Search Results (19)

Influenza D virus (IDV), an emerging orthomyxovirus with zoonotic potential, infects diverse hosts, causes respiratory disease, and remains poorly characterized in China despite its global expansion. From October 2023 to January 2025, we collected 563 nasal swabs from cattle across 28 farms in Jilin Province, Northeast China, and identified seven IDV-positive samples (1.2%), recovering two viable isolates (JL/YB2024 and JL/CC2024). Full-genome sequencing revealed complete, stable seven-segment genomes with high nucleotide identity (up to 99.9%) to contemporary Chinese D/Yamagata/2019 strains and no evidence of reassortment. Maximum-likelihood and time-resolved Bayesian phylogenies of 231 global hemagglutinin-esterase-fusion (HEF) sequences placed the Jilin isolates within the East Asian D/Yamagata/2019 clade and traced their most recent common ancestor to approximately 2017 (95% highest posterior density: 2016–2018), suggesting a cross-border introduction likely associated with regional cattle movement. No IDV was detected in parallel surveillance of swine, underscoring cattle as the principal reservoir and amplifying host. Bayesian skyline analysis demonstrated a marked decline in global IDV genetic diversity during 2020–2022, coinciding with livestock-movement restrictions imposed during the COVID-19 pandemic. Collectively, these findings indicate that IDV circulation in China is sporadic and geographically localized, dominated by the D/Yamagata/2019 lineage, and shaped by multiple independent incursions rather than a single emergence. Both the incorporation of IDV diagnostics into routine bovine respiratory disease surveillance and cattle-import quarantine programs, and the adoption of a One Health framework to monitor potential human spillover and future viral evolution, were recommend. Full article

Active and passive surveillance, followed by gene sequencing, continue to be used to identify a diverse range of novel bacteria, viruses, and other microorganisms in ticks with the potential to cause disease in vertebrate hosts following tick bite. In this study, we describe the isolation and characterization of a novel virus from Bothriocroton hydrosauri ticks collected from a blotched blue-tongue, Tiliqua nigrolutea. In an attempt to isolate rickettsia, the inoculation of Vero cell cultures with tick extracts led to the isolation of a virus, identified as a novel tick Orthomyxovirus by electron microscopy and gene sequencing. Transmission electron microscopic analysis revealed that B. hydrosauri tick virus-1 (BHTV-1) is a spherical orthomyxovirus, 85 nm in size. Multiple developmental stages of the virus were evident in vitro. Analysis of putative BHTV-1 amino acid sequences derived from a genomic analysis of virus-infected host cell extracts revealed the presence of six putative RNA segments encoding genes, sharing the closest sequence similarity to viral sequences belonging to the arthropod-borne Thogotovirus genus within the Orthomyxoviridae. Thogotoviruses are an emerging cause of disease in humans and animals following tick bite. The detection of this new thogotovirus, BHTV-1, in B. hydrosauri, a competent vector for human tick-borne infectious diseases, warrants follow-up investigation to determine its prevalence, host range, and pathogenic potential. Full article

In summer 2023, during an outbreak of highly pathogenic avian influenza (HPAI) in cats in Poland, a 16-year-old dog was presented to the veterinary clinic with persistent, debilitating, dry cough, submandibular lymphadenomegaly, mild serous nasal discharge, and left apical heart murmur. A preliminary diagnosis of kennel cough was made and the treatment with amoxicillin/clavulanic acid and dexamethasone was initiated. Due to the lack of improvement within 2 days, a blood check-up, thoracic radiography and ultrasonography, and echocardiography were performed. Moreover, a rapid test for orthomyxovirus type A antigen in a throat swab was carried out and proved positive. The result was verified using RT-qPCR, which yielded a positive result for A/H5N1 influenza virus and negative results for A/H1N1, A/H3N2, type B influenza, and SARS-CoV-2. This case indicates that HPAI should be considered as a differential diagnosis not only in cats, but also in dogs with upper respiratory tract disease, particularly in regions experiencing A/H5N1 avian influenza outbreaks. Full article

