Search Results (400)

The COVID-19 pandemic highlighted the urgent need to develop effective and broad-spectrum antiviral therapies against coronaviruses. One strategy to address this concern is a combination therapy using repurposed drugs against zoonotic viruses with pandemic potential. We previously demonstrated that the combination of Remdesivir and Ivermectin is highly potent and synergistic in inhibiting the replication of murine hepatitis virus (MHV) in RAW264.7 macrophages. This study investigated the interactions between the drug combination, coronavirus and host by proteomics and RNA sequencing of MHV-infected H2.35 murine liver epithelial cells. Time-of-addition and time-of-removal assays suggested that the drug combination likely affected the synthesis of viral RNA and viral protein. This combination drastically diminished the live virus titer greater than the respective monotherapies in MHV-infected H2.35 cells (by ~4 log₁₀), as well as in SARS-CoV-2-infected VeroE6 cells and human nasal epithelial cells. Proteomic and transcriptomic analyses revealed that viral protein and RNA levels were significantly depressed upon combination treatment. The drug combination exhibited considerable negative effects upon host RNA processes and resulted in the upregulation of host protein processes (e.g., response to unfolded protein; protein insertion into ER membrane). Molecular pathways affected by the combination treatment were markedly distinct from the monotherapies and indicated that Ivermectin enhances Remdesivir by modulating critical host processes to synergistically exert its inhibitory effect on the coronavirus replication cycle. Full article

The Chinese bahaba (Bahaba taipingensis), an endangered marine fish, is highly vulnerable to infectious spleen and kidney necrosis virus (ISKNV). In this work, we developed a gill filament-derived cell line, designated BTG, to investigate how these cells respond to ISKNV over time, specifically from 3 to 24 h post-infection (hpi). BTG cells grew steadily, displayed a diploid chromosome number of 2n = 48, demonstrated high transfection efficiency, and were highly susceptible to viral infection. Characteristic cytopathic effects (CPEs) became noticeable as early as 6 hpi at 27 °C. RNA-seq profiling showed that the number of differentially expressed genes (DEGs) steadily increased with time. Standard enrichment analysis at individual time points (3, 6, 12, and 24 hpi) highlighted pathways mainly involved in DNA replication, cell cycle control, ribosome assembly, transcription and translation, mismatch repair, and cell adhesion. Temporal clustering analysis, however, revealed hidden patterns in immune gene expression. Genes that were consistently downregulated were enriched in immune-related pathways, including ECM–receptor interaction, cytokine–receptor signaling, PI3K–AKT, and Wnt signaling, indicating prolonged suppression of host defense mechanisms. In contrast, clusters of genes transiently upregulated during the first 6 h post-infection were associated with antiviral and innate immune pathways, such as NF-κB, JNK, IRF3, IRF7, caspases, JAK, MHC I, and lysosome-related functions, suggesting a rapid but short-lived antiviral response. Genes that were continuously upregulated were primarily involved in nucleic acid replication and protein synthesis, reflecting a gradual host cell reprogramming to support viral replication. Taken together, these findings reveal a temporal shift in BTG cells from an initial burst of immune activity to immune suppression, accompanied by enhanced viral replication. The BTG cell line thus represents a valuable in vitro model for dissecting ISKNV–host interactions and offers new perspectives on the molecular strategies employed by megalocytiviruses in B. taipingensis. Full article

