RESUMO
Reston virus (RESTV), an ebolavirus, causes clinical disease in macaques but has yet only been associated with rare asymptomatic infections in humans. Its 2008 emergence in pigs in the Philippines raised concerns about food safety, pathogenicity, and zoonotic potential, questions that are still unanswered. Until today, the virulence of RESTV for pigs has remained elusive, with unclear pathogenicity in naturally infected animals and only one experimental study demonstrating susceptibility and evidence for shedding but no disease. Here we show that combined oropharyngeal and nasal infection of young (3- to 7-wk-old) Yorkshire cross pigs with RESTV resulted in severe respiratory disease, with most animals reaching humane endpoint within a week. RESTV-infected pigs developed severe cyanosis, tachypnea, and acute interstitial pneumonia, with RESTV shedding from oronasal mucosal membranes. Our studies indicate that RESTV should be considered a livestock pathogen with zoonotic potential.
Assuntos
Ebolavirus/imunologia , Insuficiência Respiratória/virologia , Doenças dos Suínos/virologia , Animais , Anticorpos Antivirais/imunologia , Causalidade , Vírus de DNA/patogenicidade , Surtos de Doenças/prevenção & controle , Ebolavirus/metabolismo , Ebolavirus/patogenicidade , Filipinas/epidemiologia , Insuficiência Respiratória/veterinária , Sus scrofa/virologia , Suínos/virologia , Doenças dos Suínos/epidemiologia , Eliminação de Partículas Virais/imunologiaRESUMO
Adaptation of prions to new species is thought to reflect the capacity of the host-encoded cellular form of the prion protein (PrPC) to selectively propagate optimized prion conformations from larger ensembles generated in the species of origin. Here we describe an alternate replicative process, termed nonadaptive prion amplification (NAPA), in which dominant conformers bypass this requirement during particular interspecies transmissions. To model susceptibility of horses to prions, we produced transgenic (Tg) mice expressing cognate PrPC Although disease transmission to only a subset of infected TgEq indicated a significant barrier to EqPrPC conversion, the resulting horse prions unexpectedly failed to cause disease upon further passage to TgEq. TgD expressing deer PrPC was similarly refractory to deer prions from diseased TgD infected with mink prions. In both cases, the resulting prions transmitted to mice expressing PrPC from the species of prion origin, demonstrating that transmission barrier eradication of the originating prions was ephemeral and adaptation superficial in TgEq and TgD. Horse prions produced in vitro by protein misfolding cyclic amplification of mouse prions using horse PrPC also failed to infect TgEq but retained tropism for wild-type mice. Concordant patterns of neuropathology and prion deposition in susceptible mice infected with NAPA prions and the corresponding prion of origin confirmed preservation of strain properties. The comparable responses of both prion types to guanidine hydrochloride denaturation indicated this occurs because NAPA precludes selection of novel prion conformations. Our findings provide insights into mechanisms regulating interspecies prion transmission and a framework to reconcile puzzling epidemiological features of certain prion disorders.
Assuntos
Especificidade de Hospedeiro/fisiologia , Proteínas PrPC/fisiologia , Doenças Priônicas/transmissão , Doenças Priônicas/veterinária , Príons/fisiologia , Animais , Cervos , Guanidina/farmacologia , Cavalos , Camundongos , Camundongos Endogâmicos C57BL , Proteínas PrPC/química , Proteínas PrPC/genética , Príons/química , Conformação Proteica , Desnaturação Proteica , Coelhos , Ovinos , Especificidade da Espécie , Relação Estrutura-AtividadeRESUMO
Vesicular stomatitis (VS) is a notifiable disease of livestock affecting cattle, horses, pigs and humans. Vesicular stomatitis virus (VSV) serotypes Indiana and New Jersey are endemic to Central America; however, they also cause sporadic and scattered outbreaks in various countries in South and North America, including the USA. In order to develop an effective experimental challenge model for VSV, we compared the pathogenicity of three VSV serotype Indiana isolates in 36 4-5 week-old pigs. Two bovine isolates of Central American origin and one equine isolate from the USA were used for the experimental infections. Each pig was inoculated with a single isolate by both the intradermal and intranasal routes. Clinical signs of VSV infection were recorded daily for 10 days post-inoculation (days p.i.). Nasal and tonsillar swab samples and blood were collected to monitor immune responses, virus replication and shedding. Post-challenge, characteristic signs of VS were observed, including vesicles on the nasal planum and coronary bands, lameness, loss of hoof walls and pyrexia. Pigs inoculated with the Central American isolates showed consistently more severe clinical signs in comparison to the pigs infected with the USA isolate. Genomic RNA was isolated from the original challenge virus stocks, sequenced and compared to VSV genomes available in GenBank. Comparative genome analysis demonstrated significant differences between the VSV isolate from the USA and the two Central American isolates. Our results indicate that the Central American isolates of VSV serotype Indiana used in this study are more virulent in swine than the USA VSV serotype Indiana isolate and represent good candidate challenge strains for future VSV studies.
