Influenza C and D Viruses Demonstrated a Differential Respiratory Tissue Tropism in a Comparative Pathogenesis Study in Guinea Pigs.

Influenza C virus (ICV) is increasingly associated with community-acquired pneumonia (CAP) in children and its disease severity is worse than the influenza B virus, but similar to influenza A virus associated CAP. Despite the ubiquitous infection landscape of ICV in humans, little is known about its replication and pathobiology in animals. The goal of this study was to understand the replication kinetics, tissue tropism, and pathogenesis of human ICV (huICV) in comparison to the swine influenza D virus (swIDV) in guinea pigs. Intranasal inoculation of both viruses did not cause clinical signs, however, the infected animals shed virus in nasal washes. The huICV replicated in the nasal turbinates, soft palate, and trachea but not in the lungs while swIDV replicated in all four tissues. A comparative analysis of tropism and pathogenesis of these two related seven-segmented influenza viruses revealed that swIDV-infected animals exhibited broad tissue tropism with an increased rate of shedding on 3, 5, and 7 dpi and high viral loads in the lungs compared to huICV. Seroconversion occurred late in the huICV group at 14 dpi, while swIDV-infected animals seroconverted at 7 dpi. Guinea pigs infected with huICV exhibited mild to moderate inflammatory changes in the epithelium of the soft palate and trachea, along with mucosal damage and multifocal alveolitis in the lungs. In summary, the replication kinetics and pathobiological characteristics of ICV in guinea pigs agree with the clinical manifestation of ICV infection in humans, and hence guinea pigs could be used to study these distantly related influenza viruses. IMPORTANCE Similar to influenza A and B, ICV infections are seen associated with bacterial and viral co-infections which complicates the assessment of its real clinical significance. Further, the antivirals against influenza A and B viruses are ineffective against ICV which mandates the need to study the pathobiological aspects of this virus. Here we demonstrated that the respiratory tract of guinea pigs possesses specific viral receptors for ICV. We also compared the replication kinetics and pathogenesis of huICV and swIDV, as these viruses share 50% sequence identity. The tissue tropism and pathology associated with huICV in guinea pigs are analogous to the mild respiratory disease caused by ICV in humans, thereby demonstrating the suitability of guinea pigs to study ICV. Our comparative analysis revealed that huICV and swIDV replicated differentially in the guinea pigs suggesting that the type-specific genetic differences can result in the disparity of the viral shedding and tissue tropism.

A fully human monoclonal antibody possesses antibody-dependent cellular cytotoxicity (ADCC) activity against the H1 subtype of influenza A virus by targeting a conserved epitope at the HA1 protomer interface.

The DiversitabTM system produces target specific high titer fully human polyclonal IgG immunoglobulins from transchromosomic (Tc) bovines shown to be safe and effective against multiple virulent pathogens in animal studies and Phase 1, 2 and 3 human clinical trials. We describe the functional properties of a human monoclonal antibody (mAb), 38C2, identified from this platform, which recognizes recombinant H1 hemagglutinins (HAs) and induces appreciable antibody-dependent cellular cytotoxicity (ADCC) activity in vitro. Interestingly, 38C2 monoclonal antibody demonstrated no detectable neutralizing activity against H1N1 virus in both hemagglutination inhibition and virus neutralization assays. Nevertheless, this human monoclonal antibody induced appreciable ADCC against cells infected with multiple H1N1 strains. The HA-binding activity of 38C2 was also demonstrated in flow cytometry using Madin-Darby canine kidney cells infected with multiple influenza A H1N1 viruses. Through further investigation with the enzyme-linked immunosorbent assay involving the HA peptide array and 3-dimensional structural modeling, we demonstrated that 38C2 appears to target a conserved epitope located at the HA1 protomer interface of H1N1 influenza viruses. A novel mode of HA-binding and in vitro ADCC activity pave the way for further evaluation of 38C2 as a potential therapeutic agent to treat influenza virus infections in humans.

