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Transmissibility of respiratory viruses is a complex viral trait that is intricately linked to tropism. Several highly transmissible viruses, including severe acute respiratory syndrome coronavirus 2 and Influenza viruses, specifically target multiciliated cells in the upper respiratory tract to facilitate efficient human-to-human transmission. In contrast, the zoonotic Middle East respiratory syndrome coronavirus (MERS-CoV) generally transmits poorly between humans, which is largely attributed to the absence of its receptor dipeptidyl peptidase 4 (DPP4) in the upper respiratory tract. At the same time, MERS-CoV epidemiology is characterized by occasional superspreading events, suggesting that some individuals can disseminate this virus effectively. Here, we utilized well-differentiated human pulmonary and nasal airway organoid-derived cultures to further delineate the respiratory tropism of MERS-CoV. We find that MERS-CoV replicated to high titers in both pulmonary and nasal airway cultures. Using single-cell messenger-RNA sequencing, immunofluorescence, and immunohistochemistry, we show that MERS-CoV preferentially targeted multiciliated cells, leading to loss of ciliary coverage. MERS-CoV cellular tropism was dependent on the differentiation of the organoid-derived cultures, and replication efficiency varied considerably between donors. Similarly, variable and focal expression of DPP4 was revealed in human nose tissues. This study indicates that the upper respiratory tract tropism of MERS-CoV may vary between individuals due to differences in DPP4 expression, providing an explanation for the unpredictable transmission pattern of MERS-CoV., Competing Interests: Competing interests statement:The authors declare no competing interest.

Identifying conserved mechanisms used by viruses to delay host innate responses can reveal potential targets for antiviral therapeutics. Here, we investigated coronavirus nonstructural protein 15 (nsp15), which encodes a highly conserved endoribonuclease (EndoU). EndoU functions as an immune antagonist by limiting the accumulation of viral replication intermediates that would otherwise be sensed by the host. Despite being a promising antiviral target, it has been difficult to develop small-molecule inhibitors that target the EndoU active site. We generated nsp15 mutants of the coronaviruses severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and mouse hepatitis virus (MHV)-A59 and identified conserved residues within the amino-terminal domain that are required for EndoU activity. Loss of EndoU activity caused the activation of host sensors, which limited viral replication in interferon-responsive cells and attenuated disease in MHV-infected mice. Using transcriptional profiling, we found that MHV EndoU mutant viruses upregulate multiple host sensors, including Z-form nucleic acid-binding protein 1 (ZBP1). We found that nsp15 mutants induced early, robust ZBP1-mediated necroptosis. EndoU mutant viruses also induced ZBP1-independent apoptosis and pyroptosis pathways, causing early, robust cell death that limits virus replication and pathogenesis. Overall, we document the importance of the amino-terminal domain for EndoU function. We also highlight the importance of nsp15/EndoU activity for evading host sensors, delaying cell death, and promoting pathogenesis., Competing Interests: Competing interests statement:The authors declare no competing interest.

Simnotrelvir is an oral small-molecule antiviral agent targeting the 3C-like protease (3CL pro ) of SARS-CoV-2, proven effective against the Delta variant with favorable pharmacokinetics and safety in preclinical study. In this study, we further evaluated the antiviral efficacy of simnotrelvir against a range of emerging Omicron variants, including BA.1, BA.4, BA.5, CH.1.1, XBB.1.5, XBB.1.16, EG.5, and JN.1. In vitro assays with Vero E6 cells confirmed that simnotrelvir exhibited robust antiviral activity across these variants, comparable to the Food and Drug Administration (FDA)-approved drug nirmatrelvir. Additionally, simnotrelvir demonstrated effective inhibition against several nirmatrelvir-resistant SARS-CoV-2 3CL pro mutants, including A260V, Y54A, (T21I + S144A), F140A, H172Y, and E166V. Importantly, simnotrelvir showed better potency against the E166V mutation compared to nirmatrelvir. Resistance selection studies revealed that BA.5 developed reduced sensitivity after 5 and 10 passages, increasing the IC 50 values by 3.2 and 4.5-fold, respectively, while HCoV-OC43 showed an 8.3-fold increase after 12 passages. Despite this, simnotrelvir's overall efficacy remains strong. Furthermore, clinical trials demonstrated that combining simnotrelvir with ritonavir significantly shortened symptom resolution in COVID-19 patients. Genomic analysis of treated patients found random nucleotide substitutions but no significant mutations linked to 3CL pro resistance. In conclusion, simnotrelvir shows strong antiviral activity against SARS-CoV-2 variants and maintains a high barrier to resistance, reinforcing its potential as an effective therapeutic option for current and future SARS-CoV-2 variants.

Background: Human immunodeficiency virus-1 (HIV-1) is the causative agent of acquired immunodeficiency syndrome, which is a major global health problem. Although combination antiretroviral therapy (cART) successfully expands the lifespan of HIV-1-infected patients, long-term cART often increases drug resistance and adverse effects. Therefore, efforts are ongoing to develop novel anti-HIV-1 drugs., Methods: The anti-HIV-1 activities of compounds were investigated using TZM-bl reporter cell line, A3.01 T cell line, and peripheral blood mononuclear cells infected with several HIV-1 strains, including wild type and drug-resistance associated mutants. Next-generation sequencing analysis and in silico molecular docking studies were employed to determine the mode of action of the compound., Results: We identified a small-molecule inhibitor consisting of a thiadiazole core appended to two pyrazoles (BPPT), which exerted a highly potent inhibitory effect on HIV-1 infectivity, with a half-maximal effective concentration (EC 50 ) of 60 nM, without causing cytotoxicity. In experiments with various HIV-1 strains and cell types, the potency of BPPT was found to be comparable to that of commercial antiretroviral agents (azidothymidine, nevirapine, and others). Further analysis of the mode of action demonstrated that BPPT is a novel type of HIV-1 non-nucleoside reverse transcriptase inhibitor (NNRTI). Analysis of viruses harboring drug-resistance-associated mutations showed that BPPT was potent against G190A (C or S) mutations in reverse transcriptase (RTase), exhibiting high-level resistance to other NNRTIs. Next-generation sequencing analysis of long-term treatment with BPPT displayed an RTase mutation profile different from that in the case of established NNRTIs. Given these data, in silico molecular docking studies demonstrated the molecular mechanism underlying the BPPT-mediated inhibition of RTase., Conclusion: Our data suggest that BPPT is a novel small-molecule inhibitor of HIV-1 RTase and could serve as a promising chemical scaffold to complement or replace conventional treatments, particularly for overcoming resistance associated with the G190 mutation., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a recently identified tick-borne virus that has emerged in the twenty-first century. Its primary clinical manifestations include fever and thrombocytopenia, and its high morbidity and mortality rates have garnered significant attention. It is crucial to have a comprehensive understanding of the spatial and temporal characteristics of SFTSV migration in order to prevent and control this disease. The SFTSV strains from East Asian countries in GenBank during 2017-2023 were collected and analyzed with phylogenetic and Bayesian methods. Phylogenetic analysis showed that SFTSV can be categorized into five genotypes (A, B, C, D, and E), with 24 recombination events and 15 reassortment events identified. This represented a higher number than previously observed. The results of our study indicated that SFTSV first diverged around 1785. We categorized the migration of SFTSV into two distinct periods, and identified the centers of spread and migration routes of SFTSV in each period. We propose that the eastern migration routes of migratory birds played a pivotal role during the initial stages of virus transmission, functioning as a primary conduit for the dispersal of the virus across the sea. The eastern and central migratory routes were similarly pivotal in subsequent phases of virus transmission. The results of the study showed that Japan was the first region where the virus originated and became endemic, and that the virus spread widely among countries. Elucidating the spatial and temporal characteristics of SFTSV migration will help prevent and control SFTS., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2025 Sheng et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The emergence of various SARS-CoV-2 variants presents challenges for antibody therapeutics, emphasizing the need for more potent and broadly neutralizing antibodies. Here, we employed an unbiased screening approach and successfully isolated two antibodies from individuals with only exposure to ancestral SARS-CoV-2. One of these antibodies, CYFN1006-1, exhibited robust cross-neutralization against a spectrum of SARS-CoV-2 variants, including the latest KP.2, KP.3 and XEC, with consistent IC 50 values ranging from ~1 to 5 ng/mL. It also displayed broad neutralization activity against SARS-CoV and related sarbecoviruses. Structural analysis revealed that these antibodies target shared hotspot but mutation-resistant epitopes, with their Fabs locking receptor binding domains (RBDs) in the "down" conformation through interactions with adjacent Fabs and RBDs, and cross-linking Spike trimers into di-trimers. In vivo studies conducted in a JN.1-infected hamster model validated the protective efficacy of CYFN1006-1. These findings suggest that antibodies with cross-neutralization activities can be identified from individuals with exclusively ancestral virus exposure., Competing Interests: Competing interests: Y.G. is a co-founder and shareholder of Antibody BioPharm, Inc. and ChangYuan FuNeng (Shanghai) Life Technology Co., Ltd. The remaining authors declare no competing interests., (© 2025. The Author(s).)