Background/Objectives: As important post-transcriptional and epigenetic regulators of gene expression, miRNAs play a pivotal role in modulating host–virus interactions. While prior reviews have addressed either direct miRNA–HIV genome interactions or miRNA-mediated immune modulation in isolation, the integrated dual functionality of these molecules has not been systematically characterized. This review aimed to comprehensively explore how miRNAs that target the HIV-1 genome simultaneously modulate key innate and adaptive host immune signaling pathways. The conceptual novelty of this study is determined not by the identification of previously unknown miRNA-target gene pairs, but by the systemic integration of two regulatory levels (direct inhibition of the viral genome and modulation of the host cell immune signaling pathways) within a unified analytical framework. Such an integrated approach reveals a proviral regulatory network that remains non-obvious when each of these levels is examined separately. Methods: A narrative review was conducted using PubMed, Scopus, Web of Science, and Google Scholar (all years through 2025). In Stage 1, publications reporting experimentally confirmed interactions between host miRNAs and the HIV-1 genome were identified, yielding a curated set of 15 miRNAs. In Stage 2, target genes for each miRNA were retrieved from miRTarBase, TarBase (experimentally validated) and TargetScan 8.0 (in silico predicted). In Stage 3, target genes were manually mapped to key immune signaling pathways (TLR, NF-κB, JAK-STAT). In Stage 4, targeted literature searches were performed for each miRNA–target gene pair to identify direct experimental evidence of interaction. All stages were performed by two independent researchers, with discrepancies resolved by a third. Results: Fifteen host miRNAs with experimentally confirmed binding to the HIV-1 genome were identified, targeting viral genes including nef, pol, vpr, gag, env, vif, and the 3′-UTR. Thirteen of these miRNAs were found to regulate components of major immune pathways. miR-92a-3p, miR-29a/b-3p, miR-150-5p, and miR-125b-5p emerged as the most pleiotropic regulators, simultaneously suppressing TLR signaling (TLR3, TLR7, TLR8, MyD88, TRAF3/6, IRAK1/4), NF-κB components (REL, RELA, NFKB1), JAK-STAT effectors (STAT1–3, STAT5A/B, JAK2), and negative regulators of cytokine signaling (SOCS and PIAS family proteins). miR-133b and miR-196b-5p were found to selectively regulate SOCS/PIAS proteins without involvement in other analyzed pathways, suggesting potential for selective therapeutic targeting. Conclusions: The analyzed miRNAs exhibit functional dualism, acting as direct post-transcriptional suppressors of the HIV-1 genome while simultaneously functioning as epigenetic modulators of host immune signaling. These two modes of action are not independent but together form a conceptual framework of a self-reinforcing proviral regulatory network that, based on the synthesis of published evidence, is proposed to promote viral latency and immune evasion. The identified miRNAs represent promising, albeit complex, targets for novel therapeutic strategies aimed at eliminating latent HIV reservoirs. Full article

Post-acute sequelae of SARS-CoV-2 infection (PASC) extend well beyond the acute respiratory phase, with accumulating virological evidence that SARS-CoV-2 RNA, viral antigens, and proteolytic fragments may persist in cardiovascular and other extrapulmonary tissues, although the extent to which such detection represents replication-competent reservoirs versus residual viral material with uncertain pathological relevance remains under active investigation. Sudden cardiac death (SCD) and fatal pulmonary thromboembolism (PTE) have emerged as forensically and epidemiologically significant outcomes in individuals with prior infection, situated at the intersection of microbiology, public health, and forensic medicine. To synthesize current evidence on the virological mechanisms by which SARS-CoV-2 may contribute to post-acute sudden cardiac death (SCD) and pulmonary thromboembolism (PTE), the population-level epidemiology of these outcomes, and their implications for public health surveillance and forensic practice, we conducted a narrative review of PubMed (MEDLINE), Scopus, and Web of Science Core Collection. The search covered publications from January 2020 to December 2025 and focused on SARS-CoV-2 cellular tropism and tissue persistence, immune-mediated and thromboinflammatory mechanisms, excess cardiovascular and thromboembolic mortality, and autopsy-based pathological findings. After de-duplication of 1837 initially identified records (412 duplicates removed) and screening of 1425 unique records, 78 studies were retained for final synthesis based on virological, epidemiological, and forensic relevance. SARS-CoV-2 enters cardiomyocytes, pericytes, and vascular endothelial cells through ACE2-dependent mechanisms, with cathepsin L compensating for the limited cardiac expression of TMPRSS2. Viral RNA and antigen have been detected in cardiovascular and other extrapulmonary tissues months after symptom onset in selected autopsy series, although persistent detection of viral components does not necessarily indicate ongoing productive infection or direct tissue injury. Endothelial dysfunction, neutrophil extracellular trap (NET) formation, complement activation, and persistent thromboinflammation have been proposed as plausible mechanistic substrates for arrhythmogenic remodelling and thromboembolic events, although definitive causal pathways remain incompletely understood. Population-based studies document persistent excess cardiovascular mortality across multiple jurisdictions, with hazard ratios for pulmonary embolism remaining elevated months after acute infection, particularly in unvaccinated individuals. Autopsy series identify mixed pathological patterns including focal lymphocytic infiltrates, microvascular thrombosis, contraction-band necrosis, and cardiomyocyte vacuolation, although fulminant lymphocytic myocarditis fulfilling Dallas criteria remains uncommon. A microbiology-informed framework uniting tissue-based viral detection, standardized cardiac and pulmonary sampling protocols, and prospective post-mortem registries is needed to better characterize the potential contribution of SARS-CoV-2 to post-acute cardiovascular mortality and to support cause-of-death certification, public health surveillance, and medicolegal practice in the post-pandemic era. Many of the proposed mechanisms remain under active investigation, and definitive causal relationships between viral persistence and adverse cardiovascular outcomes have not yet been conclusively established. Full article

