RESUMEN
The increased detection of H3 C-IVA (1990.4.a) clade influenza A viruses (IAVs) in US swine in 2019 was associated with a reassortment event to acquire an H1N1pdm09 lineage nucleoprotein (pdmNP) gene, replacing a TRIG lineage NP (trigNP). We hypothesized that acquiring the pdmNP conferred a selective advantage over prior circulating H3 viruses with a trigNP. To investigate the role of NP reassortment in transmission, we identified two contemporary 1990.4.a representative strains (NC/19 and MN/18) with different evolutionary origins of the NP gene. A reverse genetics system was used to generate wild-type (wt) strains and swap the pdm and TRIG lineage NP genes, generating four viruses: wtNC/19-pdmNP, NC/19-trigNP, wtMN/18-trigNP, and MN/18-pdmNP. The pathogenicity and transmission of the four viruses were compared in pigs. All four viruses infected 10 primary pigs and transmitted to five indirect contact pigs per group. Pigs infected via contact with MN/18-pdmNP shed virus 2 days earlier than pigs infected with wtMN/18-trigNP. The inverse did not occur for wtNC/19-pdmNP and NC/19-trigNP. This suggests that pdmNP reassortment resulted in a combination of genes that improved transmission efficiency when paired with the 1990.4.a hemagglutinin (HA). This is likely a multigenic trait, as replacing the trigNP gene did not diminish the transmission of a wild-type IAV in swine. This study demonstrates how reassortment and evolutionary change of internal genes can result in more transmissible viruses that influence HA clade detection frequency. Thus, rapidly identifying novel reassortants paired with dominant hemagglutinin/neuraminidase may improve the prediction of strains to include in vaccines.IMPORTANCEInfluenza A viruses (IAVs) are composed of eight non-continuous gene segments that can reassort during coinfection of a host, creating new combinations. Some gene combinations may convey a selective advantage and be paired together preferentially. A reassortment event was detected in swine in the United States that involved the exchange of two lineages of nucleoprotein (NP) genes (trigNP to pdmNP) that became a predominant genotype detected in surveillance. Using a transmission study, we demonstrated that exchanging the trigNP for a pdmNP caused the virus to shed from the nose at higher levels and transmit to other pigs more rapidly. Replacing a pdmNP with a trigNP did not hinder transmission, suggesting that transmission efficiency depends on interactions between multiple genes. This demonstrates how reassortment alters IAV transmission and that reassortment events can provide an explanation for why genetically related viruses with different internal gene combinations experience rapid fluxes in detection frequency.
Asunto(s)
Virus de la Influenza A , Proteínas de la Nucleocápside , Infecciones por Orthomyxoviridae , Enfermedades de los Porcinos , Animales , Hemaglutininas , Virus de la Influenza A/clasificación , Virus de la Influenza A/genética , Infecciones por Orthomyxoviridae/veterinaria , Infecciones por Orthomyxoviridae/virología , Virus Reordenados/genética , Porcinos , Estados Unidos , Proteínas de la Nucleocápside/metabolismoRESUMEN
The 2009 H1N1 pandemic (pdm09) lineage of influenza A virus (IAV) crosses interspecies barriers with frequent human-to-swine spillovers each year. These spillovers reassort and drift within swine populations, leading to genetically and antigenically novel IAV that represent a zoonotic threat. We quantified interspecies transmission of the pdm09 lineage, persistence in swine, and identified how evolution in swine impacted zoonotic risk. Human and swine pdm09 case counts between 2010 and 2020 were correlated and human pdm09 burden and circulation directly impacted the detection of pdm09 in pigs. However, there was a relative absence of pdm09 circulation in humans during the 2020-21 season that was not reflected in swine. During the 2020-21 season, most swine pdm09 detections originated from human-to-swine spillovers from the 2018-19 and 2019-20 seasons that persisted in swine. We identified contemporary swine pdm09 representatives of each persistent spillover and quantified cross-reactivity between human seasonal H1 vaccine strains and the swine strains using a panel of monovalent ferret antisera in hemagglutination inhibition (HI) assays. The swine pdm09s had variable antigenic reactivity to vaccine antisera, but each swine pdm09 clade exhibited significant reduction in cross-reactivity to one or more of the human seasonal vaccine strains. Further supporting zoonotic risk, we showed phylogenetic evidence for 17 swine-to-human transmission events of pdm09 from 2010 to 2021, 11 of which were not previously classified as variants, with each of the zoonotic cases associated with persistent circulation of pdm09 in pigs. These data demonstrate that reverse-zoonoses and evolution of pdm09 in swine results in viruses that are capable of zoonotic transmission and represent a potential pandemic threat.
Asunto(s)
Subtipo H1N1 del Virus de la Influenza A , Virus de la Influenza A , Gripe Humana , Infecciones por Orthomyxoviridae , Enfermedades de los Porcinos , Animales , Estados Unidos/epidemiología , Humanos , Porcinos , Subtipo H1N1 del Virus de la Influenza A/genética , Infecciones por Orthomyxoviridae/epidemiología , Infecciones por Orthomyxoviridae/veterinaria , Filogenia , Hurones , Zoonosis/epidemiología , Sueros Inmunes , Gripe Humana/epidemiologíaRESUMEN
We report highly pathogenic avian influenza A(H5N1) virus in dairy cattle and cats in Kansas and Texas, United States, which reflects the continued spread of clade 2.3.4.4b viruses that entered the country in late 2021. Infected cattle experienced nonspecific illness, reduced feed intake and rumination, and an abrupt drop in milk production, but fatal systemic influenza infection developed in domestic cats fed raw (unpasteurized) colostrum and milk from affected cows. Cow-to-cow transmission appears to have occurred because infections were observed in cattle on Michigan, Idaho, and Ohio farms where avian influenza virus-infected cows were transported. Although the US Food and Drug Administration has indicated the commercial milk supply remains safe, the detection of influenza virus in unpasteurized bovine milk is a concern because of potential cross-species transmission. Continued surveillance of highly pathogenic avian influenza viruses in domestic production animals is needed to prevent cross-species and mammal-to-mammal transmission.
