Virology

Okra (bhendi) is a widely cultivated food crop in warm regions of the world, with India contributing approximately 60% of global production. However, okra cultivation in India is severely constrained by viral diseases, among which infections caused by the geminiviruses bhendi yellow vein mosaic virus (BYVMV) and okra enation leaf curl virus (OELCuV), in association with their satellites, represent major limitations to crop productivity. In recent years, the geminivirus-encoded C4 protein has emerged as a key pathogenicity determinant in this viral family, with functions that include suppression of multiple layers of plant antiviral defence and induction of disease symptoms. Here, we comparatively characterize the C4 proteins of BYVMV and OELCuV by determining their targeting signals and subcellular localization, and by assessing their ability to induce developmental abnormalities and suppress the cell-to-cell spread of RNA silencing. Our results reveal that the two C4 proteins display distinct subcellular localization patterns, yet both are capable of inducing developmental alterations, likely through different mechanisms, and of suppressing the intercellular spread of RNA silencing, possibly via interaction with a common host factor. Together, these findings suggest that C4 might be a critical virulence factor in okra-infecting geminiviruses and act as a symptom determinant. The C4 proteins encoded by BYVMV and OELCuV therefore emerge as promising targets for the development of antiviral management strategies in okra.

Borealpox virus (BRPV; formerly Alaskapox) is an orthopoxvirus that has caused seven reported human infections in Alaska since 2015, including a fatal case in 2023. The natural reservoir of BRPV is unknown, although previous investigations have raised the possibility of wild small mammals transmitting the virus to humans, either through direct contact or via domestic cats and dogs. To understand which species may be involved in the maintenance and/or spillover of BRPV in Alaska, we trapped and sampled wild small mammals (including voles, shrews, and squirrels) in 2021 and 2024 near reported human case locations in Fairbanks and the Kenai Peninsula, respectively. We found evidence of previous exposure to orthopoxviruses in five species (including the House Mouse, Mus musculus) and detected BRPV DNA as well as viable virus in Northern Red-backed Voles (Clethrionomys rutilus). Further, screening of tissues from historical museum specimens revealed BRPV DNA in C. rutilus specimens collected in Denali National Park and Preserve in 1998 and 1999, 17 years before the first reported human case of BRPV. Phylogenomic analysis of all human and animal BRPV isolates strongly supports the hypothesis of local human infections through multiple spillover events. These findings suggest C. rutilus as a possible reservoir species for BRPV and indicate that BRPV has been present in Alaskan wild small-mammal populations for at least 25 years. Our study highlights the potential of museum collections to elucidate the temporal, spatial, and host ranges of emerging pathogens. Further museum- and field-based sampling will clarify the true geographic range of BRPV, which is closely related to Old World orthopoxviruses and may be circulating beyond North America.

Thogotoviruses are a group of tick-borne, six-segmented, negative-sense single-stranded RNA viruses. These viruses encode an RNA-dependent RNA polymerase that recognizes promoter sequences located at the genomic termini to initiate RNA synthesis. The 5' and 3' ends of the genome bind to the polymerase and function as a promoter. Outside the catalytic center, they base-pair with each other to form a double-stranded RNA structure. This structure is referred to as the distal duplex and plays an important role in RNA synthesis. In this study, we investigated how the RNA sequence of the distal duplex influences polymerase activity using minigenome systems of two thogotoviruses, Oz virus (OZV) and Dhori virus (DHOV). Each virus exhibits distinct activities among its six segments. In OZV, one determinant of these differences is the base pair at positions 5'12 and 3'11 within the distal duplex, where promoter activity varies depending on whether the base pair is G:C or A:U. In contrast, the DHOV polymerase is not affected by this difference. These results indicate that, even within the genus Thogotovirus, viruses differ in whether they possess a mechanism that modulates promoter activity based on subtle sequence differences within the distal duplex. Furthermore, phylogenetic analysis and comparison of promoter sequences suggest that thogotoviruses can be divided into groups that do or do not regulate intersegment promoter activity via the base pair at positions 5'12 and 3'11. HighlightsO_LIMinigenome systems of Oz virus and Dhori virus reveal segment-specific differences in promoter activity C_LIO_LIThe distal duplex sequence modulates RNA synthesis in a virus-dependent manner C_LIO_LIThe base pair at positions 5'12/3'11 determines promoter activity in Oz virus but not in Dhori virus C_LIO_LIThogotoviruses can be divided into groups that do or do not regulate promoter activity via distal duplex sequence variation at positions 5'12/3'11 C_LI