Assuntos
Modelos Animais de Doenças , Estomatite Vesicular/patologia , Estomatite Vesicular/virologia , Vesiculovirus/crescimento & desenvolvimento , Vesiculovirus/patogenicidade , Estruturas Animais/patologia , Estruturas Animais/virologia , Animais , Sangue/virologia , Sorogrupo , Suínos , Vesiculovirus/classificação , Virulência , Replicação Viral , Eliminação de Partículas ViraisRESUMO
In mammals, susceptibility to prion infection is primarily modulated by the host's cellular prion protein (PrPC) sequence. In the sheep scrapie model, a graded scale of susceptibility has been established both in vivo and in vitro based on PrPC amino acids 136, 154 and 171, leading to global breeding programmes to reduce the prevalence of scrapie in sheep. Chronic wasting disease (CWD) resistance in cervids is often characterized as decreased prevalence and/or protracted disease progression in individuals with specific alleles; at present, no PrPC allele conferring absolute resistance in cervids has been identified. To model the susceptibility of various naturally occurring and hypothetical cervid PrPC alleles in vitro, we compared the amplification rates and amyloid extension efficiencies of eight distinct CWD isolates in recombinant cervid PrPC substrates using real-time quaking-induced conversion. We hypothesized that the in vitro conversion characteristics of these isolates in cervid substrates would correlate to in vivo susceptibility - permitting susceptibility prediction for the rare alleles found in nature. We also predicted that hypothetical alleles with multiple resistance-associated codons would be more resistant to in vitro conversion than natural alleles with a single resistant codon. Our studies demonstrate that in vitro conversion metrics align with in vivo susceptibility, and that alleles with multiple amino acid substitutions, each influencing resistance independently, do not necessarily contribute additively to conversion resistance. Importantly, we found that the naturally occurring whitetail deer QGAK substrate exhibited the slowest amplification rate among those evaluated, suggesting that further investigation of this allele and its resistance in vivo is warranted.
Assuntos
Cervos , Predisposição Genética para Doença , Proteínas Priônicas/genética , Doença de Emaciação Crônica/genética , Alelos , Amiloide/metabolismo , Animais , Agregação Patológica de Proteínas , Doença de Emaciação Crônica/patologiaRESUMO
The CRISPR/Cas9 genome editing tool has increased the efficiency of creating genetically modified pigs for use as biomedical or agricultural models. The objectives were to determine if DNA editing resulted in a delay in development to the blastocyst stage or in a skewing of the sex ratio. Six DNA templates (gBlocks) that were designed to express guide RNAs that target the transmembrane protease, serine S1, member 2 (TMPRSS2) gene were in vitro transcribed. Pairs of CRISPR guide RNAs that flanked the start codon and polyadenylated Cas9 were co-injected into the cytoplasm of zygotes and cultured in vitro to the blastocyst stage. Blastocysts were collected as they formed on days 5, 6 or 7. PCR was performed to determine genotype and sex of each embryo. Separately, embryos were surgically transferred into recipient gilts on day 4 of estrus. The rate of blastocyst development was not significantly different between CRISPR injection embryos or the non-injected controls at day 5, 6 or 7 (p = 0.36, 0.09, 0.63, respectively). Injection of three CRISPR sets of guides resulted in a detectable INDEL in 92-100 % of the embryos analyzed. There was not a difference in the number of edits or sex ratio of male to female embryos when compared between days 5, 6 and 7 to the controls (p > 0.22, >0.85). There were 12 resulting piglets and all 12 had biallelic edits of TMRPSS2. Zygote injection with CRISPR/Cas9 continues to be a highly efficient tool to genetically modify pig embryos.