Spillover of Canine Parvovirus Type 2 to Pigs, South Dakota, USA, 2020.

In 1978, canine parvovirus type 2 originated from spillover of a feline panleukopenia-like virus, causing a worldwide pandemic of enteritis and myocarditis among canids. In 2020, the virus was identified in pigs in South Dakota, USA, by PCR, sequencing, in situ hybridization, and serology. Genetic analysis suggests spillover from wildlife.

Bovine rhinitis B virus is highly prevalent in acute bovine respiratory disease and causes upper respiratory tract infection in calves.

Bovine respiratory disease (BRD) is the most significant cause of cattle morbidity and mortality worldwide. This multifactorial disease has a complex aetiology. Dogma posits a primary viral infection followed by secondary bacterial pneumonia. Bovine rhinitis B virus (BRBV) is an established aetiological agent of BRD, but little is known regarding its pathogenesis. Here, a BRD PCR panel identified 18/153 (11.8 %) lung samples and 20/49 (40.8 %) nasal swabs collected from cattle with respiratory signs as positive for BRBV, which was the most prevalent virus in nasal swabs. Primary bovine tracheal epithelial cells were used to isolate BRBV that was phylogenetically related to contemporary sequences from the USA and Mexico and genetically divergent from the previous sole BRBV isolate. To investigate virus pathogenesis, 1-week-old colostrum-deprived dairy calves were inoculated intranasally with 7.0 log10 TCID50 BRBV. Virus was isolated from nasal swabs, nasal turbinates, trachea and the brain of the challenged animals. Neutralizing antibodies were detected beginning 7 days post-inoculation and peaked at day 14. In situ hybridization (ISH) localized BRBV infection in the upper respiratory ciliated epithelial and goblet cells, occasionally associated with small defects of the superficial cilia lining. Sporadically, pinpoint ISH signals were also detected in cells resembling glial cells in the cerebrum in one calf. Together, these results demonstrate the BRBV infection is highly prevalent in acute BRD samples and while the pathogenicity of BRBV is minimal with infection largely limited to the upper respiratory tract, further research is needed to elucidate a possible initiatory role in BRD.

Novel hepatoviruses in synanthropic bats in the upper Midwestern United States.

A nearly complete genome sequence of hepatovirus G was isolated from an Eptesicus fuscus bat submitted for rabies virus testing due to human exposure in South Dakota. The predicted polyprotein sequence was 78.2% and 74.4% identical to genotypes G1 and G2, respectively, recovered from bats in Ghana. Quantitative PCR on 90 E. fuscus bats showed that eight (8.9%) were positive for hepatovirus G. Targeted sequencing of the VP2 region of the genome for five positive samples showed >99% identity to hepatovirus G strain Ef15893, demonstrating that hepatovirus G commonly circulates in E. fuscus bats in the upper Midwest.

Identification of a novel statovirus in a faecal sample from a calf with enteric disease.

A novel clade of RNA viruses was identified in the mammalian gastrointestinal tract by next-generation sequencing. Phylogenetically, these viruses are related to the genera Tombusviridae (plant viruses) and Flaviviridae, which includes mammalian, avian and insect hosts. Named in line with their characterization as stool-associated Tombus-like viruses, it is unclear if statoviruses infect mammals or are dietary in origin. Here, metagenomic sequencing of faecal material collected from a 10-week-old calf with enteric disease found that 20 % of the reads mapped to a de novo-assembled 4 kb contig with homology to statoviruses. Phylogenetic analysis of the statovirus genome found a clear evolutionary relationship with statovirus A, but, with only 47 % similarity, we propose that the statovirus sequence presents a novel species, statovirus F. A TaqMan PCR targeting statovirus F performed on faecal material found a cycle threshold of 11, suggesting a high titre of virus shed from the calf with enteric disease. A collection of 48 samples from bovine enteric disease diagnostic submissions were assayed by PCR to investigate statovirus F prevalence and 6 of 48 (12.5 %) were positive. An ELISA to detect antibodies to the coat protein found that antibodies to statovirus F were almost ubiquitous in bovine serum. Combined, the PCR and ELISA results suggest that statovirus F commonly infects cattle. Further research is needed to elucidate the aetiological significance of statovirus infection.

