Viruses

15 pages, 1139 KB

Open AccessArticle

Comparative Evaluation of SARS-CoV-2 RNA Concentration and Normalization Strategies in Prison Wastewater: Implications for Viral Dynamics in Confined Environments

by Raheel Nazakat, Nabilla Athieqa Mahdzar, Amirul Haziq Azwan, Reethiya Letchumanan and Siti Aishah Rashid

Viruses 2026, 18(5), 563; https://doi.org/10.3390/v18050563 (registering DOI) - 15 May 2026

Abstract

Background: Wastewater-based epidemiology (WBE) is a valuable population-level surveillance tool for monitoring SARS-CoV-2 circulation. However, evidence on optimal viral concentration approaches in confined institutional settings such as prisons remains limited. This study aimed to compare the performance of Direct Capture (DC) and Electronegative [...] Read more.

Background: Wastewater-based epidemiology (WBE) is a valuable population-level surveillance tool for monitoring SARS-CoV-2 circulation. However, evidence on optimal viral concentration approaches in confined institutional settings such as prisons remains limited. This study aimed to compare the performance of Direct Capture (DC) and Electronegative Membrane Filtration (EMF) for SARS-CoV-2 RNA detection in wastewater from a prison facility in Selangor, Malaysia. Methods: Composite wastewater samples collected over 18 weeks (April–August 2023; n = 50) were analysed by RT-dPCR targeting the N1 and N2 gene regions, with concentrations normalized to pepper mild mottle virus (PMMoV). DC consistently outperformed EMF across both gene targets. Median concentrations obtained using DC were 11.09 × 10³ copies L⁻¹ (N1) and 3.43 × 10³ copies L⁻¹ (N2), compared with 0.70 × 10³ copies L⁻¹ (N1) and 0.48 × 10³ copies L⁻¹ (N2) using EMF. Detection frequencies were higher with DC (N1: 94%, N2: 84%) than with EMF (N1: 88%, N2: 76%). Paired statistical analysis confirmed significant differences between methods (N1: p = 2.3 × 10⁻⁷; N2: p = 9.4 × 10⁻⁵), and Bland–Altman analysis demonstrated systematic underestimation by EMF (mean bias −1.15 log₁₀ for N1; −0.87 log₁₀ for N2), indicating that the methods are not analytically interchangeable. Conclusions: Normalization reduced absolute SARS-CoV-2 RNA concentrations while preserving temporal trends, supporting its use to improve comparability across sampling periods. Overall, these findings demonstrate that DC combined with N1 detection provides a more sensitive and reliable approach for SARS-CoV-2 WBE in confined settings, underscoring the importance of methodological optimization to strengthen early-warning capacity in high-risk environments. Full article

(This article belongs to the Special Issue Wastewater-Based Epidemiology and Viral Surveillance)

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15 pages, 5132 KB

Open AccessArticle

Genetic Diversity and Evolutionary Dynamics of Feline Panleukopenia Virus in China: Phylogenetic Analysis and Substitution Patterns in NS1 and VP2 Proteins

by Zihan Ye, Danni Wu, Xueru Jiang, Lina Liu, Guoliang Luo, Zhenjun Wang, Yuening Cheng and Erkai Feng

Viruses 2026, 18(5), 562; https://doi.org/10.3390/v18050562 (registering DOI) - 15 May 2026

Abstract

Feline panleukopenia virus (FPLV) is the primary causative agent of a highly contagious and often fatal disease affecting domestic cats and other felids. The increasing isolation of species-specific FPLV variants from multiple host species has garnered considerable attention, highlighting the need to investigate [...] Read more.

Feline panleukopenia virus (FPLV) is the primary causative agent of a highly contagious and often fatal disease affecting domestic cats and other felids. The increasing isolation of species-specific FPLV variants from multiple host species has garnered considerable attention, highlighting the need to investigate their genetic diversity. In this study, three FPLV isolates were obtained and phylogenetically classified into two distinct FPLV-China groups within separate clusters. Compared to the prototype FPLV (M38246.1), these isolates exhibited seven amino acid substitutions in the NS1 (n = 6) and VP2 (n = 1) proteins. Further analysis of 157 NS1 sequences and 947 VP2 sequences retrieved from the NCBI database revealed 113 and 479 synonymous substitutions and 71 and 279 non-synonymous substitutions, respectively. Notably, the majority of these substitutions occurred as single events (57% in NS1, 40/71; 55% in VP2, 153/279) or were present in no more than five FPLV sequences (23% in NS1, 16/71; 32% in VP2, 89/279). However, three non-synonymous substitutions in the NS1 protein (Ile443Val, His595Gln, and Val596Leu) were detected in more than half of the 157 sequences analyzed. In the VP2 protein, six non-synonymous substitutions (Ala91Ser, Thr101Ile, Val232Ile, Lys93Asn, Asp323Asn, and Val562Leu) were each found in 20 to 40 FPLV sequences. Furthermore, ten sites in the NS protein and 224 sites in the VP2 protein exhibited both synonymous and non-synonymous substitutions simultaneously. Additionally, 75 sites in VP2 harbored multiple non-synonymous substitutions. These findings provide valuable insights for future research on the genetic determinants and vaccine development of FPLV. Full article

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17 pages, 2660 KB

Open AccessArticle

In Vitro Pharmacokinetic Properties of MK-2048, a Potent Drug Candidate for HIV Prevention

by Ruohui Zheng, Guru Raghavendra Valicherla, Phillip W. Graebing, Junmei Zhang, Sharon L. Hillier and Lisa Cencia Rohan

Viruses 2026, 18(5), 561; https://doi.org/10.3390/v18050561 (registering DOI) - 15 May 2026

Abstract

MK-2048 is a potent second-generation HIV integrase inhibitor that has demonstrated acceptable safety and pharmacokinetics (PKs) in clinical trials of vaginal formulations. The substrate-type interactions between MK-2048 and the transporters/metabolizing enzymes that are highly expressed in the human female reproductive tract (FRT) were [...] Read more.