Assuntos
Desenvolvimento Embrionário/genética , Marcação de Genes/métodos , Suínos/genética , Zigoto/crescimento & desenvolvimento , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/crescimento & desenvolvimento , Blastocisto/metabolismo , Sistemas CRISPR-Cas/genética , RNA Guia de Cinetoplastídeos/genética , Razão de Masculinidade , Suínos/crescimento & desenvolvimentoRESUMO
Invariant natural killer T (iNKT) cells are an "innate-like" T cell lineage that recognize glycolipid rather than peptide antigens by their semi-invariant T cell receptors. Because iNKT cells can stimulate an extensive array of immune responses, there is considerable interest in targeting these cells to enhance human vaccines against a wide range of microbial pathogens. However, long overlooked is the potential to harness iNKT cell antigens as vaccine adjuvants for domestic animal species that express the iNKT cell-CD1d system. In this review, we discuss the prospect of targeting porcine iNKT cells as a strategy to enhance the efficiency of swine influenza vaccines. In addition, we compare the phenotype and tissue distribution of porcine iNKT cells. Finally, we discuss the challenges that must be overcome before iNKT cell agonists can be contemplated for veterinary use in livestock.
Assuntos
Vacinas contra Influenza/imunologia , Células T Matadoras Naturais/imunologia , Infecções por Orthomyxoviridae/veterinária , Doenças dos Suínos/prevenção & controle , Suínos/imunologia , Animais , Antígenos CD1d/imunologia , Imunidade Inata , Vacinas contra Influenza/uso terapêutico , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/prevenção & controle , Doenças dos Suínos/imunologia , Linfócitos T Auxiliares-Indutores/imunologiaRESUMO
Chronic wasting disease (CWD) is a fatal neurodegenerative disease, classified as a prion disease or transmissible spongiform encephalopathy (TSE) similar to bovine spongiform encephalopathy (BSE). Cervids affected by CWD accumulate an abnormal protease-resistant prion protein throughout the central nervous system (CNS), as well as in both lymphatic and excretory tissues - an aspect of prion disease pathogenesis not observed in cattle with BSE. Using seeded amplification through real-time quaking-induced conversion, we investigated whether the bovine host or prion agent was responsible for this aspect of TSE pathogenesis. We blindly examined numerous central and peripheral tissues from cattle inoculated with CWD for prion seeding activity. Seeded amplification was readily detected in the CNS, though rarely observed in peripheral tissues, with a limited distribution similar to that of BSE prions in cattle. This seems to indicate that prion peripheralization in cattle is a host-driven characteristic of TSE infection.
Assuntos
Sistema Nervoso Central/patologia , Cervos/metabolismo , Proteínas Priônicas/metabolismo , Príons/metabolismo , Doença de Emaciação Crônica/patologia , Doença de Emaciação Crônica/transmissão , Animais , Bovinos/metabolismo , Encefalopatia Espongiforme Bovina/patologia , Dobramento de ProteínaRESUMO
Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of cervids, was first documented nearly 50 years ago in Colorado and Wyoming and has since spread to cervids in 23 states, two Canadian provinces, and the Republic of Korea. The expansion of this disease makes the development of sensitive diagnostic assays and antemortem sampling techniques crucial for the mitigation of its spread; this is especially true in cases of relocation/reintroduction of farmed or free-ranging deer and elk or surveillance studies of private or protected herds, where depopulation is contraindicated. This study sought to evaluate the sensitivity of the real-time quaking-induced conversion (RT-QuIC) assay by using recto-anal mucosa-associated lymphoid tissue (RAMALT) biopsy specimens and nasal brush samples collected antemortem from farmed white-tailed deer (n= 409). Antemortem findings were then compared to results from ante- and postmortem samples (RAMALT, brainstem, and medial retropharyngeal lymph nodes) evaluated by using the current gold standardin vitroassay, immunohistochemistry (IHC) analysis. We hypothesized that the sensitivity of RT-QuIC would be comparable to IHC analysis in antemortem tissues and would correlate with both the genotype and the stage of clinical disease. Our results showed that RAMALT testing by RT-QuIC assay had the highest sensitivity (69.8%) compared to that of postmortem testing, with a specificity of >93.9%. These data suggest that RT-QuIC, like IHC analysis, is an effective assay for detection of PrP(CWD)in rectal biopsy specimens and other antemortem samples and, with further research to identify more sensitive tissues, bodily fluids, or experimental conditions, has potential for large-scale and rapid automated testing for CWD diagnosis.