Asunto(s)
Enfermedades de los Gatos , Enfermedades de los Bovinos , Subtipo H5N1 del Virus de la Influenza A , Infecciones por Orthomyxoviridae , Animales , Gatos , Bovinos , Enfermedades de los Gatos/virología , Enfermedades de los Gatos/epidemiología , Enfermedades de los Bovinos/virología , Enfermedades de los Bovinos/epidemiología , Enfermedades de los Bovinos/transmisión , Infecciones por Orthomyxoviridae/virología , Infecciones por Orthomyxoviridae/veterinaria , Infecciones por Orthomyxoviridae/epidemiología , Subtipo H5N1 del Virus de la Influenza A/patogenicidad , Subtipo H5N1 del Virus de la Influenza A/genética , Estados Unidos/epidemiología , Gripe Aviar/virología , Gripe Aviar/epidemiología , Gripe Aviar/transmisión , Leche/virología , FemeninoRESUMEN
Influenza A viruses (FLUAV) cause respiratory diseases in many host species, including humans and pigs. The spillover of FLUAV between swine and humans has been a concern for both public health and the swine industry. With the emergence of the triple reassortant internal gene (TRIG) constellation, establishment of human-origin FLUAVs in pigs has become more common, leading to increased viral diversity. However, little is known about the adaptation processes that are needed for a human-origin FLUAV to transmit and become established in pigs. We generated a reassortant FLUAV (VIC11pTRIG) containing surface gene segments from a human FLUAV strain and internal gene segments from the 2009 pandemic and TRIG FLUAV lineages and demonstrated that it can replicate and transmit in pigs. Sequencing and variant analysis identified three mutants that emerged during replication in pigs, which were mapped near the receptor binding site of the hemagglutinin (HA). The variants replicated more efficiently in differentiated swine tracheal cells compared to the virus containing the wildtype human-origin HA, and one of them was present in all contact pigs. These results show that variants are selected quickly after replication of human-origin HA in pigs, leading to improved fitness in the swine host, likely contributing to transmission. IMPORTANCE Influenza A viruses cause respiratory disease in several species, including humans and pigs. The bidirectional transmission of FLUAV between humans and pigs plays a significant role in the generation of novel viral strains, greatly impacting viral epidemiology. However, little is known about the evolutionary processes that allow human FLUAV to become established in pigs. In this study, we generated reassortant viruses containing human seasonal HA and neuraminidase (NA) on different constellations of internal genes and tested their ability to replicate and transmit in pigs. We demonstrated that a virus containing a common internal gene constellation currently found in U.S. swine was able to transmit efficiently via the respiratory route. We identified a specific amino acid substitution that was fixed in the respiratory contact pigs that was associated with improved replication in primary swine tracheal epithelial cells, suggesting it was crucial for the transmissibility of the human virus in pigs.
Asunto(s)
Glicoproteínas Hemaglutininas del Virus de la Influenza , Virus de la Influenza A , Gripe Humana , Infecciones por Orthomyxoviridae , Enfermedades de los Porcinos , Animales , Humanos , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Virus de la Influenza A/genética , Gripe Humana/transmisión , Mutación , Infecciones por Orthomyxoviridae/transmisión , Virus Reordenados/genética , Porcinos , Enfermedades de los Porcinos/virologíaRESUMEN
Influenza A viruses (IAV) sporadically transmit from swine to humans, typically associated with agricultural fairs in the United States. A human seasonal H3 virus from the 2010-2011 IAV season was introduced into the U.S. swine population and termed H3.2010.1 to differentiate it from the previous swine H3 virus. This H3N2 lineage became widespread in the U.S. commercial swine population, subsequently spilling over into exhibition swine, and caused a majority of H3N2 variant (H3N2v) cases in humans in 2016 and 2017. A cluster of human H3N2v cases were reported at an agricultural fair in 2017 in Ohio, where 2010.1 H3N2 IAV was concurrently detected in exhibition swine. Genomic analysis showed that the swine and human isolates were nearly identical. In this study, we evaluated the propensity of a 2010.1 H3N2 IAV (A/swine/Ohio/A01354299/2017 [sw/OH/2017]) isolated from a pig in the agricultural fair outbreak to replicate in ferrets and transmit from swine to ferret. sw/OH/2017 displayed robust replication in the ferret respiratory tract, causing slight fever and moderate weight loss. Further, sw/OH/2017 was capable of efficient respiratory droplet transmission from infected pigs to contact ferrets. These findings establish a model for evaluating the propensity of swine IAV to transmit from pig to ferret as a measure of risk to the human population. The identification of higher-risk swine strains can then be targeted for control measures to limit the dissemination at human-swine interfaces to reduce the risk of zoonotic infections and to inform pandemic planning.IMPORTANCE A recently emerged lineage of human-like H3N2 (H3.2010.1) influenza A virus (IAV) from swine has been frequently detected in commercial and exhibition swine in recent years and has been associated with H3N2 variant cases in humans from 2016 and 2017. To demonstrate a model for characterizing the potential for zoonotic transmission associated with swine IAV, we performed an in vivo study of transmission between pigs infected with an H3.2010.1 H3N2 IAV and aerosol contact ferrets. The efficient interspecies transmission demonstrated for the H3.2010.1 IAV in swine emphasizes the need for further characterization of viruses circulating at the swine-human interface for transmission potential prior to human spillover and the development and implementation of more robust vaccines and control strategies to mitigate human exposure to higher-risk swine strains.