Avian influenza virus (AIV) epidemiology is well-documented in temperate regions but remains poorly understood in isolated ecosystems like tropical oceanic islands. On these islands, seabirds nest in dense interspecific colonies where the role of different species as reservoirs and dispersers of AIV may vary greatly. Here, we examine the role of noddies (Anous spp.) as potential reservoirs for low pathogenic AIV and evaluate their potential as sentinel species for highly pathogenic AIV introduction on tropical oceanic islands. We analyzed blood samples from 11 seabird species across eight islands in the southwestern Indian Ocean (2015-2020). Noddies exhibited high, stable seroprevalence (30-45%), comparable to reservoir host species in temperate regions. The detection of two N7-positive noddies, sampled the same year on two distinct islands, provided direct molecular evidence that AIV actively circulates on these island colonies. While most other species showed low exposure, Bridled Terns (Onychoprion anaethetus) had exceptionally high seroprevalence (80%), though their reservoir status requires further investigation due to limited sampling. Given noddies consistent exposure and regional distribution, we recommend prioritizing islands with large noddy populations for AIV surveillance. Continued investigation of viral dynamics within and among islands is now called for to elucidate the ecological drivers of AIV maintenance and transmission.

In the present study, we assessed the pathogenicity of H5N8 avian influenza viruses belongs to the clade 2.3.4.4b in chicken. Birds of three different dose groups, 102, 104, and 106 EID50 were used in the study. No mortality was observed in 102 EID0 group. Percent cumulative mortality of 104 and 106 EID50 group was 66.67 and 100 %, respectively. Varying duration of MDT of 3.2 and 2 days was observed in 104 and 106 EID50 group, respectively. The CID50 of virus was found to be 104.5 EID50. High no. of viral RNA copies were found both in oropharyngeal and cloacal swabs and in various organs of birds infected in 104 and 106 EID50 group. Significant gross and histological changes and presence of viral antigen in various organs were observed in 104 and 106 EID50 group. So, the study concludes that Indian HPAI, H5N8 isolates are highly pathogenic in nature to chicken by affecting most organs systemically. CID50 of this H5N8 virus indicates poor adaption in chicken and it implies poor transmission possibility of this virus for host species in field condition. Though this virus is highly pathogenic in nature as that of HPAI, H5N1 viruses, absence of endothelial staining in most organ attributes variation in replication process and pathogenesis from HPAI, H5N1 viruses. Hence, further studies need to be done to elucidate the pathobiology of this virus in various bird species. HighlightsO_LIH5N8 virus belong to the clade 2.3.4.4b, Indian isolate is highly pathogenic in nature as that of HPAIV, H5N1. C_LIO_LIThe dose inocula, 102 EID50 is noninfectious to chicken. C_LIO_LIThe dose inocula, 104 and 106 EID50 had caused significant mortality in the inoculated chicken with MDT of 2 and 3.2 days, respectively. C_LIO_LIH5N8 virus was detected with high viral titres in clocal and oral shedding and in multiple organ with the dose inocula, 104 and 106 EID50. C_LIO_LI104 and 106 EID50 of H5N8 inocula virus caused significant gross and histological changes in multiple organs and viral antigens were detected in respective organs. C_LI