Assuntos
Testes Diagnósticos de Rotina/métodos , Mucosa Intestinal/química , Tecido Linfoide/química , Mucosa Nasal/química , Patologia Molecular/métodos , Príons/análise , Doença de Emaciação Crônica/diagnóstico , Animais , Biópsia , Cervos , Sensibilidade e Especificidade , Fatores de TempoRESUMO
Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of cervids, was first documented nearly 50 years ago in Colorado and Wyoming and has since been detected across North America and the Republic of Korea. The expansion of this disease makes the development of sensitive diagnostic assays and antemortem sampling techniques crucial for the mitigation of its spread; this is especially true in cases of relocation/reintroduction or prevalence studies of large or protected herds, where depopulation may be contraindicated. This study evaluated the sensitivity of the real-time quaking-induced conversion (RT-QuIC) assay of recto-anal mucosa-associated lymphoid tissue (RAMALT) biopsy specimens and nasal brushings collected antemortem. These findings were compared to results of immunohistochemistry (IHC) analysis of ante- and postmortem samples. RAMALT samples were collected from populations of farmed and free-ranging Rocky Mountain elk (Cervus elaphus nelsoni;n= 323), and nasal brush samples were collected from a subpopulation of these animals (n= 205). We hypothesized that the sensitivity of RT-QuIC would be comparable to that of IHC analysis of RAMALT and would correspond to that of IHC analysis of postmortem tissues. We found RAMALT sensitivity (77.3%) to be highly correlative between RT-QuIC and IHC analysis. Sensitivity was lower when testing nasal brushings (34%), though both RAMALT and nasal brush test sensitivities were dependent on both thePRNPgenotype and disease progression determined by the obex score. These data suggest that RT-QuIC, like IHC analysis, is a relatively sensitive assay for detection of CWD prions in RAMALT biopsy specimens and, with further investigation, has potential for large-scale and rapid automated testing of antemortem samples for CWD.
Assuntos
Testes Diagnósticos de Rotina/métodos , Mucosa Intestinal/química , Tecido Linfoide/química , Mucosa Nasal/química , Patologia Molecular/métodos , Príons/análise , Doença de Emaciação Crônica/diagnóstico , Animais , Biópsia , Feminino , Masculino , Ruminantes , Sensibilidade e Especificidade , Fatores de TempoRESUMO
UNLABELLED: At least 10 different genotypes of novel reassortant H3N2 influenza viruses with 2009 pandemic H1N1 [A(H1N1)pdm09] gene(s) have been identified in U.S. pigs, including the H3N2 variant with a single A(H1N1)pdm09 M gene, which has infected more than 300 people. To date, only three genotypes of these viruses have been evaluated in animal models, and the pathogenicity and transmissibility of the other seven genotype viruses remain unknown. Here, we show that three H3N2 reassortant viruses that contain 3 (NP, M, and NS) or 5 (PA, PB2, NP, M, and NS) genes from A(H1N1)pdm09 were pathogenic in pigs, similar to the endemic H3N2 swine virus. However, the reassortant H3N2 virus with 3 A(H1N1)pdm09 genes and a recent human influenza virus N2 gene was transmitted most efficiently among pigs, whereas the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes was transmitted less efficiently than the endemic H3N2 virus. Interestingly, the polymerase complex of reassortant H3N2 virus with 5 A(H1N1)pdm09 genes showed significantly higher polymerase activity than those of endemic and reassortant H3N2 viruses with 3 A(H1N1)pdm09 genes. Further studies showed that an avian-like glycine at position 228 at the hemagglutinin (HA) receptor binding site is responsible for inefficient transmission of the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes. Taken together, our results provide insights into the pathogenicity and transmissibility of novel reassortant H3N2 viruses in pigs and suggest that a mammalian-like serine at position 228 in the HA is critical for the transmissibility of these reassortant H3N2 viruses. IMPORTANCE: Swine influenza is a highly contagious zoonotic disease that threatens animal and public health. Introduction of 2009 pandemic H1N1 virus [A(H1N1)pdm09] into swine herds has resulted in novel reassortant influenza viruses in swine, including H3N2 and H1N2 variants that have caused human infections in the United States. We showed that reassortant H3N2 influenza viruses with 3 or 5 genes from A(H1N1)pdm09 isolated from diseased pigs are pathogenic and transmissible in pigs, but the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes displayed less efficient transmissibility than the endemic and reassortant H3N2 viruses with 3 A(H1N1)pdm09 genes. Further studies revealed that an avian-like glycine at the HA 228 receptor binding site of the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes is responsible for less efficient transmissibility in pigs. Our results provide insights into viral pathogenesis and the transmission of novel reassortant H3N2 viruses that are circulating in U.S. swine herds and warrant future surveillance.
