The epidermal growth factor receptor (EGFR) signaling pathway is an important regulator of insect reproduction, yet its function in parasitoid wasps remains largely unknown. Here, we investigated the role of EGFR in the reproductive development of the parasitoid wasp Leptopilina drosophilae, an effective natural enemy of the globally invasive pest Drosophila suzukii. We cloned the LdEGFR gene and confirmed its structural conservation through phylogenetic analysis. Using RNA interference (RNAi), we demonstrated that LdEGFR is essential for ovarian development but dispensable for testis development. Intriguingly, this sexually dimorphic functional requirement contrasts with its expression pattern, as LdEGFR transcript levels were approximately 171-fold higher in testes than in ovaries. Transcriptomic analysis of ovaries following LdEGFR knockdown revealed significant down-regulation of genes involved in transporter, DNA replication, cell cycle, and oocyte meiosis pathways, providing a molecular basis for the observed ovarian defects. Our findings establish LdEGFR as a critical regulator of female reproduction in L. drosophilae and highlight a notable decoupling between transcript abundance and functional essentiality. This study not only advances the understanding of reproductive regulation in parasitoid wasps but also identifies LdEGFR as a potential target for enhancing the mass rearing of this promising biocontrol agent.

Although terahertz (THz) metasurfaces based on bound state in the continuum (BIC) have garnered significant attention in biomedical applications, their technical implementation in high-sensitivity cancer cells detection remains a critical challenge. In this work, we present a THz biosensor employing dual split-ring resonator (DSRR) arrays based on quasi-bound state in the continuum (Q-BIC). Numerical simulations reveal a high-Q resonance dip at 2.35 THz with a detection sensitivity of 522 GHz/RIU. Experimentally, the performance was validated by detecting normal cells (murine splenocytes) and three cancer cell lines (LLC, LoVo, and MC38). In addition, analysis of cell type discrimination was achieved by integrating machine learning algorithms to project high-dimensional spectral data into a low-dimensional space. This study establishes a label-free approach for long-term cellular monitoring, advancing THz technology as an innovative platform for practical biomedical applications.

Inhalation-induced lung injury, caused by harmful factors like chemical fumes and dust, leads to acute and chronic inflammation and fibrosis. Traditional treatments, such as mechanical ventilation and anti-inflammatory drugs, can relieve symptoms but fail to promote tissue regeneration. Stem cells and their extracellular vesicles (EVs) offer new treatment possibilities due to their anti-inflammatory and regenerative properties. However, the specific pathological environment of these lung injuries limits the effectiveness and targeting of EVs, challenging their clinical use. This review outlines stem cell EVs' mechanisms in treating inhalation-induced lung injury, examines recent engineering advancements, and addresses challenges in moving from research to clinical application. It highlights the importance of interdisciplinary collaboration in carrier design, production, and regulation, offering a theoretical foundation for developing precision EV-based treatments.

While parasitoid wasps have been extensively reported to rely on plant-derived volatiles for host location, host habitats such as fermented fruits often harbor not only plants but also diverse microbial communities that are essential for hosts. This raises the question of whether parasitoid wasps exploit microbial volatiles for host location. Although some ecological evidence exists for such microbe-mediated attraction, the underlying molecular mechanisms remain largely unknown. Here, using the Leptopilina boulardi-Drosophila melanogaster system, it is reported that yeast (a ubiquitous microorganism in host habitats) attracts both Drosophila hosts and L. boulardi female parasitoids. Four yeast-derived ethyl ester volatiles that elicit olfactory attraction in female wasps are identified. Through integrated genomic and transcriptomic analyses, the complete odorant receptor repertoire of L. boulardi is characterized, and two female-biased receptors (LbouOR167 and LbouOR136) are identified as necessary for the detection of these ethyl esters. Structural and functional analyses revealed that a conserved residue, Leu159, in both receptors is essential for odorant ligand binding. These findings uncover a microbial volatile-mediated mechanism underlying parasitoid host-seeking behavior and provide novel insights into the chemical ecology of a four-party interaction involving fruit, fungi, Drosophila hosts, and their parasitoid wasps.