Rio Negro virus (RNV) is an enzootic alphavirus and a member of the Venezuelan equine encephalitis virus (VEEV) complex. Despite its wide circulation in South America, RNV remains a neglected pathogen with no established wild-type animal model to study its pathogenesis. In this study, we developed a lethal mouse model using 18-day-old wild-type C57BL/6 mice to characterize the systemic and neurological features of RNV infection. Following subcutaneous inoculation, RNV exhibited rapid systemic dissemination with a brief and low-titer viremic phase and high viral loads in lymphoid tissues, pancreas, brains, and lungs. Notably, infected mice developed progressive neurological signs, including ataxia and hindlimb paralysis, culminating in 100% lethality. Histopathological analysis revealed significant damage, highlighted by a striking collapse of the splenic architecture, inflammatory and remodeling changes in the lungs, and prominent inflammatory infiltrates with neurodegenerative changes in the brain. The splenic disruption was further evaluated by immunofluorescence analysis of the spleen, which showed a consistent loss of compartmentalization, characterized by an atypical infiltration of CD8+ T cells into B-cell follicles. The terminal stage of disease was characterized by extensive neuroinflammation and neurodegeneration. Histological examination of the brain revealed meningoencephalitis, robust astrogliosis, and widespread somatodendritic and terminal degeneration, particularly clustered around blood vessels. These findings were supported by cytokine analysis of brain homogenates, which showed a significant upregulation of IFN-{gamma}, IL-6, and MCP-1/CCL2 during symptomatic stages. Collectively, these findings establish a reproducible, non-genetically modified animal model that reveals the pathogenic potential of RNV in the context of immune immaturity characteristic of early life. By identifying these specific pathological and neuroinflammatory markers, our study provides a foundational experimental framework to investigate the mechanisms underlying RNV emergence and host-pathogen interactions within the VEEV complex. Author summaryMany viruses circulate silently in nature, hidden within animal populations, until environmental or social changes bring them into contact with humans. The Rio Negro virus is one such example. Despite being closely related to the better-known Venezuelan equine encephalitis virus and showing evidence of circulation in South America, RNV has remained largely overlooked by the scientific community and public health authorities. In this study, we established a new experimental model using infant mice that allowed us to observe how the virus spreads and causes damage. We found that the virus rapidly reaches the brain and other vital organs, causing severe inflammation in the brain, inflammatory changes in the lungs, and a breakdown of the immune systems organization in the spleen. By using a genetically unmodified model, we were able to observe the infection in a host with an intact but naturally immature immune system. This approach avoids the artificial conditions of genetic engineering while providing a more realistic window into how host age and developmental stages can influence the outcome of neglected viral infections. We believe our work is a first step toward understanding how this overlooked virus emerges and highlights the need to monitor and prepare for "silent" pathogens that could pose a risk to public health in an era of intensifying human activity and ecological pressure.

Insects are associated with diverse RNA viruses, including vertically transmitted viruses that form persistent infections without apparent symptoms. One of the first documented vertically transmitted viruses is sigmavirus (Rhabdoviridae) affecting fitness of Drosophila. Sigmaviruses and related rhabdoviruses have also been detected in pest fruit flies and other arthropods. However, their prevalence, transmission, tissue localisation and fitness effects remain poorly known, despite their potentially common infections in diverse hosts. We investigated Sigmavirus tryoni (BtSV) prevalence, load, transmission across multiple generations and host effects in Queensland fruit fly (Bactrocera tryoni), Australias most significant horticultural pest, which carries BtSV at low prevalence (13.7%) across field populations. We detected BtSV in 6 of 12 laboratory populations (prevalence 12.5% to 80.4%) where it was transmitted biparentally within embryos. Although incomplete, maternal transmission was more reliable and resulted in higher BtSV load than paternal transmission. Paternally transmitted BtSV was almost entirely lost after two generations. BtSV became detectable in most uninfected individuals cohabiting with infected flies, but this resulted in a low load that was subsequently transmitted to only few offspring. BtSV occurred across developmental stages, digestive and reproductive tissues, albeit its viral load was lower in reproductive tissues when received paternally than maternally, and lower in testes than ovaries. Furthermore, BtSV-infected individuals suffered paralysis and mortality when exposed to high CO2 concentrations, a Rhabdoviridae effect previously reported for several Drosophila species, a muscid fly and mosquitoes. Our study suggests that sigmavirus transmission dynamics and fitness effects may apply broadly to arthropod hosts and affect their management.