Assuntos
Vírus da Influenza A Subtipo H3N2/fisiologia , Vírus da Influenza A Subtipo H3N2/patogenicidade , Infecções por Orthomyxoviridae/veterinária , Vírus Reordenados/fisiologia , Vírus Reordenados/patogenicidade , Doenças dos Suínos/transmissão , Doenças dos Suínos/virologia , Animais , Modelos Animais de Doenças , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/isolamento & purificação , Infecções por Orthomyxoviridae/transmissão , Infecções por Orthomyxoviridae/virologia , Vírus Reordenados/genética , Vírus Reordenados/isolamento & purificação , Suínos , Estados UnidosRESUMO
The challenge of increasing swine production and a rising number of novel and known swine influenza viruses has prompted a considerable boost in research into how and why pigs have become such significant hosts for influenza viruses. The ecology of influenza A viruses is rather complicated, involving multiple host species and a segmented genome. Wild aquatic birds are the reservoir for the majority of influenza A viruses, but novel influenza viruses were recently identified in bats. Occasionally, influenza A viruses can be transmitted to mammals from avian species and this event could lead to the generation of human pandemic strains. Swine are thought to be "mixing vessels" because they are susceptible to infection with both avian and mammalian influenza viruses; and novel influenza viruses can be generated in pigs by reassortment. At present, it is difficult to predict which viruses might cause a human pandemic. Therefore, both human and veterinary research needs to give more attention to the potential cross-species transmission capacity of influenza A viruses.
Assuntos
Vírus da Influenza A/fisiologia , Influenza Humana/virologia , Doenças dos Suínos/virologia , Animais , Pesquisa Biomédica , Humanos , Vírus da Influenza A/genética , Influenza Humana/transmissão , Suínos , Doenças dos Suínos/transmissãoRESUMO
The matrix 1 (M1) protein is a multifunctional protein in the life cycle of influenza virus. It plays an important role in virus budding and intracellular trafficking of viral ribonucleoproteins (vRNPs). The M1 protein consists of three domains based on the structure: N-terminal domain, Middle domain, and C-terminal domain. However, the functions of different domains of the M1 protein remain largely unclear. In this study, using bimolecular fluorescence complementation assays (BIFC) we demonstrated that swine importin α1 interacts with the M1 protein and transports it to the nucleus. Interestingly, M1 with mutated nuclear localization signal (NLS; 101-RKLKR-105 to 101-AALAA-105) still interacts with swine importin α1 and is localized in the nucleus, suggesting that the NLS located at residues 101-105 is not the only NLS within M1 recombinant protein containing 1-160 residues of M1 with mutated nuclear localization signal is able to interact with swine importin α1, but M1/60-252 domains cannot bind importin α1. Further mapping showed that the deletion of residues 1-20 impaired the interaction between N terminus of M1 and importin α1. Collectively, our data suggested that the N-terminal domain of M1 protein is critical for binding swine importin α1 and for nuclear localization.
Assuntos
Interações Hospedeiro-Patógeno , Mapeamento de Interação de Proteínas , Proteínas da Matriz Viral/metabolismo , alfa Carioferinas/metabolismo , Animais , Linhagem Celular , Vírus da Influenza A Subtipo H1N1/genética , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Sinais de Localização Nuclear , Ligação Proteica , Transporte Proteico , Deleção de Sequência , Suínos , Proteínas da Matriz Viral/genéticaRESUMO
IMPORTANCE: Although the role of bovine coronavirus (BCoV) in calf diarrhea and respiratory disorders is well documented, its contribution to neurological diseases is unclear. OBJECTIVE: This study conducted virological investigations of calves showing diarrhea and respiratory and neurological signs. METHODS: An outbreak of diarrhea, respiratory, and neurological disorders occurred among the 12 calves in July 2022 in Istanbul, Türkiye. Two of these calves exhibited neurological signs and died a few days after the appearance of symptoms. One of these calves was necropsied and analyzed using molecular and histopathological tests. RESULTS: BCoV RNA was detected in the brain, lung, spleen, liver, and intestine of the calf that had neurological signs by real-time reverse transcription polymerase chain reaction. Immunostaining was also observed in the intestine and brain. A 622 bp S1 gene product was noted on gel electrophoresis only in the brain. Phylogenetic analysis indicated that the BCoV detected in this study had a high proximity to the BCoV strain GIb with 99.19% nucleotide sequence homology to the strains detected in Poland, Israel, Türkiye, and France. No distinct genetic lineages were observed when the brain isolate was compared with the respiratory and enteric strains reported to GenBank. In addition, the highest identity (98,72%) was obtained with the HECV 4408 and L07748 strains of human coronaviruses. CONCLUSIONS AND RELEVANCE: The strain detected in a calf brain belongs to the GIb-European lineage and shares high sequence homology with BCoV strains detected in Europe and Israel. In addition, the similarity between the human coronaviruses (4408 and L07748) raises questions about the zoonotic potential of the strains detected in this study.