MK-2048 is a potent second-generation HIV integrase inhibitor that has demonstrated acceptable safety and pharmacokinetics (PKs) in clinical trials of vaginal formulations. The substrate-type interactions between MK-2048 and the transporters/metabolizing enzymes that are highly expressed in the human female reproductive tract (FRT) were evaluated. The interactions between MK-2048 and P-gp/BCRP were investigated using a cellular bidirectional permeability assay, while those between MK-2048 and MRP4 were assessed using a vesicular uptake assay. Reaction phenotyping was performed to characterize the interactions between MK-2048 and CYP1A1 and CYP1B1. Using human cervicovaginal fluids (CVFs), MK-2048’s solubility was determined using a thermodynamic solubility method and its protein binding was determined using a rapid equilibrium dialysis method. Our study shows an efflux of MK-2048 in P-gp/BCRP-overexpressing MDCKII cells, which was reduced by a P-gp/BCRP inhibitor. Uptake of MK-2048 in MRP4/control vesicles was found to be ATP-independent. MK-2048 was metabolized by the CYP1A1 enzyme but not by CYP1B1. These data confirm that MK-2048 is a substrate of P-gp, BCRP, and CYP1A1, but is not a substrate of MRP4 or CYP1B1. MK-2048 displays low solubility and high protein binding in human CVF. This data suggests that MK-2048 may potentially interact with drugs that modulate the activity of P-gp, BCRP, and CYP1A1. Full article

(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)

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18 pages, 1024 KB

Open AccessReview

Biological Trajectory of Virophage Research and the Emergence of Marine Virophages: A Scoping Review

by Min-Jeong Kim, Yu-Jin Kim, Hyun-Ju Ha, Joon-Sang Park, Ika Agus Rini, Suk-Chan Lee and Taek-Kyun Lee

Viruses 2026, 18(5), 560; https://doi.org/10.3390/v18050560 (registering DOI) - 14 May 2026

Abstract

Virophages are satellite viruses that depend on the replication machinery of giant double-stranded DNA viruses and influence the structure and dynamics of viral communities through multilayered interactions among giant viruses, their hosts, and virophages. Since the discovery of the Sputnik virophage in 2008, [...] Read more.

Virophages are satellite viruses that depend on the replication machinery of giant double-stranded DNA viruses and influence the structure and dynamics of viral communities through multilayered interactions among giant viruses, their hosts, and virophages. Since the discovery of the Sputnik virophage in 2008, virophages have been increasingly recognized for their roles in regulating giant virus replication, contributing to host defense mechanisms, and shaping the evolution of mobile genetic elements. However, quantitative syntheses examining how virophage research has developed over time, particularly in marine environments, remain limited. Here, we conducted a bibliometric analysis of virophage research published between 2008 and 2025 using the Web of Science Core Collection. By comparing an overall virophage research corpus with a marine virophage sub-corpus, we assessed publication and citation trends, collaboration structures, and keyword-based intellectual and thematic evolution. Our results show that virophage research has gradually transitioned from an early phase dominated by landmark discoveries and experimental model systems to a data-intensive stage driven by genome- and metagenome-based analyses and computational approaches. Although marine virophage studies represent a relatively small proportion of the total literature, they exhibit sustained citation impact and form a distinct research axis within the field. In particular, marine-focused studies emphasize metagenomic discovery, genome sequence alignment, and the analysis of mobile genetic elements such as polinton-like viruses, highlighting the role of marine environments in accelerating the intellectual transition of virophage research. Collectively, these findings demonstrate that virophage research has moved beyond a “discovery and definition” phase toward data-driven integrative interpretation, with marine virophage research emerging as a key domain for understanding the structure and evolutionary dynamics of marine viral ecosystems. Full article

(This article belongs to the Section Bacterial Viruses)

15 pages, 3194 KB

Open AccessArticle

Sphingosine-1-Phosphate Receptor and Kinase Expression in the Reproductive Tract Is Associated with HIV Infection and Preterm Birth in a Cohort of Pregnant Women in Zambia

by Rachel S. Resop, Innocent Mwape, Yuri V. Sebastião, Katelyn J. Rittenhouse, Ntazana Sindano, Humphrey Mwape, Margaret P. Kasaro, Bellington Vwalika, Joan T. Price, Jeffrey S. A. Stringer and Kristina De Paris

Viruses 2026, 18(5), 559; https://doi.org/10.3390/v18050559 (registering DOI) - 14 May 2026

Abstract

Women living with HIV face an increased burden of spontaneous preterm birth (sPTB); however, the underlying immunological mechanisms of sPTB and its association with HIV infection are poorly understood. Although the limited earlier literature implicates sphingosine-1-phosphate (S1P), a lysosphingolipid signaling molecule, in reproductive [...] Read more.