Asunto(s)
Subtipo H3N2 del Virus de la Influenza A/metabolismo , Infecciones por Orthomyxoviridae/transmisión , Zoonosis/transmisión , Aerosoles , Animales , Reacciones Cruzadas/inmunología , Hurones/virología , Glicoproteínas Hemaglutininas del Virus de la Influenza/metabolismo , Humanos , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/patogenicidad , Virus de la Influenza A/metabolismo , Virus de la Influenza A/patogenicidad , Gripe Humana/virología , Infecciones por Orthomyxoviridae/virología , Porcinos/virología , Enfermedades de los Porcinos/virología , Estados Unidos , Zoonosis/virologíaRESUMEN
Porcine reproductive and respiratory syndrome virus (PRRSV), an important pathogen that affects the pig industry, is a highly genetically diverse RNA virus. However, the phylogenetic and genomic recombination properties of this virus have not been completely elucidated. In this study, comparative analyses of all available genomic sequences of North American (NA)-type PRRSVs (n = 355, including 138 PRRSV genomes sequenced in this study) in China and the United States during 2014-2018 revealed a high frequency of interlineage recombination hot spots in nonstructural protein 9 (NSP9) and the GP2 to GP3 regions. Lineage 1 (L1) PRRSV was found to be susceptible to recombination among PRRSVs both in China and the United States. The recombinant major parent between the 1991-2013 data and the 2014-2018 data showed a trend from complex to simple. The major recombination pattern changed from an L8 to L1 backbone during 2014-2018 for Chinese PRRSVs, whereas L1 was always the major backbone for US PRRSVs. Intralineage recombination hot spots were not as concentrated as interlineage recombination hot spots. In the two main clades with differential diversity in L1, NADC30-like PRRSVs are undergoing a decrease in population genetic diversity, NADC34-like PRRSVs have been relatively stable in population genetic diversity for years. Systematic analyses of insertion and deletion (indel) polymorphisms of NSP2 divided PRRSVs into 25 patterns, which could generate novel references for the classification of PRRSVs. The results of this study contribute to a deeper understanding of the recombination of PRRSVs and indicate the need for coordinated epidemiological investigations among countries.IMPORTANCE Porcine reproductive and respiratory syndrome (PRRS) is one of the most significant swine diseases. However, the phylogenetic and genomic recombination properties of the PRRS virus (PRRSV) have not been completely elucidated. In this study, we systematically compared differences in the lineage distribution, recombination, NSP2 polymorphisms, and evolutionary dynamics between North American (NA)-type PRRSVs in China and in the United States. Strikingly, we found high frequency of interlineage recombination hot spots in nonstructural protein 9 (NSP9) and in the GP2 to GP3 region. Also, intralineage recombination hot spots were scattered across the genome between Chinese and US strains. Furthermore, we proposed novel methods based on NSP2 indel patterns for the classification of PRRSVs. Evolutionary dynamics analysis revealed that NADC30-like PRRSVs are undergoing a decrease in population genetic diversity, suggesting that a dominant population may occur and cause an outbreak. Our findings offer important insights into the recombination of PRRSVs and suggest the need for coordinated international epidemiological investigations.
Asunto(s)
Polimorfismo Genético , Virus del Síndrome Respiratorio y Reproductivo Porcino/genética , Recombinación Genética , Proteínas Virales/genética , Animales , China/epidemiología , Filogeografía , Síndrome Respiratorio y de la Reproducción Porcina/epidemiología , Síndrome Respiratorio y de la Reproducción Porcina/genética , Porcinos , Estados Unidos/epidemiologíaRESUMEN
In the past decade, different members of the genus Mamastrovirus have been associated with outbreaks of neurologic disease in humans, cattle, sheep, mink, and, most recently, porcine astrovirus 3 (PoAstV3) in swine. We performed a retrospective analysis of 50 cases of porcine neurologic disease of undetermined cause but with microscopic lesions compatible with a viral encephalomyelitis to better understand the role and pathogenesis of PoAstV3 infection. Nucleic acid was extracted from formalin-fixed paraffin-embedded (FFPE) tissue for reverse transcription quantitative polymerase chain reaction (RT-qPCR) testing for PoAstV3. In addition, 3 cases with confirmed PoAstV3-associated disease were assayed by RT-qPCR to investigate PoAstV3 tissue distribution. PoAstV3 was detected in central nervous system (CNS) tissue via RT-qPCR and in situ hybridization in 13 of 50 (26%) FFPE cases assayed. PoAstV3 was rarely detected in any tissues outside the CNS. Positive cases from the retrospective study included pigs in various production categories beginning in 2010, the earliest year samples were available. Based on these results, PoAstV3 appears to be a recurring putative cause of viral encephalomyelitis in swine that is rarely detected outside of the CNS at the time of clinical neurologic disease, unlike other common viral causes of neurologic disease in swine.