Human Monoclonal Antibody Derived from Transchromosomic Cattle Neutralizes Multiple H1 Clades of Influenza A Virus by Recognizing a Novel Conformational Epitope in the Hemagglutinin Head Domain.

Influenza remains a global health risk and challenge. Currently, neuraminidase (NA) inhibitors are extensively used to treat influenza, but their efficacy is compromised by the emergence of drug-resistant variants. Neutralizing antibodies targeting influenza A virus surface glycoproteins are critical components of influenza therapeutic agents and may provide alternative strategies to the existing countermeasures. However, the major hurdle for the extensive application of antibody therapies lies in the difficulty of generating nonimmunogenic antibodies in large quantities rapidly. Here, we report that one human monoclonal antibody (MAb), 53C10, isolated from transchromosomic (Tc) cattle exhibits potent neutralization and hemagglutination inhibition titers against different clades of H1N1 subtype influenza A viruses. In vitro selection of antibody escape mutants revealed that 53C10 recognizes a novel noncontinuous epitope in the hemagglutinin (HA) head domain involving three amino acid residues, glycine (G), serine (S), and glutamic acid (E) at positions 172, 207, and 212, respectively. The results of our experiments supported a critical role for substitution of arginine at position 207 (S207R) in mediating resistance to 53C10, while substitutions at either G172E or E212A did not alter antibody recognition and neutralization. The E212A mutation may provide structural stability for the epitope, while the substitution G172E probably compensates for loss of fitness introduced by S207R. Our results offer novel insights into the mechanism of action of MAb 53C10 and indicate its potential role in therapeutic treatment of H1 influenza virus infection in humans.IMPORTANCE Respiratory diseases caused by influenza viruses still pose a serious concern to global health, and neutralizing antibodies constitute a promising area of antiviral therapeutics. However, the potential application of antibodies is often hampered by the challenge in generating nonimmunogenic antibodies in large scale. In the present study, transchromosomic (Tc) cattle were used for the generation of nonimmunogenic monoclonal antibodies (MAbs), and characterization of such MAbs revealed one monoclonal antibody, 53C10, exhibiting a potent neutralization activity against H1N1 influenza viruses. Further characterization of the neutralization escape mutant generated using this MAb showed that three amino acid substitutions in the HA head domain contributed to the resistance. These findings emphasize the importance of Tc cattle in the production of nonimmunogenic MAbs and highlight the potential of MAb 53C10 in the therapeutic application against H1 influenza virus infection in humans.

Identification of boosepivirus B in U.S. calves.

Bovine enteric disease has a complex etiology that can include viral, bacterial, and parasitic pathogens and is a significant source of losses due to morbidity and mortality. Boosepivirus was identified in calves with enteric disease with unclear etiology in Japan in 2009 and has not been reported elsewhere. Metagenomic sequencing and PCR here identified boosepivirus in bovine enteric disease diagnostic submissions from six states in the USA with 98% sequence identity to members of the species Boosepivirus B. In all cases, boosepivirus was identified as a coinfection with the established pathogens bovine coronavirus, bovine rotavirus, and cryptosporidia. Further research is needed to determine the clinical significance of boosepivirus infection.

Genetic and antigenic characteristics of a human influenza C virus clinical isolate.