Assuntos
Encéfalo , Doenças dos Bovinos , Infecções por Coronavirus , Coronavirus Bovino , Filogenia , Animais , Bovinos , Coronavirus Bovino/genética , Coronavirus Bovino/isolamento & purificação , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Infecções por Coronavirus/patologia , Doenças dos Bovinos/virologia , Doenças dos Bovinos/patologia , Encéfalo/virologia , Encéfalo/patologia , Turquia/epidemiologia , Surtos de Doenças/veterinária , Pneumonia Viral/veterinária , Pneumonia Viral/virologia , Pneumonia Viral/patologia , Doenças do Sistema Nervoso/veterinária , Doenças do Sistema Nervoso/virologia , Doenças do Sistema Nervoso/patologiaRESUMO
Triple reassortant swine influenza viruses (SIVs) and 2009 pandemic H1N1 (pH1N1) virus contain an avian-origin PB2 with 271A, 590S, 591R, and 627E. To evaluate the role of PB2 271A, 590S, and 591R in the replication and virulence of SIV, single (1930-TX98-PB2-271T)-, double (1930-TX98-PB2-590A591A)-, and triple (1930-TX98-PB2-271T590A591A)-mutated viruses were generated in the background of the H1N1 A/swine/Iowa/15/30 (1930) virus with an avian-origin PB2 from the triple-reassortant A/swine/Texas/4199-2/98 (TX98) virus, called the parental 1930-TX98-PB2. Compared to parental virus and single- and double-mutated viruses, the triple-mutated virus replicated less efficiently in cell cultures and was attenuated in mice. These results suggest that a combination of 271A with the 590/591 SR polymorphism is critical for pH1N1 and triple-reassortant SIVs for efficient replication and adaptation in mammals.
Assuntos
Vírus da Influenza A Subtipo H1N1/patogenicidade , Infecções por Orthomyxoviridae/veterinária , Polimorfismo de Nucleotídeo Único , RNA Polimerase Dependente de RNA/química , RNA Polimerase Dependente de RNA/genética , Doenças dos Suínos/virologia , Proteínas Virais/química , Proteínas Virais/genética , Replicação Viral , Animais , Linhagem Celular , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/fisiologia , Influenza Humana/virologia , Camundongos , Mutação , Infecções por Orthomyxoviridae/virologia , RNA Polimerase Dependente de RNA/metabolismo , Vírus Reordenados/genética , Vírus Reordenados/patogenicidade , Vírus Reordenados/fisiologia , Suínos , Doenças dos Suínos/mortalidade , Proteínas Virais/metabolismo , VirulênciaRESUMO
Control of swine influenza A virus (IAV) in the United States is hindered because inactivated vaccines do not provide robust cross-protection against the multiple antigenic variants cocirculating in the field. Vaccine efficacy can be limited further for vaccines administered to young pigs that possess maternally derived immunity. We previously demonstrated that a recombinant A/sw/Texas/4199-2/1998 (TX98) (H3N2) virus expressing a truncated NS1 protein is attenuated in swine and has potential for use as an intranasal live attenuated influenza virus (LAIV) vaccine. In the present study, we compared 1 dose of intranasal LAIV with 2 intramuscular doses of TX98 whole inactivated virus (WIV) with adjuvant in weanling pigs with and without TX98-specific maternally derived antibodies (MDA). Pigs were subsequently challenged with wild-type homologous TX98 H3N2 virus or with an antigenic variant, A/sw/Colorado/23619/1999 (CO99) (H3N2). In the absence of MDA, both vaccines protected against homologous TX98 and heterologous CO99 shedding, although the LAIV elicited lower hemagglutination inhibition (HI) antibody titers in serum. The efficacy of both vaccines was reduced by the presence of MDA; however, WIV vaccination of MDA-positive pigs led to dramatically enhanced pneumonia following heterologous challenge, a phenomenon known as vaccine-associated enhanced respiratory disease (VAERD). A single dose of LAIV administered to MDA-positive pigs still provided partial protection from CO99 and may be a safer vaccine for young pigs under field conditions, where dams are routinely vaccinated and diverse IAV strains are in circulation. These results have implications not only for pigs but also for other influenza virus host species.
