Exploring the impact of persistent mutations in SARS-CoV-2 variants and reduced immunity on breakthrough infections (BTIs) is crucial, particularly in understanding how antigen-specific memory B cells (MBCs) respond to new variants. We followed 107 participants who received the ancestral inactivated vaccine and experienced one or two Omicron BTIs over six months. Using flow cytometry, SARS-CoV-2 antigen probes, single-cell RNA sequencing, and B cell receptor (BCR) profiling, we assessed MBCs and immune diversity. Our findings revealed that although neutralizing antibody levels decreased over time, the number of specific MBCs remained stable and matured progressively. Notably, pre-existing Omicron-specific MBCs played a key role in preventing secondary Omicron infections. Differential gene analysis showed enrichment in antigen processing and immune regulation pathways, while clonal lineage analysis revealed more B cell expansion and V(D)J gene-specific rearrangements in high neutralization samples. These results emphasize MBCs' critical role in long-term immunity and inform future vaccination strategies.
© 2025 The Author(s).

Schistosomiasis affects over 250 million people, predominantly in Sub-Saharan Africa. In Schistosoma -endemic regions a lack of natural sterilizing immunity means individuals are repeatedly infected, treated and reinfected. Due to difficulties in tracking infection in endemic areas, the kinetics of the host immune response during these reinfections have not been elucidated. Here, we use repeated (3x) controlled human Schistosoma mansoni ( Sm ) infection (repeated CHI) to directly study how antigen-specific T cells develop during reinfection. We compared these responses to natural Sm -infected endemic Ugandan individuals. We report that a mixed Th1/Th2/regulatory CD4 + T cell response develops in repeated CHI, primarily against adult worm antigens. Adult worm-specific responses after repeated CHI were similar to those observed in naturally infected endemic participants. However, endemic participants showed differential responses to antigens from the egg and cercariae life-cycle stages. Repeated CHI (3x) with sequential exposure to Sm cercariae of different sexes (male-female-male) revealed an elevated CD4 + T cell cytokine response to adult worm and egg antigens. Our findings demonstrate that reinfection with single-sex schistosomes elicits adult worm-specific T cell cytokine responses that reflect endemic-natural infection, highlighting the translatability of the CHI model to natural infection in endemic settings. Overall, this study advances our understanding of how schistosome immune responses develop over reinfection cycles in the human host, thereby increasing our understanding of the immunology of natural schistosome (re)infection.

B cells are crucial components of the immune system, responsible for producing specific antibodies in response to infections and vaccines. Despite their uniform appearance, B cells display diverse surface molecules and functional properties, which are not yet fully understood. Apart from antibody production, B cells also play roles in antigen presentation and cytokine secretion, essential for initiating T-cell immune responses. Their significance as disease biomarkers and therapeutic targets has led to increased research focus. However, the lack of standardized protocols for B-cell identification and the variability in defining B-lymphocyte subpopulations pose some challenges. This paper proposes a B-cell identification panel throughout the evaluation of previous cytometry panels and nomenclature heterogeneity for B-cell subpopulations. Major subpopulations recognized in human peripheral blood include transitional, naive, switched memory, unswitched memory, double negative, and plasmablasts, characterized based on their functional and phenotypic features. We present a standardized flow cytometry protocol utilizing surface phenotypic markers (CD3, CD19, IgD, CD27, CD38, and CD24) to differentiate and analyze B-cell subpopulations. This practical and cost-effective panel can be used in various research and laboratory settings. The challenges of standardizing names and markers for classifying B-lymphocyte subpopulations are discussed, along with protocols utilizing multiple markers and gating strategies, allied with the importance of considering viability markers. In summary, this standardized protocol and panel provide a comprehensive approach to identifying B-cell subpopulations to enhance the reproducibility and comparability of B-cell subpopulation studies.
© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.

Severe defects in human IFNγ immunity predispose individuals to both Bacillus Calmette-Guérin disease and tuberculosis, whereas milder defects predispose only to tuberculosis1. Here we report two adults with recurrent pulmonary tuberculosis who are homozygous for a private loss-of-function TNF variant. Neither has any other clinical phenotype and both mount normal clinical and biological inflammatory responses. Their leukocytes, including monocytes and monocyte-derived macrophages (MDMs) do not produce TNF, even after stimulation with IFNγ. Blood leukocyte subset development is normal in these patients. However, an impairment in the respiratory burst was observed in granulocyte-macrophage colony-stimulating factor (GM-CSF)-matured MDMs and alveolar macrophage-like (AML) cells2 from both patients with TNF deficiency, TNF- or TNFR1-deficient induced pluripotent stem (iPS)-cell-derived GM-CSF-matured macrophages, and healthy control MDMs and AML cells differentiated with TNF blockers in vitro, and in lung macrophages treated with TNF blockers ex vivo. The stimulation of TNF-deficient iPS-cell-derived macrophages with TNF rescued the respiratory burst. These findings contrast with those for patients with inherited complete deficiency of the respiratory burst across all phagocytes, who are prone to multiple infections, including both Bacillus Calmette-Guérin disease and tuberculosis3. Human TNF is required for respiratory-burst-dependent immunity to Mycobacterium tuberculosis in macrophages but is surprisingly redundant otherwise, including for inflammation and immunity to weakly virulent mycobacteria and many other infectious agents.
© 2024. The Author(s).

T cells are the most common immune cells in atherosclerotic plaques, and the function of T cells can be altered by fatty acids. Here, we show that pre-exposure of CD4+ T cells to oleic acid, an abundant fatty acid linked to cardiovascular events, upregulates core metabolic pathways and promotes differentiation into interleukin-9 (IL-9)-producing cells upon activation. RNA sequencing of non-activated T cells reveals that oleic acid upregulates genes encoding key enzymes responsible for cholesterol and fatty acid biosynthesis. Transcription footprint analysis links these expression changes to the differentiation toward TH9 cells, a pro-atherogenic subset. Spectral flow cytometry shows that pre-exposure to oleic acid results in a skew toward IL-9+-producing T cells upon activation. Importantly, pharmacological inhibition of either cholesterol or fatty acid biosynthesis abolishes this effect, suggesting a beneficial role for statins beyond cholesterol lowering. Taken together, oleic acid may affect inflammatory diseases like atherosclerosis by rewiring T cell metabolism.
© 2024 The Author(s).