Asunto(s)
Infecciones por Astroviridae/veterinaria , Encefalomielitis/veterinaria , Mamastrovirus/aislamiento & purificación , Enfermedades de los Porcinos/virología , Animales , Infecciones por Astroviridae/patología , Infecciones por Astroviridae/virología , Encefalomielitis/patología , Encefalomielitis/virología , Femenino , Hibridación in Situ/veterinaria , Masculino , Mamastrovirus/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Estudios Retrospectivos , Porcinos , Enfermedades de los Porcinos/patologíaRESUMEN
Influenza A viruses in swine (IAV-S) circulating in the United States of America are phylogenetically and antigenically distinct. A human H3 hemagglutinin (HA) was introduced into the IAV-S gene pool in the late 1990s, sustained continued circulation, and evolved into five monophyletic genetic clades, H3 clades IV-A to -E, after 2009. Across these phylogenetic clades, distinct antigenic clusters were identified, with three clusters (cyan, red, and green antigenic cluster) among the most frequently detected antigenic phenotypes (Abente EJ, Santos J, Lewis NS, Gauger PC, Stratton J, et al. J Virol 90:8266-8280, 2016, https://doi.org/10.1128/JVI.01002-16). Although it was demonstrated that antigenic diversity of H3N2 IAV-S was associated with changes at a few amino acid positions in the head of the HA, the implications of this diversity for vaccine efficacy were not tested. Using antigenically representative H3N2 viruses, we compared whole inactivated virus (WIV) and live-attenuated influenza virus (LAIV) vaccines for protection against challenge with antigenically distinct H3N2 viruses in pigs. WIV provided partial protection against antigenically distinct viruses but did not prevent virus replication in the upper respiratory tract. In contrast, LAIV provided complete protection from disease and virus was not detected after challenge with antigenically distinct viruses.IMPORTANCE Due to the rapid evolution of the influenza A virus, vaccines require continuous strain updates. Additionally, the platform used to deliver the vaccine can have an impact on the breadth of protection. Currently, there are various vaccine platforms available to prevent influenza A virus infection in swine, and we experimentally tested two: adjuvanted-whole inactivated virus and live-attenuated virus. When challenged with an antigenically distinct virus, adjuvanted-whole inactivated virus provided partial protection, while live-attenuated virus provided effective protection. Additional strategies are required to broaden the protective properties of inactivated virus vaccines, given the dynamic antigenic landscape of cocirculating strains in North America, whereas live-attenuated vaccines may require less frequent strain updates, based on demonstrated cross-protection. Enhancing vaccine efficacy to control influenza infections in swine will help reduce the impact they have on swine production and reduce the risk of swine-to-human transmission.
Asunto(s)
Hemaglutininas Virales/genética , Hemaglutininas Virales/inmunología , Subtipo H3N2 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/veterinaria , Vacunas Atenuadas/inmunología , Vacunas de Productos Inactivados/inmunología , Animales , Protección Cruzada/inmunología , Subtipo H3N2 del Virus de la Influenza A/genética , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Infecciones por Orthomyxoviridae/virología , Sistema Respiratorio/inmunología , Sistema Respiratorio/virología , Porcinos , Replicación Viral/inmunologíaRESUMEN
BACKGROUND: Influenza A Virus (IAV) causes respiratory disease in swine and is a zoonotic pathogen. Uncontrolled IAV in swine herds not only affects animal health, it also impacts production through increased costs associated with treatment and prevention efforts. The Iowa State University Veterinary Diagnostic Laboratory (ISU VDL) diagnoses influenza respiratory disease in swine and provides epidemiological analyses on samples submitted by veterinarians. DESCRIPTION: To assess the incidence of IAV in swine and inform stakeholders, the ISU FLUture website was developed as an interactive visualization tool that allows the exploration of the ISU VDL swine IAV aggregate data in the clinical diagnostic database. The information associated with diagnostic cases has varying levels of completeness and is anonymous, but minimally contains: sample collection date, specimen type, and IAV subtype. Many IAV positive samples are sequenced, and in these cases, the hemagglutinin (HA) sequence and genetic classification are completed. These data are collected and presented on ISU FLUture in near real-time, and more than 6,000 IAV positive diagnostic cases and their epidemiological and evolutionary information since 2003 are presented to date. The database and web interface provides rapid and unique insight into the trends of IAV derived from both large- and small-scale swine farms across the United States of America. CONCLUSION: ISU FLUture provides a suite of web-based tools to allow stakeholders to search for trends and correlations in IAV case metadata in swine from the ISU VDL. Since the database infrastructure is updated in near real-time and is integrated within a high-volume veterinary diagnostic laboratory, earlier detection is now possible for emerging IAV in swine that subsequently cause vaccination and control challenges. The access to real-time swine IAV data provides a link with the national USDA swine IAV surveillance system and allows veterinarians to make objective decisions regarding the management and control of IAV in swine. The website is publicly accessible at http://influenza.cvm.iastate.edu .
Asunto(s)
Virus de la Influenza A/clasificación , Internet , Laboratorios/normas , Técnicas de Diagnóstico Molecular/métodos , Infecciones por Orthomyxoviridae/veterinaria , Enfermedades de los Porcinos/diagnóstico , Enfermedades de los Porcinos/virología , Animales , Virus de la Influenza A/genética , Virus de la Influenza A/aislamiento & purificación , Infecciones por Orthomyxoviridae/epidemiología , Infecciones por Orthomyxoviridae/virología , Filogenia , Porcinos , Enfermedades de los Porcinos/epidemiología , Estados Unidos/epidemiologíaRESUMEN
Repeated spillovers of the H1N1 pandemic virus (H1N1pdm09) from humans to pigs resulted in substantial evolution of influenza A viruses infecting swine, contributing to the genetic and antigenic diversity of influenza A viruses (IAV) currently circulating in swine. The reassortment with endemic swine viruses and maintenance of some of the H1N1pdm09 internal genes resulted in the circulation of different genomic constellations in pigs. Here, we performed a whole-genome phylogenetic analysis of 368 IAV circulating in swine from 2009 to 2016 in the United States. We identified 44 different genotypes, with the most common genotype (32.33%) containing a clade IV-A HA gene, a 2002-lineage NA gene, an M-pdm09 gene, and remaining gene segments of triple reassortant internal gene (TRIG) origin. To understand how different genetic constellations may relate to viral fitness, we compared the pathogenesis and transmission in pigs of six representative genotypes. Although all six genotypes efficiently infected pigs, they resulted in different degrees of pathology and viral shedding. These results highlight the vast H3N2 genetic diversity circulating in U.S. swine after 2009. This diversity has important implications in the control of this disease by the swine industry, as well as a potential risk for public health if swine-adapted viruses with H1N1pdm09 genes have an increased risk to humans, as occurred in the 2011-2012 and 2016 human variant H3N2v cases associated with exhibition swine. IMPORTANCE: People continue to spread the 2009 H1N1 pandemic (H1N1pdm09) IAV to pigs, allowing H1N1pdm09 to reassort with endemic swine IAV. In this study, we determined the 8 gene combinations of swine H3N2 IAV detected from 2009 to 2016. We identified 44 different genotypes of H3N2, the majority of which contained at least one H1N1pdm09 gene segment. We compared six representative genotypes of H3N2 in pigs. All six genotypes efficiently infected pigs, but they resulted in different degrees of lung damage and viral shedding. These results highlight the vast genetic diversity of H3N2 circulating in U.S. swine after 2009, with important implications for the control of IAV for the swine industry. Because H1N1pdm09 is also highly adapted to humans, these swine viruses pose a potential risk to public health if swine-adapted viruses with H1N1pdm09 genes also have an increased risk for human infection.