Background: Encephalitis is rarely caused by nontuberculous mycobacteria (NTM), which is generally not considered a highly virulent pathogen. However, NTM encephalitis in immunocompromised hosts occurs with varied clinical presentations, posing a diagnostic challenge in clinical practice. This study aims to describe an atypical case of NTM encephalitis caused by Mycobacterium celatum, which has not previously been reported to infect the central nervous system of immunocompromised hosts, mimicking autoimmune striatal encephalitis (ASE)., Case Presentation: A 35-year-old immunosuppressed woman presented with prolonged fever for 4 months and rapidly progressive cognitive decline for 3 months. Neurological examination showed impaired cognition and parkinsonism. Laboratory testing was unremarkable. Her brain imaging on T2-weighted fluid-attenuated inversion recovery (T2/FLAIR) exhibited lesions involving basal ganglia and subcortical white matter in both hemispheres, mimicking ASE. Cerebrospinal fluid (CSF) analysis revealed mild pleocytosis with normal glucose and protein levels. CSF comprehensive microbiological studies and autoimmune panels were negative. ASE was suspected, and immunotherapies were given. Despite immunotherapies, her condition worsened with seizures, warranting a stereotactic brain biopsy to achieve a definite diagnosis. Her brain tissue pathology result was non-specific. However, we identified M. celatum from her brain tissue. Thus, the final diagnosis was M. celatum encephalitis. Therefore, we discontinued immunotherapies and started anti-NTM treatment, including isoniazid, rifampicin, ethambutol, and levofloxacin. After completing a 16-month treatment course, her clinical condition was stable, afebrile, and seizure-free., Conclusions: We proposed that NTM invades the central nervous system and also triggers immune dysregulation, developing features resembling ASE. In case of suspicious autoimmune encephalitis with poor response to immunotherapies, a tissue biopsy should be performed to exclude chronic infection., Competing Interests: Declarations. Ethics approval and consent to participate: This case report was ethically approved by the institutional review board from Faculty of Medicine, Chulalongkorn University (COE No. 031/2024 IRB No. 0434/67). Written informed consent to participate was obtained from the patient’s surrogate. Consent for publication: Written informed consent for the publication of this case report and any accompanying images was obtained from the patient’s surrogate. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Non-pharmaceutical interventions (NPIs) are critical for influenza control and prevention, however, the data about such interventions are insufficient among students in grades below middle school. Hence, this study aims to explore the effectiveness of NPIs (class-suspension and symptom-based isolation) on the control of influenza spread among children in primary school and kindergarten. Findings will support evidence-based strategies for influenza control among children enrolled in primary schools and kindergartens., Methods: We purposively selected two regions (Zigong and Nanan) in Southwest China as study places, and class-suspension and symptom-based isolation was conducted in the two regions, respectively. RT (effective reproduction number) value, incidence cases, and attack rate were considered as outcome indicators. R4.1.2 software was used to conduct statistical analysis, and p < 0.05 (two-tailed) was considered as statistically significant., Results: In total, 593 students in the Nanan district received symptom-based isolation while 1340 students in Zigong city received class-suspension. The vaccination rate of Zigong (21.27%) was much greater than Nanan (8.26%) (p < 0.001). Parents in the Nanan were more highly educated, with undergraduate, master and above degrees, compared to parents in the Zigong (p < 0.05). Though there was no statistical significance difference in RT between Nanan (1.23) and Zigong (1.16) after quarantine and control measures were conducted, the RT value in the two regions was sharply decreased. The incident cases after class-suspension was much higher than symptom-based isolation in both kindergarten and primary school. The attack rate had a significant difference between class-suspension and symptom-based isolation in primary school (p < 0.05)., Conclusions: Both symptom-based isolation and class-suspension are effective measures in control of influenza spread, and symptom-based isolation are more effective than class-suspension in primary school. Health education and daily surveillance are needed in the control and prevention of influenza., Competing Interests: Declarations. Ethics approval and consent to participate: The project proposal was approved by the Institutional Review Board of the Nanan District Center for Disease Control and Prevention (Grant No. 2023-001). This study was conducted in accordance with the Declaration of Helsinki. All participants gave their written informed consent to participate. For participants who are under age 16, informed consent was obtained from a parent and/or legal guardian. No patients who were not literate participated in this study. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Purpose: Yangyin Fuzheng Jiedu Prescription (YFJP) is a traditional Chinese medicine (TCM) used for the treatment of hepatocellular carcinoma (HCC). However, the potential mechanisms remain unclear. The objective of this study is to clarify the mechanism of action of YFJP in treating HCC., Methods: By constructing networks, the active components and molecular targets of YFJP in the treatment of HCC were explored. The TCGA database was utilized to analyze the correlation between the core target and the overall survival (OS) of patients with HCC. The regulatory effect of YFJP on T cell was evaluated by detecting samples from patients with HCC. The molecular mechanism of YFJP in treating HCC was validated through in vivo and in vitro experiments., Results: Constructing networks and analyse indicated that the key targets of YFJP in the treatment of HCC is FoxO1. Analysis of the HCC patient cohort in the TCGA database demonstrated that FoxO1 is an independent protective factor for overall survival in patients with HCC. Pathway enrichment analysis enriched FoxO signaling pathway and Cellular senescence pathway. Prospectively collecting samples from patients with HCC suggested that YFJP treatment increased the proportion of CD8 + T cells. In vivo experiments showed that YFJP treatment ameliorated CD8 + T cell senescence in tumor-bearing mice. Western blot, flow cytometry and multi-color immunofluorescence co-staining showed that YFJP treatment increased the expression of FoxO1 in CD8 + T cells. The primary CD8 + T cells were sorted and co-cultured with an HCC cell line in vitro. Inhibiting the expression of FoxO1 in CD8 + T cells confirmed that FoxO1 is a key target for YFJP to improve the senescence of CD8 + T cell., Conclusion: FoxO1 is the key molecular target of YFJP in improving CD8 + T cell senescence in HCC. This study preliminarily clarified the mechanism of YFJP in regulating immunosenescence of HCC., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2025 Xie et al.)

Nano-plastics (NPs) are emerging hazardous environmental contaminants that pose health risks with size-dependent toxic effects and are potential risk factors for hepatocellular carcinoma (HCC) and lipid metabolism disorders including non-alcoholic fatty liver disease (NAFLD). However, their underlying molecular mechanisms remain unclear. To shed more light on the causes of these risks, we developed a digestive system microphysiological platform (DS-MPP) for simulating dynamic internal-exposure of multi-size NPs in the gastrointestinal tract and liver. Multi-omics analysis based on DS-MPP revealed hepatic cells are more sensitive to 72 μg/day NPs than gastrointestinal mucosa cells. Specifically, 50 nm NPs disrupt phospholipid metabolism, promote diacylglycerol (DG) accumulation, convert more DG to phosphatidic acid (PA) than triacylglycerol (TG), thus facilitating endocytic vesicles production. Meanwhile, it can active tumorigenesis related pathway mTOR, inducing HCC marked by CAB39. Moreover, 500 nm NPs promote NAFLD by inducing insulin resistance pathways and decreasing PLD1 expression. Our results demonstrate the mechanism of disease and metabolic disorders induced by NPs vary depending on particle size. DS-MPP is a reliable platform for evaluating risk of dynamic NPs exposure and elucidating mechanisms of related metabolic diseases. This platform provides a promising method for health risk assessment caused by environmental pollutants., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: In late 2023, the first human case caused by an H10N5 avian influenza virus (AIV) was diagnosed in China. H10Ny AIVs have been identified in various poultry and wild birds in Eurasia, the Americas, and Oceania., Methods: We analyzed the clinical data of the H10N5 AIV-infected patient, isolated the virus, and evaluated the virus receptor-binding properties together with the H10N8 and H10N3 AIVs identified in humans and poultry. The genomic data of the human-infecting H10N5 strain and avian H10Ny AIVs (n = 48, including 16 strains of H10N3 and 2 strains of H10N8) from live poultry markets in China, during 2019-2021, were sequenced. We inferred the genetic origin and spread pattern of the H10N5 AIV using the phylodynamic methods. In addition, given all available nucleotide sequences, the spatial-temporal dynamics, host distribution, and the maximum-likelihood phylogenies of global H10 AIVs were reconstructed., Findings: The first H10N5 AIV-infected human case co-infected with seasonal influenza H3N2 virus was identified. Unfortunately, the patient died after systematic treatments. The H10N5 virus predominantly bound avian-type receptor, without any known mammalian-adapted mutations. Phylodynamic inference indicated that the H10N5 AIV was generated by multiple reassortments among viruses from Korea and Japan, central Asia, and China in late 2022, acquiring the seven gene segments from H10N7 or other low pathogenic AIVs in wild Anseriformes, except for the PA gene from H5N2 AIVs in Domestic Anseriformes. The HA gene of the H10N5 virus belongs to the North American lineage, which was probably introduced into Asia by migratory birds, subsequently forming local circulation., Interpretation: Unlike the human-infecting H10N3 and H10N8 AIVs acquiring six internal protein-coding genes from H9N2 AIVs in domestic poultry, the human-infecting H10N5 AIV was generated through multiple reassortments among viruses mainly carried by wild Anseriformes. Furthermore, worldwide distribution, inter-continental transmission, and genetic exchanges between Eurasian and North American lineages call for more concerns about influenza surveillance on H10Ny AIVs, especially in the flyway overlapping areas., Competing Interests: Declaration of Interests The authors declare no conflicts of interest. Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Previous work from our lab indicates that the heterotrimeric Gβ subunit 1 (Gpb1) enhances hyphal development and virulence in Mucor lusitanicus. In this study, three Gγ- and two additional Gβ-encoding genes were deleted to identify which ones might have a similar role as Gpb1. Deletion of gpg1 reduces hyphal growth, virulence, cyclic adenosine monophosphate (cAMP) levels, and protein kinase A (PKA) activity, similar to gpb1 deletion, suggesting that gpg1 participates in the same regulatory pathway as gpb1. The defects observed in Δgpg1 or Δgpb1 were suppressed by overexpression of the gene pkaR1 encoding the regulatory subunit 1 of PKA, indicating that this pathway is controlled by Gpg1 and Gpb1. Moreover, Δgpg1 and Δgpb1 show a downregulation of the transcription factors tec1 and tec2. Furthermore, tec-overexpression in Δgpg1, Δgpb1, and ΔpkaR1 restores the wild-type phenotype, indicating that both Tec are under control by the Gpb1, Gpg1, and PKA pathway. Moreover, the Δgpb1/Δgbg1 (+)(-) exhibits lower aerobic germination, hyphal growth and downregulates NAD + -glutamate dehydrogenases (gdh2a/b), whereas virulence is similar to that of the wild-type (WT) strain. These alterations in Δgpb1/Δgbg1 (+)(-) were reversed by the presence of glutamate during growth, suggesting that NAD + -Gdh2 could be under control of these subunits. Compared to the WT and Δgpb1/Δgbg1 (+)(-) strains under aerobic growth, single deletion strains showed lower rhizoferrin levels, respiration and reactive oxygen species levels. Our results suggest that Gpg1 interacts with Gpb1 to positively control the hyphal development and virulence by repressing the PKA pathway, thereby regulating the mitochondrial oxidative metabolism in M. lusitanicus., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The author is an Editorial Board Member/Editor-in-Chief/Associate Editor/Guest Editor for this journal and was not involved in the editorial., (Copyright © 2025. Published by Elsevier Inc.)