Women living with HIV face an increased burden of spontaneous preterm birth (sPTB); however, the underlying immunological mechanisms of sPTB and its association with HIV infection are poorly understood. Although the limited earlier literature implicates sphingosine-1-phosphate (S1P), a lysosphingolipid signaling molecule, in reproductive biology, the association of S1P signaling with HIV and sPTB has not been investigated. We examined whether two S1P signaling components, S1P receptors and sphingosine kinases, are expressed in the female reproductive tract and whether levels are associated with HIV status or spontaneous preterm birth. We quantified the mRNA expression of sphingosine-1-phosphate receptors 1 and 3 (S1PR1/S1PR3) and sphingosine kinases 1 and 2 (SPHK1/SPHK2) in 167 banked vaginal swab specimens collected between 14 and 26 weeks of gestation in a longitudinal pregnancy cohort in Lusaka, Zambia. We evaluated the expression of S1PR1, S1PR3, SPHK1, and SPHK2 by real-time quantitative reverse transcription PCR (RT-qPCR) in four groups (n = 41–42 each): women without HIV (WWoH) with term birth (≥37 weeks of gestation; TB), WWoH with spontaneous preterm birth (<37 weeks of gestation, sPTB), women with HIV (WWH) with TB, and WWH with sPTB. We found that S1P receptors and sphingosine kinases are expressed in the female reproductive tract. SPHK1 and SPHK2 mRNA expression were generally comparable among women independent of HIV status or birth outcome, though SPHK2 trended toward higher expression in women with HIV and women with sPTB. In contrast, S1PR1 mRNA trended toward higher expression in WWH vs. WWoH overall, as well as in WWH vs. WWoH among women with sPTB. Similarly, S1PR3 mRNA expression was greater in women with HIV than in women without HIV, and WWH, both with TB and sPTB, had higher S1PR3 mRNA expression than WWoH with TB. Perturbations in S1PR1 and S1PR3 mRNA expression may be associated with inflammation related to HIV infection and spontaneous preterm birth, suggesting that further studies of S1P signaling in pregnancy, especially among women with HIV, are warranted. Full article

(This article belongs to the Special Issue Viruses in the Reproductive Tract)

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14 pages, 1671 KB

Open AccessArticle

Reassortant High Pathogenicity Avian Influenza A(H5N1) Viruses During the Reemergence in Uruguay Suggest Increasing Genetic Diversity in South America

by Ana Marandino, Gonzalo Tomás, Yanina Panzera, Valeria Uriarte, Virginia Russi, Ramiro Pérez, Lucía Bassetti, Raúl Negro, Sirley Rodríguez and Ruben Pérez

Viruses 2026, 18(5), 558; https://doi.org/10.3390/v18050558 (registering DOI) - 14 May 2026

Abstract

Highly pathogenic avian influenza (HPAI) H5N1 viruses of the goose/Guangdong (Gs/GD) lineage have driven a global panzootic since 2020, with clade 2.3.4.4b establishing sustained transmission in wild birds. In South America, early outbreaks were largely associated with the North American-derived B3.2 genotype, which [...] Read more.

Highly pathogenic avian influenza (HPAI) H5N1 viruses of the goose/Guangdong (Gs/GD) lineage have driven a global panzootic since 2020, with clade 2.3.4.4b establishing sustained transmission in wild birds. In South America, early outbreaks were largely associated with the North American-derived B3.2 genotype, which showed limited diversification after its introduction. Here, we report the genomic characterization of eight H5N1 viruses detected in Uruguay during the reemergence of avian influenza in February–March 2026. Complete genomes were obtained from wild birds exhibiting neurological signs, predominantly Coscoroba coscoroba. All viruses belong to clade 2.3.4.4b but exhibit a reassortant genomic constellation distinct from B3.2. The HA, NA, and MP segments retain the Eurasian backbone, whereas internal genes derive from both South American and North American low-pathogenicity avian influenza lineages. PB2 variation distinguishes two closely related viral groups differing in PB2 origin, whereas the remaining genomic segments retain a shared background. Sequence variation in the neuraminidase gene reduced the sensitivity of a widely used N1-specific RT-qPCR assay, highlighting limitations of existing diagnostic tools during viral evolution. These findings confirm the presence of reassortant H5N1 viruses in Uruguay and, together with recent reports from Argentina and Brazil, support an emerging pattern of genomic diversification in southern South America. Full article

(This article belongs to the Special Issue Advances in Research on Emerging and Zoonotic Diseases)

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9 pages, 654 KB

Open AccessBrief Report

Spent Medium Inhibits rVSV Infection

by Rebecca Habisch, Johannes Georg Wieland, Sophia Kessler, Peter Neubauer, Jorge Soza-Ried and Eva Puschmann

Viruses 2026, 18(5), 557; https://doi.org/10.3390/v18050557 (registering DOI) - 13 May 2026

Abstract

The cell density effect, defined as reduced cell-specific productivity above a critical cell density, remains a major limitation in virus manufacturing processes. While medium exchange prior to infection has been reported to mitigate this effect, the role of spent medium during the early [...] Read more.