Asunto(s)
Variación Genética , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/genética , Infecciones por Orthomyxoviridae/virología , Virus Reordenados , Enfermedades de los Porcinos/virología , Animales , Reacciones Cruzadas/inmunología , Genoma Viral , Genotipo , Pruebas de Inhibición de Hemaglutinación , Subtipo H1N1 del Virus de la Influenza A/clasificación , Subtipo H3N2 del Virus de la Influenza A/clasificación , Infecciones por Orthomyxoviridae/epidemiología , Infecciones por Orthomyxoviridae/transmisión , Filogenia , ARN Viral , Porcinos , Enfermedades de los Porcinos/epidemiología , Estados Unidos/epidemiología , Esparcimiento de VirusRESUMEN
BACKGROUND: Emergence and re-emergence of porcine epidemic diarrhea virus (PEDV) in North America, Asia and Europe has caused severe economic loss to the global swine industry. However, the virome of PEDV infected pigs and its effect on disease severity remains unknown. The advancements of sequencing technology have made it possible to characterize the entire microbiome of different body sites for any host. METHODS: The objective of this study was to characterize the RNA virome in PEDV-positive pigs using the hypothesis-free metagenomics approach based on next-generation sequencing. Specifically, 217 PEDV-positive swine fecal swab samples collected from diarrheic piglets over 17 US states during 2015-2016 were analyzed. RESULTS: A Kraken algorithm-based bioinformatics analysis revealed the presence of up to 9 different RNA genera besides PEDV (Alphacoronavirus genus), including Mamastrovirus (52%, 113/217), Enterovirus (39%, 85/217), Sapelovirus (31%, 67/217), Posavirus (30%, 66/217), Kobuvirus (23%, 49/217), Sapovirus (13%, 28/217), Teschovirus (10%, 22/217), Pasivirus (9%, 20/217), and Deltacoronavirus (3%, 6/217). There were 58 out of 217 piglets (27%) have PEDV infection alone whereas the remaining 159 (73%) shed 2 up to 9 different viruses. CONCLUSION: These findings demonstrated that PEDV infected diarrheic pigs had an extensive RNA viral flora consisting of four different families: Astroviridae, Picornaviridae, Caliciviridae, and Coronaviridae.
Asunto(s)
Astroviridae/genética , Caliciviridae/genética , Coronaviridae/genética , Infecciones por Coronavirus/veterinaria , Picornaviridae/genética , Virus de la Diarrea Epidémica Porcina/genética , Enfermedades de los Porcinos/epidemiología , Algoritmos , Secuencia de Aminoácidos , Animales , Astroviridae/clasificación , Astroviridae/aislamiento & purificación , Caliciviridae/clasificación , Caliciviridae/aislamiento & purificación , Coinfección , Biología Computacional , Coronaviridae/clasificación , Coronaviridae/aislamiento & purificación , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/virología , Granjas , Heces/virología , Secuenciación de Nucleótidos de Alto Rendimiento , Metagenómica/métodos , Filogenia , Picornaviridae/clasificación , Picornaviridae/aislamiento & purificación , Virus de la Diarrea Epidémica Porcina/clasificación , Virus de la Diarrea Epidémica Porcina/aislamiento & purificación , ARN Viral/genética , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Porcinos , Enfermedades de los Porcinos/virología , Estados Unidos/epidemiologíaRESUMEN
In 2013, PED emerged for the first time in the United States (US). The porcine epidemic diarrhea virus (PEDV) spread quickly throughout North America. Infection with PEDV causes watery diarrhea and up to 100% mortality in piglets, particularly for highly pathogenic non-InDel strains circulating in the US. PEDV is mainly transmitted by the fecal-oral route. Transmission via the venereal route has been suspected but not previously investigated. The aim of the study was to determine if PEDV could be detected in semen from infected specific pathogen-free (SPF) boars inoculated with a PEDV US non-InDel strain suggesting venereal transmission may occur. Two boars orally inoculated with PEDV showed clinical signs and virus shedding in feces. Transient presence of the PEDV genome was detected by RT-qPCR in the seminal (5.06 × 102 to 2.44 × 103 genomic copies/mL) and sperm-rich fraction of semen (5.64 × 102 to 3.40 × 104 genomic copies/mL) and a longer duration of viral shedding was observed in the sperm-rich fraction. The evidence of PEDV shedding in semen raises new questions in term of disease spread within the pig population with the use of potentially contaminated semen.
