RESUMO
Porcine epidemic diarrhea virus (PEDV) has affected the pork industry worldwide and during outbreaks the mortality of piglets has reached 100%. Lipid nanocarriers are commonly used in the development of immunostimulatory particles due to their biocompatibility and slow-release delivery properties. In this study, we developed a lipid nanoparticle (LNP) complex based on glycyrrhizinic acid (GA) and tested its efficacy as an adjuvant in mice immunized with the recombinant N-terminal domain (NTD) of porcine epidemic diarrhea virus (PEDV) spike (S) protein (rNTD-S). The dispersion stability analysis (Z-potential -27.6 mV) confirmed the size and charge stability of the LNP-GA, demonstrating that the particles were homogeneously dispersed and strongly anionic, which favors nanoparticles binding with the rNTD-S protein, which showed a slightly positive charge (2.11 mV) by in silico analysis. TEM image of LNP-GA revealed nanostructures with a spherical-bilayer lipid vesicle (~100 nm). The immunogenicity of the LNP-GA-rNTD-S complex induced an efficient humoral response 14 days after the first immunization (p < 0.05) as well as an influence on the cellular immune response by decreasing serum TNF-α and IL-1ß concentrations, which was associated with an anti-inflammatory effect.
Assuntos
Infecções por Coronavirus , Lipossomos , Nanopartículas , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Vacinas Virais , Animais , Suínos , Camundongos , Anticorpos Antivirais , Vírus da Diarreia Epidêmica Suína/genética , Ácido Glicirrízico/farmacologia , Glicoproteína da Espícula de Coronavírus , Adjuvantes Imunológicos , Imunidade , Proteínas Recombinantes , LipídeosRESUMO
The characteristics of the whole PEDV genome that has circulated in Mexico from the first outbreak to the present are unknown. We chose samples obtained from 2013 to 2017 and sequenced them, which enabled us to identify the genetic variation and phylogeny in the virus during the first four years that it circulated in Mexico. A 99% identity was found among the analyzed pandemic strains; however, the 1% difference affected the structure of the S glycoprotein, which is essential for the binding of the virus to the cellular receptor. The S protein induces the most efficacious antibodies; hence, these changes in structure could be implicated in the clinical antecedents of the outbreaks. Antigenic changes could also help PEDV avoid neutralization, even in the presence of previous immunity. The characterization of the complete genome enabled the identification of three circulating strains that have a deletion in ORF1a, which is present in attenuated Asian vaccine strains. The phylogenetic analysis of the complete genome indicates that the first PEDV outbreaks in Mexico were caused by INDEL strains and pandemic strains related to USA strains; however, the possibility of the entry of European strains exists, which may have caused the 2015 and 2016 outbreaks.
Assuntos
Infecções por Coronavirus , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Animais , Suínos , Vírus da Diarreia Epidêmica Suína/genética , Filogenia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , México/epidemiologia , Surtos de Doenças , Doenças dos Suínos/epidemiologia , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/química , DiarreiaRESUMO
Porcine epidemic diarrhea virus (PEDV) is a virus that can cause diarrhea in pigs, resulting in significant economic losses to the pig industry. The mutation of the virus and its co-infection with other enteroviruses leads to poor control of PEDV infection. In this study, we found that the diarrhea outbreak in a pig farm in Shandong Province was mainly caused by PEDV infection. Through high-throughput sequencing, we also detected one other diarrhea-related virus (porcine kobuvirus). In the phylogenetic analysis and molecular characterization of the detected PEDV S gene and PKV, it was found that the S gene of the PEDV strain detected in this study (named SD22-2) had more mutations than the CV777 strain. The highest homology between PKV (named SD/2022/China) detected in this study and other strains was only 89.66%. Based on polyprotein, we divided SD/2022/China strains into a new grouping (designated group 4) and detected recombination signals. In summary, SD22-2 detected in this study is a new PEDV variant strain, and SD/2022/China strain might be a novel PKV strain. We also found the co-infection of the new PEDV variant and the novel PKV isolated from piglets with diarrhea. Our data suggested the importance of continuous surveillance of PEDV and PKV.