Unlike influenza A and B viruses that infect humans and cause severe diseases in seasonal epidemics, influenza C virus (ICV) is a ubiquitous childhood pathogen typically causing mild respiratory symptoms. ICV infections are rarely diagnosed and less research has been performed on it despite the virus being capable of causing severe disease in infants. Here we report on the isolation of a human ICV from a child with acute respiratory disease, provisionally designated C/Victoria/2/2012 (C/Vic). The full-length genome sequence and phylogenetic analysis revealed that the hemagglutinin-esterase-fusion (HEF) gene of C/Vic was derived from C/Sao Paulo lineage, while its PB2 and P3 genes evolved separately from all characterized historical ICV isolates. Furthermore, antigenic analysis using the hemagglutination inhibition (HI) assay found that 1947 C/Taylor virus (C/Taylor lineage) was antigenically more divergent from1966 C/Johannesburg (C/Aichi lineage) than from 2012 C/Vic. Structure modeling of the HEF protein identified two mutations in the 170-loop of the HEF protein around the receptor-binding pocket as a possible antigenic determinant responsible for the discrepant HI results. Taken together, results of our studies reveal novel insights into the genetic and antigenic evolution of ICV and provide a framework for further investigation of its molecular determinants of antigenic property and replication.

A Novel Porcine Circovirus Distantly Related to Known Circoviruses Is Associated with Porcine Dermatitis and Nephropathy Syndrome and Reproductive Failure.

Porcine circovirus-associated disease (PCVAD) is clinically manifested by postweaning multisystemic wasting syndrome (PMWS), respiratory and enteric disease, reproductive failure, and porcine dermatitis and nephropathy syndrome (PDNS). Porcine circovirus 2 (PCV2) is an essential component of PCVAD, although an etiologic role in PDNS is not well established. Here, a novel circovirus, designated porcine circovirus 3 (PCV3), was identified in sows that died acutely with PDNS-like clinical signs. The capsid and replicase proteins of PCV3 are only 37% and 55% identical to PCV2 and bat circoviruses, respectively. Aborted fetuses from sows with PDNS contained high levels of PCV3 (7.57 × 107 genome copies/ml), and no other viruses were detected by PCR and metagenomic sequencing. Immunohistochemistry (IHC) analysis of sow tissue samples identified PCV3 antigen in skin, kidney, lung, and lymph node samples localized in typical PDNS lesions, including necrotizing vasculitis, glomerulonephritis, granulomatous lymphadenitis, and bronchointerstitial pneumonia. Further study of archived PDNS tissue samples that were negative for PCV2 by IHC analysis identified 45 of 48 that were PCV3 positive by quantitative PCR (qPCR), with 60% of a subset also testing positive for PCV3 by IHC analysis. Analysis by qPCR of 271 porcine respiratory disease diagnostic submission samples identified 34 PCV3-positive cases (12.5%), and enzyme-linked immunosorbent assay detection of anti-PCV3 capsid antibodies in serum samples found that 46 (55%) of 83 samples tested were positive. These results suggest that PCV3 commonly circulates within U.S. swine and may play an etiologic role in reproductive failure and PDNS. Because of the high economic impact of PCV2, this novel circovirus warrants further studies to elucidate its significance and role in PCVAD. IMPORTANCE: While porcine circovirus 2 (PCV2) was first identified in sporadic cases of postweaning multisystemic wasting syndrome in Canada in the early 1990s, an epidemic of severe systemic disease due to PCV2 spread worldwide in the ensuing decade. Despite being effectively controlled by commercial vaccines, PCV2 remains one of the most economically significant viruses of swine. Here, a novel porcine circovirus (PCV3) that is distantly related to known circoviruses was identified in sows with porcine dermatitis and nephropathy syndrome (PDNS) and reproductive failure. PCV2, which has previously been associated with these clinical presentations, was not identified. High levels of PCV3 nucleic acid were observed in aborted fetuses by quantitative PCR, and PCV3 antigen was localized in histologic lesions typical of PDNS in sows by immunohistochemistry (IHC) analysis. PCV3 was also identified in archival PDNS diagnostic samples that previously tested negative for PCV2 by IHC analysis. The emergence of PCV3 warrants further investigation.