Asunto(s)
Infecciones por Coronavirus/veterinaria , Virus de la Diarrea Epidémica Porcina/fisiología , Enfermedades de los Porcinos/virología , Esparcimiento de Virus , Animales , Infecciones por Coronavirus/virología , Masculino , Semen , Organismos Libres de Patógenos Específicos , PorcinosRESUMEN
Hepatitis E virus (HEV) is a nonenveloped, positive-sense, single-stranded RNA virus that has been detected in a wide variety of animals. In 2017, an avian-like HEV was identified in sparrow feces sampled from around a pig farm in the midwestern United States. Sequence analysis revealed that the sparrow isolate represents a novel HEV that is distantly related to chicken and little egret HEVs.
Asunto(s)
Enfermedades de las Aves/virología , Virus de la Hepatitis E/aislamiento & purificación , Hepatitis E/veterinaria , Gorriones/virología , Animales , Pollos/virología , Heces/virología , Genómica , Hepatitis E/virología , Virus de la Hepatitis E/clasificación , Virus de la Hepatitis E/genética , Filogenia , Enfermedades de las Aves de Corral/virología , Estados UnidosRESUMEN
This study investigated the pathogenicity and transmissibility of a reverse-genetics-derived highly pathogenic avian influenza (HPAI) H5N1 lineage influenza A virus that was isolated from a human, A/Iraq/755/06. We also examined surface gene reassortant viruses composed of the haemagglutinin and neuraminidase from A/Iraq/755/06 and the internal genes of a 2009 pandemic H1N1 virus, A/New York/18/2009 (2Iraq/06 : 6NY/09 H5N1), and haemagglutinin and neuraminidase from A/New York/18/2009 with the internal genes of A/Iraq/755/06 (2NY/09 : 6Iraq/06 H1N1). The parental A/Iraq/755/06 caused little to no lesions in swine, limited virus replication was observed in the upper respiratory and lower respiratory tracts and transmission was detected in 3/5 direct-contact pigs based on seroconversion, detection of viral RNA or virus isolation. In contrast, the 2Iraq/06 : 6NY/09 H5N1 reassortant caused mild lung lesions, demonstrated sustained virus replication in the upper and lower respiratory tracts and transmitted to all contacts (5/5). The 2NY/09 : 6Iraq/06 H1N1 reassortant also caused mild lung lesions, there was evidence of virus replication in the upper respiratory and lower respiratory tracts and transmission was detected in all contacts (5/5). These studies indicate that an HPAI-derived H5N1 reassortant with pandemic internal genes may be more successful in sustaining infection in swine and that HPAI-derived internal genes were marginally compatible with pandemic 2009 H1N1 surface genes. Comprehensive surveillance in swine is critical to identify a possible emerging HPAI reassortant in all regions with HPAI in wild birds and poultry and H1N1pdm09 in pigs or other susceptible hosts.
Asunto(s)
Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H1N1 del Virus de la Influenza A/fisiología , Subtipo H5N1 del Virus de la Influenza A/genética , Subtipo H5N1 del Virus de la Influenza A/fisiología , Virus Reordenados/genética , Virus Reordenados/fisiología , Replicación Viral , Animales , Humanos , Subtipo H1N1 del Virus de la Influenza A/aislamiento & purificación , Subtipo H5N1 del Virus de la Influenza A/aislamiento & purificación , Gripe Humana/virología , Infecciones por Orthomyxoviridae/patología , Infecciones por Orthomyxoviridae/veterinaria , Infecciones por Orthomyxoviridae/virología , Virus Reordenados/aislamiento & purificación , Sistema Respiratorio/patología , Sistema Respiratorio/virología , Genética Inversa , Porcinos , Enfermedades de los Porcinos/patología , Enfermedades de los Porcinos/virologíaRESUMEN
UNLABELLED: Influenza A virus (IAV) of the H3 subtype is an important respiratory pathogen that affects both humans and swine. Vaccination to induce neutralizing antibodies against the surface glycoprotein hemagglutinin (HA) is the primary method used to control disease. However, due to antigenic drift, vaccine strains must be periodically updated. Six of the 7 positions previously identified in human seasonal H3 (positions 145, 155, 156, 158, 159, 189, and 193) were also indicated in swine H3 antigenic evolution. To experimentally test the effect on virus antigenicity of these 7 positions, substitutions were introduced into the HA of an isogenic swine lineage virus. We tested the antigenic effect of these introduced substitutions by using hemagglutination inhibition (HI) data with monovalent swine antisera and antigenic cartography to evaluate the antigenic phenotype of the mutant viruses. Combinations of substitutions within the antigenic motif caused significant changes in antigenicity. One virus mutant that varied at only two positions relative to the wild type had a >4-fold reduction in HI titers compared to homologous antisera. Potential changes in pathogenesis and transmission of the double mutant were evaluated in pigs. Although the double mutant had virus shedding titers and transmissibility comparable to those of the wild type, it caused a significantly lower percentage of lung lesions. Elucidating the antigenic effects of specific amino acid substitutions at these sites in swine H3 IAV has important implications for understanding IAV evolution within pigs as well as for improved vaccine development and control strategies in swine. IMPORTANCE: A key component of influenza virus evolution is antigenic drift mediated by the accumulation of amino acid substitutions in the hemagglutinin (HA) protein, resulting in escape from prior immunity generated by natural infection or vaccination. Understanding which amino acid positions of the HA contribute to the ability of the virus to avoid prior immunity is important for understanding antigenic evolution and informs vaccine efficacy predictions based on the genetic sequence data from currently circulating strains. Following our previous work characterizing antigenic phenotypes of contemporary wild-type swine H3 influenza viruses, we experimentally validated that substitutions at 6 amino acid positions in the HA protein have major effects on antigenicity. An improved understanding of the antigenic diversity of swine influenza will facilitate a rational approach for selecting more effective vaccine components to control the circulation of influenza in pigs and reduce the potential for zoonotic viruses to emerge.