Influenza D virus (IDV) is a novel orthomyxovirus initially isolated from pigs exhibiting influenza-like disease in the USA. Since then, IDV has been detected worldwide in several host species, including livestock animals, whilst specific antibodies have been identified in humans, raising concerns about interspecies transmission and zoonotic risks. Few data regarding the seroprevalence of IDV in small ruminants have been available to date. In this study, we assessed the prevalence of antibodies against IDV in ovine serum samples in Sicily, Southern Italy. Six hundred serum samples, collected from dairy sheep herds located in Sicily in 2022, were tested by haemagglutination inhibition (HI) and virus neutralization (VN) assays using reference strains, D/660 and D/OK, representative of two distinct IDV lineages circulating in Italy. Out of 600 tested samples, 168 (28.0%) tested positive to either IDV strain D/660 or D/OK or to both by HI whilst 378 (63.0%) tested positive to either IDV strain D/660 or D/OK or to both by VN. Overall, our findings demonstrate that IDV circulates in ovine dairy herds in Sicily. Since IDV seems to have a broad host range and it has zoonotic potential, it is important to collect epidemiological information on susceptible species. Full article

Equine influenza virus (EIV) causes acute respiratory disease in horses and belongs to the influenza A virus family Orthomyxoviridae, genus Orthomyxovirus. This virus may have severe financial implications for the horse industry owing to its highly contagious nature and rapid transmission. In the Republic of Korea, vaccination against EIV has been practiced with the active involvement of the Korea Racing Authority since 1974. In this study, we monitored the viral RNA for EIV using PCR, as well as the antibody levels against ‘A/equine/South Africa/4/03 (H3N8, clade 1)’, from 2020 to 2022. EIV was not detected using RT-PCR. The seropositivity rates detected using a hemagglutination inhibition assay were 90.3% in 2020, 96.7% in 2021, and 91.8% in 2022. The geometric mean of antibody titer (GMT) was 83.4 in 2020, 135.7 in 2021, and 95.6 in 2022. Yearlings and two-year-olds in training exhibited lower positive rates (59.1% in 2020, 38.9% in 2021, and 44.1% in 2022) than the average. These younger horses may require more attention for vaccination and vaccine responses against EIV. Continuous surveillance of EIV should be performed to monitor the prevalence and spread of this disease. Full article

Infectious salmon anaemia virus (ISAV) binds circulating Atlantic salmon erythrocytes, but the relevance of this interaction for the course of infection and development of disease remains unclear. We here characterise ISAV-erythrocyte interactions in experimentally infected Atlantic salmon and show that ISAV-binding to erythrocytes is common and precedes the development of disease. Viral RNA and infective particles were enriched in the cellular fraction of blood. While erythrocyte-associated ISAV remained infectious, erythrocytes dose-dependently limited the infection of cultured cells. Surprisingly, immunostaining of blood smears revealed expression of ISAV proteins in a small fraction of erythrocytes in one of the examined trials, confirming that ISAV can be internalised in this cell type and engage the cellular machinery in transcription and translation. However, viral protein expression in erythrocytes was rare and not required for development of disease and mortality. Furthermore, active transcription of ISAV mRNA was higher in tissues than in blood, supporting the assumption that ISAV replication predominantly takes place in endothelial cells. In conclusion, Atlantic salmon erythrocytes bind ISAV and sequester infective virus particles during infection, but do not appear to significantly contribute to ISAV replication. We discuss the implications of our findings for infection dynamics and pathogenesis of infectious salmon anaemia. Full article

Broad-spectrum antiviral therapies hold promise as a first-line defense against emerging viruses by blunting illness severity and spread until vaccines and virus-specific antivirals are developed. The nucleobase favipiravir, often discussed as a broad-spectrum inhibitor, was not effective in recent clinical trials involving patients infected with Ebola virus or SARS-CoV-2. A drawback of favipiravir use is its rapid clearance before conversion to its active nucleoside-5′-triphosphate form. In this work, we report a synergistic reduction of flavivirus (dengue, Zika), orthomyxovirus (influenza A), and coronavirus (HCoV-OC43 and SARS-CoV-2) replication when the nucleobases favipiravir or T-1105 were combined with the antimetabolite 6-methylmercaptopurine riboside (6MMPr). The 6MMPr/T-1105 combination increased the C-U and G-A mutation frequency compared to treatment with T-1105 or 6MMPr alone. A further analysis revealed that the 6MMPr/T-1105 co-treatment reduced cellular purine nucleotide triphosphate synthesis and increased conversion of the antiviral nucleobase to its nucleoside-5′-monophosphate, -diphosphate, and -triphosphate forms. The 6MMPr co-treatment specifically increased production of the active antiviral form of the nucleobases (but not corresponding nucleosides) while also reducing levels of competing cellular NTPs to produce the synergistic effect. This in-depth work establishes a foundation for development of small molecules as possible co-treatments with nucleobases like favipiravir in response to emerging RNA virus infections. Full article