Pneumonia virus of mice (PVM), the mouse homolog to respiratory syncytial virus (RSV), is increasingly used as surrogate model to study pneumovirus pathogenesis in a more natural pathogen-host relation. Two major strains of PVM, strain 15 and J3666 are currently used in laboratories, with preferences for either one or the other based on the well-documented isolation history of strain 15 or the suggested higher virulence of strain J3666. Using conventional and long read sequencing, we found that the PVM strain J3666 represents two distinct virus populations, which are defined by sequence and structure of the G and SH genes encoding the putative attachment and small hydrophobic proteins, in addition to further nucleotide polymorphisms. Specifically, a nucleotide polymorphism at position 65 in the G gene results in either an upstream open reading frame (uORF) preceding the main ORF in frame, or an extension of the major G ORF by 18 codons. The impact of the different forms of the J3666-G genes on PVM was examined by generating recombinant PVMs differing exclusively in the distinctive 5 portion of the respective G gene. This revealed that the population expressing a G protein with an extended main G ORF was more virulent, whereas the presence of a uORF attenuated virulence. The virulence of PVM correlated with increased expression levels of G, whereas attenuation was rather associated with downregulated expression of G due to the presence of a uORF. Thus, modulation of G protein levels may be an important mechanism by which pneumoviruses modulate virulence. ImportanceThe pneumonia virus of mice strain J3666 is considered a more virulent and more suitable model for severe lower respiratory tract infections. The organization of the gene for the attachment protein G is reported to contain a small upstream open reading frame (uORF) preceding the main G ORF in frame. The translated G protein is predicted to comprise 396 amino acids. We report that this virus strain may be a mixture of two different populations, each with differing virulence. The more virulent population encodes a G protein of potentially 414 amino acids instead of a small uORF. This G gene organization is associated with an increased G protein expression. Importantly, this organization of the G gene is in line with that of several newly identified pneumoviruses, i.e., canine and swine pneumoviruses. These viruses may comprise a distinct group within the Pneumoviridae family.

Dengue infections are considered an increasing threat to mankind due to their rapid global spread rate. The development of a widely accepted drug/vaccine is hindered due to an incomplete understanding of the virus lifecycle. Present data suggest that a cytoskeleton protein, called MYH9 binds to the 3UTR, at A4 region, a highly conserved part of the UTR across the serotypes. The levels of this protein were found to be elevated in the cells infected with the virus and the above increase is commensurate with the virus load. This protein is found to accumulate at the endoplasmic reticulum (site of virus replication) and interacts with dsRNA (a replicative intermediate), suggesting its involvement in replication. Inhibition of this proteins expression by its siRNA reduced viral load, supporting its role in viral replication. Immunofluorescence studies indicate that this protein accumulates at the cell periphery and pulldown studies suggest that this protein interacts with the viral envelope protein, suggesting a role in the dengue viruss cellular entry, possibly by acting as a receptor. Use of an anti-MYH9 drug, ML-7 indicated the reduction of the virus load, prevented the accumulation at the periphery and aided in regaining the cell morphology of virus infected cells, confirming its role in replication and entry. Collectively, these studies demonstrate a dual function of MHY9 in the virus life cycle, which may serve as a general paradigm for the other viruses and hence to develop specific drugs.