Assuntos
Coinfecção , Infecções por Coronavirus , Kobuvirus , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Animais , Suínos , Filogenia , Vírus da Diarreia Epidêmica Suína/genética , Kobuvirus/genética , Infecções por Coronavirus/epidemiologia , Diarreia/epidemiologia , China/epidemiologiaRESUMO
Porcine epidemic diarrhoea virus (PEDV) infects pigs of all ages causing vomiting and diarrhoea. PEDV is transmitted via the oral-faecal route, and a very low dose is enough to infect susceptible pigs, resulting in significant production losses. This short communication aims to describe the introduction of PEDV into a 10,000-sow farrow-to-wean farm located in northwest Mexico. Following the onset of clinical signs, an outbreak investigation was conducted to determine the most probable route of introduction. Based on data collected from interviews, construction of a timeline of events, and the detection of PEDV RNA in feed samples and samples collected from various surfaces of feed transport vehicles, it was concluded that the most probable route for PEDV incursion into this breeding herd was contaminated feed or a contaminated feed transport vehicle. This paper describes how feed or feed transport could serve as potential routes of PEDV infection to a farm and highlights the importance of establishing biosecurity programs to mitigate these risks.
Assuntos
Ração Animal/virologia , Infecções por Coronavirus , Contaminação de Alimentos , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Animais , Biosseguridade , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/transmissão , Infecções por Coronavirus/veterinária , Diarreia/epidemiologia , Diarreia/veterinária , Surtos de Doenças/veterinária , Feminino , México/epidemiologia , Vírus da Diarreia Epidêmica Suína/genética , Suínos , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/transmissão , Doenças dos Suínos/virologiaRESUMO
Swine enteric viral infections are responsible for substantial economic losses in the pork industry worldwide. Porcine epidemic diarrhea (PEDV) is one of the main causative agents of diarrhea in lactating pigs, and reports of PEDV coinfection with other enteric viruses highlight the importance of viral interactions for disease presentation and outcomes. Using next-generation sequencing (NGS) and sequence analyses from samples taken from piglets with acute diarrhea, we explored the possible interactions between PEDV and other less reported pathogens. PEDV coinfection with porcine kobuvirus (PKV) was detected in 36.4% (27/74) of samples. Full genomes from porcine coronavirus and kobuvirus were obtained, as was a partial porcine sapovirus genome (PSaV). The phylogenetic results show the clustering of these strains corresponding to the geographical relationship. To our knowledge, this is the first full genome and isolation report for porcine kobuvirus in México, as well as the first phylogenetic analysis for porcine sapovirus in the country. The NGS approach provides a better perspective of circulating viruses and other pathogens in affected production units.
Assuntos
Coinfecção/virologia , Infecções por Coronavirus/virologia , Kobuvirus/genética , Kobuvirus/isolamento & purificação , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/isolamento & purificação , Animais , Coinfecção/epidemiologia , Infecções por Coronavirus/epidemiologia , Diarreia/virologia , Fezes/virologia , Genoma Viral , Kobuvirus/classificação , México/epidemiologia , Técnicas de Diagnóstico Molecular , Filogenia , Vírus da Diarreia Epidêmica Suína/classificação , Sapovirus/genética , Análise de Sequência , Suínos , Doenças dos Suínos/virologiaRESUMO
Porcine epidemic diarrhea virus (PEDV) is a significant global, enteric coronavirus in pigs and was first reported in Colombia in 2014. However, the epidemiology, genetic and antigenic characteristics of the virus have yet to be investigated. In this study, we investigated the dissemination of PEDV by testing 536 samples by RT-PCR over a 33-month period. The 35.8% of positive samples (n = 192) was significantly different (p < .01) between months over time, with a higher number of positives samples occurring at the beginning of the epidemic and during the second epidemic wave within the main pork producing region. The complete PEDV genomes were generated for 21 strains, which shared a high nucleotide and amino acid sequence identity, except for the spike (S) gene. Recombinant regions were identified within the Colombian strains and between Colombian and Asian PEDV strains. Phylogenetic analysis of the 21 Colombian strains demonstrated the presence of 7 lineages that shared common ancestors with PEDV strains from the United States. Moreover, the antigenic analysis demonstrated residue differences in the neutralizing epitopes in the spike and nucleocapsid proteins. Our results illustrated the simultaneous introduction of the two PEDV genotypes (GIIa American pandemic and S-INDEL) into the Colombian swine industry during the 2014 PEDV epidemic and enhanced our understanding of the epidemiology and molecular diversity of PEDV in Colombia.