Comparative epidemiology of porcine circovirus type 3 in pigs with different clinical presentations.

BACKGROUND: Porcine circovirus type 3 (PCV3), as an emerging circovirus species, was reported to be widely circulating in the United States, China, South Korea and Poland. Previous studies revealed that PCV3 was mainly concentrated in sick animals with respiratory disease, skin disease, reproductive disorders and so on. However, the circulating status of PCV3 in pigs with other clinical presentations (especilly asymptomatic or diarrhea) was not well established. FINDINGS: In this study, to conduct a comparative epidemiological survey of PCV3, 80 weaned pig serum samples with severe respiratory disease (SRD), 175 weaned pig serum samples with mild respiratory disease (MRD), 216 asymptomatic weaned pig serum samples, 35 diarrheal weaned pig samples and 35 non-diarrheal weaned pig samples were collected from eight provinces of China. Via qPCR testing, PCV3 was circulating in all sampling provinces, with total positive rates varying from 1.04% to 100%. Interestingly, the PCV3-positive rate was significantly higher in weaned pigs with SRD (63.75%, 51/80) than in those weaned pigs with MRD (13.14%, 23/175) and asymptomatic pigs (1.85%, 4/216) (P < 0.01). Similarly, the PCV3-positive rate was significantly higher in diarrheal weaned pigs (17.14%, 6/35) than in non-diarrheal weaned pigs (2.86%, 1/35) (P < 0.05). Moreover, the lower Ct values of qPCR were frequently found in those weaned pigs or fattening pigs with respiratory disease and diarrhea rather than that in asymptomatic pigs. Sequence analysis showed that low genetic diversity existed among those PCV3 sequences collected from pigs with different clinical presentations. CONCLUSIONS: The present study further extends evidence that newly described PCV3 widely circulates in six additional provinces of Southern and Northern China and has high similarity to previously reported isolates. As an emerging virus of swine, although the present case-control study reveals that PCV3 has a potential association with swine respiratory disease and diarrhea, further investigations into the pathogenesis are needed to ascertain the role of PCV3 in swine health.

Highly diverse posaviruses in swine faeces are aquatic in origin.

Posaviruses are a group of highly divergent viruses identified in swine faeces that are distantly related to other members of the order Picornavirales. Eighteen posavirus genomes were assembled from 10 out of 25 (40 %) faecal-swab pools collected from healthy adult swine. Phylogenetic analysis of the conserved RNA-dependent RNA polymerase (Pol) domain found that posaviruses form a large, highly diverse, monophyletic clade, which includes similar viruses identified in human (husavirus) and fish (fisavirus) faeces or intestinal contents, respectively. Quantitative reverse transcription PCR analysis of water samples collected from commercial swine barns identified four out of 19 (21 %) samples were positive using a 5'-nuclease assay targeting the Pol region of posavirus 1. ICPD (immunoprecipitation coupled to PCR detection) assays to explore serological evidence of posavirus infection found only a single positive sample, suggesting posaviruses do not commonly infect swine, and together these results suggests a likely aquatic host.

Feral swine virome is dominated by single-stranded DNA viruses and contains a novel Orthopneumovirus which circulates both in feral and domestic swine.

Feral swine are known reservoirs for various pathogens that can adversely affect domestic animals. To assess the viral ecology of feral swine in the USA, metagenomic sequencing was performed on 100 pooled nasal swabs. The virome was dominated by small, ssDNA viruses belonging to the families Circoviridae, Anelloviridae and Parvovirinae. Only four RNA viruses were identified: porcine kobuvirus, porcine sapelovirus, atypical porcine pestivirus and a novel Orthopneumovirus, provisionally named swine orthopneumovirus (SOV). SOV shared ~90 % nucleotide identity to murine pneumonia virus (MPV) and canine pneumovirus. A modified, commercially available ELISA for MPV found that approximately 30 % of both feral and domestic swine sera were positive for antibodies cross-reactive with MPV. Quantitative reverse transcription-PCR identified two (2 %) and four (5.0 %) positive nasal swab pools from feral and domestic swine, respectively, confirming that SOV circulates in both herds.