Emamectin benzoate (EB) is widely used in agricultural pest management, but its toxic effects on non-target natural enemies and the underlying mechanisms remain poorly understood. Here, we investigated the toxicity of EB to the parasitoid wasp Leptopilina drosophilae Huang, a promising biological control agent of Drosophila suzukii Matsumura and other drosophilid pests, and explored the role of gut microbiota in mediating host physiological responses. Bioassay results showed that EB exhibited high toxicity to L. drosophilae in a dose-dependent manner, with LC5, LC10, and LC50 values of 0.57 mg/L, 0.69 mg/L, and 1.32 mg/L, respectively. Transcriptomic analysis further revealed that sublethal EB exposure significantly suppressed genes related to detoxification and immunity. Consistent with these molecular changes, 16S rRNA sequencing demonstrated that EB induced severe gut dysbiosis, characterized by reduced microbial diversity and richness. Moreover, integrated correlation analysis revealed significant positive associations between the abundance of declined bacterial genera and the expression of detoxification genes, suggesting a functional link between gut microbiota disruption and impaired host detoxification capacity. Collectively, our findings demonstrate that sublethal EB exposure disrupts gut microbiota homeostasis through suppression of gut antimicrobial peptide expression, which in turn leads to downregulation of host detoxification genes and reduced insecticide resistance. This microbially-mediated mechanism contributes to the compromised fitness and increased mortality of parasitoid wasps, highlighting the ecological risks of EB to biological control agents and emphasizing the need to consider gut microbiota-mediated effects in insecticide safety assessments.

BACKGROUND:The use of alternative hosts is a key strategy for enhancing the efficacy of parasitoid wasps in pest control. Pachycrepoideus vindemmiae is an important pupal ectoparasitoid of numerous agricultural and public-health pests, including Drosophila suzukii, Bactrocera dorsalis, and Megaselia scalaris. However, a suitable alternative host for the mass rearing of P. vindemmiae has yet to be established. RESULTS:In this study, we compared the biocontrol traits of P. vindemmiae reared on Drosophila melanogaster (PVm) with those reared on the larger alternative host Drosophila virilis (PVv). PVv exhibited a developmental duration similar to that of PVm but attained a significantly larger body size. PVv also showed higher survival under temperature stress (4 °C and 35 °C) and during starvation. Furthermore, PVv achieved significantly higher parasitism rates against Drosophila suzukii, B. dorsalis, and M. scalaris, likely attributable to its greater number of mature eggs and enhanced reproductive capacity. Transcriptomic analysis revealed up-regulation of genes associated with stress resistance, growth and development, and venom function, which may collectively contribute to the improved biocontrol performance of PVv. CONCLUSION:In summary, rearing P. vindemmiae on D. virilis enhances its biocontrol potential, offering a promising strategy to mitigate public-health risks and reduce agricultural economic losses. © 2026 Society of Chemical Industry.

The application of copper-based nanoparticles (Cu-based NPs, e.g., CuO-NPs and Cu-NPs) as nanopesticides represents a promising avenue for sustainable agriculture. However, their potential ecotoxicological effects on beneficial arthropods, particularly parasitoid wasps which are crucial for biological pest control, remain poorly understood. Here, we investigated the chronic toxicity of CuO-NPs and Cu-NPs to the endoparasitoid wasp Asobara japonica and elucidated the underlying mechanisms. We found that dietary exposure induced concentration-dependent lethality, primarily driven by released copper ions rather than the particles themselves. Sublethal exposure severely impaired wasp fitness, with reduced host-searching locomotion thereby mediating a decline in parasitism efficiency. Mechanistically, integrated transcriptomic and metabolomic analyses revealed a coordinated downregulation of some key genes in fatty acid homeostasis pathways and a consequent decline in free fatty acid levels. This disruption in energy mobilization likely compromised the energy supply necessary for sustained activity, thereby explaining the observed locomotion behavioral deficits. Furthermore, Cu-based NPs exposure selectively altered the rare gut microbiota, and shifts in specific bacterial genera correlated with the suppression of host fatty acid metabolism genes. Our results demonstrate that Cu-based NPs impair parasitoid wasp fitness through the disruption of gut microbiome homeostasis and energy metabolism, highlighting the need to consider these impacts in nanomaterial risk assessment for sustainable agriculture.

