Humanos , Criança , Células B de Memória , Imunoglobulina G , Alérgenos , Imunoglobulina ERESUMO
Age is a major risk factor for severe coronavirus disease-2019 (COVID-19), yet the mechanisms responsible for this relationship have remained incompletely understood. To address this, we evaluated the impact of aging on host and viral dynamics in a prospective, multicenter cohort of 1,031 patients hospitalized for COVID-19, ranging from 18 to 96 years of age. We performed blood transcriptomics and nasal metatranscriptomics, and measured peripheral blood immune cell populations, inflammatory protein expression, anti-SARS-CoV-2 antibodies, and anti-interferon (IFN) autoantibodies. We found that older age correlated with an increased SARS-CoV-2 viral load at the time of admission, and with delayed viral clearance over 28 days. This contributed to an age-dependent increase in type I IFN gene expression in both the respiratory tract and blood. We also observed age-dependent transcriptional increases in peripheral blood IFN-γ, neutrophil degranulation, and Toll like receptor (TLR) signaling pathways, and decreases in T cell receptor (TCR) and B cell receptor signaling pathways. Over time, older adults exhibited a remarkably sustained induction of proinflammatory genes (e.g., CXCL6) and serum chemokines (e.g., CXCL9) compared to younger individuals, highlighting a striking age-dependent impairment in inflammation resolution. Augmented inflammatory signaling also involved the upper airway, where aging was associated with upregulation of TLR, IL17, type I IFN and IL1 pathways, and downregulation TCR and PD-1 signaling pathways. Metatranscriptomics revealed that the oldest adults exhibited disproportionate reactivation of herpes simplex virus and cytomegalovirus in the upper airway following hospitalization. Mass cytometry demonstrated that aging correlated with reduced naïve T and B cell populations, and increased monocytes and exhausted natural killer cells. Transcriptional and protein biomarkers of disease severity markedly differed with age, with the oldest adults exhibiting greater expression of TLR and inflammasome signaling genes, as well as proinflammatory proteins (e.g., IL6, CXCL8), in severe COVID-19 compared to mild/moderate disease. Anti-IFN autoantibody prevalence correlated with both age and disease severity. Taken together, this work profiles both host and microbe in the blood and airway to provide fresh insights into aging-related immune changes in a large cohort of vaccine-naïve COVID-19 patients. We observed age-dependent immune dysregulation at the transcriptional, protein and cellular levels, manifesting in an imbalance of inflammatory responses over the course of hospitalization, and suggesting potential new therapeutic targets.
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Post-acute sequelae of SARS-CoV-2 (PASC) is a significant public health concern. We describe Patient Reported Outcomes (PROs) on 590 participants prospectively assessed from hospital admission for COVID-19 through one year after discharge. Modeling identified 4 PRO clusters based on reported deficits (minimal, physical, mental/cognitive, and multidomain), supporting heterogenous clinical presentations in PASC, with sub-phenotypes associated with female sex and distinctive comorbidities. During the acute phase of disease, a higher respiratory SARS-CoV-2 viral burden and lower Receptor Binding Domain and Spike antibody titers were associated with both the physical predominant and the multidomain deficit clusters. A lower frequency of circulating B lymphocytes by mass cytometry (CyTOF) was observed in the multidomain deficit cluster. Circulating fibroblast growth factor 21 (FGF21) was significantly elevated in the mental/cognitive predominant and the multidomain clusters. Future efforts to link PASC to acute anti-viral host responses may help to better target treatment and prevention of PASC.