The newly emerged avian influenza A H5N1 Clade 2.3.4.4b can infect dairy cows and shed live virus in their milk. Sporadic cattle-to-human infections have been reported, highlighting the urgent need to understand its pathogenesis in mammals. Using both non-lactating and lactating BALB/c mice, we examined the viral tissue tropism, histopathological damages, and host immune responses upon intranasal inoculation with a reverse-genetic virus constructed based on A/dairy cattle/Texas/24-008749-003/2024 (Cattle-H5N1) and comparing with an older reference Clade 1 virus, A/Vietnam/1194/2004 virus (VNM1194-H5N1). Cattle-H5N1 was highly lethal in mice (mLD 50 =1.48PFU) with broad tissue tropism and produced higher titer in respiratory tissue and multiple extrapulmonary organs than VNM1194-H5N1. In the lungs, Cattle-H5N1 infection of airway epithelium, type II pneumocytes and CD45 + immune cells were at a higher frequency than those of VNM1194-H5N1-infected mice, resulting in severe epithelial destruction and diffuse alveolar damage accompanied by elevated lung and serum pro-inflammatory cytokine/chemokines. Although both H5N1 viruses showed lactating mammary gland tropism, the gland tissue was more severely damaged after Cattle-H5N1 infection with abundant viral antigens expression in glandular cells, associated fat and lymphoid tissues. Furthermore, more suckling mice co-housed with Cattle-H5N1 infected lactating mice were virus-positive (7/30 pups) than VNM1194-H5N1. Brains were heavily infected by Cattle-H5N1, and neurological signs such as body-rolling/spinning, trembling and/or limb paralysis were seen only in Cattle-H5N1 infected mice. The spleen was more severely damaged by Cattle-H5N1 infection, which showed massive viral antigen expression accompanied by severe apoptosis and splenic atrophy, concluding that Cattle-H5N1 is more virulent in mice than VNM1194-H5N1.

Background: Wild rodents and shrews serve as vital sentinel species for monitoring zoonotic viruses due to their close interaction with human environments and role as natural reservoirs for diverse viral pathogens. Although several studies have explored viral diversity and assessed pathogenic risks in wild rodents and shrews, the full extent of this diversity remains insufficiently understood., Results: We conducted high-throughput sequencing on 1113 small mammals collected from 97 townships across seven cities in Hubei Province during 2021, supplemented by publicly available data from 2014 and 2016-2017. This analysis revealed a diverse array of novel viruses spanning several viral families, including Arenaviridae, Hepeviridae, Chuviridae, Paramyxoviridae, Arteriviridae, Nodaviridae, Rhabdoviridae, Dicistroviridae, Astroviridae, and Picornaviridae. Phylogenetic analysis and genome structure characterization highlighted the discovery of these novel viruses, enhancing our understanding of viral diversity and evolution. Key host species such as Chodsigoa smithii, Anourosorex squamipes, Niviventer niviventer, and Apodemus agrarius were identified as significant contributors to viral circulation, making them crucial targets for future surveillance. Additionally, the central Plain of Hubei Province was recognized as a critical geographic hub for viral transmission, underscoring its importance in monitoring and controlling viral spread. Machine learning models were employed to assess the zoonotic potential of the identified viruses, revealing that families such as Arenaviridae, Coronaviridae, Hantaviridae, Arteriviridae, Astroviridae, Hepeviridae, Lispiviridae, Nairoviridae, Nodaviridae, Paramyxoviridae, Rhabdoviridae, Picornaviridae, and Picobirnaviridae possess a high likelihood of infecting humans. Notably, rodent-derived Rotavirus A, HTNV, and SEOV displayed almost complete amino acid identity with their human-derived counterparts, indicating a significant risk for human outbreaks., Conclusion: This study provides a comprehensive virome landscape for wild rodents and shrews in Central China, highlighting novel viruses and the critical roles of specific host species and regions in viral transmission. By identifying key species and hotspots for viral spread and assessing the zoonotic potential of the discovered viruses, this research enhances our understanding of virus ecology and the factors driving zoonotic disease emergence. The findings emphasize the need for targeted surveillance and proactive strategies to mitigate the risks of zoonotic spillovers, contributing to global public health preparedness. Video Abstract., Competing Interests: Declarations. Ethics approval and consent to participate: This study received approval from the Experimental Ethics Committee of the Hubei Provincial Center for Disease Control and Prevention (Project No. 2021–034–02). Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background and Aims: The impact of HBsAg reduction from small interfering RNA (siRNA) treatments on HBV-specific immunity of CHB participants has not been adequately analyzed in human. We conducted a phase 2a study treating CHB participants with 9 4-weekly doses of HBV-targeted siRNA elebsiran (BRII-835), either alone (n=10) or in combination with a VLP-based therapeutic vaccine (BRII-179) containing Pre-S1, Pre-S2, and S antigens, co-administered with (n=39) or without (n=41) IFNα., Methods: We analyzed longitudinally for 72 weeks virological, clinical, and immunological parameters including HBsAg, ALT, anti-HBs, the neutralizing activity of representative sera, and frequency and cytokine secretion ability of T cells specific for Pre-S1, Pre-S2, and S both directly ex vivo and after in vitro expansion., Results: Combination therapy of elebsiran and BRII-179 was well tolerated. While no sustained HBsAg seroclearance or notable difference in mean HBsAg reduction at the group level was observed, we detected marked heterogeneity in immunological responses among groups. HBsAg reduction mediated by siRNA alone was associated with minimal HBV-specific immune response recovery. In contrast, combination of elebsiran with BRII-179 induced a significant modification of immune responses demonstrated by anti-HBs antibody production and an expansion of IL-2-producing CD4+ T cells specific for Pre-S1/Pre-S2 antigens only. Importantly, anti-HBs antibodies persisted ≥100 IU/L in approximately 40% of the participants for at least 32 weeks after combinatory treatment. Moreover, the neutralizing ability of the anti-HBs positive sera was associated with HBsAg reduction., Conclusions: SiRNA-induced HBsAg reduction may contribute to the persistence and efficacy of the humoral arm of HBV-specific adaptive immunity in CHB participants receiving therapeutic vaccine BRII-179., (Copyright © 2025 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: Difficulties in microbiologically confirming childhood tuberculosis (TB) can result in delayed treatment and increased disease severity., Methods: In this study, we for the first time used whole genome next-generation sequencing (NGS) to detect cell-free DNA (cfDNA) from Mycobacterium tuberculosis (MTB) in plasma from children., Results: We enrolled 94 children with active TB and 32 children with other respiratory infections. Combining NGS with probe capture enrichment (targeted cfNGS) showed higher coverage and detecting capability than did NGS alone. The targeted cfNGS showed slightly lower sensitivity (31.9% vs. 44.7%, P = 0.072) and specificity (96.9% vs. 100.0%, P = 0.236) to those of sputum tested using Xpert. Agreement between cfNGS-plasma and Xpert-sputum was weak (κ = 0.217). Concordant results were obtained for only 85 children (67.5%; 16 cases positive by both tests and 69 cases negative by both tests). A total of 40 children with MTB culture negative results were tested to have positive cfNGS-plasma or Xpert-sputum outcomes, yielding a significantly increased percentage of children with bacteriological evidence (20.2% [19/94] for MTB culture-positive only vs. 62.8% [59/94] for cfNGS-plasma, Xpert-sputum or culture positive)., Conclusions: These data suggest that cfNGS performed well for diagnosing TB using plasma from children. cfNGS may be a new method for diagnosing patients with paucibacillary TB., Competing Interests: Declarations. Ethics approval and consent to participate: This study was approved by the Ethics Committee of Beijing Children’s Hospital affiliated with Capital Medical University (ID:2018-96). The patients’ guardians provided written informed consent. In cases of illiteracy, a witness attested to informed oral consent. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Oropouche virus (OROV), first detected in Trinidad and Tobago in 1955, was historically confined to the Brazilian Amazon Basin. However, since late 2022, an increasing number of OROV cases have been reported across various regions ofBrazilas well as in urban centers in Bolivia, Ecuador, Guyana, Colombia, Cuba, Panama and Peru. In collaboration with Central Public Health Laboratories across Brazil, we integrated epidemiological metadata with genomic analyses from recent cases, generating 133 whole-genome sequences covering the virus's three genomic segments (L, M, and S). These include the first genomes from regions outside the Amazon and from the first recorded fatal cases. Phylogenetic analyses show that the 2024 OROV genomes form a monophyletic group with sequences from the Amazon Basin sampled since 2022, revealing a rapid north-to-south viral movement into historically non-endemic areas. We identified 21 reassortment events, though it remains unclear whether these genomic changes have facilitated viral adaptation to local ecological conditions or contributed to phenotypic traits of public health significance. Our findings demonstrate how OROV has evolved through reassortment and spread rapidly across multiple states in Brazil, leading to the largest outbreak ever recorded outside the Amazon and the first confirmed fatalities. Additionally, by analysing travel-related cases, we provide the first insights into the international spread of OROV beyond Brazil, further highlighting the role of human mobility in its dissemination. The virus's recent rapid geographic expansion and the emergence of severe cases emphasize the urgent need for enhanced surveillance across the Americas. In the absence of significant human population changes over the past two years, factors such as viral adaptation, deforestation, and climate shifts-either individually or in combination-may have facilitated the spread of OROV beyond the Amazon Basin through both local and travel-associated transmission., (© The Author(s) 2025. Published by Oxford University Press on behalf of International Society of Travel Medicine.)