Asunto(s)
Anticuerpos Antivirales/inmunología , Antígenos Virales/inmunología , Flujo Genético , Glicoproteínas Hemaglutininas del Virus de la Influenza/inmunología , Virus de la Influenza A/inmunología , Sustitución de Aminoácidos , Animales , Antígenos Virales/genética , Pruebas de Inhibición de Hemaglutinación , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Virus de la Influenza A/genética , Pulmón/patología , Mutagénesis Sitio-Dirigida , Proteínas Mutantes/genética , Proteínas Mutantes/inmunología , Infecciones por Orthomyxoviridae/patología , Infecciones por Orthomyxoviridae/transmisión , Infecciones por Orthomyxoviridae/veterinaria , Infecciones por Orthomyxoviridae/virología , Genética Inversa , Porcinos , Enfermedades de los Porcinos/patología , Enfermedades de los Porcinos/virología , Virulencia , Esparcimiento de VirusRESUMEN
At least two genetically different porcine epidemic diarrhoea virus (PEDV) strains have been identified in the USA: US PEDV prototype and S-INDEL-variant strains. The objective of this study was to compare the pathogenicity differences of the US PEDV prototype and S-INDEL-variant strains in conventional neonatal piglets under experimental infections. Fifty PEDV-negative 5-day-old pigs were divided into five groups of ten pigs each and were inoculated orogastrically with three US PEDV prototype isolates (IN19338/2013, NC35140/2013 and NC49469/2013), an S-INDEL-variant isolate (IL20697/2014), and virus-negative culture medium, respectively, with virus titres of 104 TCID50 ml- 1, 10âml per pig. All three PEDV prototype isolates tested in this study, regardless of their phylogenetic clades, had similar pathogenicity and caused severe enteric disease in 5-day-old pigs as evidenced by clinical signs, faecal virus shedding, and gross and histopathological lesions. Compared with pigs inoculated with the three US PEDV prototype isolates, pigs inoculated with the S-INDEL-variant isolate had significantly diminished clinical signs, virus shedding in faeces, gross lesions in small intestines, caeca and colons, histopathological lesions in small intestines, and immunohistochemistry staining in ileum. However, the US PEDV prototype and the S-INDEL-variant strains induced similar viraemia levels in inoculated pigs. Whole genome sequences of the PEDV prototype and S-INDEL-variant strains were determined, but the molecular basis of virulence differences between these PEDV strains remains to be elucidated using a reverse genetics approach.
Asunto(s)
Infecciones por Coronavirus/veterinaria , Virus de la Diarrea Epidémica Porcina/patogenicidad , Enfermedades de los Porcinos/virología , Animales , Animales Recién Nacidos , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/virología , Variación Genética , Virus de la Diarrea Epidémica Porcina/genética , ARN Viral , Porcinos , Enfermedades de los Porcinos/epidemiología , Estados Unidos/epidemiología , Virulencia , Esparcimiento de VirusRESUMEN
Multiple subtypes and many antigenic variants of influenza A virus (IAV) co-circulate in swine in the USA, complicating effective use of commercial vaccines to control disease and transmission. Whole inactivated virus (WIV) vaccines may provide partial protection against IAV with substantial antigenic drift, but have been shown to induce vaccine-associated enhanced respiratory disease (VAERD) when challenged with an antigenic variant of the same haemagglutinin (HA) subtype. This study investigated the role the immune response against HA, neuraminidase (NA) and nucleoprotein (NP) may play in VAERD by reverse engineering vaccine and challenge viruses on a common backbone and using them in a series of vaccination/challenge trials. Mismatched HA between vaccine and challenge virus was necessary to induce VAERD. However, vaccines containing a matched NA abrogated the VAERD phenomenon induced by the HA mismatch and this was correlated with NA-inhibiting (NI) antibodies. Divergence between the two circulating swine N2 lineages (92 % identity) resulted in a loss of NI cross-reactivity and also resulted in VAERD with the mismatched HA. The NP lineage selected for use in the WIV vaccine strains did not affect protection or pathology. Thus the combination of HA and NA in the vaccine virus strains played a substantial role in vaccine protection versus immunopathology, suggesting that vaccines that target the HA protein alone could be more prone to VAERD due to the absence of cross-protective NI antibodies.
Asunto(s)
Subtipo H1N1 del Virus de la Influenza A/inmunología , Subtipo H1N2 del Virus de la Influenza A/inmunología , Subtipo H3N2 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/efectos adversos , Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/virología , Enfermedades de los Porcinos/virología , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Glicoproteínas Hemaglutininas del Virus de la Influenza/inmunología , Neuraminidasa/inmunología , Nucleoproteínas/inmunología , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Sistema Respiratorio/virología , Porcinos , Enfermedades de los Porcinos/inmunología , Enfermedades de los Porcinos/prevención & control , Vacunación , Vacunas de Productos Inactivados/efectos adversos , Vacunas de Productos Inactivados/inmunologíaRESUMEN
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.