Host plasma membrane protein SERINC5 is incorporated into budding retrovirus particles where it blocks subsequent entry into susceptible target cells. Three structurally unrelated proteins encoded by diverse retroviruses, human immunodeficiency virus type 1 (HIV-1) Nef, equine infectious anemia virus (EIAV) S2, and ecotropic murine leukemia virus (MLV) GlycoGag, disrupt SERINC5 antiviral activity by redirecting SERINC5 from the site of virion assembly on the plasma membrane to an internal RAB7+ endosomal compartment. Pseudotyping retroviruses with particular glycoproteins, e.g., vesicular stomatitis virus glycoprotein (VSV G), renders the infectivity of particles resistant to inhibition by virion-associated SERINC5. To better understand viral determinants for SERINC5-sensitivity, the effect of SERINC5 was assessed using HIV-1, MLV, and Mason-Pfizer monkey virus (M-PMV) virion cores, pseudotyped with glycoproteins from Arenavirus, Coronavirus, Filovirus, Rhabdovirus, Paramyxovirus, and Orthomyxovirus genera. SERINC5 restricted virions pseudotyped with glycoproteins from several retroviruses, an orthomyxovirus, a rhabdovirus, a paramyxovirus, and an arenavirus. Infectivity of particles pseudotyped with HIV-1, amphotropic-MLV (A-MLV), or influenza A virus (IAV) glycoproteins, was decreased by SERINC5, whether the core was provided by HIV-1, MLV, or M-PMV. In contrast, particles pseudotyped with glycoproteins from M-PMV, parainfluenza virus 5 (PIV5), or rabies virus (RABV) were sensitive to SERINC5, but only with particular retroviral cores. Resistance to SERINC5 did not correlate with reduced SERINC5 incorporation into particles, route of viral entry, or absolute infectivity of the pseudotyped virions. These findings indicate that some non-retroviruses may be sensitive to SERINC5 and that, in addition to the viral glycoprotein, the retroviral core influences sensitivity to SERINC5. Full article

Influenza A Viruses (IAV) in domestic swine (IAV-S) are associated with sporadic zoonotic transmission at the human–animal interface. Previous pandemic IAVs originated from animals, which emphasizes the importance of characterizing human immunity against the increasingly diverse IAV-S. We analyzed serum samples from healthy human donors (n = 153) using hemagglutination-inhibition (HAI) assay to assess existing serologic protection against a panel of contemporary IAV-S isolated from swine in the United States (n = 11). Age-specific seroprotection rates (SPR), which are the proportion of individuals with HAI ≥ 1:40, corresponded with lower or moderate pandemic risk classifications for the multiple IAV-S examined (one H1-δ1, one H1-δ2, three H3-IVA, one H3-IVB, one H3-IVF). Individuals born between 2004 and 2013 had SPRs of 0% for the five classified H3 subtype IAV-S, indicating youth may be particularly predisposed to infection with these viruses. Expansion of existing immunologic gaps over time could increase likelihood of future IAV-S spillover to humans and facilitate subsequent sustained human-to-human transmission resulting in disease outbreaks with pandemic potential. Full article

Pilchard orthomyxovirus (POMV) is an emerging pathogen of concern to the salmon industry in Australia. To explore the molecular events that underpin POMV infection, we challenged Atlantic salmon (Salmo salar) post-smolts in seawater via cohabitation. Tissue samples of the head kidney and liver were collected from moribund and surviving individuals and analyzed using transcriptome sequencing. Viral loads were higher in the head kidney compared to the liver, yet the liver presented more upregulated genes. Fish infected with POMV showed a strong innate immune response that included the upregulation of pathogen recognition receptors such as RIG-I and Toll-like receptors as well as the induction of interferon-stimulated genes (MX, ISG15). Moribund fish also presented a dramatic induction of pro-inflammatory cytokines, contributing to severe tissue damage and morbidity. An induction of major histocompatibility complex (MHC) class I genes (B2M) and markers of T cell-mediated immunity (CD8-alpha, CD8-beta, Perforin-1, Granzyme-A) was observed in both moribund fish and survivors. In addition, differential connectivity analysis showed that three key regulators (RELA/p65, PRDM1, and HLF) related to cell-mediated immunity had significant differences in connectivity in “clinically healthy” versus “clinically affected” or moribund fish. Collectively, our results show that T cell-mediated immunity plays a central role in the response of Atlantic salmon to the infection with POMV. Full article