The cell density effect, defined as reduced cell-specific productivity above a critical cell density, remains a major limitation in virus manufacturing processes. While medium exchange prior to infection has been reported to mitigate this effect, the role of spent medium during the early phase of infection is poorly understood. Here, we show that spent medium conditioned by high-density HEK293 cultures inhibits infection with recombinant vesicular stomatitis virus (rVSV), even when infection is performed at low cell density. The strength of inhibition increased with the density and conditioning time of the donor culture and resulted in slower replication kinetics, thereby delaying the optimal harvest time and potentially reducing overall yield. Notably, the inhibitory effect was reversible when the virus was added to cells maintained in fresh medium, indicating that inhibition is mediated by the medium rather than intrinsic changes in the cells. We excluded pH effects within 7.1–8.0, nutrient depletion, and lactate/ammonium accumulation as primary causes. Removal of cell debris and extracellular vesicles by filtration (down to 0.02 µm) and size-based retention down to 3 kDa did not restore infection, and AUC indicated no major differences in particle distributions between fresh and conditioned media. Together, our data suggest an unidentified <3 kDa inhibitor in spent medium that partially suppresses rVSV infection and slows replication kinetics. Full article

(This article belongs to the Section General Virology)

12 pages, 3245 KB

Open AccessArticle

Immunization of Foals with Equine Rotavirus A Vaccine Can Stabilize or Increase Rotavirus-Specific Neutralizing Antibody Titers

by Lianne G. Eertink, Olivia Jacob, Emma N. Adam, Allen E. Page, Dan Wang and Feng Li

Viruses 2026, 18(5), 556; https://doi.org/10.3390/v18050556 (registering DOI) - 13 May 2026

Abstract

Despite a monovalent G3P[¹²] (‘G3’) vaccine being available for horses, equine rotavirus A (ERVA) is still the predominant infectious pathogen causing diarrhea in foals in the United States of America (U.S.). Previous research has shown that maternal neutralizing antibody (NAb) titers [...] Read more.

Despite a monovalent G3P[¹²] (‘G3’) vaccine being available for horses, equine rotavirus A (ERVA) is still the predominant infectious pathogen causing diarrhea in foals in the United States of America (U.S.). Previous research has shown that maternal neutralizing antibody (NAb) titers are too high and will interfere with the vaccination of foals at 30 and 45 days of age. We aimed to determine if it is possible to increase NAb titers in foals through vaccination before they are vulnerable to ERVA infection. We immunized two foals with the commercially available vaccine (G3) at solely three months of age and seven foals at both two and three months of age, respectively. Two mock foals were vaccinated with saline buffer in this study. The dams of these foals were not vaccinated during their gestation period. All pre-vaccination G3 and G14P[¹²] (‘G14’) NAb titers in this foal cohort were 256 or lower. Following vaccination, NAb titers in foals were increased up to 1024 against G3 and up to 512 against G14 viruses, respectively. Interestingly, ERVA NAb titers either increased or stabilized in immunized foals depending on the pre-vaccination NAb titer, which contrasts with unvaccinated foals showing a rapid decline in NAb titers over time. Full article

(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)

22 pages, 2370 KB

Open AccessCommunication

The Largest Outbreak of Acute Gastroenteritis of Mixed Norovirus Genogroups in the Coast of São Paulo State, Brazil

by Rita de Cássia Carmona, Simone Guadagnucci, Mayara Esteves, Carla Costa, Simone Blotta, Daniele de Morais, Bráulio Machado, Cecilia Martins, Christiane Ristori, Ruth Rowlands, Damaris Pinto, Vitória de Souza, Bernadete Liphaus, Alessandra Xavier, Maria Inês Sato, Mikaela Barbosa, Ronalda de Araújo, Vanessa Cardoso, Luciano Candido, Renan Silva and Audrey Cilli Show full author list Hide full author list

Viruses 2026, 18(5), 555; https://doi.org/10.3390/v18050555 (registering DOI) - 13 May 2026

Abstract

During the Brazilian summer, from 29 December 2024, to 6 March 2025, a large cluster of acute gastroenteritis (AGE) outbreaks was reported along the coast of São Paulo State, Brazil, peaking in January 2025. Overall, 55 outbreaks involving 755 cases were officially notified, [...] Read more.

During the Brazilian summer, from 29 December 2024, to 6 March 2025, a large cluster of acute gastroenteritis (AGE) outbreaks was reported along the coast of São Paulo State, Brazil, peaking in January 2025. Overall, 55 outbreaks involving 755 cases were officially notified, while more than 76,000 medical consultations for AGE were recorded across the region during the same period. A total of 50 stool samples were analyzed by RT-qPCR for group A rotavirus (RVA) and norovirus (NoV). NoV was detected in 27 samples (54.0%), confirming it as the main etiological agent, while RVA was identified in one sample (2.0%). Among NoV-positive cases, genogroup II (GII) predominated (59.0%), followed by genogroup I (GI) (19.0%) and mixed infections (22.0%). Genomic sequencing successfully genotyped 23 strains (95.8%), revealing six distinct genotypes. The recombinant GII.17[P17] was predominant (48.0%), followed by GI.3[P3], GI.3[P13], GI.5[P5], GII.4 Sydney_2012[P16], GII.3[P30], as well as mixed infections. No enteric viruses were detected in drinking water. However, seawater samples showed high concentrations of NoV GI and GII (up to 10⁴ GC L⁻¹) at beaches unsuitable for bathing. Wastewater surveillance revealed high viral loads, particularly NoV GII (up to 10⁸ GC L⁻¹), consistent with reported cases. To our knowledge, this is the first report in Brazil of a NoV-associated AGE outbreak investigated through an integrated approach combining clinical, environmental, and epidemiological surveillance data. Findings highlight genotype diversity and reinforcing the importance of integrated surveillance. Full article