Pathogens can have major impacts on ecosystem function, especially when they infect foundational taxa such as habitat-forming kelp. Understanding which potentially pathogenic microbes are in an ecosystem is increasingly important as environmental change intensifies ecosystem imbalances, yet few studies have examined which viruses are associated with key habitat-forming macroalgae. Here, we screened multiple populations of intertidal / shallow-subtidal macroalgae in New Zealand for viruses. Samples were collected from three southern bull kelp species: Durvillaea antarctica, Durvillaea poha and Durvillaea willana. Total RNA was extracted and sequenced from both obviously unhealthy kelp tissue and apparently healthy tissue. We identified several novel viruses belonging to the families Mitoviridae, Narnaviridae, Partitiviridae, Totiviridae, and the recently proposed Ormycovirus group. Viral abundance and diversity were notably higher at Lawyers Head (the site closest to a city) compared to other sites, especially in unhealthy kelp samples. The elevated viral presence at Lawyers Head may indicate that human impacts, such as pollution, are influencing viral prevalence. This study reveals that coastal macroalgae in New Zealand host diverse viral lineages, some likely pathogenic, and hints at the potential influence of anthropogenic pollution on marine virome composition.

RNA viruses are common in tephritid fruit flies including the Queensland fruit fly, Australias most significant horticultural pest. For many their transmission, tissue tropism and load across host development remain unexplored. Yet these factors are important for host biology, ecology and pest management. We investigated Bactrocera tryoni orbivirus (OV), Bactrocera tryoni xinmovirus (XV), Bactrocera tryoni toti-like virus (TLV) and Bactrocera tryoni iflavirus species 2 (IVsp.2) that commonly coinfect B. tryoni laboratory populations. OV and XV transmission was vertical within and on eggs, while TLV transmission was vertical within eggs. IVsp.2 was not detected in eggs but was present in adults; however, IVsp.2 was horizontally transmitted, with viral load increasing with cohabitation time with infected flies. Horizontal transmission was not observed for the other viruses. OV had a similar load across all tissues, while XV was consistently more abundant in ovaries. TLV had a high viral load in the brain whereas IVsp.2 was abundant in the thorax, foregut and midgut. Besides differences in eggs, the viruses were detected in all other developmental stages, but viral load patterns differed: viral load remained constant for TLV, fluctuated for OV and XV, and was low in pre-adult stages and high in adults for IVsp.2. Our findings demonstrate distinct transmission strategies and tissue tropism among the viruses, providing new insights into their epidemiology and role in host biology. Furthermore, contrary to prevailing views that viruses are generally horizontally transmitted, most known RNA viruses of B. tryoni are vertically transmitted affecting the evolution of host-virus interactions.

Defective interfering particles (DIPs) derived from the influenza A virus (IAV) are a promising antiviral agent due to their strong antiviral efficacy demonstrated in various animal models. OP7 is an unconventional IAV DIP with multiple point mutations in the viral RNA (vRNA) of genome segment 7, as opposed to the large internal genomic deletions typically found in conventional IAV DIPs. Further, OP7 showed an even higher interfering efficacy than conventional DIPs. However, the inhibitory effect of OP7 on standard virus (STV) replication has primarily been investigated in Madin-Darby Canine Kidney (MDCK) cells, which lack a functional myxovirus resistance (Mx)-mediated antiviral activity against IAV. In this study, we examined the antiviral activity and mechanism of antiviral action of OP7 in an interferon (IFN)-competent human lung carcinoma cell line (Calu-3) in vitro. We performed STV and OP7 co-infection experiments using a variety of infection conditions and measured the time-resolved dynamics in viral titer, vRNA, protein level, and host cell gene expression. We observed that OP7 co-infection results in enhanced type I IFN responses and markedly reduced infectious virus release, even at low doses. Additionally, we found that at a high STV multiplicity of infection (MOI), the replication interference of OP7, suppressing the replication of STV vRNA, appears to be the dominant mechanism of its antiviral action. At a low MOI, however, IFN induction seems to be more important. Furthermore, we examined the efficacious co-infection time window for potential prophylactic and therapeutic antiviral treatment. We observed an antiviral effect exerted by OP7 infection for up to seven days before STV infection and up to 24 hours after STV infection. Together, these findings demonstrate that OP7 is a potent antiviral DIP. Therefore, this work supports the further development of OP7 as a therapeutic and prophylactic antiviral agent.