Coronaviruses use discontinuous transcription to generate subgenomic RNAs (sgRNAs) that encode structural and accessory proteins. However, the factors regulating sgRNA abundance in SARS-CoV-2 remain unclear. Here, we combined strand-specific RNA sequencing, RNA–RNA interaction mapping, prediction of RNA folding energies, and targeted mutagenesis to define the regulation of (–) sgRNA synthesis in SARS-CoV-2 infection. We demonstrated that the relative (–) sgRNA abundance across viral genes is stable throughout infection and largely correlates with corresponding (+) sgmRNA levels. Through meta-analysis of published SPLASH data, we found that the frequency of long-range interactions between the 5′ genomic transcription regulatory sequence TRS-Leader and downstream TRS-Body sequences correlates with sgRNA abundance. Notably, the folding energy (ΔG) of these duplexes quantitatively predicts (–) sgRNA transcript levels. Mutations in non-coding regulatory regions that altered the ΔG resulted in corresponding changes in (–) sgRNA expression, suggesting a causal role for TRS duplex stability in transcriptional regulation. Analysis of naturally occurring mutations near regulatory sites further suggests that modulation of duplex stability may also serve as an evolutionary mechanism to fine-tune viral gene expression. Together, our findings identify the pairing stability of TRS-Leader:TRS-Body as a determinant of discontinuous transcription and reveal how RNA pairing potential contributes to the regulation of (–) sgRNA synthesis in SARS-CoV-2. Full article

The liver circadian clock coordinates hepatic lipid metabolism, bile acid synthesis, and glucose homeostasis through interlocking transcription–translation feedback loops. Disruption of this temporal organization is increasingly recognized as a shared pathological feature across the chronic liver disease spectrum. Transcriptomic profiling alone cannot capture the full scope of circadian dysregulation. Approximately half of rhythmically abundant hepatic proteins lack correspondingly rhythmic mRNAs. Roughly 25% of hepatic phosphosites oscillate with a 24-h period. Integrating transcriptomics, proteomics, post-translational modification profiling, metabolomics, and emerging single-cell and spatial approaches is therefore necessary for an accurate account of how circadian programs are remodeled in disease. This narrative review delineates the multi-omics landscape of circadian clock dysregulation across six chronic liver disease categories. These encompass metabolic dysfunction-associated fatty liver disease (MAFLD), alcoholic liver disease (ALD), viral hepatitis, hepatocellular carcinoma (HCC), liver fibrosis, and cholestatic disease. Four molecular features recur across these contexts. BMAL1 functional downregulation, REV-ERBα oscillatory output attenuation, NAD⁺ oscillatory amplitude reduction, and gut–liver axis circadian desynchronization together constitute an inferential framework for hepatic circadian failure. These features represent recurring disease-associated motifs rather than an established pan-disease mechanism. The upstream mechanisms and evidence depth differ substantially by disease category. Oncogenic kinase-driven CLOCK post-translational modifications in HCC, phosphoproteomic remodeling in MAFLD, and epigenomic clock disruption persisting after HCV clearance represent findings that transcriptomics alone would not resolve. The near-complete absence of temporally resolved human tissue data remains the principal barrier to translational progress. This evidence gap limits the clinical actionability of current mechanistic findings across all disease categories. Circadian phase inference algorithms and prospective temporally designed cohort studies offer a methodologically grounded path toward clinically actionable circadian hepatology. Full article

Endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) represent fundamental cellular adaptive mechanisms that maintain protein homeostasis and metabolic balance. Many RNA viruses, particularly flaviviruses such as dengue virus (DENV), Zika virus (ZIKV), West Nile virus (WNV), yellow fever virus (YFV), and Japanese encephalitis virus (JEV), extensively remodel the ER to establish replication compartments and assemble progeny virions. This massive reorganization disrupts ER homeostasis, leading to UPR activation. Emerging evidence reveals that flaviviruses not only trigger but also manipulate the three UPR branches—PERK, IRE1, and ATF6—to optimize viral translation, replication, and egress. In parallel, flavivirus infection profoundly alters host lipid metabolism and promotes dynamic changes in lipid droplets (LDs), key organelles that mediate lipid storage and serve as scaffolds for viral replication and assembly. The UPR intimately connects to LD biogenesis through transcriptional and translational programs mediated by XBP1, ATF4, and ATF6, thereby coupling ER stress responses to lipid remodeling and energy homeostasis. This intricate crosstalk between UPR and LDs creates a metabolic and structural niche favorable for viral replication but detrimental to host cell integrity. This review provides a comprehensive analysis of the molecular mechanisms by which flaviviruses exploit ER stress and the UPR to reprogram lipid metabolism and LD dynamics. We highlight the dual role of UPR signaling in promoting adaptive lipid synthesis and initiating cell death under prolonged stress, discuss recent insights into ER–LD interactions during flavivirus infection, and explore therapeutic opportunities targeting UPR–lipid metabolic pathways as broad-spectrum antiviral strategies. Understanding this interconnected network will advance our knowledge of viral pathogenesis and identify new avenues for host-directed antiviral intervention. Full article

Nipah virus (NiV) is a highly pathogenic zoonotic paramyxovirus responsible for sporadic outbreaks of severe disease with high case fatality rates in South and Southeast Asia. Human infection occurs through spillover from natural reservoirs, primarily fruit bats, or via human-to-human transmission, and is characterized by a broad clinical spectrum ranging from asymptomatic infection to acute respiratory disease and fatal encephalitis. Following entry via ephrin-B2 and ephrin-B3 receptors, NiV exhibits marked endothelial and neuronal tropism, leading to systemic vasculitis, disruption of the blood–brain barrier, and direct infection of the central nervous system. Disease progression is driven by a complex interplay between viral replication strategies and host immune responses. NiV effectively counteracts innate immunity through multiple viral proteins that inhibit interferon signaling, while simultaneously inducing dysregulated inflammatory responses that contribute to tissue damage and multi-organ failure. Neurological involvement represents the most severe manifestation, often resulting in acute or relapsing encephalitis with long-term sequelae among survivors. Despite the severity of the disease, no licensed antiviral therapies or human vaccines are currently available. Therapeutic development has focused on neutralizing monoclonal antibodies targeting viral glycoproteins and small-molecule antivirals that inhibit viral RNA synthesis, both of which show promising results in preclinical models, but remain limited by timing and translational challenges. In parallel, several vaccine platforms—including viral vectors, mRNA-based constructs, and recombinant protein subunits—have advanced to early-phase clinical trials, demonstrating encouraging immunogenicity. Beyond biomedical interventions, effective outbreak containment relies on integrated public health strategies. The “Kerala model” highlights the importance of rapid case identification, isolation, contact tracing, and community engagement within a One Health framework to mitigate transmission and reduce mortality. This review synthesizes the current knowledge on NiV pathogenesis, immune evasion, clinical manifestations, and emerging therapeutic and vaccine strategies, while highlighting critical gaps and future directions for improving the preparedness and response to this high-consequence emerging pathogen. Full article

Nucleocapsids protect viral genomes and play fundamental roles in viral assembly and infection. While many viruses adopt icosahedral or helical symmetries, negative-strand RNA viruses (NSVs) assemble their nucleocapsids with a distinct translation-based symmetry that is often considered helical because of their curvature. Our study analyzes the structural basis, assembly principles, and functional implications of the linear nucleocapsids. Structural coordinates of viruses were obtained from the Protein Data Bank (PDB) and examined using PyMOL version 1.3 to compare protein folds, RNA–protein interactions, inter-subunit contacts, and curvature properties across multiple nucleocapsids. We found that linear nucleocapsids share a similar 5H + 3H fold in their capsid proteins and encapsidate a fixed number of nucleotides per subunit, though the degree of nucleotide sequestration varies. Their architecture differs in inter-subunit interactions, determining whether empty capsids can assemble and influencing RNase sensitivity. Although these nucleocapsids may appear helical, they lack strict helical symmetry and instead display variable curvature that is modulated by environmental conditions. Relaxation of this curvature is likely required for viral RNA-dependent RNA polymerase to access the sequestered RNA genome during transcription/replication. In conclusion, linear nucleocapsids constitute a class of RNA–protein assemblies with variable curvature. The topologically conserved fold of the capsid protein enables genome protection while regulating exposure of RNA during viral RNA synthesis. Full article