This review addresses the significant advancements in the identification of blood-based prognostic biomarkers for tuberculosis (TB), highlighting the importance of early detection to prevent disease progression. The manuscript discusses various biomarker categories, including transcriptomic, proteomic, metabolomic, immune cell-based, cytokine-based, and antibody response-based markers, emphasizing their potential in predicting TB incidence. Despite promising results, practical application is hindered by high costs, technical complexities, and the need for extensive validation across diverse populations. Transcriptomic biomarkers, such as the Risk16 signature, show high sensitivity and specificity, while proteomic and metabolic markers provide insights into protein-level changes and biochemical alterations linked to TB. Immune cell and cytokine markers offer real-time data on the body's response to infection. The manuscript also explores the role of single-nucleotide polymorphisms in TB susceptibility and the challenges of implementing novel RNA signatures as point-of-care tests in low-resource settings. The review concludes that, while significant progress has been made, further research and development are necessary to refine these biomarkers, improve their practical application, and achieve the World Health Organization's TB elimination goals., Competing Interests: Conflict of interest The authors declare no competing interests., (Copyright © 2025 Elsevier GmbH. All rights reserved.)

Background: Crimean-Congo hemorrhagic fever is a tick-borne zoonotic disease that may be severe and is present in many African countries. We aimed to understand the seroprevalence and risk for Crimean-Congo hemorrhagic fever virus in Tanzania by testing archived serum samples from patients enrolled in a prospective cohort study., Methods: We prospectively enrolled febrile inpatients and outpatients from 2012 through 2014 at two referral hospitals in northern Tanzania. Archived serum samples were tested for Crimean-Congo hemorrhagic fever virus antibodies initially by a Luminex assay screen followed by confirmation with immunofluorescence assay. Evidence of exposure to Crimean-Congo hemorrhagic fever virus was defined as antibody detection by Luminex and confirmed by immunofluorescence assay. Questionnaire data were used to construct logistic regression models to understand factors associated with prior exposure to Crimean-Congo hemorrhagic fever virus. Domains of predictor variables included sociodemographics, livestock-rearing activities, and environmental factors., Results: Of 735 participants included, antibodies to Crimean-Congo hemorrhagic fever virus nucleocapsid protein were detected by Luminex assay in 23 (3.1%) and confirmed by immunofluorescence assay in 13 (1.8%). In multivariable logistic regression, prior exposure to Crimean-Congo hemorrhagic fever virus was associated with self-report of milking livestock in the past month (adjusted OR [aOR]: 12.6, 95% CI 1.6-99.8) and natural log increase in goat density (head/km 2 ; aOR: 1.7, 95% CI: 1.1-2.7)., Conclusions: We show serologic evidence of prior exposure to Crimean-Congo hemorrhagic fever virus among humans in northern Tanzania. Similar to other settings, our results suggest that exposure is closely linked to livestock activities. Additional research is warranted to understand reservoirs and modes of transmission of Crimean-Congo hemorrhagic fever virus to humans in northern Tanzania., (© 2025 John Wiley & Sons Ltd.)

In recent years, outbreak of the novel duck reovirus (NDRV) disease has occurred frequently in duck populations. Due to its rapid spreading, absence of effective control methods, and high treatment costs, the NDRV disease has caused huge losses to waterfowl breeding in China. As reported, four non-structural (NS) proteins are encoded by the NDRV genome, among which the σNS protein is an RNA-binding protein that can improve the stability of bound RNA by forming oligomers (Adams and Cory, 1998). Nevertheless, the mechanism by which it facilitates reovirus replication remains ambiguous. According to previous studies, the NS protein 11 of the porcine reproductive and respiratory syndrome virus (PRRSV) can interact with tripartite motif-containing 59 (TRIM59) to regulate viral infection. However, the specific role of TRIM59 in NDRV infection remains unclear. This study focused on full-length amplification of duTRIM59, the mRNA distribution of duTRIM59 in Cherry Valley duck and successive biological examinations. The homology with Anas platyrhynchos TRIM59 was 98.6 %. The mRNA distribution level of duTRIM59 showed that duTRIM59 was widely expressed in bursae and thymus of the immune organs. Nevertheless, TRIM59 comprises three domains, including the transmembrane (TM), B-box (B), and RING-finger (R) domains. It also has the activity of ubiquitin-protein ligase (E3). It has been demonstrated that NDRV replication is inhibited by TRIM59 overexpression in duck embryonic fibroblasts (DEF) cells, particularly when the R domain is intact, suggesting that the R domain plays a key role in the spreading of the NDRV virus. In contrast, NDRV infection in DEF cells increased when TRIM59 was depleted by using small interfering RNA. Moreover, the σNS protein can be co-localized with duTRIM59 and stimulate NDRV replication in DEF cells in cases of NDRV infection. This study clarifies the correlation of NDRV infection and TRIM59-mediated antiviral innate immunity, and provides a sound theoretical basis for further understanding this disease., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest., (Copyright © 2025. Published by Elsevier B.V.)

Background: Acute febrile illness is a common reason for seeking healthcare in low- and middle-income countries. We describe the diagnostic utility of a TaqMan Array Card (TAC) real-time polymerase chain reaction (PCR) panel for pathogen detection in paediatric and adult inpatients admitted with febrile illness., Methods: In this prospective cohort study, we screened medical admissions for a tympanic temperature ≥38.0°C or reported fever within 72 h and used a PCR panel to detect pathogens, including bacteria, viruses, fungi and protozoa, in 697 participants. We compared PCR results to conventional diagnostic methods and considered PCR detections as the cause of fever, except for Plasmodium spp. and Schistosoma spp. Participants for PCR testing was consecutively selected from the end of enrolment., Results: Of 1132 participants enrolled in the cohort, 697 (61.6%) were tested by PCR. Median (IQR) age was 29.6 (4.6-46.4) years. Three hundred seventy-eight (54.2%) were male. The PCR method improved illness identification, increasing diagnostic yield from 73 (10.5%) by conventional methods to 124 (17.8%) of 697 participants. PCR detections included four viral pathogens: dengue (n = 1), enterovirus (n = 7), measles (n = 1) and Rift Valley Fever Virus (RVFV) (n = 3). Forty-six bacterial pathogens were detected in 44 (6.3%) participants, including fastidious bacteria such as Bartonella spp. (n = 2), Brucella spp. (n = 3), Coxiella burnetii (n = 2), Leptospira spp. (n = 1), M. tuberculosis (n = 7) and Rickettsia spp. (n = 9)., Conclusion: The PCR panel improved pathogen detection in febrile inpatients, providing clinically actionable results for fastidious bacteria and epidemiologically relevant findings like RVFV detections, when combined with conventional methods., (© 2024 John Wiley & Sons Ltd.)