(This article belongs to the Special Issue Enteric Viruses in Environment and Humans: Identification, Surveillance and Control)

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13 pages, 327 KB

Open AccessArticle

Emerging Trends in HIV-1 Sub-Subtype A6 in Belgium: Transmission Dynamics, Drug Resistance, and Subtyping Tool Evaluation

by Virginie Mortier, Laurent Debaisieux, Deborah De Geyter, Marie-Luce Delforge, Melissa Depypere, Géraldine Dessilly, Benoît Kabamba-Mukadi, Khalid El Moussaoui, Samy Mzougui, Ben Serrien, Karolien Stoffels, Dominique Van Beckhoven, Ellen Van Cutsem, Dorien Van den Bossche, Sigi Van den Wijngaert, Fien Vanroye, Elizaveta Padalko, Chris Verhofstede and Kristel Van Laethem

Viruses 2026, 18(5), 554; https://doi.org/10.3390/v18050554 (registering DOI) - 12 May 2026

Abstract

The international spread of HIV-1 sub-subtype A6 raises concerns due to its association with contraindications for long-acting injectable formulations of cabotegravir (LA-CAB) and rilpivirine (LA-RPV). This study investigated its increasing proportion in Belgium, assessing transmission dynamics and potential migration links. Additionally, genotypic drug [...] Read more.

The international spread of HIV-1 sub-subtype A6 raises concerns due to its association with contraindications for long-acting injectable formulations of cabotegravir (LA-CAB) and rilpivirine (LA-RPV). This study investigated its increasing proportion in Belgium, assessing transmission dynamics and potential migration links. Additionally, genotypic drug resistance in the Belgian HIV-1 sub-subtype A6 population were analyzed and four automatic subtyping tools were compared. A dataset of 4764 HIV-1 protease and reverse transcriptase (RT) sequences from newly diagnosed, treatment-naïve individuals in Belgium (2013–2022) was analyzed. A combination of phylogenetic analysis and online subtyping tools identified 136 sub-subtype A6 sequences. The increase in the proportion of HIV-1 sub-subtype A6 observed in Belgium since 2020 reflects changing transmission patterns, especially among Belgium-born men having sex with men, and cannot be solely linked to the recent influx of Ukrainian migrants. Of these sub-subtype A6 sequences, less than 10% showed LA-CAB + LA-RPV resistance, mainly due to E138A within RT. HIVdb and ANRS reliably assessed resistance in this therapy-naïve cohort, and HIVdb, COMET, and SmartGene^® produced concordant subtyping results. While algorithm choice has little impact at low resistance prevalence, further research is necessary and HIVdb and ANRS remain more suitable for ongoing clinical and research use. Full article

(This article belongs to the Section Human Virology and Viral Diseases)

24 pages, 803 KB

Open AccessReview

Gut Microbiota and Probiotics in Influenza: A Narrative Review of Mechanisms and Emerging Evidence

by Feihu Guan, Jie Zhang, Ye Tian, Bofan Fu, Ji Liu, Yafen Song, Aoyang Yan, Bing Zhang, Ling Chen, Min Zhang, Pengfei Du, Lei Wang, Xiaoyue Yang, Sifan Guo, Chenghuai Yang, Hui Zhang and Qianyi Zhang

Viruses 2026, 18(5), 553; https://doi.org/10.3390/v18050553 (registering DOI) - 12 May 2026

Abstract

The gut microbiota, often referred to as the “forgotten organ”, plays an indispensable role in maintaining host physiological metabolism, immune function, and nutrient absorption. Moreover, the gut microbiome serves as a critical biological barrier against viral infections and is increasingly recognized as a [...] Read more.

The gut microbiota, often referred to as the “forgotten organ”, plays an indispensable role in maintaining host physiological metabolism, immune function, and nutrient absorption. Moreover, the gut microbiome serves as a critical biological barrier against viral infections and is increasingly recognized as a potential target to augment antiviral therapies. Recent studies have revealed that microbial ligands and metabolites derived from the gut microbiota are pivotal in modulating respiratory immune responses, providing compelling evidence of the complex interaction network between microorganisms and the host, particularly the signaling pathways linking the gut to distal organs such as the lungs. This review examines the communication and regulatory mechanisms between the gut microbiota and pulmonary mucosal surfaces during influenza virus infection, emphasizing how gut microbial communities and probiotics influence host immune responses, promote the production of immune-related molecules, and enhance antiviral defenses. The aim is to provide comprehensive insights into the gut–lung axis and its implications for respiratory health. Full article

(This article belongs to the Special Issue Advances in Respiratory Viruses Research: From Basic Studies to Public Health)

18 pages, 3149 KB

Open AccessArticle

Dynamics of Paraspeckle Components in Herpes Simplex Virus 1 (HSV-1)-Infected Human Neuronal Cells

by Carolina Filipponi, David C. Bloom, Carlo Gambotto, Callen T. Wallace, Jadranka Milosevic, Simon C. Watkins, Shane Buckley, Maribeth A. Wesesky, Vishwajit L. Nimgaonkar and Leonardo D’Aiuto

Viruses 2026, 18(5), 552; https://doi.org/10.3390/v18050552 (registering DOI) - 12 May 2026

Abstract

Paraspeckles are subnuclear ribonucleoprotein condensates that regulate host stress responses, including those triggered by viral infection. In vitro studies using non-neuronal cells have shown the involvement of specific paraspeckle components in facilitating the replication of certain viruses, including Herpes Simplex Virus 1 (HSV-1), [...] Read more.