Host receptors can detect traces of non-self-pathogenic RNAs within a sea of cellular mRNA molecules. In host cells, mRNA cap methylation occurs in the nucleus, generating Cap1 and Cap2 structures (m⁷GpppNm and m⁷GpppNmNm, respectively). By contrast, alphavirus genomes carry a Cap0 structure (m⁷GpppN), which lacks 2′-O-methylation. This difference in the structure of the host and viral caps serves as a molecular signature that enables discrimination between self and non-self RNAs. Several host immune sensors, such as RIG-I and IFIT1, recognize the alphavirus Cap0 structure and trigger an antiviral response to restrict viral replication. It has been proposed that IFIT1 sequesters aberrant RNAs, preventing their translation by host ribosomes and blocking viral protein synthesis. However, alphaviruses have evolved molecular strategies to circumvent IFIT1-mediated restriction and facilitate infection in mammalian cells. One such strategy involves the folding of a 5′ RNA structure that hides the cap from host immune sensors. This highlights the dynamic interplay between viral evasion tactics and host immune defenses. This review will discuss how specific modifications at the 5′ end of alphavirus RNA modulate host defenses and how a deeper understanding of the virus–host interaction may inform the development of novel vaccine strategies. Full article

Antimicrobial resistance (AMR) poses a significant global health threat, with multidrug-resistant pathogens undermining the effectiveness of conventional antibiotics. Natural alkaloids, a diverse group of nitrogen-containing compounds mainly derived from plants, are gaining attention as potential antimicrobial agents due to their broad-spectrum activity, structural variety, and unique mechanisms of action. This review examines the antimicrobial properties of natural alkaloids, classifying them by chemical structure (e.g., quinoline, isoquinoline, pyridine, indole, and imidazole alkaloids). Their antibacterial, antifungal, and antiviral activities are discussed, along with the mechanisms by which they target pathogenic microorganisms, including disruption of cell walls and membranes, inhibition of protein synthesis, interference with DNA replication, and viral assembly. The review also explores the synergistic effects of alkaloids when combined with conventional antimicrobial agents. Alkaloids demonstrate potent antimicrobial activity against various pathogens. Quinoline alkaloids, such as quinine, inhibit DNA replication and damage cell membranes. Isoquinoline alkaloids like berberine and sanguinarine exhibit broad-spectrum antibacterial effects. Pyridine alkaloids, including nicotine, disrupt bacterial membranes. In fungi, alkaloids such as sanguinarine and indole derivatives prevent cell wall synthesis and spore germination. Antiviral alkaloids like lycorine target viral RNA polymerases. Additionally, alkaloids enhance the activity of traditional antibiotics by overcoming resistance. Natural alkaloids represent a promising source of antimicrobial agents with diverse mechanisms to combat AMR. Future research should focus on optimizing alkaloid structures, ensuring safety and efficacy, and exploring combination therapies to address the escalating AMR challenge. Full article

Background: Infectious hematopoietic necrosis virus (IHNV), a rhabdovirus classified within the genus Novirhabdovirus, continues to be one of the most detrimental pathogens impacting salmonid aquaculture on a global scale. Notable for inducing high mortality rates among fry and fingerlings, IHNV represents a substantial threat to the economic stability of the aquaculture industry. This review offers an in-depth analysis of the contemporary advancements in IHNV vaccine development. Methods: We assess the efficacy and immunological mechanisms of traditional vaccine platforms, including inactivated and live-attenuated vaccines, while emphasizing the groundbreaking success of DNA vaccines, particularly those encoding the viral glycoprotein (G). Although nucleic acid-based therapies provide high levels of protection, they face logistical challenges related to delivery and regulatory obstacles associated with Genetically Modified Organisms (GMOs). Additionally, we examine emerging “next-generation” platforms, such as viral vector vaccines, subunit proteins produced in yeast or plant systems, and RNA-based technologies. We critically analyze technical bottlenecks, including the lack of efficient mucosal delivery systems and the limited understanding of long-term cellular memory in teleosts. Results: We propose future research directions that emphasize the development of multivalent formulations and the incorporation of molecular adjuvants to augment mucosal immunity. Conclusions: This synthesis seeks to integrate fundamental viral pathogenesis with applied immunology to develop a strategic framework for the sustainable, long-term management of IHNV in global salmonid populations. Full article