Background: Usutu virus (USUV) and West Nile virus (WNV) are zoonotic arthropod-borne orthoflaviviruses. The enzootic transmission cycles of both include Culex mosquitoes as vectors and birds as amplifying hosts. For more than 10 years, these viruses have been monitored in birds in Germany by a multidisciplinary network. While USUV is present nationwide, WNV used to be restricted to the central-east., Methods: In 2021 and 2022, over 2300 live bird blood samples and organs from over 3000 deceased birds were subjected to molecular and serological analysis regarding the presence of WNV and USUV. The samples were collected at sites all over Germany., Results: Circulation of both viruses increased in 2022. For USUV, the nationwide presence of lineages Africa 3 and Europe 3 reported in previous years was confirmed. Lineage Europe 2, formerly restricted to the German east, was able to expand westward. Nonetheless, USUV neutralizing antibody (nAb) detection rates remained low (< 9%). Years 2021 and 2022 were characterized by stable enzootic circulation of WNV lineage 2, dominated by one previously identified subcluster (95% of generated sequences). In 2022, >20% of birds in the endemic region in eastern Germany carried nAb against WNV. Serological data also indicate expanding WNV circulation west and south of the known hotspots in Germany., Conclusions: USUV circulates enzootically nationwide. Emergence of WNV at several new locations in Germany with a potential increase in human infections may be imminent. In this context, wild bird monitoring serves as a capable early warning system in a One Health setting.

Rationale: The global burden of sepsis is concentrated in sub-Saharan Africa, where inciting pathogens are diverse and HIV coinfection is a major driver of poor outcomes. Biological heterogeneity inherent to sepsis in this setting is poorly defined. Objectives: To identify dominant pathobiological signatures of sepsis in sub-Saharan Africa and their relationship to clinical phenotypes, patient outcomes, and biological classifications of sepsis identified in high-income countries (HICs). Methods: We analyzed two prospective cohorts of adults hospitalized with sepsis (severe infection with quick Sepsis-related Organ Failure Assessment score ⩾1) at disparate settings in Uganda (discovery cohort [Entebbe, urban], n  = 242; validation cohort [Tororo, rural], n  = 253). To identify pathobiological signatures in the discovery cohort, we applied unsupervised clustering to 173 soluble proteins reflecting key domains of the host response to severe infection. A random forest-derived classifier was used to predict signature assignment in the validation cohort. Measurements and Main Results: Two signatures (Uganda Sepsis Signature [USS]-1 and USS-2) were identified in the discovery cohort, distinguished by expression of proteins involved in myeloid cell and inflammasome activation, T-cell costimulation and exhaustion, and endothelial barrier dysfunction. A five-protein classifier (area under the receiver operating characteristic curve, 0.97) reproduced two signatures in the validation cohort with similar biological profiles. In both cohorts, USS-2 mapped to a more severe clinical phenotype associated with HIV and related immunosuppression, severe tuberculosis, and increased risk of 30-day mortality. Substantial biological overlap was observed between USS-2 and hyperinflammatory and reactive sepsis phenotypes identified in HICs. Conclusions: We identified prognostically enriched pathobiological signatures among patients with sepsis with diverse infections and high HIV prevalence in Uganda. Globally inclusive investigations are needed to define generalizable and context-specific mechanisms of sepsis pathobiology, with the goal of improving access to precision medicine treatment strategies.

Titanium dioxide nanoparticles (nano-TiO 2 ) can cause a reduction in sperm counts, and lactate production in Sertoli cells plays a key role in spermatogenesis. The aim of this study was to evaluate the effect of nano-TiO 2 on lactate production in mouse Sertoli cell line (TM4 cells) and to investigate whether the effect is mediated through the PI3K/AKT/HIF-1α pathway. After TM4 cells were treated with different concentrations of nano-TiO 2 for 48 h, cell viability, contents of glucose and lactate, and the expression levels of GLUT3 and key enzymes (HK1, HK2, PFKM, ENO1, LDH) during lactate production were detected. PI3K/AKT/HIF-1α pathway proteins were measured. In addition, PI3K agonist (IGF-1) was added to explore whether nano-TiO 2 regulates HIF-1α through PI3K/AKT pathway, thereby affecting the production of lactate in TM4 cells. The results showed that nano-TiO 2 significantly inhibited TM4 cell viability, increased glucose content, decreased lactate content, and downregulated the expression levels of GLUT3 and key enzymes during lactate production. Meanwhile, nano-TiO 2 decreased the expression of PI3K/AKT pathway phosphorylated proteins and HIF-1α, and IGF attenuated this effect of nano-TiO 2 . Collectively, nano-TiO 2 downregulated the expression level of proteins and enzymes related to lactate production in TM4 cells by inhibiting PI3K/AKT/HIF-1α pathway, resulting in the decrease of lactate production in TM4 cells., (© 2024 John Wiley & Sons Ltd.)

Acute respiratory infections (ARIs) stand as a significant cause of morbidity and mortality among children worldwide, contributing substantially to paediatric hospitalisation rates. ARIs stem from various pathogens, including bacteria, viruses, among others. With the advent of novel diagnostic techniques like molecular detection methods, the identification rate of multiple pathogens in paediatric ARIs is steadily rising. However, there is currently no consensus on the impact of mixed infections on the severity of respiratory infections in children. This narrative review summarises existing research indicating that the co-detection rate of multiple viruses among paediatric patients with ARIs ranged from 0.07% to 55%. Multi-virus coinfections did not appear to increase the severity of the disease in children because of viral interference, immune modulation, etc. Conversely, mixed infection of virus and bacteria may exacerbate disease severity through many mechanisms, such as synergistic activation of inflammation, diminished repair efficiency, increased transmission and release and so on. The insights provide aim to improve diagnostic precision and treatment strategies for paediatric ARIs, ultimately reducing complications and mortality rates associated with ARIs in children., (© 2025 John Wiley & Sons Ltd.)

Organoids were successfully established from primary tumor and its metastatic lymph nodes of a patient. These organoids faithfully replicated tumor pathology and genetic characteristics. Organoid-based drug screening was conducted, which revealed significant difference in sensitivity to drugs between organoids dervived from primary tumor versus metastatic lymph nodes. The results guided clinical decisions for personalized treatment for the patient. This approach provides an insightful strategy for advancing treatment for gastric cancer., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The author Leli Zeng is the editorial board of Drug Resistance Updates. The other authors have no competing interests. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier Ltd.)

The α-Klotho is crucial for human health and longevity. However, the relationship between trace elements and α-Klotho levels needs further investigation. We aimed to explore the relationship between serum levels of selenium (Se), copper (Cu), and zinc (Zn), and serum α-Klotho levels. We analyzed 2138 samples from the 2011-2016 National Health and Nutrition Examination Survey, and the weighted linear regression, WQS, and qgcomp models were utilized to evaluate the effects of these elements on serum α-Klotho levels, individually and combined. A negative correlation was observed between serum Cu concentration and serum α-Klotho levels (β =  - 0.128, 95% CI - 0.196, - 0.059), with each increase in Cu concentration grade showing a gradual decrease in serum α-Klotho levels (P trend  = 0.002). The WQS model exhibited a negative correlation between the combined effect of Se, Cu, and Zn and serum α-Klotho levels (β =  - 0.035, 95%CI - 0.060, - 0.010), consistently in males (β =  - 0.038 (- 0.059, - 0.017)) and in the 40-49 age group (β =  - 0.059, 95% CI - 0.119, - 0.012). The qgcomp model mirrored these findings, showing a negative correlation in the combined effect index of Se, Cu, and Zn with serum α-Klotho levels (β =  - 0.027, 95% CI - 0.047, - 0.006), consistent in females (β =  - 0.032, 95% CI - 0.061, - 0.004) and in individuals with BMI ≥ 25 (β =  - 0.030, 95% CI - 0.054, - 0.006), and in the 40-49 age group (β =  - 0.047, 95% CI - 0.088, - 0.006). Elevated serum Cu levels may be associated with lower serum α-Klotho levels. The combined effect of serum Se, Cu, and Zn shows a negative correlation with serum α-Klotho levels, with Cu contributing the most. Our findings provide significant insights into assessing the role of trace nutrients in maintaining human health., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: Streptococcus suis serotype 14 is the second most prevalent serotype being highly prevalent in Southeast Asia. This study aimed to characterize genetic background, population structure, virulent genes, antimicrobial-resistant genes, and virulence of human S. suis serotype 14., Methods: Genomes of 11 S. suis serotype 14 were sequenced by short- and long-read sequencing platforms. The genomes were analyzed for genetic relationship, virulence-associated genes, and antimicrobial-resistant genes. Antimicrobial susceptibility was conducted and the virulence was tested based on cell assay., Results: All isolates belonged to clonal complex (CC) 1, with nine sequence type (ST) 105 isolates and each isolate of ST1 and ST237. They were susceptible to penicillin, whereas tetracycline and macrolide were resistance due to tetO and ermB. Genomic analysis revealed that the serotype 14-ST105 isolates were closely related to zoonotic serotype 14-ST105 isolates from Vietnam and the serotype 1-ST105 Thai strain. The serotype 14-ST1 isolate was closely related to pig-diseased serotype 1-ST1 isolates from UK and USA, whereas the serotype 14-ST237 isolate was related to serotype 1-ST237 strains recovered from healthy pig from Thailand. Of 150 virulence-associated genes, 13 were absent from the serotype 14 isolates, including atl1, atlAss, hhly3, nisK, nisR, pnuC, salK, salR, sp1, srtG, virB4, virD4, and zmp. The virulence of strain 32481, a representative S. suis serotype 14-ST105 isolate showed reduced adhesion and invasion of two epithelial cell lines (A549 and HeLa) when compared to the serotype 2-ST1 strain P1/7, whereas apoptosis was similar., Conclusion: This study highlighted the pathogenic potential of virulent serotype 14-ST105 strains and the need for increased monitoring of S. suis serotypes other than for serotype 2., Competing Interests: Declarations. Ethical approval: The stored bacterial isolates were used in the current study that were not considered to be involved in human subject research. Therefore, ethical review and approval were exempted because no human specimens or data were used in the current study. The study was carried out in compliance with relevant laws and guidelines. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The comprehensive antibiotic resistance of pathogens signifies the oneset of the "post-antibiotic era", and the myriad treatment challenges posed by "superbugs" have emerged as the primary threat to human health. Recent studies indicate that bacterial resistance and tolerance development are mediated at the metabolic level by various signaling networks (e.g., quorum sensing systems, second messenger systems, and two-component systems), resulting in metabolic rearrangements and alterations in bacterial community behavior. This review focuses on current research, highlighting the intrinsic link between signaling and metabolic networks in bacterial resistance and tolerance., (Copyright © 2025. Published by Elsevier Ltd.)