Paraspeckles are subnuclear ribonucleoprotein condensates that regulate host stress responses, including those triggered by viral infection. In vitro studies using non-neuronal cells have shown the involvement of specific paraspeckle components in facilitating the replication of certain viruses, including Herpes Simplex Virus 1 (HSV-1), but these processes have not been investigated in human neuronal cells, which represent a relevant target of the virus. We employed human neural precursor cells (NPCs), neurons, and brain organoids derived from hiPSCs to investigate the previously unexplored dynamics of paraspeckle components in HSV-1-infected human neuronal cells. Our results reveal cell-type-specific differences in the expression of paraspeckle genes in response to HSV-1 infection. Unlike other viruses, HSV-1 orchestrates a previously unreported redistribution of paraspeckle proteins, leading to their accumulation in viral replication compartments (VRCs). Importantly, the expression of the paraspeckle proteins NONO and SFPQ correlates with HSV-1 permissiveness in human neuronal cells and may be required to establish a nuclear environment favoring viral transcription/replication. This enhances our understanding of how stress-response pathways in cells can be exploited by viruses in a cell-type-specific manner. Full article

(This article belongs to the Special Issue 3D Models in Viral Pathogenesis)

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2 pages, 114 KB

Open AccessEditorial

Special Issue: Microscopy Methods for Virus Research

by Krishanu Ray

Viruses 2026, 18(5), 551; https://doi.org/10.3390/v18050551 (registering DOI) - 12 May 2026

Abstract

Significant advances in microscopy methods for biomedical applications have been made in the past two decades, and these methods are now being actively pursued for virus research [...] Full article

(This article belongs to the Special Issue Microscopy Methods for Virus Research)

28 pages, 18934 KB

Open AccessArticle

Microglial-Derived IGF-1 Serves as a Regulator for Neuroimmune Homeostasis During Viral-Induced Demyelination

by Vanessa M. Scarfone, Collin Pachow, Pauline U. Nguyen, Anita Lakatos, Jamie-Jean De La Torre, Alisa Xie, Kellie Fernandez, Charlene Collado, Kaitlin Murray, Roberto Tinoco, Craig M. Walsh, Trevor Owens, Agnieszka Wlodarczyk and Thomas E. Lane

Viruses 2026, 18(5), 550; https://doi.org/10.3390/v18050550 (registering DOI) - 9 May 2026

Abstract

This study investigated the role of microglia-derived insulin-like growth factor 1 (IGF-1) in modulating host defense and disease progression in a viral model of neuroinflammation and demyelination. Intracranial infection of susceptible mice with the glial-tropic JHM strain of mouse hepatitis virus (JHMV) induces [...] Read more.

This study investigated the role of microglia-derived insulin-like growth factor 1 (IGF-1) in modulating host defense and disease progression in a viral model of neuroinflammation and demyelination. Intracranial infection of susceptible mice with the glial-tropic JHM strain of mouse hepatitis virus (JHMV) induces acute encephalomyelitis, followed by an immune-mediated demyelinating disease that mimics many clinical and histologic features of multiple sclerosis (MS). Utilizing an inducible fractalkine receptor (Cx3cr1) promoter-driven Cre-loxP recombinant system, we performed timed ablation of Igf1 in microglia to assess its impact on the central nervous system (CNS) response to JHMV. While the loss of microglial IGF-1 did not impair the control of viral replication, it significantly exacerbated spinal cord demyelination. CyTOF and imaging mass cytometry analysis of spinal cords indicated increased myelin damage was associated with increased accumulation of CD8⁺Ly6C⁺ effector T cells and reduced expression of TREM2 that impaired transition into a disease-associated microglia (DAM) phenotype capable of sensing and potentially mitigating myelin damage. Collectively, these findings argue that microglial IGF-1 is a non-redundant coordinator of the CNS immune responses that occur in response to CNS viral infection. Full article

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27 pages, 2179 KB

Open AccessArticle

Intersecting Epidemics: A Multilevel Syndemic Analysis of a Chikungunya Virus Epidemic in Colombia Through Clinical, Biological, and Socioeconomic Factors

by Juan C. Rueda, Ana María Santos, Ignacio Angarita, Ingris Peláez-Ballesta, Alfonso Gastelum, Igor Rueda, Jaime Cortés-Ramos, Cristian Astudillo, Daniel Rincón-Sierra, Karina Guzmán, Jesús Giovanny Ballesteros, Juan Manuel Bello and John Londono

Viruses 2026, 18(5), 549; https://doi.org/10.3390/v18050549 (registering DOI) - 9 May 2026

Abstract

This study applied a syndemic framework to chikungunya virus (CHIKV) infection during the 2014–2015 Colombian epidemic, integrating biological and social determinants. Methods: A community-based cohort of 279 serologically confirmed adults from six cities was analyzed. Clinical, sociodemographic, and cytokine data were evaluated using [...] Read more.