Amdoparvoviruses have historically been documented almost exclusively in carnivores, with recent detections in bats. However, endogenous viral elements in rodent genomes suggest a more ancient and taxonomically broader evolutionary history. Despite this, small mammals have never been systematically surveyed for extant amdoparvovirus infections. In this study, we used whole genome sequencing to screen four different shrew species and wild American mink in the UK, which may act as a reservoir host for amdoparvoviruses. We identified a highly divergent amdoparvovirus in native common shrews (Sorex araneus) from northern England, tentatively named Shrew parvovirus 1(SP 1). Classical amdoparvovirus sequences were also detected in wild American mink (Neogale vison), confirming the presence of known amdoparvovirus strains in UK mustelids. Phylogenetic analysis revealed that the shrew virus, SP 1, forms a distinct clade, suggesting ancient divergence or long-term cryptic circulation in small mammal reservoirs. These findings fundamentally challenge the hypothesis that amdoparvoviruses are carnivore-restricted pathogens and underscore the importance of systematic wildlife surveillance for understanding viral host range evolution and assessing spillover risks.

Dairy cattle have emerged as a prolific amplifying host for highly pathogenic avian influenza virus (HPAIV) H5N1 clade 2.3.4.4b and a new source for cross-species and zoonotic transmission. Independent introductions of H5N1 with unclear exposure routes have been reported in several dairy herds across the U.S. These events escalate the pandemic potential of HPAIV H5N1 as transmission within and between mammalian species present opportunities for mammalian adapted H5N1 viruses to emerge. Although more than 1000 herds have been infected, bovine H5N1 influenza virus pathogenesis, transmission, and evolution in dairy cattle remains not well characterized. Working with H5N1-infected lactating cattle in high containment has been a major challenge due to the required infrastructure and logistics associated with housing, husbandry, and waste management for this model. Thus, developing alternative bovine models that maintain biological relevance while reducing operational complexity is warranted. Here we evaluate the susceptibility of lactating Jersey cattle in the dry-off period and characterize the effect of inoculation dose on the mammary pathogenicity of HPAIV H5N1 genotype B3.13. The results of this study demonstrate that dairy cows 21 days into the dry-off period are highly susceptible to HPAIV H5N1, recapitulating the severe clinical and pathological outcomes observed in infected lactating cows under experimental conditions and in field cases. We also observed an association between virus dose and the onset and severity of mastitis in individual udder-quarters and compartmentalized clonal expansion of variant populations. Overall, this study demonstrates that dry cows can provide a feasible model to study H5N1 virology, pathology, and humoral immunology in dairy cows.

Reptarenaviruses are viruses belonging to the genus Reptarenavirus within the family Arenaviridae, which infect snakes and cause inclusion body disease (IBD), a fatal condition characterized by behavioral abnormalities and wasting. Although many reptarenaviruses have been identified thus far, the phylogenetic gaps between reptarenaviruses and the other arenaviruses suggest the existence of yet-to-be-identified reptarenaviruses filling the gaps. In this study, we identified a novel reptarenavirus from publicly available RNA-seq data derived from Amazon coral snake (Micrurus spixii) and tentatively named it Amazon coral snake virus 1 (ACSV-1). We identified four ACSV-1 contigs containing the putative full-length open reading frames of the NP, GP, and L genes, as well as the partial Z gene. Phylogenetic analyses showed that ACSV-1 is highly divergent from known reptarenaviruses. The NP, GP, and L genes showed 48.3%, 42.3%, and 45.7% nucleotide sequence identities, respectively, with those of the closest relatives. Based on the International Committee on Taxonomy of Viruses (ICTV) species demarcation criteria, ACSV-1 can be assigned to a novel species of virus within the genus Reptarenavirus. This study expands our understanding of the diversity and evolution of reptarenaviruses.