Emerging influenza D viruses (IDVs), the newest member in the genus Orthomyxovirus family, which can infect and transmit in multiple mammalian species as its relatives the influenza A viruses (IAVs). Additional studies of biological characteristics of IDVs are needed; here, we studied the characteristics of IDV nonstructural protein 2 (NS2), which shares the lowest homology to known influenza proteins. First, we generated reassortant viruses via reverse genetics to analyze the segment compatibility and gene interchangeability between IAVs and IDVs. Next, we investigated the locations and exact sequences of nuclear export signals (NESs) of the IDV NS2 protein. Surprisingly, three separate NES regions were found to contribute to the nuclear export of an eGFP fusion protein. Alanine scanning mutagenesis identified critical amino acid residues within each NES, and co-immunoprecipitation experiments demonstrated that their nuclear export activities depend on the CRM1-mediated pathway, particularly for the third NES (136-146aa) of IDV NS2. Interestingly, the third NES was important for the interaction of NS2 protein with CRM1. The findings in this study contribute to the understanding of IDV NS2 protein’s role during nucleocytoplasmic transport of influenza viral ribonucleoprotein complexes (vRNPs) and will also facilitate the development of novel anti-influenza drugs targeting nuclear export signals of IDV NS2 protein. Full article

Although seasonal influenza vaccines block most predominant influenza types and subtypes, humans still remain vulnerable to waves of seasonal and new potential pandemic influenza viruses for which no immunity may exist because of viral antigenic drift and/or shift. Previously, we described a human monoclonal antibody (hMAb), KPF1, which was produced in human embryonic kidney 293T cells (KPF1-HEK) with broad and potent neutralizing activity against H1N1 influenza A viruses (IAV) in vitro, and prophylactic and therapeutic activities in vivo. In this study, we produced hMAb KPF1 in tobacco plants (KPF1-Antx) and demonstrated how the plant-produced KPF1-Antx hMAb possesses similar biological activity compared with the mammalian-produced KPF1-HEK hMAb. KPF1-Antx hMAb showed broad binding to recombinant HA proteins and H1N1 IAV, including A/California/04/2009 (pH1N1) in vitro, which was comparable to that observed with KPF1-HEK hMAb. Importantly, prophylactic administration of KPF1-Antx hMAb to guinea pigs prevented pH1N1 infection and transmission in both prophylactic and therapeutic experiments, substantiating its clinical potential to prevent and treat H1N1 infections. Collectively, this study demonstrated, for the first time, a plant-produced influenza hMAb with in vitro and in vivo activity against influenza virus. Because of the many advantages of plant-produced hMAbs, such as rapid batch production, low cost, and the absence of mammalian cell products, they represent an alternative strategy for the production of immunotherapeutics for the treatment of influenza viral infections, including emerging seasonal and/or pandemic strains. Full article