This study applied a syndemic framework to chikungunya virus (CHIKV) infection during the 2014–2015 Colombian epidemic, integrating biological and social determinants. Methods: A community-based cohort of 279 serologically confirmed adults from six cities was analyzed. Clinical, sociodemographic, and cytokine data were evaluated using multilevel and multivariate statistical approaches. Results: Among 279 patients, 141 (50. 5%) met World Health Organization (WHO) criteria for acute CHIKV infection. The cohort was predominantly female and of lower socioeconomic status (SES). The most frequent manifestations were arthralgia (91%), fatigue (58%), fever (50.5%), myalgia (45.9%), and rash (45.2%). Multivariate models identified IL-15, IL-17A, IL-12p40, MCP-1, and MIP-1α as significant correlates of fever, rash, and myalgia. Socioeconomic and ethnic factors influenced cytokine expression; Caucasian patients showed higher proinflammatory cytokine levels than Afro-American patients. Lower SES was associated with greater symptom burden. Network analyses revealed distinct immune signatures linking biological responses with clinical and demographic variables. Conclusion: Immune responses, clinical manifestations, and social disadvantages interact significantly in CHIKV infection. These findings support a syndemic model in which socioeconomic vulnerability amplifies disease impact, highlighting the need for integrated biosociological public health strategies, particularly targeting populations with low socioeconomic status. Full article

(This article belongs to the Section Human Virology and Viral Diseases)

25 pages, 7474 KB

Open AccessArticle

Application of a Blood–Brain Barrier Organ-on-a-Chip Model for Assessment of Countermeasure Efficiency Against Eastern Equine Encephalitis Virus

by Niloufar A. Boghdeh-Olson, Michael D. Barrera, Clayton M. Britt, David K. Schaffer, Jacquelyn A. Brown, John P. Wikswo and Aarthi Narayanan

Viruses 2026, 18(5), 548; https://doi.org/10.3390/v18050548 (registering DOI) - 9 May 2026

Abstract

Infection by neurotropic alphaviruses such as the Eastern equine encephalitis virus (EEEV) causes extensive inflammation in the central nervous system and tissue damage, including disruption of the blood–brain barrier (BBB). Neuroinflammation and BBB disruption following infection are critical pathological considerations for the development [...] Read more.

Infection by neurotropic alphaviruses such as the Eastern equine encephalitis virus (EEEV) causes extensive inflammation in the central nervous system and tissue damage, including disruption of the blood–brain barrier (BBB). Neuroinflammation and BBB disruption following infection are critical pathological considerations for the development of robust countermeasure strategies. Encephalitic disease resulting from EEEV infection currently lacks FDA-approved therapeutic intervention strategies, thus exposing a major capability gap in the ability to address the global health burden that could result from alphavirus infections. In this manuscript, we present a gravity-flow Neurovascular Unit (gNVU) model of the human BBB that may be used for modeling EEEV-induced neuropathology and evaluating countermeasures. The data generated using this model show that EEEV infection causes a time-dependent disruption of BBB integrity and increases the inflammatory load in a manner that correlates with an increase in the viral load. The data also show that the route of introduction of the pathogen has an impact on the pathology measured, with infection through the brain side eliciting a greater inflammatory outcome than infection through the vascular route. Overall, the included data support the utility of this organ-on-a-chip (OOC) platform of the human BBB in understanding encephalitic disease caused by neurotropic viruses and evaluation of therapeutic intervention strategies. Full article

(This article belongs to the Section Human Virology and Viral Diseases)

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23 pages, 6978 KB

Open AccessArticle

Hypoxia-Enhanced N110 Glycosylation of Hemagglutinin Promotes H3N2 Influenza Virus Fitness by Modulating Receptor Binding and Immune Evasion

by Ting Zhang, Yihui Fang, Jie Liu, Ao Guo, Bin Yuan, Yanan Zhang, Lihua Ding and Qinong Ye

Viruses 2026, 18(5), 547; https://doi.org/10.3390/v18050547 - 8 May 2026

Abstract

The hemagglutinin (HA) of influenza A/H3N2 virus evolves rapidly, with glycosylation driving immune evasion. However, how host microenvironmental cues influence this process remains poorly understood. We identified a novel N-linked glycosylation site at position 110 (N110) in contemporary H3N2 viruses (NSS genotype) that [...] Read more.

The hemagglutinin (HA) of influenza A/H3N2 virus evolves rapidly, with glycosylation driving immune evasion. However, how host microenvironmental cues influence this process remains poorly understood. We identified a novel N-linked glycosylation site at position 110 (N110) in contemporary H3N2 viruses (NSS genotype) that enhances viral fitness by increasing receptor-binding signal, HA cleavage, and replication. Remarkably, hypoxia, which mimics the respiratory tract microenvironment, significantly augments N110 glycosylation. Mechanistically, we identified the B4GAT1-B4GALT1 complex as the key mediator of this modification. Hypoxia upregulates their expression and strengthens their interaction with HA. In ferret models, N110-glycosylated viruses exhibit heightened pathogenicity and evade ancestral antibodies. Furthermore, immunization with N110-containing HA confers broad-spectrum protection, whereas reciprocal immunization is ineffective. Our findings reveal hypoxia-driven glycosylation as a previously unrecognized mechanism of H3N2 adaptation, providing critical insights for vaccine efficacy and highlighting the importance of integrating microenvironmental factors into future antiviral strategies. Full article

(This article belongs to the Section Human Virology and Viral Diseases)

20 pages, 999 KB

Open AccessReview

NLR Inflammasomes in Viral Infections: From Molecular Mechanisms to Therapeutic Interventions

by Shiyuan Hou, Xing Shen, Danni Sun, Yulin An, Yuxuan Zhou, Xing Sun, Shuhan Wang, Xinyue Liu, Mengting Zhu, Shuai Zhao, Ziyu Liu, Xingan Wu and Rongrong Liu

Viruses 2026, 18(5), 546; https://doi.org/10.3390/v18050546 - 8 May 2026

Abstract

The innate immune system serves as the primary barrier against viral invasion, utilizing pattern recognition receptors (PRRs) to orchestrate a rapid defense. Among these, the nucleotide-binding domain and leucine-rich repeat (NLR) containing proteins function as central signaling scaffolds, assembling into multiprotein complexes known [...] Read more.