Porcine reproductive and respiratory syndrome (PRRS) is a globally distributed disease caused by Betaarterivirus europensis (PRRSV-1) and Betaarterivirus americense (PRRSV-2). Its clinical presentation ranges from subclinical infection to severe disease, depending on viral evolution and the emergence of novel variants. The aim of this study was to characterize the genetic diversity and identify recombination events in the ORF7 (nucleocapsid, N) gene of ten PRRSV-2 strains circulating in pig farms in Lima, Peru. Bioinformatic analyses were performed using DNAMAN v10.0, MEGA 6, BepiPred-2.0, DnaSP v6, and RDP v4.101. Phylogenetic analysis revealed two well-defined lineages: eight strains clustered within lineage 1A (NADC34-like), and two within lineage 5A (VR2332-like), demonstrating the co-circulation of genetically distinct variants in the region. Comparative sequence analysis identified significant amino acid substitutions in eight strains (15, 16, 17, 20, 21, 22, 23, and 24), with strain 24 being the most divergent, accumulating multiple substitutions, including T81I, R109S, I115F, R116S, and A119K within the C-terminal region encompassing antigenic domains I-V. B-cell epitope prediction using BepiPred-2.0 identified six epitope patterns (A-F) comprising nine potential B-cell epitope regions (positions 5-19, 33-72, 33-73, 84-85, 87-98, 84-98, 87-97, 84, and 119). Patterns B, E, and F exhibited four to five predicted epitope sites and corresponded to strains 21, 22, 23, and 24. Recombination analysis using RDP v4.101 detected a statistically robust recombination event in strain 18_montana2020-R (lineage 5A), with strain 24_montana2020-WT (lineage 1A) identified as the putative major parent (100% similarity) and the vaccine-like VR2332 strain (lineage 5A) as the minor parent (99.3% similarity). Secondary evidence of the same recombination event was observed in strain 19_montana2020-R. Genetic diversity analysis of the ORF7 gene identified 50 polymorphic nucleotide sites and 52 mutations. Overall, these findings demonstrate substantial genetic variability in the ORF7 gene of PRRSV-2 circulating in Lima, Peru, characterized by lineage co-circulation and inter-lineage recombination. Continuous molecular surveillance is warranted to monitor viral evolution, assess potential antigenic implications, and support effective PRRS control strategies in the Peruvian swine industry.

The emerging tick-borne flaviviruses Alkhumra haemorrhagic fever virus (AHFV) and Kyasanur Forest disease virus (KFDV) can cause severe haemorrhagic disease, yet the mechanisms driving vascular leakage remain unclear. The microvascular capillaries and post-capillary venues affected during orthoflavivirus-related haemorrhagic disease consist of an endothelial cell barrier ensheathed in and supported by perivascular cells (pericytes), and we recently identified a critical role for pericytes in amplifying dengue microvascular dysfunction. The orthoflavivirus non-structural protein 1 (NS1) has been implicated in endothelial dysfunction and vascular leakage for several tick- and mosquito-borne flaviviruses. Here, we examined whether NS1 from AHFV and KFDV disrupts microvascular barrier integrity in pericyte-endothelial cell cocultures. Under our experimental conditions, we found AHFV and KFDV NS1 to disrupt the ability of pericytes to support endothelial cell function, which is required for maintenance of the microvascular barrier, but saw no endothelial hyperpermeability when NS1-treated endothelial cells were cultured alone or in combination with pericytes. Our findings suggest that a direct effect of NS1 on endothelial dysfunction is required for pericyte-driven amplification of hyperpermeability, and that effects on pericytes alone are insufficient to trigger microvascular leakage. Our work highlights the synergistic effect of microvascular cells during orthoflavivirus haemorrhage and emphasises the need for further work into the mechanisms of vascular dysfunction during AHFV and KFDV infection.