Tilapia lake virus (TiLV) is a negative-sense single-stranded RNA (-ssRNA) icosahedral virus classified to be the only member in the family Amnoonviridae. Although TiLV segment-1 shares homology with the influenza C virus PB1 and has four conserved motifs similar to influenza A, B, and C polymerases, it is unknown whether there are other properties shared between TiLV and orthomyxovirus. In the present study, we wanted to determine whether TiLV agglutinated avian and piscine erythrocytes, and whether its replication was inhibited by lysosomotropic agents, such as ammonium chloride (NH₄Cl), as seen for orthomyxoviruses. Our findings showed that influenza virus strain A/Puerto Rico/8 (PR8) was able to hemagglutinate turkey (Meleagris gallopavo), Atlantic salmon (Salmo salar L), and Nile tilapia (Oreochromis niloticus) red blood cells (RBCs), while infectious salmon anemia virus (ISAV) only agglutinated Atlantic salmon, but not turkey or tilapia, RBCs. In contrast to PR8 and ISAV, TiLV did not agglutinate turkey, Atlantic salmon, or tilapia RBCs. qRT-PCR analysis showed that 30 mM NH₄Cl, a basic lysosomotropic agent, neither inhibited nor enhanced TiLV replication in E-11 cells. There was no difference in viral quantities in the infected cells with or without NH₄Cl treatment during virus adsorption or at 1, 2, and 3 h post-infection. Given that hemagglutinin proteins that bind RBCs also serve as ligands that bind host cells during virus entry leading to endocytosis in orthomyxoviruses, the data presented here suggest that TiLV may use mechanisms that are different from orthomyxoviruses for entry and replication in host cells. Therefore, future studies should seek to elucidate the mechanisms used by TiLV for entry into host cells and to determine its mode of replication in infected cells. Full article

Varroa destructor is an ectoparasitic mite of Asian or Eastern honeybees Apis cerana (A. cerana) which has become a serious threat to European subspecies of Western honeybees Apis mellifera (A. mellifera) within the last century. V. destructor and its vectored honeybee viruses became serious threats for colony survival. This is a short period for pathogen- and host-populations to adapt. To look for possible variation in the composition of viral populations we performed RNA metagenomic analysis of the Western honeybee subspecies A. m. ligustica, A. m. syriaca, A. m. intermissa, and A. cerana and their respective V. destructor mites. The analysis revealed two novel viruses: Varroa orthomyxovirus-1 (VOV-1) in A. mellifera and V. destructor and a Hubei like-virga virus-14 homolog in V. destructor. VOV-1 was more prevalent in V. destructor than in A. mellifera and we found evidence for viral replication in both hosts. Interestingly, we found differences in viral loads of A. cerana and their V. destructor, A. m. intermissa, and its V. destructor showed partial similarity, while A. m. ligustica and A. m. syriaca and their varroa where very similar. Deformed wing virus exhibited 82.20%, 99.20%, 97.90%, and 0.76% of total viral reads in A. m. ligustica, A. m. syriaca, A. m. intermissa, and A. cerana, respectively. This is the first report of a complete segmented-single-stranded negative-sense RNA virus genome in honeybees and V. destructor mites. Full article

A crucial step in the molecular detection of viruses in clinical specimens is the efficient extraction of viral nucleic acids. The total yield of viral nucleic acid from a clinical specimen is dependent on the specimen's volume, the initial virus concentration and the effectiveness provided by the extraction method. Recent next generation sequencing (NGS)-based diagnostic approaches (i.e. metagenomics) provide a molecular 'open view' into the sample, as they theoretically generate sequence reads of any nucleic acid present in a specimen in a statistically representative manner. However, since a higher virus-related read output promises better sensitivity in the subsequent bioinformatic analysis, the extraction method selected determines the reliability of diagnostic NGS. In this study nine commercially available kits for nucleic acid extraction were compared regarding the simultaneous isolation of DNA and RNA by real-time PCR, four of which were selected for subsequent comparison by NGS (QIAamp Viral RNA Mini Kit, QIAamp DNA Blood Mini Kit, QIAamp cador Pathogen Mini Kit and QIAamp MinElute Virus Spin Kit). The nucleic acid yields and the sequence read output were compared for four different model viruses–reovirus, orthomyxovirus, orthopoxvirus and paramyxovirus–each at defined but varying concentrations in the same sample. The total amount of nucleic acid was processed to sequence the RNA (as cDNA) and the DNA with quantification by Qubit and virus-specific quantitative real-time PCRs. NGS libraries were prepared for sequencing on the Illumina HiSeq 1500 system. Finally, the percentage of reads assignable to each virus was determined via mapping. Evaluation of different commercial nucleic acid extraction kits with four different viruses indicates little variation in the read numbers obtained for transcribed RNA or DNA by NGS. Since NGS is increasingly being used as a tool in diagnostics of infectious diseases, the individual steps of the complete process have to be validated carefully. Here we could show that for virus identification in liquid clinical specimens, any nucleic acid extraction kit that is performing well for PCR diagnostics can be used for NGS diagnostics as well and that the selection of the kit has only a minor impact on the yield of viral reads. Full article