Nitrogen exchange between plants and insects is a major component of ecosystem nitrogen cycling. Endophytic insect pathogenic fungi transfer insect-derived nitrogen to plants through symbiotic associations mediated by fungal mycelia, enabling plants to thrive even after losing nitrogen to insects. However, the mechanisms underlying this process remain unexplored. Here, we show that the widespread endophytic entomopathogen Metarhizium robertsii degrades the common root-derived antifungal compound caulilexin C to produce the volatile 1-methoxyindole. This compound is recognized by the Or74a olfactory receptor in Drosophila melanogaster larvae and attracts multiple Dipteran species to the plant-Metarhizium consortium. The recruited insects are subsequently infected and consumed, resulting in enhanced insect-derived nitrogen transfer to the plants. This self-reinforcing mechanism strengthens the plant-fungus symbiosis and reveals a pathway contributing to ecosystem nitrogen flux.

AmisChemotherapy-induced hepatotoxicity (CIH) is a significant concern in colorectal cancer (CRC) patients treated with the CAPEOX (capecitabine and oxaliplatin) regimen. Identifying predictive factors for CIH is crucial for clinical management.Patients and MethodsThis study analyzed colorectal tissue (CRT), plasma, and urine samples from CRC patients. Differentially expressed metabolites (DEMs) across these tissues were integrated for multi-omics analysis, and predictive models for CIH susceptibility were developed. An independent set of 75 plasma samples was used for validation.ResultsA total of 492 differentially expressed compounds were identified in samples from 63 CRC patients, including 105, 149, and 238 DEMs in CRT, plasma, and urine, respectively. Lipids and lipid-like molecules were predominant in all samples. Among these, urine samples exhibited the highest variability and provided the strongest predictive power for CIH susceptibility. Principal component analysis (PCA) effectively differentiated normal patients from those with CIH. The study revealed steatosis as the primary pathological feature of CIH, with disrupted lipid metabolism emerging as a key characteristic. Predictive models constructed from multi-tissue metabolites profile exhibited high accuracy, with the plasma model achieving an AUC of 0.933 in external validation set. Our study underscores the importance of individual metabolic variations in CIH susceptibility, reflecting the complex interplay of genetic, environmental, and lifestyle factors.ConclusionThis study emphasizes the critical role of alterations in lipid, polyamine, and purine metabolism, as well as impaired tissue repair mechanisms, were identified as key endogenous factors underlying CIH susceptibility. The developed predictive models demonstrate potential for clinical application in assessing CIH risk in CRC patients undergoing CAPEOX chemotherapy.

RSV and seasonal influenza are two of the most prevalent causes of respiratory infection in the U.S. In this study, we used weekly positive case reports and genetic surveillance data to characterize the circulation of these viruses in the United States between 2011 and 2019 and a mathematical modeling approach to explore their potential interaction at a regional level. Our analyses showed that RSV and seasonal influenza co-circulate with different relative epidemic sizes and seasonal overlaps across regions and seasons. We found that RSV had a different evolutionary dynamic compared to seasonal influenza and that local persistence may play a role in underlying annual epidemics. Our analysis supports a potential competitive interaction between RSV and seasonal influenza in most regions across the United States. The multiple-pathogen modeling framework suggests that cross-immunity following infection of either virus might be one of the key drivers of viral competition. However, this finding is based on model-derived inferences and limited surveillance data; further investigation is needed to confirm its robustness and gain a better understanding of the underlying mechanisms. These findings underscore the importance of continued research into the immunological and ecological mechanisms of viral inference, which might be important for the development of more effective protective strategies against co-circulating respiratory viruses.

Breast cancer (BC) exhibits extensive heterogeneity, complicating personalized treatment. Glycosyl transferases (GTs) are potential biomarkers for cancer treatment. Consensus clustering was performed on GT-related genes (GTRGs) in the The Cancer Genome Atlas Breast Cancer cohort. Clinical characteristics, survival outcomes, and molecular pathways were analyzed. Differences in the tumor microenvironment were assessed. Machine learning techniques (Lasso, random forest, XGBoost) identified hub GTRGs. A prognostic nomogram was constructed integrating hub GTRGs and clinical features. The cell validation experiments were finally performed. Two subtypes (C1 and C2) were identified, and C1 showed better survival than C2 (P = .0029). C1 had higher CD8 T cells and activated NK cells, while macrophage fractions were lower. In addition, GTs-related mTORC1 and PI3K/AKT/mTOR pathways were suppressed, whereas p53, Wnt/β-Catenin, and Notch pathways were activated. 4 hub GTRGs (ATP5F1B, CS, EPB41L4B, and LIMCH1) were identified. High expression of these genes correlated with poorer prognosis. The nomogram demonstrated high accuracy of this hub GTRGs for predicting 1-, 3-, and 5-year overall survival. Additionally, expression levels of 4 hub genes were higher in the BC cell line MDA-MB-415, and CS knockdown significantly inhibited the proliferation and migration of MDA-MB-415 cells. GT-related subtypes in BC are linked to distinct survival outcomes and immune landscapes. Hub GTRGs are valuable prognostic markers, aiding personalized treatment strategies.