The innate immune system serves as the primary barrier against viral invasion, utilizing pattern recognition receptors (PRRs) to orchestrate a rapid defense. Among these, the nucleotide-binding domain and leucine-rich repeat (NLR) containing proteins function as central signaling scaffolds, assembling into multiprotein complexes known as inflammasomes. These complexes drive the maturation of pro-inflammatory cytokines IL-1β and IL-18, and initiate gasdermin D (GSDMD)-mediated pyroptosis, a lytic cell death pathway that eliminates intracellular replication niches. This comprehensive review synthesizes the diversified landscape of inflammasome activation during viral infections, extending beyond the canonical NLRP3 inflammasome to include specialized sensors such as NLRP6, NLRP9, NLRP1, NLRP12, and NLRC4. We critically evaluate the evolutionary “arms race” between host defenses and viral pathogens, detailing the sophisticated immune evasion strategies employed by viruses—ranging from the expression of decoy proteins and direct proteolytic cleavage of immune sensors to the manipulation of post-translational modifications (PTMs). Furthermore, we discuss the dual nature of inflammasome activation, which balances protective viral clearance against pathological hyperinflammation, and provide an exhaustive analysis of novel therapeutic strategies, including direct NLR inhibitors and downstream cytokine blockers, currently navigating clinical transition. Full article

(This article belongs to the Special Issue Viral Mechanisms of Immune Evasion)

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20 pages, 2135 KB

Open AccessArticle

Identification of Cyclin L1 as a Host Factor Regulating Hepatitis B Virus Replication

by Collins Oduor Owino, Balakrishnan Chakrapani Narmada, Gian Yi Lin, Pauline Poh Kim Aw, Nivrithi Ganesh, Jovi Tan Siying, Marie-Laure Plissonnier, Thangavelu Thangavelu Matan, Niranjan Shirgaonkar, Pablo Bifani, Massimo Levrero, Giridharan Periyasamy, Seng Gee Lim and Ramanuj DasGupta

Viruses 2026, 18(5), 545; https://doi.org/10.3390/v18050545 (registering DOI) - 8 May 2026

Abstract

Background and Aims: Understanding regulatory interactions between hepatitis B virus (HBV) and host factors is essential for the development of next generation host-directed antiviral therapies and the achievement of a functional HBV cure. Here, we investigated HBV-induced alterations in host gene expression in [...] Read more.

Background and Aims: Understanding regulatory interactions between hepatitis B virus (HBV) and host factors is essential for the development of next generation host-directed antiviral therapies and the achievement of a functional HBV cure. Here, we investigated HBV-induced alterations in host gene expression in primary human hepatocytes (PHH) to identify host factors exploited by the virus for replication and persistence. Whole-transcriptome sequencing (WTS) of HBV-infected PHH identified host pathways with potential roles in the HBV life cycle. RNA interference-based functional screening of dysregulated candidate genes identified cyclin L1 (CCNL1) as a key host factor. RNAi-mediated knockdown of CCNL1 reduced HBV gene expression, including hepatitis B surface antigen (HBsAg). Mechanistically, CCNL1 regulates phosphorylation of the C-terminal domain (CTD) of RNA polymerase II (RNAPII) at serine 2 (S2), consistent with a role in transcriptional regulation. CCNL1 knockdown further reduced the binding of total and phospho- (Ser2/Ser5) RNAPII, pan-acetylated histone H3 (H3ac), and H3K27ac to HBV covalently closed circular DNA (cccDNA), indicating impaired cccDNA-dependent transcription. In addition, CCNL1 expression was elevated in chronic hepatitis B patients compared with those with resolved infection. Collectively, these data demonstrate that CCNL1 promotes HBV transcription and replication through modulation of RNAPII phosphorylation and chromatin-associated transcriptional activity, identifying CCNL1 as a potential host susceptibility factor for HBV. Importance: Hepatitis B virus infection remains a major threat to human health in areas with high prevalence. There is need to fully understand the complex interactions between the virus and human host factors/processes to support ongoing efforts to develop anti-HBV therapies that can be used with existing therapies to achieve a better cure. HBV relies on host cellular factors and biological processes to establish and maintain efficient infection, making host–virus interactions attractive targets for therapeutic intervention. Thus, identifying host factors that support and/or restrict HBV infection is essential for understanding the molecular basis of chronic HBV infection and for developing host-targeting anti-HBV drugs. This study identifies cyclin L1 (CCNL1) as a host susceptibility factor that promotes HBV transcription and replication through regulation of RNA polymerase II activity and or post-transcriptional mechanisms. Full article

(This article belongs to the Section Human Virology and Viral Diseases)

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