Metagenomic characterization of the virome associated with bovine respiratory disease in feedlot cattle identified novel viruses and suggests an etiologic role for influenza D virus.

Bovine respiratory disease (BRD) is the most costly disease affecting the cattle industry. The pathogenesis of BRD is complex and includes contributions from microbial pathogens as well as host, environmental and animal management factors. In this study, we utilized viral metagenomic sequencing to explore the virome of nasal swab samples obtained from feedlot cattle with acute BRD and asymptomatic pen-mates at six and four feedlots in Mexico and the USA, respectively, in April-October 2015. Twenty-one viruses were detected, with bovine rhinitis A (52.7 %) and B (23.7 %) virus, and bovine coronavirus (24.7 %) being the most commonly identified. The emerging influenza D virus (IDV) tended to be significantly associated (P=0.134; odds ratio=2.94) with disease, whereas viruses commonly associated with BRD such as bovine viral diarrhea virus, bovine herpesvirus 1, bovine respiratory syncytial virus and bovine parainfluenza 3 virus were detected less frequently. The detection of IDV was further confirmed using a real-time PCR assay. Nasal swabs from symptomatic animals had significantly more IDV RNA than those collected from healthy animals (P=0.04). In addition to known viruses, new genotypes of bovine rhinitis B virus and enterovirus E were identified and a newly proposed species of bocaparvovirus, Ungulate bocaparvovirus 6, was characterized. Ungulate tetraparvovirus 1 was also detected for the first time in North America to our knowledge. These results illustrate the complexity of the virome associated with BRD and highlight the need for further research into the contribution of other viruses to BRD pathogenesis.

Widespread detection and characterization of porcine parainfluenza virus 1 in pigs in the USA.

Porcine parainfluenza virus 1 (PPIV1) was first identified in 2013 in slaughterhouse pigs in Hong Kong, China. Here, two near-complete genomes were assembled from swine exhibiting acute respiratory disease that were 90.0-95.3% identical to Chinese PPIV1. Analysis of the HN gene from ten additional PPIV1-positive samples found 85.0-95.5% identity, suggesting genetic diversity between strains. Molecular analysis identified 17 out of 279 (6.1%) positive samples from pigs with respiratory disease. Eleven nursery pigs from a naturally infected herd were asymptomatic; however, nasal swabs from six pigs and the lungs of a single pig were quantitative reverse transcriptase (qRT)-PCR positive. Histopathology identified PPIV1 RNA in the nasal respiratory epithelium and trachea. Two serological assays demonstrated seroconversion of infected pigs and further analysis of 59 swine serum samples found 52.5% and 66.1% seropositivity, respectively. Taken together, the results confirm the widespread presence of PPIV1 in the US swine herd.

Cocirculation of two distinct genetic and antigenic lineages of proposed influenza D virus in cattle.