We previously discovered that HOXC6 was the most significantly upregulated gene in right-sided colon cancer compared to left-sided colon cancer according to our previous study; however, the role of HOXC6 in microsatellite instability-high (MSI-H) tumors remains poorly understood. Here, multiple public datasets, and in-house cohorts were used to analyze the differential expression and prognostic role of HOXC6 in colorectal cancer (CRC). Immunohistochemistry and immunofluorescence were performed to evaluate the correlation between HOXC6 expression and M2 macrophage infiltration. CCK8 and Transwell assays were used to evaluate the proliferation and migration of tumor cells in vitro. BALB/c nude mice were utilized to construct a humanized immune system model to evaluate the efficacy of ruxolitinib in vivo. We found that HOXC6 was overexpressed in MSI-H CRC and associated with a poor prognosis. Upregulation of CCL2 by HOXC6 increased M2 macrophage infiltration. IL6 secreted by M2 macrophages induced the epithelial-mesenchymal transition of tumor cells by upregulating HOXC6. M2 macrophages promoted effector T cell exhaustion by downregulating 4-1BB. Thus, inhibition of the IL6/JAK pathway in M2 macrophages restored 4-1BB expression and T-cell cytotoxicity offering a promising therapeutic target for the treatment of HOXC6-overexpressing MSI-H CRC.

Animals often exhibit increased aggression in response to starvation, while parasites often manipulate host behavior. In contrast, underlying molecular mechanisms for these behavioral changes are mostly unknown. The diamondback moth, Plutella xylostella, is an agricultural pest that feeds on cruciferous plants as larvae, while Cotesia vestalis is a parasitoid wasp that parasitizes diamondback moth larvae. In this study, we determined that unparasitized diamondback moth larvae exhibit increased aggression and cannibalism when starved, while starved larvae parasitized by C. vestalis were more aggressive than unparasitized larvae. C. vestalis harbors a domesticated endogenized virus named Cotesia vestalis bracovirus (CvBV) that wasps inject into parasitized hosts. Starvation increased octopamine (OA) levels in the central nervous system (CNS) of diamondback moth larvae while a series of experiments identified a CvBV-encoded gene product named Assailant that further increased aggression in starved diamondback moth larvae. We determined that Assailant increases OA levels by activating tyramine beta-hydroxylase (PxTβh), which is a key enzyme in the OA biosynthesis pathway. Ectopic expression of assailant in Drosophila melanogaster likewise upregulated expression of DmTβh and OA, which increased aggressive behavior in male flies as measured by a well-established assay. While parasitized hosts are often thought to be at a competitive disadvantage to nonparasitized individuals, our results uncover how a parasitoid uses an endogenized virus to increase host aggression and enhance survival of offspring when competing against unparasitized hosts.

BACKGROUND:Previous studies have shown that WZC can increase tacrolimus blood concentration when co-administered. However, limited knowledge exists regarding the pharmacokinetics of both tacrolimus and the bioactive lignans in WZC when administered simultaneously in renal transplantation patients. AIMS:This study aimed to investigate the pharmacokinetics of tacrolimus and multiple bioactive lignans in Wuzhi capsule (WZC) when co-administered with 5 bioactive components in renal transplantation recipients. OBJECTIVES:The objective of this study was to develop a method for simultaneous quantification of tacrolimus and multiple bioactive lignans in WZC using liquid-liquid extraction followed by LC-MS/MS analysis. METHODS:A liquid-liquid extraction method combined with LC-MS/MS analysis was developed for simultaneous quantification of tacrolimus and multiple bioactive lignans in WZC. Human whole blood samples were analyzed, and the accuracy and precision of the method were evaluated. RESULTS:The developed method showed good linearity and accuracy for the quantification of tacrolimus and bioactive lignans in WZC. Pharmacokinetic analysis revealed significant effects of WZC co-administration on both V/F and CL/F in renal transplantation patients. CONCLUSION:This study demonstrated that simultaneous administration of WZC had notable effects on the pharmacokinetics of tacrolimus and bioactive lignans in renal transplantation patients. The developed method proved to be reliable and sensitive for determining the whole blood concentrations of tacrolimus and WZC, making it suitable for pharmacokinetic studies in transplant patients.