Background: Immune senescence can occur in untreated HIV infection and is partially reversible with effective antiretroviral therapy (ART). Here, we investigated the differences in immune senescence of T cell subsets among Chinese men who have sex with men (MSM) living with HIV virologically suppressed on ART compared to those ART-naive., Methods: A cohort of MSM living with HIV with different disease courses was included, untreated viral non-controllers (n = 26) and those on ART (n = 30). The percentages of naive, central memory (TCM), effector memory (TEM), and terminally differentiated memory (TemRA) subsets of CD4 and CD8 T cells were studied, along with markers of senescence (CD28-CD57+) and activation (HLA-DR). Telomere length of naive and memory CD8 T cells was quantified by real-time PCR. The correlation between senescent CD4 and CD8 T cell subsets and CD4 and CD8 cell counts was analyzed with the Spearman rank correlation., Results: Compared with the ART-naive group, the percentage of senescent cells (CD28- CD57+) in total CD8 T cells was significantly lower in the ART group (P < 0.01). Significant differences were observed among CD8 T cell subsets, but not in CD4 T cell subsets (P < 0.05). In the ART group, the percentage of senescent cells (CD28-CD57+) in TN and TCM subsets of both CD4 and CD8 T cells was lower (all P < 0.05). HLA-DR expression was significantly lower in all CD4 and CD8 T cell subsets except TEMRA subset (P < 0.05). The telomere length of CD8 T cell subsets did not differ significantly between the two groups. The percentage of senescent naive CD4 T cells was inversely correlated with CD4 T cell counts (r = -0.42, P = 0.0343), while the percentage of senescent naive CD8 T cells was positively correlated with CD8 T cell counts (r = 0.47, P = 0.0161) in the ART-naive group, but not in the ART group., Conclusions: Virologically suppressed MSM living with HIV exhibit lower immune senescence of T cell subsets, which is more pronounced for CD8 cell subsets. The percentage of senescent naive T cells is significantly correlated with clinical immunity based on CD4 and CD8 T cell counts., Competing Interests: Declarations. Ethics approval and consent to participate: Written informed consent was obtained from each participant. The ethics committee of Beijing Ditan Hospital of Capital Medical University approved the study (Approval number: JDL034-002). The study was carried out in accordance with the tenets of the Declaration of Helsinki. Consent to publish: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

This study identifies avian tuberculosis as a potential cause of mass mortality in wild migratory birds in Inner Mongolia, China. Combining meta-transcriptomic sequencing and histopathological analysis, it reveals one of the rare instances of tuberculosis-associated outbreaks in avian populations. These findings underscore the importance of surveillance on wildlife diseases to mitigate the risk of interspecies transmission of the disease associated pathogens and their broader implications for biodiversity and public health., Competing Interests: The authors declare no conflict of interests., (© 2025 The Author(s). mLife published by John Wiley & Sons Australia, Ltd on behalf of Institute of Microbiology, Chinese Academy of Sciences.)

Objectives: Human parainfluenza viruses (hPIV) are a common cause of acute respiratory infections, especially in children under five years and the elderly. hPIV can be subclassified as types 1-4: these showed various seasonality patterns worldwide, and it is unclear how climatic factors might consistently explain their global epidemiology., Methods: This study collected time-series incidence data from the literature and hPIV surveillance programs worldwide (47 locations). Wavelet analysis and circular statistics were used to detect the seasonality and the months of peak incidence for each hPIV type. Relationships between climatic drivers and incidence peaks were assessed using a generalized estimating equation., Results: The average positive rate of hPIV among patients with respiratory symptoms was 5.6% and ranged between 0.69-3.48% for different types. In the northern temperate region, the median peak incidence months for hPIV1, hPIV2, and hPIV4 were from September to October, while for hPIV3, it was in late May. Seasonal peaks of hPIV3 were associated with higher monthly temperatures and lower diurnal temperatures range throughout the year; hPIV4 peaks appeared to correlate with lower monthly temperatures and higher precipitation throughout the year. Different hPIV types exhibit different patterns of global epidemiology and transmission., Conclusions: Climate drivers may play a role in hPIV transmission. More comprehensive and coherent surveillance of hPIV types would enable more in-depth analyses and inform the timing of preventive measures., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier Ltd.)

Background and Aims: The objective was to describe the dynamics of noninvasive tests (NITs) in cirrhotic patients following sustained virological response (SVR) and to assess their correlation with hepatocellular carcinoma (HCC) risk., Methods: The dynamics of NITs (Fib4, APRI and LSM) were described in patients with cirrhosis after SVR included between 2006 and 2015 in two prospective French multicentre cohorts (ANRS CO22 Hepather and CO12 CirVir). To assess their relationship with the risk of HCC, a joint modeling approach was employed using both standard and flexible models adjusted for age and sex. The impacts of NIT current value and slope during follow-up on HCC risk were assessed taking into account competing risks of death., Results: 3067 patients with cirrhosis who achieved SVR were analyzed, among whom 228 (7.4%) developed HCC and 210 (6.9%) died during a 26-month follow-up. All NITs were increased at baseline in patients who ultimately developed HCC, whereas platelet counts were lower. All NITs improved in patients who did not develop HCC. More contrasted changes were observed during the follow-up of patients who ultimately developed HCC. Joint model analyses showed that current values of Fib4, APRI and platelet count at any time impacted HCC risk. Only Fib4 and APRI slopes influenced the same outcome. When considering NIT current value and slope simultaneously, only the current value of NITs impacted HCC risk while the slopes were not informative., Conclusions: The dynamics of NITs following SVR do not identify cirrhotic patients who could be safely excluded from surveillance programmes. NIT current value is more informative than slope which would necessitate to regularly re-assess HCC risk to design individualized surveillance strategies., Impact and Implications: It has been postulated that that monitoring noninvasive tests (NIT) dynamics following HCV cure may inform on HCC residual risk in patients with cirrhosis, and may allow for the discontinuation of surveillance in certain patient subsets. We analyzed data from over 3,000 patients and found that while all NITs improved in cirrhotic patients who did not develop HCC, those who eventually developed liver cancer showed more contrasted changes in these tests. Specifically, the current values of tests like Fib4 and APRI were linked to an increased risk of HCC. while their slopes did not provide additional useful information, suggesting that dedicated prospective studies are warranted to define how repeated measurement of NIT could be combined with other variables into HCC risk stratification algorithms. Until then, HCC surveillance should be maintained in all patients with cirrhosis following HCV eradication, even in case of decreased NIT., Competing Interests: Declaration of Competing Interest None to declare., (Copyright © 2025 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Emerging tick-borne orthonairovirus infections pose a growing global concern, with limited understanding of the viral ovarian tumor-like cysteine proteases (vOTUs) encoded by novel orthonairoviruses. These vOTUs, a group of deubiquinylases (DUBs), disrupt the innate immune response. Yezo virus (YEZV), a recently discovered pathogenic orthonairovirus, was first reported in Japan in 2021. In this study, we successfully isolated and identified YEZV and a new orthonairovirus, Jiànchuān tick virus (JCTV), from Ixodes persulcatus and Haemaphysalis montgomeryi ticks, respectively, in China. We found that the vOTU domains encoded by YEZV and JCTV exhibited both DUB and deISGylase activities, though with potentially less broad deISGylation compared to that of Crimean-Congo hemorrhagic fever virus (CCHFV) during natural infection. Phylogenetic analysis of global vOTUs, including 83 new sequences, revealed a high diversity of this domain. Interestingly, retrospective screening of tick-bite patients from 2012 to 2016 in northeastern China traced YEZV infections as far back as 2012, identifying four cases. Additionally, YEZV primarily infected I. persulcatus (31.4%) and Dermacentor nuttalli (10.5%) in northern China, while JCTV exhibited high infection rates in H. montgomeryi (81.3%) in southern China. In summary, our work emphasizes the active surveillance of orthonairovirus infections and the imperative need for the development of vOTU domain-targeted anti-virals, offering potential therapeutic solutions for a broad spectrum of orthonairoviruses.IMPORTANCEThe vOTUs, a group of DUBs, mimic the functions of host DUBs to enhance viral infectivity and may serve as potential drug targets. vOTUs from different orthonairoviruses exhibit distinct preferences toward ubiquitin (Ub) and ubiquitin-like protein interferon stimulated gene 15 (ISG15). In this study, we investigated the deubiquitinase and deISGylase functions of various orthonairoviral vOTUs using both an overexpression system and natural viral infections in vitro . Our findings illustrate that the vOTUs from YEZV and JCTV can cleave both Ub and ISG15 in an overexpression system, but these viruses exhibit potentially narrower deISGylation capacity than CCHFV during natural infection. This suggests that the diversity of vOTUs may have a potential relationship with the pathogenesis., Competing Interests: The authors declare no conflict of interest.