Chikungunya virus (CHIKV) infection is one of the major public health concerns in several countries around the world. CHIKV non-structural protein 2 (nsP2) is a promising drug design target due to the enzymes multifunctional properties that facilities viral replication and propagation. To date, there is an evident lack of preventative and therapeutic developments that can be used against CHIKV. Drug repurposing is a time saving and cost-effective method used for the development of new drugs. In this study, drug repurposing was implemented with the use of HIV/HCV protease inhibitors to inhibit the active site of nsP2. Molecular dynamics simulations and analysis revealed the stability of two drugs, Indinavir and Paritaprevir. Indinavir forms a hydrogen bond with a major residue, which closes the flexible loop, situated in close proximity to the active site. This conformational shift in the orientation of the enzyme prevents accessibility to the active site thus disrupting the nsP2 protein from functioning effectively in viral replication. In conclusion, the findings of this study identified Indinavir was identified as a promising CHIKV nsP2 inhibitor. This study will provide the basis to further facilitate the drug repurposing strategy as an alternative approach for drug design of CHIKV inhibitors as well as other viral families.

Sudan virus (SUDV) is a member of the family Filoviridae, which comprises highly pathogenic viruses associated with unusually high case fatality rates. The development of medical countermeasures against filoviruses, including antivirals, vaccines, and therapeutic antibodies, requires preclinical evaluation in suitable animal models. C57BL/6J IFNAR-/- mice, which lack the type I interferon (IFN-/{beta}) receptor, have been reported to be susceptible to filovirus infections, although their impaired innate immune response may represent a potential limitation of the model. Here, we show that IFNAR-/- mice constitute a suitable model for SUDV infection. Following infection, animals developed a clear clinical disease characterized by significant weight loss and pronounced changes in behaviour and appearance. Mice reached the predefined clinical endpoint 3-5 days post infection. Post mortem analysis of terminal samples revealed high viral loads and viral genome copies in all tested organs as well as in serum, indicating widespread systemic dissemination. Importantly, infection was associated with a marked increase in several key chemokines and cytokines linked to systemic inflammation, consistent with the development of a cytokine storm-like response. Together, these findings demonstrate that SUDV infection in IFNAR-/- mice induces systemic viral dissemination and a pronounced inflammatory response, supporting the suitability of this model for investigating filovirus pathogenesis and infection-associated immune dysregulation.

The yerba mate psyllid (Gyropsylla spegazziniana) poses a significant threat to yerba mate crops, causing extensive economic losses. While some ecological aspects as well as control strategies have been studied, its associated viral diversity remains unexplored. Here, by generating the first RNA high-throughput analysis (HTS) of this pest, we explored the G. spegazziniana virome, revealing novel and diverse RNA viruses. We characterized five new viral members belonging to distinct families, with evolutionary cues of beny-like viruses (Benyviridae), picorna-like viruses (Picornaviridae), and sobemo-like viruses (Solemoviridae); which were tentatively named Gyropsylla spegazziniana beny-like virus 1 (GSBlV1), Gyropsylla spegazziniana picorna-like virus 1 (GSPlV1), and Gyropsylla spegazziniana sobemo-like virus 1-3 (GSSlV1-3), respectively. Phylogenetic analysis of the bi-segmented and highly divergent sobemo-like viruses showed a distinctive evolutionary trajectory of its encoding proteins at the periphery of recently reported invertebrate Sobelivirales. The beny-like virus belonged to a cluster of insect-associated beny-like viruse; while the picorna-like virus clustered together with psyllid-associated picorna-like viruses. These results highlight the existence of a complex virome within G. spegazziniana and establish the basis for future studies investigating the ecological roles, evolutionary dynamics, and potential biocontrol applications of these viruses in the G. spegazziniana -yerba mate eco-systems.