Severe high temperature (HT) climate significantly impacts cotton quality and yield. Consequently, it is essential to mine thermal-responsive genes and explore the underlying mechanisms of HT response in cotton. In this study, we employed a high-throughput cDNA-library method in conjunction with the ALRS system to screen thermotolerant genes in Upland cotton. As a result, a total of 16,120, 13,216 and 172 effective survival genes were filtered after HT stress exposure (42 °C, 220 rpm) for 48 h, 60 h and 72 h, respectively. Functional annotation and enrichment analysis revealed that 170 common genes were involved in regulatory processes associated with HT stress, and the relevant transcriptome data indicated that the majority of these genes responded to temperature fluctuations. Twenty-one genes were randomly selected for verification, and it was found that these genes could enhance yeast resistance to HT stress. Additionally, we selected mutants of homologous Arabidopsis genes for four candidate genes to validate plant thermotolerance during flowering; the thermotolerances of SALK_201915 and SALK_120540.1 were significantly worse. The results demonstrate that numerous candidate genes identified from the cDNA-library contribute to the highly complex molecular network that governs the response and resistance to HT stress in Upland cotton. The high-throughput heat-screening method utilized in this study was optimized for mining thermotolerant genes including improvement in yeast library construction, screening system, gradient reverse pressure, and sequencing library construction. We hope that this new method can be applied in future studies on stress in cotton and other species.

BACKGROUND:Plants differ more than threefold in seed oil contents (SOCs). Soybean (Glycine max), cotton (Gossypium hirsutum), rapeseed (Brassica napus), and sesame (Sesamum indicum) are four important oil crops with markedly different SOCs and fatty acid compositions.RESULTS:Compared to grain crops like maize and rice, expanded acyl-lipid metabolism genes and relatively higher expression levels of genes involved in seed oil synthesis (SOS) in the oil crops contributed to the oil accumulation in seeds. Here, we conducted comparative transcriptomics on oil crops with two different SOC materials. In common, DIHYDROLIPOAMIDE DEHYDROGENASE, STEAROYL-ACYL CARRIER PROTEIN DESATURASE, PHOSPHOLIPID:DIACYLGLYCEROL ACYLTRANSFERASE, and oil-body protein genes were both differentially expressed between the high- and low-oil materials of each crop. By comparing functional components of SOS networks, we found that the strong correlations between genes in "glycolysis/gluconeogenesis" and "fatty acid synthesis" were conserved in both grain and oil crops, with PYRUVATE KINASE being the common factor affecting starch and lipid accumulation. Network alignment also found a conserved clique among oil crops affecting seed oil accumulation, which has been validated in Arabidopsis. Differently, secondary and protein metabolism affected oil synthesis to different degrees in different crops, and high SOC was due to less competition of the same precursors. The comparison of Arabidopsis mutants and wild type showed that CINNAMYL ALCOHOL DEHYDROGENASE 9, the conserved regulator we identified, was a factor resulting in different relative contents of lignins to oil in seeds. The interconnection of lipids and proteins was common but in different ways among crops, which partly led to differential oil production.CONCLUSIONS:This study goes beyond the observations made in studies of individual species to provide new insights into which genes and networks may be fundamental to seed oil accumulation from a multispecies perspective.

Influenza A/H9 viruses circulate worldwide in wild and domestic avian species, continuing to evolve and posing a zoonotic risk. A substantial increase in human infections with A/H9N2 subtype avian influenza viruses (AIVs) and the emergence of novel reassortants carrying A/H9N2-origin internal genes has occurred in recent years. Different names have been used to describe the circulating and emerging A/H9 lineages. To address this issue, an international group of experts from animal and public health laboratories, endorsed by the WOAH/FAO Network of Expertise on Animal Influenza, has created a practical lineage classification and nomenclature system based on the analysis of 10,638 hemagglutinin sequences from A/H9 AIVs sampled worldwide. This system incorporates phylogenetic relationships and epidemiologic characteristics designed to trace emerging and circulating lineages and clades. To aid in lineage and clade assignment, an online tool has been created. This proposed classification enables rapid comprehension of the global spread and evolution of A/H9 AIVs.