Mosquito-borne flaviviruses, such as dengue virus (DENV), Zika virus (ZIKV), West Nile virus, and yellow fever virus, pose significant public health threats globally. Extensive efforts have led to the development of promising highly active compounds against DENV targeting viral non-structural protein 4B (NS4B) protein. However, due to the cocirculation of flaviviruses and to prepare for emerging flaviviruses, there is a need for more broadly acting antivirals. Host-directed therapy where one targets a host factor required for viral replication may be active against multiple viruses that use similar replication strategies. Here, we used a CRISPR-Cas9 library that we designed to target the druggable genome and identified signal peptide peptidase (SPP, encoded by Histocompatibility Minor 13, HM13), as a critical host factor in DENV infection. Genetic knockout or introducing mutations that disrupt the proteolytic activity of SPP markedly reduced the replication of multiple flaviviruses. Although their substrates differ, SPP has structural homology with γ-secretase, which has been pursued as a pharmacological target for Alzheimer's disease. Notably, SPP-targeting compounds exhibited potent anti-DENV activity at low nanomolar concentrations across multiple primary and disease-relevant cell types, acting specifically through SPP inhibition rather than γ-secretase inhibition. Importantly, SPP inhibitors were active at low nanomolar concentrations against flaviviruses other than DENV including ZIKV while DENV NS4B inhibitors lost activity. This study emphasizes the strong potential of SPP as a pan-flaviviral target and provides a framework for identifying host druggable targets to screen for broad-spectrum antivirals., Competing Interests: Competing interests statement:The authors declare no competing interest.

The role of mesenchymal cells during respiratory infection is not well defined, including whether, which, and how the different types of mesenchymal cells respond. We collected all mesenchymal cells from lung single-cell suspensions of mice that were naïve (after receiving only saline vehicle), pneumonic (after intratracheal instillation of pneumococcus 24 hours previously), or resolved from infection (after non-lethal pneumococcal infections 6 weeks previously) and performed single-cell RNA sequencing. Cells clustered into five well-separated groups based on their transcriptomes: matrix fibroblasts, myofibroblasts, pericytes, smooth muscle cells, and mesothelial cells. Fibroblasts were the most abundant and could be further segregated into Pdgfra+Npnt+Ces1d+Col13a1+ alveolar fibroblasts and Cd9+Pi16+Sca1+Col14a1+ adventitial fibroblasts. The cells from naïve and resolved groups overlapped in dimension reduction plots, suggesting the mesenchymal cells returned to baseline transcriptomes after resolution. During pneumonia, all mesenchymal cells responded with altered transcriptomes, revealing a core response that had been conserved across cell types as well as distinct mesenchymal cell type-specific responses. The different subsets of fibroblasts induced similar gene sets, but the alveolar fibroblasts responded more strongly than the adventitial fibroblasts. These data demonstrated diverse and specialized immune activities of lung mesenchymal cells during pneumonia.

Influenza A virus transcribes viral mRNAs from the eight segmented viral genome when it infects. The kinetics of viral transcription, nuclear export of viral transcripts, and their potential variation between the eight segments are poorly characterised. Here, we introduce a statistical framework for estimating the nuclear export rate of each segment from a snapshot of in situ mRNA localisation. This exploits the cell-to-cell variation at a single time point observed by an imaging-based in situ transcriptome assay. Using our model, we revealed the variation in the mRNA nuclear export rate of the eight viral segments. Notably, the two influenza viral antigens hemagglutinin and neuraminidase were the slowest segments in the nuclear export, suggesting the possibility that influenza A virus uses the nuclear retention of viral transcripts to delay the expression of antigenic molecules. Our framework presented in this study can be widely used for investigating the nuclear retention of nascent transcripts produced in a transcription burst., Competing Interests: MM, NK, SL, BM, HU, TN, HC, MS No competing interests declared, (© 2023, Miura et al.)

Background: Bats and rodents have been identified as reservoirs for several highly pathogenic and zoonotic viruses including henipaviruses, a genus within the Paramyxoviridae family. A number of studies have revealed the circulation of henipaviruses at the wildlife-human-livestock interface in Cameroon. In this study, we describe the molecular analysis as well as the development and evaluation of a Bead-based Multiplex Binding Assay (BMBA) using an in-house Indirect Enzyme Linked Immunosorbent Assay (ELISA) to confirm the detection of henipavirus infection in wildlife species., Results: A total of 600 fruit bats and 600 rodents were sampled between March 2018 and June 2020. Samples were analyzed using a semi-nested RT-PCR assay followed by sequencing of the PCR fragments. Transudates (754) were screened for the presence of henipavirus-specific antibodies in a BMBA and confirmed by ELISA using Hendra virus (HeV), Nipah virus (NiV) and Ghana virus (GhV) glycoproteins expressed in Leishmania tarentolae, and commercially available HeV G and NiV G glycoproteins. Henipavirus-specific antibodies were detected in 19/531 (3.6%) bat transudates screened by BMBA and confirmed by ELISA. Seroprevalence rates in the Centre and North Regions were 12/291 (4.1%) and 7/240 (2.9%) respectively. All rodents and shrews were serologically negative. Henipavirus RNA sequences were not detected in any of the samples screened in this work., Conclusion: This study provides further data supporting the circulation of Henipaviruses in fruit bats (Eidolon helvum) which are roosting and reproducing in proximity to human and livestock populations in the Centre and North Regions of Cameroon. This also establishes the first detection of Henipavirus specific antibodies in Eidolon helvum populations in the North Region of Cameroon., Competing Interests: Declarations. Ethics approval and consent to participate: Approvals for bat and rodent sampling were obtained from the Cameroon Ministry of Research and Scientific Innovation – MINRESI with reference number of permit No000000311/MINRESI/B00/C00/C10/C12 and the Ministry of Wildlife and Forestry – MINFOF with reference number of permit No0183 PRS/MINFOF/SG/DFAP/SDVEF/SC/MC, and were renewed yearly during the sampling period from March 2018 to June 2020. Assessment and review of the project protocol prior to approval was done by the Division of Scientific Policy and Planning of the MINRESI and the Department of Wildlife and Protected Areas of the MINFOF. This study was conducted in strict accordance with the Cameroon’s national guidelines and regulations. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Recent avian-origin H3N8 influenza A virus (IAV) that have infected humans pose a potential public health concern. Alterations in the viral surface glycoprotein, hemagglutinin (HA), are typically required for IAVs to cross the species barrier for adaptation to a new host, but whether H3N8 has adapted to infect humans remains elusive. The observation of a degenerative codon in position 228 of HA in human H3N8 A/Henan/4-10/2022 protein sequence, which could be residue G or S, suggests a dynamic viral adaptation for human infection. Previously, we found this human-isolated virus has shown the ability to transmit between ferrets via respiratory droplets, with the HA-G228S substitution mutation emerging as a critical determinant for the airborne transmission of the virus in ferrets. Here, we investigated the receptor-binding properties of these two H3N8 HAs. Our results showed H3N8 HAs have dual receptor-binding properties with a preference for avian receptor binding, and G228S slightly increased binding to human receptors. Cryo-electron microscopy structures of the two H3N8 HAs with avian and human receptor analogs revealed the basis for dual receptor binding. Mutagenesis studies reveal that the Q226L mutation shifts H3N8 HA's receptor preference from avian to human, while the G228S substitution enhances binding to both receptor types. H3N8 exhibits distinct antigenic sites compared to H3N2, prompting concerns regarding vaccine efficacy. These findings suggest that the current H3N8 human isolates are yet to adapt for efficient human-to-human transmission and further continuous surveillance should be implemented.IMPORTANCEInfluenza virus transmission remains a public health concern currently. H3N8 subtype influenza A viruses infect humans and their HAs acquire the ability to bind to both human and avian receptors, posing a threat to human health. We have solved and analyzed the structural basis of dual receptor binding of recently human-infecting H3N8 HA, and we demonstrate that the G228S enhances human receptor binding and adaptation. We also found that HN/4-10 H3N8 HA has distinct antigenic sites, which challenges vaccine efficacy. Taken together, our work is critical to the prevention and control of human H3 influenza virus infection.

Geckos are generally small, predominantly nocturnal reptiles, with several species commonly found close to human habitations. However, little is known about viral diversity in geckos. Using meta-transcriptomic sequencing we identified four novel RNA viruses - provisionally denoted Gecko astrovirus, Gecko parechovirus, Gecko reptillovirus, and Gecko hartmanivirus - in geckos sampled in October 2019 from Hainan Province, China. The presence of these viruses was confirmed by reverse transcription (RT)-PCR. Phylogenetic analyses revealed that these viruses were most closely related to those identified in various gecko species from China and Australia, such that they represent gecko-specific lineages, yet were also genetically distinct, with amino acid sequence identities to their closest relatives ranging from 38.6 % to 74.2 %. A co-phylogeny analysis revealed a complex interplay between long-term virus-host co-divergence and more recent host jumping, which differed in frequency among groups. In sum, we demonstrate the presence of four novel gecko-associated RNA viruses, expanding our understanding of viral diversity in these common animal species., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier B.V.)