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Líquidos Corporais , COVID-19 , Feminino , Humanos , SARS-CoV-2 , COVID-19/complicações , Linfócitos B , Progressão da Doença , FenótipoRESUMO
Adaptive Immune Receptor Repertoire sequencing (AIRR-seq) is a valuable experimental tool to study the immune state in health and following immune challenges such as infectious diseases, (auto)immune diseases, and cancer. Several tools have been developed to reconstruct B cell and T cell receptor sequences from AIRR-seq data and infer B and T cell clonal relationships. However, currently available tools offer limited parallelization across samples, scalability or portability to high-performance computing infrastructures. To address this need, we developed nf-core/airrflow, an end-to-end bulk and single-cell AIRR-seq processing workflow which integrates the Immcantation Framework following BCR and TCR sequencing data analysis best practices. The Immcantation Framework is a comprehensive toolset, which allows the processing of bulk and single-cell AIRR-seq data from raw read processing to clonal inference. nf-core/airrflow is written in Nextflow and is part of the nf-core project, which collects community contributed and curated Nextflow workflows for a wide variety of analysis tasks. We assessed the performance of nf-core/airrflow on simulated sequencing data with sequencing errors and show example results with real datasets. To demonstrate the applicability of nf-core/airrflow to the high-throughput processing of large AIRR-seq datasets, we validated and extended previously reported findings of convergent antibody responses to SARS-CoV-2 by analyzing 97 COVID-19 infected individuals and 99 healthy controls, including a mixture of bulk and single-cell sequencing datasets. Using this dataset, we extended the convergence findings to 20 additional subjects, highlighting the applicability of nf-core/airrflow to validate findings in small in-house cohorts with reanalysis of large publicly available AIRR datasets. nf-core/airrflow is available free of charge, under the MIT license on GitHub (https://github.com/nf-core/airrflow). Detailed documentation and example results are available on the nf-core website at (https://nf-co.re/airrflow).
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The widespread availability of single-cell RNA sequencing (scRNA-seq) has led to the development of new methods for understanding immune responses. Single-cell transcriptome data can now be paired with B cell receptor (BCR) sequences. However, RNA from BCRs cannot be analyzed like most other genes because BCRs are genetically diverse within individuals. In humans, BCRs are shaped through recombination followed by mutation and selection for antigen binding. As these processes co-occur with cell division, B cells can be studied using phylogenetic trees representing the mutations within a clone. B cell trees can link experimental timepoints, tissues, or cellular subtypes. Here, we review the current state and potential of how B cell phylogenetics can be combined with single-cell data to understand immune responses.
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Linfócitos B , Receptores de Antígenos de Linfócitos B , Humanos , Filogenia , Receptores de Antígenos de Linfócitos B/genética , Imunidade Adaptativa , Mutação/genéticaRESUMO
High throughput sequencing of B cell receptors (BCRs) is increasingly applied to study the immense diversity of antibodies. Learning biologically meaningful embeddings of BCR sequences is beneficial for predictive modeling. Several embedding methods have been developed for BCRs, but no direct performance benchmarking exists. Moreover, the impact of the input sequence length and paired-chain information on the prediction remains to be explored. We evaluated the performance of multiple embedding models to predict BCR sequence properties and receptor specificity. Despite the differences in model architectures, most embeddings effectively capture BCR sequence properties and specificity. BCR-specific embeddings slightly outperform general protein language models in predicting specificity. In addition, incorporating full-length heavy chains and paired light chain sequences improves the prediction performance of all embeddings. This study provides insights into the properties of BCR embeddings to improve downstream prediction applications for antibody analysis and discovery.
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Processamento de Linguagem Natural , Receptores de Antígenos de Linfócitos B , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Imunoglobulinas , Receptores de Antígenos de Linfócitos B/química , Receptores de Antígenos de Linfócitos B/genética , Sequência de Aminoácidos , HumanosRESUMO
Infection with West Nile virus (WNV) drives a wide range of responses, from asymptomatic to flu-like symptoms/fever or severe cases of encephalitis and death. To identify cellular and molecular signatures distinguishing WNV severity, we employed systems profiling of peripheral blood from asymptomatic and severely ill individuals infected with WNV. We interrogated immune responses longitudinally from acute infection through convalescence employing single-cell protein and transcriptional profiling complemented with matched serum proteomics and metabolomics as well as multi-omics analysis. At the acute time point, we detected both elevation of pro-inflammatory markers in innate immune cell types and reduction of regulatory T cell activity in participants with severe infection, whereas asymptomatic donors had higher expression of genes associated with anti-inflammatory CD16+ monocytes. Therefore, we demonstrated the potential of systems immunology using multiple cell-type and cell-state-specific analyses to identify correlates of infection severity and host cellular activity contributing to an effective anti-viral response.