Assuntos
Infecções por Coronavirus , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Animais , Colômbia/epidemiologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , Epidemias/veterinária , Filogenia , Vírus da Diarreia Epidêmica Suína/genética , Suínos , Doenças dos Suínos/epidemiologia , Estados UnidosRESUMO
As an emerging disease, the porcine epidemic diarrhoea virus has caused substantial economic losses to the pork industry in Mexico, leading to piglet mortality rates of up to 100%. For detection, sequencing and genetic characterization of the virus, 68 samples of one-week-old piglets from pork farms in 17 states of Mexico were analysed. In total, 53 samples were positive by real-time RT-PCR, confirming the presence of the virus in 15 states. Twenty-eight samples from 10 states were amplified by endpoint RT-PCR, and 20 sequences of the spike gene were obtained. A phylogenetic analysis based on the spike gene demonstrated that all Mexican strains are in Group II and are classified as non-Indel-S emerging variants. Three strains showed amino acid insertions: PEDv/MEX/GTO/LI-DMZC15/2015 and PEDv/MEX/QRO/LI-DMZC45/2016 showed one amino acid insertion (424 Y425 and 447 D448 , respectively), and PEDv/MEX/QRO/LI-DMZC49/2019 showed one and two amino acid insertions (422 C423 and 537 SQ538 ), with the second insertion in the COE region. These results provide evidence of the prevalence of emerging, non-Indel-S strains of the virus are currently circulating in Mexico during 2016-2018, when three of which have amino acid insertions: PEDv/MEX/GTO/IN-DMZC15/2015 and PEDv/MEX/QRO/IN-DMZC45/2016 have one amino acid insertion each (424 Y425 and 447 D448 , respectively), and PEDv/MEX/QRO/IN-DMZC49/2019 has one (422 C423 ) and two amino acid insertions (537 SQ538 ), the latter being in the COE region, which could generate new antigenic variants.
Assuntos
Infecções por Coronavirus/veterinária , Variação Genética , Vírus da Diarreia Epidêmica Suína/genética , Substituição de Aminoácidos , Animais , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Fazendas , Geografia , México/epidemiologia , Filogenia , Vírus da Diarreia Epidêmica Suína/isolamento & purificação , SuínosRESUMO
Porcine deltacoronavirus has caused great economic losses in the swine industry worldwide. In this study, we carried out the first detection, sequencing and characterization of this virus in Mexico. We analysed 885 rectal samples by multiplex RT-PCR to determine coinfections. In addition, the Spike gene was amplified, sequenced and analysed phylogenetically. We found 85 positive samples for porcine deltacoronavirus, representing 9.6% of the total samples, and we determined that the most frequent coinfection was with porcine epidemic diarrhoea virus (54.1%). Four sequences of Mexican isolates were most closely related to those of the United States. The antigenic regions and the glycosylation site of the strains obtained coincide with those previously reported. This relationship is probably related to the commercial exchange of pigs between the US and Mexico and the geographical proximity of these two countries.