Assuntos
Anticorpos/química , Vacinas contra Influenza/metabolismo , Infecções Respiratórias/imunologia , Vacinas Atenuadas/metabolismo , Animais , Líquido da Lavagem Broncoalveolar , Linhagem Celular , Cães , Testes de Inibição da Hemaglutinação , Vírus da Influenza A Subtipo H3N2/metabolismo , Pulmão/metabolismo , Mucosa/metabolismo , SuínosRESUMO
The 2009 pandemic H1N1 virus (pH1N1) contains neuraminidase (NA) and matrix (M) genes from Eurasian avian-like swine influenza viruses (SIVs), with the remaining six genes from North American triple-reassortant SIVs. To characterize the role of the pH1N1 NA and M genes in pathogenesis and transmission, their impact was evaluated in the background of an H1N1 triple-reassortant (tr1930) SIV in which the HA (H3) and NA (N2) of influenza A/swine/Texas/4199-2/98 virus were replaced with those from the classical H1N1 A/swine/Iowa/15/30 (1930) virus. The laboratory-adapted 1930 virus did not shed nor transmit in pigs, but tr1930 was able to shed in infected pigs. The NA, M or both genes of the tr1930 virus were then substituted by those of pH1N1. The resulting virus with both NA and M from pH1N1 grew to significantly higher titre in cell cultures than the viruses with single NA or M from pH1N1. In a pig model, only the virus containing both NA and M from pH1N1 was transmitted to and infected sentinels, whereas the viruses with single NA or M from pH1N1 did not. These results demonstrate that the right combination of NA and M genes is critical for the replication and transmissibility of influenza viruses in pigs.
Assuntos
Vírus da Influenza A Subtipo H1N1/fisiologia , Neuraminidase/metabolismo , Infecções por Orthomyxoviridae/veterinária , Doenças dos Suínos/transmissão , Doenças dos Suínos/virologia , Proteínas da Matriz Viral/metabolismo , Proteínas Virais/metabolismo , Replicação Viral , Animais , Vírus da Influenza A Subtipo H1N1/enzimologia , Vírus da Influenza A Subtipo H1N1/genética , Iowa/epidemiologia , Neuraminidase/genética , Infecções por Orthomyxoviridae/epidemiologia , Infecções por Orthomyxoviridae/transmissão , Infecções por Orthomyxoviridae/virologia , Suínos , Doenças dos Suínos/epidemiologia , Texas/epidemiologia , Proteínas da Matriz Viral/genética , Proteínas Virais/genéticaRESUMO
Both H9N2 avian influenza and 2009 pandemic H1N1 viruses (pH1N1) are able to infect humans and swine, which has raised concerns that novel reassortant H9 viruses with pH1N1 genes might be generated in these hosts by reassortment. Although previous studies have demonstrated that reassortant H9 viruses with pH1N1 genes show increased virulence in mice and transmissibility in ferrets, the virulence and transmissibility of reassortant H9 viruses in natural hosts such as chickens and swine remain unknown. This study generated two reassortant H9 viruses (H9N2/CA09 and H9N1/CA09) in the background of the pH1N1 A/California/04/2009 (CA09) virus by replacing either both the haemagglutinin (HA) and neuraminidase (NA) genes or only the HA gene with the respective genes from the A/quail/Hong Kong/G1/1997 (H9N2) virus and evaluated their replication, pathogenicity and transmission in chickens and pigs compared with the parental viruses. Chickens that were infected with the parental H9N2 and reassortant H9 viruses seroconverted. The parental H9N2 and reassortant H9N2/CA09 viruses were transmitted to sentinel chickens, but H9N1/CA09 virus was not. The parental H9N2 replicated poorly and was not transmitted in pigs, whereas both H9N2/CA09 and H9N1/CA09 viruses replicated and were transmitted efficiently in pigs, similar to the pH1N1 virus. These results demonstrated that reassortant H9 viruses with pH1N1 genes show enhanced replication and transmissibility in pigs compared with the parental H9N2 virus, indicating that they may pose a threat for humans if such reassortants arise in swine.