Bluetongue virus (BTV), with a genome of ten double-stranded RNA segments (S1–S10), is an emerging animal pathogen causing major economic losses in livestock worldwide. BTV replication involves RNA-RNA and RNA–protein interactions, with RNA-binding proteins, VP6 and NS2 playing key roles in genome assembly and RNA packaging. To explore the dynamics of RNA segment interactions and the roles of VP6 and NS2 in RNA complex formation, we used RNA fluorescence in situ hybridization chain reaction (HCR), along with site-specific mutagenesis and reverse genetics. We found that RNA segments interact sequentially, from the smallest (S10) to the largest (S1), forming a single complex that includes the entire genome. This process is independent of VP6 or NS2, although NS2 enhances the assembly of larger segments. Additionally, we show that VP6 binds to +ssRNAs before their incorporation into viral assembly factories (inclusion bodies/VIBs). These findings reveal that RNA-RNA interactions, rather than primary replicase proteins, govern the sorting and recruitment of genome segments. Our data offer new insights into BTV RNA packaging, showing that genome segments destined for packaging and dsRNA synthesis are segregated through complex formation, distinct from +ssRNAs used in protein synthesis, including those encoding the replicase complex. Full article

Aging is associated with a high prevalence of insomnia, which is linked to somatic and neuropsychiatric diseases, as well as metabolic and immunological dysfunction. This study aims to identify alterations in the transcriptome profiles and functional metabolic pathways in older adults with different types of sleep disorders. This cross-sectional study included 1002 participants (60–90 years) who were screened for sleep disorders using the Pittsburgh Sleep Quality Index (PSQI) questionnaire. Two types of sleep disorders were identified in the study cohort, i.e., sleep onset insomnia and sleep maintenance insomnia. Both types of insomnia were further analyzed for associations with clinical characteristics, laboratory testing results, and socioeconomic backgrounds. The transcriptomic profiles of peripheral blood samples were examined in 236 individuals, supplemented with differential gene and dsRNA expression analyses (DESeq2). Both sleep onset insomnia and middle insomnia were associated with depression, chronic pain syndrome, and osteoarthritis, while only middle insomnia was associated with cardiometabolic diseases. No associations were observed between sleep onset insomnia or reduced sleep duration and transcriptomic profiles. In contrast, 244 genes were differentially expressed in patients with middle insomnia, indicating the activation of pathways related to viral infection response and inhibition of protein synthesis. Additionally, differential expression analysis of double-stranded RNA (dsRNA) identified 2139 significant changes. Middle insomnia in older adults is associated with transcriptomic changes indicative of an activated antiviral immune response, likely resulting from changes in dsRNA expression levels. The chronic inflammation arising from these transcriptomic alterations may underlie the observed association between middle insomnia and cardiometabolic conditions. Full article

Background: Persistent infection with high-risk human papillomavirus (hr-HPV) is the necessary agent of cervical cancer, yet its molecular definition remains heterogeneous. Multiple molecular approaches have been developed to characterize HPV persistence, including repeated detection of viral DNA, assessment of viral oncogene expression, and analysis of HPV-related DNA methylation. These approaches originate from different scientific traditions and reflect distinct conceptualizations of persistence. Objective: To synthesize and compare molecular methods used to detect persistent HPV infection through a narrative review and to clarify how different biomarkers conceptualize HPV persistence and disease progression. Methods: We conducted a narrative review in accordance with the RAMESES guidelines. Medline, Scopus, and the Cochrane Library were searched for original studies published between 2016 and 2025 investigating molecular markers of HPV persistence. An interpretive synthesis was performed to identify research traditions, underlying assumptions, and clinical implications. Results: Three major molecular narratives were identified. Persistent DNA positivity defines persistence as repeated detection of the same HR-HPV genotype over time and reflects an epidemiological–virological perspective with high sensitivity but limited specificity. Persistent oncogene expression, assessed by E6/E7 mRNA detection, conceptualizes persistence as active viral oncogenic activity and shows improved specificity for clinically relevant lesions. Persistent epigenetic imprint, measured by DNA methylation of viral and host genes, captures cumulative biological effects of long-term infection and is strongly associated with high-grade lesions and cervical cancer. These narratives represent complementary stages along a continuum of molecular persistence. Conclusions: Molecular markers of HPV persistence reflect the evolving understanding of cervical carcinogenesis, progressing from repeated viral DNA detection to oncogenic activity and stable epigenetic alterations. These complementary biomarkers represent different biological stages of persistent infection and may improve risk stratification in HPV-based screening and triage strategies. Full article