Asunto(s)
Virus de la Parainfluenza 1 Humana/aislamiento & purificación , Infecciones por Paramyxoviridae/veterinaria , Infecciones del Sistema Respiratorio/veterinaria , Enfermedades de los Porcinos/epidemiología , Enfermedades de los Porcinos/virología , Animales , Análisis por Conglomerados , Genoma Viral , Histocitoquímica , Datos de Secuencia Molecular , Mucosa Nasal/patología , Mucosa Nasal/virología , Infecciones por Paramyxoviridae/epidemiología , Infecciones por Paramyxoviridae/virología , Filogenia , ARN Viral/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Infecciones del Sistema Respiratorio/epidemiología , Infecciones del Sistema Respiratorio/virología , Análisis de Secuencia de ADN , Homología de Secuencia , Porcinos , Tráquea/patología , Tráquea/virología , Estados Unidos/epidemiología , Virología/métodosRESUMEN
UNLABELLED: Human-like swine H3 influenza A viruses (IAV) were detected by the USDA surveillance system. We characterized two novel swine human-like H3N2 and H3N1 viruses with hemagglutinin (HA) genes similar to those in human seasonal H3 strains and internal genes closely related to those of 2009 H1N1 pandemic viruses. The H3N2 neuraminidase (NA) was of the contemporary human N2 lineage, while the H3N1 NA was of the classical swine N1 lineage. Both viruses were antigenically distant from swine H3 viruses that circulate in the United States and from swine vaccine strains and also showed antigenic drift from human seasonal H3N2 viruses. Their pathogenicity and transmission in pigs were compared to those of a human H3N2 virus with a common HA ancestry. Both swine human-like H3 viruses efficiently infected pigs and were transmitted to indirect contacts, whereas the human H3N2 virus did so much less efficiently. To evaluate the role of genes from the swine isolates in their pathogenesis, reverse genetics-generated reassortants between the swine human-like H3N1 virus and the seasonal human H3N2 virus were tested in pigs. The contribution of the gene segments to virulence was complex, with the swine HA and internal genes showing effects in vivo. The experimental infections indicate that these novel H3 viruses are virulent and can sustain onward transmission in pigs, and the naturally occurring mutations in the HA were associated with antigenic divergence from H3 IAV from humans and swine. Consequently, these viruses could have a significant impact on the swine industry if they were to cause more widespread outbreaks, and the potential risk of these emerging swine IAV to humans should be considered. IMPORTANCE: Pigs are important hosts in the evolution of influenza A viruses (IAV). Human-to-swine transmissions of IAV have resulted in the circulation of reassortant viruses containing human-origin genes in pigs, greatly contributing to the diversity of IAV in swine worldwide. New human-like H3N2 and H3N1 viruses that contain a mix of human and swine gene segments were recently detected by the USDA surveillance system. The human-like viruses efficiently infected pigs and resulted in onward airborne transmission, likely due to the multiple changes identified between human and swine H3 viruses. The human-like swine viruses are distinct from contemporary U.S. H3 swine viruses and from the strains used in swine vaccines, which could have a significant impact on the swine industry due to a lack of population immunity. Additionally, public health experts should consider an appropriate assessment of the risk of these emerging swine H3 viruses for the human population.
Asunto(s)
Subtipo H3N2 del Virus de la Influenza A/inmunología , Subtipo H3N2 del Virus de la Influenza A/patogenicidad , Infecciones por Orthomyxoviridae/transmisión , Virus Reordenados/inmunología , Virus Reordenados/patogenicidad , Enfermedades de los Porcinos/transmisión , Animales , Antígenos Virales/genética , Antígenos Virales/inmunología , Línea Celular , Reacciones Cruzadas/inmunología , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Humanos , Subtipo H3N2 del Virus de la Influenza A/genética , Gripe Humana/virología , Neuraminidasa/clasificación , Neuraminidasa/inmunología , Infecciones por Orthomyxoviridae/virología , Virus Reordenados/genética , Porcinos/virología , Enfermedades de los Porcinos/virología , Estados UnidosRESUMEN
BACKGROUND: At least two genetically different porcine epidemic diarrhea virus (PEDV) strains have been identified in the United States (U.S. PEDV prototype and S-INDEL-variant strains). The current serological assays offered at veterinary diagnostic laboratories for detection of PEDV-specific antibody are based on the U.S. PEDV prototype strain. The objectives of this study were: 1) isolate the U.S. PEDV S-INDEL-variant strain in cell culture; 2) generate antisera against the U.S. PEDV prototype and S-INDEL-variant strains by experimentally infecting weaned pigs; 3) determine if the various PEDV serological assays could detect antibodies against the U.S. PEDV S-INDEL-variant strain and vice versa. RESULTS: A U.S. PEDV S-INDEL-variant strain was isolated in cell culture in this study. Three groups of PEDV-negative, 3-week-old pigs (five pigs per group) were inoculated orally with a U.S. PEDV prototype isolate (previously isolated in our lab), an S-INDEL-variant isolate or virus-negative culture medium. Serum samples collected at 0, 7, 14, 21 and 28 days post inoculation were evaluated by the following PEDV serological assays: 1) indirect fluorescent antibody (IFA) assays using the prototype and S-INDEL-variant strains as indicator viruses; 2) virus neutralization (VN) tests against the prototype and S-INDEL-variant viruses; 3) PEDV prototype strain whole virus based ELISA; 4) PEDV prototype strain S1-based ELISA; and 5) PEDV S-INDEL-variant strain S1-based ELISA. The positive antisera against the prototype strain reacted to and neutralized both prototype and S-INDEL-variant viruses, and the positive antisera against the S-INDEL-variant strain also reacted to and neutralized both prototype and S-INDEL-variant viruses, as examined by IFA antibody assays and VN tests. Antibodies against the two PEDV strains could be detected by all three ELISAs although detection rates varied to some degree. CONCLUSIONS: These data indicate that the antibodies against U.S. PEDV prototype and S-INDEL-variant strains cross-reacted and cross-neutralized both strains in vitro. The current serological assays based on U.S. PEDV prototype strain can detect antibodies against both U.S. PEDV strains.