UNLABELLED: Viruses with approximately 50% homology to human influenza C virus (ICV) have recently been isolated from swine and cattle. The overall low homology to ICV, lack of antibody cross-reactivity to ICV in hemagglutination inhibition (HI) and agar gel immunodiffusion assays, and inability to productively reassort with ICV led to the proposal that these viruses represented a new genus of influenza virus, influenzavirus D (IDV). To further our understanding of the epidemiology of IDV, real-time reverse transcription-PCR was performed on a set of 208 samples from bovines with respiratory disease. Ten samples (4.8%) were positive and six viruses were successfully isolated in vitro. Phylogenetic analysis of full-genome sequences of these six new viruses and four previously reported viruses revealed two distinct cocirculating lineages represented by D/swine/Oklahoma/1334/2011 (D/OK) and D/bovine/Oklahoma/660/2013 (D/660), which frequently reassorted with one another. Antigenic analysis using the HI assay and lineage-representative D/OK and D/660 antiserum found up to an approximate 10-fold loss in cross-reactivity against heterologous clade antiserum. One isolate, D/bovine/Texas/3-13/2011 (D/3-13), clustered with the D/660 lineage, but also had high HI titers to heterologous (D/OK) clade antiserum. Molecular modeling of the hemagglutinin esterase fusion protein of D/3-13 identified a mutation at position 212 as a possible antigenic determinant responsible for the discrepant HI results. These results suggest that IDV is common in bovines with respiratory disease and that at least two genetic and antigenically distinct clades cocirculate. IMPORTANCE: A novel bovine influenza virus was recently identified. Detailed genetic and antigenic studies led to the proposal that this virus represents a new genus of influenza, influenzavirus D (IDV). Here, we show that IDV is common in clinical samples of bovine respiratory disease complex (BRDC), with a prevalence similar to that of other established BRDC etiological agents. These results are in good agreement with the near-ubiquitous seroprevalence of IDV previously found. Phylogenetic analysis of complete genome sequences found evidence for two distinct cocirculating lineages of IDV which freely reassort. Significant antigenic differences, which generally agreed with the surface glycoprotein hemagglutinin esterase phylogeny, were observed between the two lineages. Based on these results, and on the ability of IDV to infect and transmit in multiple mammalian species, additional studies to determine the pathogenic potential of IDV are warranted.

Replication and Transmission of the Novel Bovine Influenza D Virus in a Guinea Pig Model.

UNLABELLED: Influenza D virus (FLUDV) is a novel influenza virus that infects cattle and swine. The goal of this study was to investigate the replication and transmission of bovine FLUDV in guinea pigs. Following direct intranasal inoculation of animals, the virus was detected in nasal washes of infected animals during the first 7 days postinfection. High viral titers were obtained from nasal turbinates and lung tissues of directly inoculated animals. Further, bovine FLUDV was able to transmit from the infected guinea pigs to sentinel animals by means of contact and not by aerosol dissemination under the experimental conditions tested in this study. Despite exhibiting no clinical signs, infected guinea pigs developed seroconversion and the viral antigen was detected in lungs of animals by immunohistochemistry. The observation that bovine FLUDV replicated in the respiratory tract of guinea pigs was similar to observations described previously in studies of gnotobiotic calves and pigs experimentally infected with bovine FLUDV but different from those described previously in experimental infections in ferrets and swine with a swine FLUDV, which supported virus replication only in the upper respiratory tract and not in the lower respiratory tract, including lung. Our study established that guinea pigs could be used as an animal model for studying this newly emerging influenza virus. IMPORTANCE: Influenza D virus (FLUDV) is a novel emerging pathogen with bovine as its primary host. The epidemiology and pathogenicity of the virus are not yet known. FLUDV also spreads to swine, and the presence of FLUDV-specific antibodies in humans could indicate that there is a potential for zoonosis. Our results showed that bovine FLUDV replicated in the nasal turbinate and lungs of guinea pigs at high titers and was also able to transmit from an infected animal to sentinel animals by contact. The fact that bovine FLUDV replicated productively in both the upper and lower respiratory tracts of guinea pigs, similarly to virus infection in its native host, demonstrates that guinea pigs would be a suitable model host to study the replication and transmission potential of bovine FLUDV.

Domestic pigs are susceptible to infection with influenza B viruses.