Assuntos
Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H9N2/genética , Infecções por Orthomyxoviridae/veterinária , Pandemias , Vírus Reordenados/genética , Animais , Linhagem Celular , Galinhas , Cães , Humanos , Infecções por Orthomyxoviridae/epidemiologia , Infecções por Orthomyxoviridae/virologia , Doenças das Aves Domésticas/transmissão , Doenças das Aves Domésticas/virologia , Distribuição Aleatória , Organismos Livres de Patógenos Específicos , Suínos , Doenças dos Suínos/transmissão , Doenças dos Suínos/virologiaRESUMO
A remarkable feature of the 2009 pandemic H1N1 influenza virus is its efficient transmissibility in humans compared to that of precursor strains from the triple-reassortant swine influenza virus lineage, which cause only sporadic infections in humans. The viral components essential for this phenotype have not been fully elucidated. In this study, we aimed to determine the viral factors critical for aerosol transmission of the 2009 pandemic virus. Single or multiple segment reassortments were made between the pandemic A/California/04/09 (H1N1) (Cal/09) virus and another H1N1 strain, A/Puerto Rico/8/34 (H1N1) (PR8). These viruses were then tested in the guinea pig model to understand which segment of Cal/09 virus conferred transmissibility to the poorly transmissible PR8 virus. We confirmed our findings by generating recombinant A/swine/Texas/1998 (H3N2) (sw/Tx/98) virus, a representative triple-reassortant swine virus, containing segments of the Cal/09 virus. The data showed that the M segment of the Cal/09 virus promoted aerosol transmissibility to recombinant viruses with PR8 and sw/Tx/98 virus backgrounds, suggesting that the M segment is a critical factor supporting the transmission of the 2009 pandemic virus.
Assuntos
Vírus da Influenza A Subtipo H1N1/patogenicidade , Infecções por Orthomyxoviridae/transmissão , Infecções por Orthomyxoviridae/virologia , Proteínas da Matriz Viral/metabolismo , Fatores de Virulência/metabolismo , Animais , Modelos Animais de Doenças , Cobaias , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/patogenicidade , Vírus Reordenados/genética , Vírus Reordenados/patogenicidade , Proteínas da Matriz Viral/genética , Fatores de Virulência/genéticaRESUMO
The first influenza pandemic of the new millennium was caused by a newly emerged swine-origin influenza virus (SOIV) (H1N1). This new virus is characterized by a previously unknown constellation of gene segments derived from North American and Eurasian swine lineages and the absence of common markers predictive of human adaptation. Overall, human infections appeared to be mild, but an alarming number of young individuals presented with symptoms atypical for seasonal influenza. The new SOIV also showed a sustained human-to-human transmissibility and higher reproduction ratio than common seasonal viruses, altogether indicating a higher pathogenic potential for this newly emerged virus. To study the virulence of the SOIV, we used a recently established cynomolgus macaque model and compared parameters of clinical disease, virology, host responses, and pathology/histopathology with a current seasonal H1N1 virus. We here show that infection of macaques with two genetically similar but clinically distinct SOIV isolates from the early stage of the pandemic (A/Mexico/4108/2009 and A/Mexico/InDRE4487/2009) resulted in upper and lower respiratory tract infections and clinical disease ranging from mild to severe pneumonia that was clearly advanced over the mild infection caused by A/Kawasaki/UTK-4/2009, a current seasonal strain. Unexpectedly, we observed heterogeneity among the two SOIV isolates in virus replication, host transcriptional and cytokine responses, and disease progression, demonstrating a higher pathogenic potential for A/Mexico/InDRE4487/2009. Differences in virulence may explain more severe disease, as was seen with certain individuals infected with the emerged pandemic influenza virus. Thus, the nonhuman primate model closely mimics influenza in humans.
Assuntos
Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Vírus da Influenza A Subtipo H1N1/patogenicidade , Infecções por Orthomyxoviridae/patologia , Infecções por Orthomyxoviridae/virologia , Doenças dos Primatas/patologia , Doenças dos Primatas/virologia , Animais , Pré-Escolar , Citocinas/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Perfilação da Expressão Gênica , Variação Genética , Humanos , Influenza Humana/virologia , Macaca , Masculino , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Infecções Respiratórias/patologia , Infecções Respiratórias/virologia , Índice de Gravidade de Doença , VirulênciaRESUMO
Reassortant H1 swine influenza viruses (SIVs) carrying 2009 pandemic H1N1 virus (pH1N1) genes have been isolated from pigs worldwide. Seven novel reassortant H3N2 SIVs were identified from diseased pigs in the USA from winter 2010 to spring 2011. These novel viruses contain three or five internal genes from pH1N1 and continue to circulate in swine herds. The emergence of novel reassortant H3N2 SIVs demonstrates reassortment between pH1N1 and endemic SIVs in pigs and justifies continuous surveillance.