The methylation of N6-methyladenosine (m6A) involves writers, erasers, and readers, acting synergistically in posttranscriptional regulation. These processes influence various biological processes, including plant floral transition. However, the specific role of m6A modifications in photoperiod sensitivity in cotton (Gossypium hirsutum) remains obscure. To elucidate this, in this study, we conducted transcriptome-wide m6A sequencing during critical flowering transition stages in the photoperiod-sensitive wild G. hirsutum var. yucatanense (yucatanense) and the photoperiod-insensitive cultivated cotton G. hirsutum acc. TM-1 (TM-1). Our results revealed significant variations in m6A methylation of 2 cotton varieties, with yucatanense exhibiting elevated m6A modification levels compared with TM-1 under long-day conditions. Notably, distinct m6A peaks between TM-1 and yucatanense correlated significantly with photoperiod sensitivity. Moreover, our study highlighted the role of the demethylase G. hirsutum ALKB homolog 5 (GhALKBH5) in modulating m6A modification levels. Silencing GhALKBH5 led to a decreased mRNA level of key photoperiodic flowering genes (GhADO3, GhAGL24, and GhFT1), resulting in delayed bud emergence and flowering. Reverse transcription quantitative PCR analyses confirmed that silencing GhADO3 and GhAGL24 significantly downregulated the expression of the floral integrator GhFT1. Collectively, our findings unveiled a transcriptional regulatory mechanism in which GhALKBH5-mediated m6A demethylation of crucial photoperiodic flowering transcripts modulated photoperiod sensitivity in cotton.

Background Apatinib (YN968D1) is the first small-molecule-targeting drug with anti-tumor activity created in China for the treatment of advanced gastric cancer (GC) and hepatocellular carcinoma (HCC). It showed significant variation in the efficacy for treating cancers, including advanced non-squamous non-small-cell lung cancer (NSCLC). Whether its efficacy could be optimized by subgrouping patients with certain genetic variation remains elusive. Methods Here, we firstly used kinase screening to identify any possible target of apatinib against 138 kinases. The effects of apatinib on proliferation rates, cell cycle, cell apoptosis, and cell migration on cancer cell lines were analyzed; the in vitro potential pathways of apatinib on cancer cell lines were screened. The effect of apatinib on mouse cancer models in vivo was also analyzed. Results Based on HCC364 cells with BRAF V600E mutation, we have shown that apatinib could inhibit their growth, migration, cell cycle, and induce their apoptosis. Based on mice with transplanted HCC364 cells, we have also shown that apatinib could inhibit the tumor growth. Based on immunohistochemistry, we have demonstrated that apatinib could suppress the phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase and extracellular regulated protein kinases. This may account at least part of the apatinib’s inhibitory effect on HCC364 cancer cells. Conclusions BRAF V600E protein kinase is a target of apatinib by kinase screening. We have demonstrated that apatinib can effectively inhibit tumor cells with BRAF V600E mutation by in vitro and in vivo experiments. Our results have demonstrated that targeting BRAF V600E mutation, apatinib appears to be effective and safe for treating NSCLC and possibly other cancers with the same mutation.

Human respiratory syncytial virus (RSV) is a major cause of lower respiratory infection. Despite more than 60 years of research, there is no licensed vaccine. While B cell response is a major focus for vaccine design, the T cell epitope profile of RSV is also important for vaccine development. Here, we computationally predicted putative T cell epitopes in the Fusion protein (F) and Glycoprotein (G) of RSV wild circulating strains by predicting Major Histocompatibility Complex (MHC) class I and class II binding affinity. We limited our inferences to conserved epitopes in both F and G proteins that have been experimentally validated. We applied multidimensional scaling (MDS) to construct T cell epitope landscapes to investigate the diversity and evolution of T cell profiles across different RSV strains. We find the RSV strains are clustered into three RSV-A groups and two RSV-B groups on this T epitope landscape. These clusters represent divergent RSV strains with potentially different immunogenic profiles. In addition, our results show a greater proportion of F protein T cell epitope content conservation among recent epidemic strains, whereas the G protein T cell epitope content was decreased. Importantly, our results suggest that RSV-A and RSV-B have different patterns of epitope drift and replacement and that RSV-B vaccines may need more frequent updates. Our study provides a novel framework to study RSV T cell epitope evolution. Understanding the patterns of T cell epitope conservation and change may be valuable for vaccine design and assessment.