Assuntos
Infecções por Coronavirus/veterinária , Vírus da Diarreia Epidêmica Suína/isolamento & purificação , Glicoproteína da Espícula de Coronavírus/análise , Doenças dos Suínos/epidemiologia , Animais , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , México/epidemiologia , Vírus da Diarreia Epidêmica Suína/genética , Prevalência , Suínos , Doenças dos Suínos/virologiaRESUMO
Porcine rotavirus (PoRV) and porcine epidemic diarrhea virus (PEDV) usually co-infect pigs in modern large-scale piggery, which both can cause severe diarrhea in newborn piglets and lead to significant economic losses to the pig industry. The VP7 protein is the main coat protein of PoRV, and the S protein is the main structural protein of PEDV, which are capable of inducing neutralizing antibodies in vivo. In this study, a DNA vaccine pPI-2.EGFP.VP7.S co-expressing VP7 protein of PoRV and S protein of PEDV was constructed. Six 8-week-old mice were immunized with the recombinant plasmid pPI-2.EGFP.VP7.S. The high humoral immune responses (virus specific antibody) and cellular immune responses (IFN-γ, IL-4, and spleen lymphocyte proliferation) were evaluated. The immune effect through intramuscular injection increased with plasmid dose when compared with subcutaneous injection. The immune-enhancing effect of IFN-α adjuvant was excellent compared with pig spleen transfer factor and IL-12 adjuvant. These results demonstrated that pPI-2.EGFP.VP7.S possess the immunological functions of the VP7 proteins of PoRV and S proteins of PEDV, indicating that pPI-2.EGFP.VP7.S is a candidate vaccine for porcine rotaviral infection (PoR) and porcine epidemic diarrhea (PED).
Assuntos
Antígenos Virais/imunologia , Proteínas do Capsídeo/imunologia , Infecções por Coronavirus/veterinária , Plasmídeos/imunologia , Infecções por Rotavirus/veterinária , Rotavirus/imunologia , Doenças dos Suínos/prevenção & controle , Proteínas Virais de Fusão/imunologia , Vacinas Virais/imunologia , Animais , Antígenos Virais/administração & dosagem , Antígenos Virais/genética , Proteínas do Capsídeo/administração & dosagem , Proteínas do Capsídeo/genética , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/virologia , DNA Recombinante/administração & dosagem , DNA Recombinante/genética , DNA Recombinante/imunologia , Avaliação Pré-Clínica de Medicamentos , Camundongos , Plasmídeos/administração & dosagem , Plasmídeos/genética , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/imunologia , Rotavirus/genética , Infecções por Rotavirus/imunologia , Infecções por Rotavirus/prevenção & controle , Infecções por Rotavirus/virologia , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/virologia , Proteínas Virais de Fusão/administração & dosagem , Proteínas Virais de Fusão/genética , Vacinas Virais/administração & dosagem , Vacinas Virais/genéticaRESUMO
From the severe porcine epidemic diarrhoea (PED) epidemics that struck in 2013 in the United States of America and other countries of North and South America, two types of porcine epidemic diarrhoea virus (PEDV) were isolated, namely the InDel and the non-InDel strains. They are differentiated by insertions/deletions in the S1 nucleotide sequence of the S gene, and differences in virulence were observed from the clinical cases. In 2014, a PED outbreak occurred in a pig farm in France, from which an InDel strain was isolated. This study aimed at comparing, under experimental conditions, the pathogenicity and the direct and indirect transmissions between a non-InDel strain isolated from a PED-affected piglet in 2014 in the USA and the French InDel strain. All infected pigs showed clinical signs with the non-InDel strain although only the inoculated and direct contact pigs showed clinical signs in the InDel strain group. Although viral RNA was detected in air samples with both strains, the indirect contact pigs remained free from infection with the InDel strain in contrast to the non-InDel group in which airborne transmission occurred in the indirect contact pigs. All infected pigs shed virus in faeces regardless of PEDV strain with 9 of 30 pigs showing intermittent faecal shedding. The transmission rate by direct contact was found to be 2.17-fold higher than the non-InDel strain compared with the InDel. In conclusion, the InDel strain was less pathogenic than the non-InDel strain in our experimental conditions. The transmission route differed between the two strains. Direct contact was the main transmission route for the InDel strain, although the non-InDel strain was transmitted through direct contact and indirectly through the air.