Sexually Transmitted Infections (STIs) seriously affect population morbidity, mortality, and are a major public health problem worldwide. A cross-sectional study was conducted using the latest Demographic and Health Survey (DHS) data published between 2005 and 2022 for five Southeast Asian countries. The study's focus was on STI risk factors in the DHS of Timor-Leste, Philippines, Indonesia, Myanmar, and Cambodia. The study included 139,062 women of reproductive age, aged 15 to 49 years, who reported whether they had experienced an STI or its symptoms and had comprehensive information on the variables of interest, including socio-demographics, sexual behaviour and knowledge. The report shows that the prevalence of STIs or symptoms of STIs in 5 Southeast Asian (SEA) countries included in the study is 11.6%, with the highest prevalence found in Cambodia, and the lowest in Philippines. Women aged 15-24 years in Indonesia (17.08%) and Philippines (9.81%), had a higher risk of STIs than those aged 25-49 years. In most studied countries of SEA (Cambodia, Indonesia and Myanmar), poorer women had higher risk of STIs. Also, in most SEA countries (Cambodia, Indonesia, and the Philippines), knowledge of STIs was significantly associated with having an STI in women aged 15-49 years. Sexual behaviour factors showed that acceptance of beating a wife for refusing to have sex with her husband was associated with STIs in all countries except Myanmar. The association between women's characteristics in reproductive age including socio-demography, sexual knowledge and behaviour, and STIs or symptoms of STIs is varied in countries of SEA. STIs interventions can be tailored to the specific characteristics of women in each of the 5 countries of SEA., Competing Interests: Declarations. Ethical approval: Procedures and questionnaires for standard DHS surveys have been reviewed and approved by ICF Institutional Review Board (IRB). Additionally, country-specific DHS survey protocols are reviewed by the ICF IRB and typically by an IRB in the host country. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Sustaining and expanding genomic surveillance capacity requires broader investment in genomics that target both novel and pandemic pathogens. Currently, there is no standardized methodology to evaluate the cost and benefit of a multi-pathogen surveillance system. We propose a framework for pathogen genomic surveillance that links public health and systems considerations to a stepwise approach., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2025 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: This study updates the COVID-19 pandemic surveillance in East Asia and the Pacific region that we first conducted in 2020 with 2 additional years of data for the region., Objective: First, we aimed to measure whether there was an expansion or contraction of the pandemic in East Asia and the Pacific region when the World Health Organization (WHO) declared the end of the COVID-19 public health emergency of international concern on May 5, 2023. Second, we used dynamic and genomic surveillance methods to describe the dynamic history of the pandemic in the region and situate the window of the WHO declaration within the broader history. Finally, we aimed to provide historical context for the course of the pandemic in East Asia and the Pacific region., Methods: In addition to updates of traditional surveillance data and dynamic panel estimates from the original study, this study used data on sequenced SARS-CoV-2 variants from the Global Initiative on Sharing All Influenza Data to identify the appearance and duration of variants of concern. We used Nextclade nomenclature to collect clade designations from sequences and Pangolin nomenclature for lineage designations of SARS-CoV-2. Finally, we conducted a 1-sided t test to determine whether the regional weekly speed was greater than an outbreak threshold of 10. We ran the test iteratively with 6 months of data across the sample period., Results: Several countries in East Asia and the Pacific region had COVID-19 transmission rates above an outbreak threshold at the point of the WHO declaration (Brunei, New Zealand, Australia, and South Korea). However, the regional transmission rate had remained below the outbreak threshold for 4 months. In the rolling 6-month window t test for regional outbreak status, the final P value ≤.10 implies a rejection of the null hypothesis (at the α=.10 level) that the region as a whole was not in an outbreak for the period from November 5, 2022, to May 5, 2023. From January 2022 onward, nearly every sequenced SARS-CoV-2 specimen in the region was identified as the Omicron variant., Conclusions: While COVID-19 continued to circulate in East Asia and the Pacific region, transmission rates had fallen below outbreak status by the time of the WHO declaration. Compared to other global regions, East Asia and the Pacific region had the latest outbreaks driven by the Omicron variant. COVID-19 appears to be endemic in the region, no longer reaching the threshold for a pandemic definition. However, the late outbreaks raise uncertainty about whether the pandemic was truly over in the region at the time of the WHO declaration., (©Alexander L Lundberg, Alan G Soetikno, Scott A Wu, Egon Ozer, Sarah B Welch, Yingxuan Liu, Claudia Hawkins, Maryann Mason, Robert Murphy, Robert J Havey, Charles B Moss, Chad J Achenbach, Lori Ann Post. Originally published in JMIR Public Health and Surveillance (https://publichealth.jmir.org), 21.02.2025.)

The ongoing circulation of highly pathogenic avian influenza (HPAI) A (H5N1) viruses, particularly clade 2.3.4.4b strains, poses a significant threat to animal and public health. Recent outbreaks in cattle highlight concerns about cross-species transmission and zoonotic spillover. Here, we found that the hemagglutinin (HA) protein from a cattle-infecting H5N1 virus has acquired slight binding to human-like α2-6-linked receptors while still exhibiting a strong preference for avian-like α2-3-linked sialic acid receptors. Immunohistochemical staining revealed HA binding to bovine pulmonary and mammary tissues, aligning with clinical observations. HA also binds effectively to human conjunctival, tracheal, and mammary tissues, indicating a risk for human transmission, notably in cases of conjunctivitis. High-resolution cryo-electron microscopy (cryo-EM) structures of this H5 HA in complex with either α2-3 or α2-6 receptors elucidate the molecular mechanisms underlying its receptor-binding properties. These findings provide critical insights into the tropism and transmission potential of this emerging pathogen., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2025 Elsevier Inc. All rights reserved.)

To detect multiple protein markers, here we present a protocol for imaging pulmonary tuberculosis (PTB) lesions from murine lungs using Opal-tyramide signal amplification (TSA) conjugated fluorescent dyes in free-floating formalin-fixed thick lung sections (50-100 μm). We describe steps for preparing tissue sections, permeabilization and antigen retrieval, and multiplexing with antibodies raised in the same species. This protocol has been optimized to preserve tissue integrity and endogenously expressed fluorescent reporter signals and nuclear staining. It enhances the signal-to-background ratio and enables 3D image rendering., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2025 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: The emergence of new SARS-CoV-2 variants, the resulting reinfections, and post-COVID-19 condition continue to impact many people's lives. Tracking websites like the one at Johns Hopkins University no longer report the daily confirmed cases, posing challenges to accurately determine the true extent of infections. Many COVID-19 cases with mild symptoms are self-assessed at home and reported on social media, which provides an opportunity to monitor and understand the progression and evolving trends of the disease., Objective: We aim to build a publicly available database of COVID-19-related tweets and extracted information about symptoms and recovery cycles from self-reported tweets. We have presented the results of our analysis of infection, reinfection, recovery, and long-term effects of COVID-19 on a visualization website that refreshes data on a weekly basis., Methods: We used Twitter (subsequently rebranded as X) to collect COVID-19-related data, from which 9 native English-speaking annotators annotated a training dataset of COVID-19-positive self-reporters. We then used large language models to identify positive self-reporters from other unannotated tweets. We used the Hibert transform to calculate the lead of the prediction curve ahead of the reported curve. Finally, we presented our findings on symptoms, recovery, reinfections, and long-term effects of COVID-19 on the Covlab website., Results: We collected 7.3 million tweets related to COVID-19 between January 1, 2020, and April 1, 2024, including 262,278 self-reported cases. The predicted number of infection cases by our model is 7.63 days ahead of the official report. In addition to common symptoms, we identified some symptoms that were not included in the list from the US Centers for Disease Control and Prevention, such as lethargy and hallucinations. Repeat infections were commonly occurring, with rates of second and third infections at 7.49% (19,644/262,278) and 1.37% (3593/262,278), respectively, whereas 0.45% (1180/262,278) also reported that they had been infected >5 times. We identified 723 individuals who shared detailed recovery experiences through tweets, indicating a substantially reduction in recovery time over the years. Specifically, the average recovery period decreased from around 30 days in 2020 to approximately 12 days in 2023. In addition, geographic information collected from confirmed individuals indicates that the temporal patterns of confirmed cases in states such as California and Texas closely mirror the overall trajectory observed across the United States., Conclusions: Although with some biases and limitations, self-reported tweet data serves as a valuable complement to clinical data, especially in the postpandemic era dominated by mild cases. Our web-based analytic platform can play a significant role in continuously tracking COVID-19, finding new uncommon symptoms, detecting and monitoring the manifestation of long-term effects, and providing necessary insights to the public and decision-makers., (©Jiacheng Xie, Ziyang Zhang, Shuai Zeng, Joel Hilliard, Guanghui An, Xiaoting Tang, Lei Jiang, Yang Yu, Xiufeng Wan, Dong Xu. Originally published in the Journal of Medical Internet Research (https://www.jmir.org), 20.02.2025.)