UNLABELLED: Influenza B virus (IBV) causes seasonal epidemics in humans. Although IBV has been isolated from seals, humans are considered the primary host and reservoir of this important pathogen. It is unclear whether other animal species can support the replication of IBV and serve as a reservoir. Swine are naturally infected with both influenza A and C viruses. To determine the susceptibility of pigs to IBV infection, we conducted a serological survey for U.S. Midwest domestic swine herds from 2010 to 2012. Results of this study showed that antibodies to IBVs were detected in 38.5% (20/52) of sampled farms, and 7.3% (41/560) of tested swine serum samples were positive for IBV antibodies. Furthermore, swine herds infected with porcine reproductive and respiratory syndrome virus (PRRSV) showed a higher prevalence of IBV antibodies in our 2014 survey. In addition, IBV was detected in 3 nasal swabs collected from PRRSV-seropositive pigs by real-time RT-PCR and sequencing. Finally, an experimental infection in pigs, via intranasal and intratracheal routes, was performed using one representative virus from each of the two genetically and antigenically distinct lineages of IBVs: B/Brisbane/60/2008 (Victoria lineage) and B/Yamagata/16/1988 (Yamagata lineage). Pigs developed influenza-like symptoms and lung lesions, and they seroconverted after virus inoculation. Pigs infected with B/Brisbane/60/2008 virus successfully transmitted the virus to sentinel animals. Taken together, our data demonstrate that pigs are susceptible to IBV infection; therefore, they warrant further surveillance and investigation of swine as a potential host for human IBV. IMPORTANCE: IBV is an important human pathogen, but its ability to infect other species, for example, pigs, is not well understood. We showed serological evidence that antibodies to two genetically and antigenically distinct lineages of IBVs were present among domestic pigs, especially in swine herds previously infected with PRRSV, an immunosuppressive virus. IBV was detected in 3 nasal swabs from PRRSV-seropositive pigs by real-time reverse transcription-PCR and sequencing. Moreover, both lineages of IBV were able to infect pigs under experimental conditions, with transmissibility of influenza B/Victoria lineage virus among pigs being observed. Our results demonstrate that pigs are susceptible to IBV infections, indicating that IBV is a swine pathogen, and swine may serve as a natural reservoir of IBVs. In addition, pigs may serve as a model to study the mechanisms of transmission and pathogenesis of IBVs.

Detection and characterization of a novel genotype of porcine astrovirus 4 from nasal swabs from pigs with acute respiratory disease.

Metagenomic sequencing of three nasal swabs collected from 10- to 21-day-old pigs exhibiting unexplained acute respiratory disease from two different commercial production facilities in Oklahoma identified a novel genotype of porcine astrovirus 4 (PAstV-4). The genomes had only ~75 % nucleotide sequence identity to previously characterized PAstV-4 isolates, while the capsid-encoding ORF2 had only ~53 % amino acid sequence identity to reference strains. A TaqMan assay targeting the novel ORF2 gene found 21 % and 19 % incidence in nasal and fecal swabs, respectively, submitted for unrelated diagnostic testing. PAstV-4 RNA levels were significantly higher (P = 0.04) in nasal swabs, suggesting a possible atypical respiratory tropism.

Discovery of a novel Parvovirinae virus, porcine parvovirus 7, by metagenomic sequencing of porcine rectal swabs.

Parvoviruses are a diverse group of viruses containing some of the smallest known species that are capable of infecting a wide range of animals. Metagenomic sequencing of pooled rectal swabs from adult pigs identified a 4103-bp contig consisting of two major open reading frames encoding proteins of 672 and 469 amino acids (aa) in length. BLASTP analysis of the 672-aa protein found 42.4 % identity to fruit bat (Eidolon helvum) parvovirus 2 (EhPV2) and 37.9 % to turkey parvovirus (TuPV) TP1-2012/HUN NS1 proteins. The 469-aa protein had no significant similarity to known proteins. Genetic and phylogenetic analyses suggest that PPV7, EhPV2, and TuPV represent a novel genus in the family Parvoviridae. Quantitative PCR screening of 182 porcine diagnostic samples found a total of 16 positives (8.6 %). Together, these data suggest that PPV7 is a highly divergent novel parvovirus prevalent within the US swine.