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1.
Arch Virol ; 169(9): 179, 2024 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-39150476

RESUMO

Seven novel porcine parvoviruses (PPV2 to PPV8) have been discovered in the last two decades. The last one reported was PPV8 in China in 2022, which was proposed to be a member of the genus Protoparvovirus. Here, we report the first detection of PPV8 outside China - in two provinces from Colombia. Six out of 146 (4.1%) pigs showing porcine respiratory disease (PRD) tested positive for PPV8. Sequencing and phylogenetic analysis of two Colombian PPV8 isolates (GenBank database accession numbers PP335559 and PP335560) showed them to be members of the genus Protoparvovirus. Furthermore, PPV8 was detected in coinfections with porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV), which are associated with PRD.


Assuntos
Infecções por Parvoviridae , Parvovirus Suíno , Doenças dos Suínos , Animais , Coinfecção/virologia , Coinfecção/veterinária , Coinfecção/epidemiologia , Colômbia/epidemiologia , Infecções por Parvoviridae/veterinária , Infecções por Parvoviridae/virologia , Infecções por Parvoviridae/epidemiologia , Parvovirus Suíno/genética , Parvovirus Suíno/isolamento & purificação , Parvovirus Suíno/classificação , Filogenia , Síndrome Respiratória e Reprodutiva Suína/virologia , Síndrome Respiratória e Reprodutiva Suína/epidemiologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Vírus da Síndrome Respiratória e Reprodutiva Suína/classificação , Suínos , Doenças dos Suínos/virologia , Doenças dos Suínos/epidemiologia
2.
Biologicals ; 86: 101763, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38641502

RESUMO

This study aimed to investigate the prevalence of viral agents causing reproductive failure in pigs in Korea. In addition, two types of multiplex real-time PCR (mqPCR) were developed for the simultaneous detection of Aujeszky's disease virus (ADV) and porcine parvovirus (PPV) in mqPCR and encephalomyocarditis virus (EMCV) and Japanese encephalitis virus (JEV) in reverse transcription mqPCR (mRT-qPCR). A total of 150 aborted fetus samples collected from 2020 to 2022 were analyzed. Porcine reproductive and respiratory syndrome virus was the most prevalent (49/150 32.7%), followed by porcine circovirus type 2 (31/150, 20.7%), and PPV1 (7/150, 4.7%), whereas ADV, EMCV, and JEV were not detected. The newly developed mqPCR and mRT-qPCR could simultaneously detect and differentiate with high sensitivities and specificities. When applied to aborted fetuses, the newly developed mqPCR for PPV was 33.3% more sensitivities than the previously established diagnostic method. Amino acid analysis of the VP2 sequences of PPV isolates revealed considerable similarity to the highly pathogenic Kresse strain. This study successfully evaluated the prevalence of viral agents causing reproductive failure among swine in Korea, the developed mqPCR and mRT-qPCR methods could be utilized as effective and accurate diagnostic methods for the epidemiological surveillance of ADV, PPV, EMCV, and JEV.


Assuntos
Reação em Cadeia da Polimerase Multiplex , Doenças dos Suínos , Animais , Suínos , República da Coreia/epidemiologia , Doenças dos Suínos/virologia , Doenças dos Suínos/diagnóstico , Doenças dos Suínos/epidemiologia , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Prevalência , Feminino , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Gravidez , Parvovirus Suíno/genética , Parvovirus Suíno/isolamento & purificação , Aborto Animal/virologia , Aborto Animal/epidemiologia , Viroses/diagnóstico , Viroses/epidemiologia , Viroses/virologia
3.
Biotechnol Bioeng ; 118(8): 3251-3262, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34129733

RESUMO

Due to the high variation in viral surface properties, a platform method for virus purification is still lacking. A potential alternative to the high-cost conventional methods is aqueous two-phase systems (ATPSs). However, optimizing virus purification in ATPS requires a large experimental design space, and the optimized systems are generally found to operate at high ATPS component concentrations. The high concentrations capitalize on hydrophobic and electrostatic interactions to obtain high viral particle yields. This study investigated using osmolytes as driving force enhancers to reduce the high concentration of ATPS components while maintaining high yields. The partitioning behavior of porcine parvovirus (PPV), a nonenveloped mammalian virus, and human immunodeficiency virus-like particle (HIV-VLP), a yeast-expressed enveloped VLP, were studied in a polyethylene glycol (PEG) 12 kDa-citrate system. The partitioning of the virus modalities was enhanced by osmoprotectants glycine and betaine, while trimethylamine N-oxide was ineffective for PPV. The increased partitioning to the PEG-rich phase pertained only to viruses, resulting in high virus purification. Recoveries were 100% for infectious PPV and 92% for the HIV-VLP, with high removal of the contaminant proteins and more than 60% DNA removal when glycine was added. The osmolyte-induced ATPS demonstrated a versatile method for virus purification, irrespective of the expression system.


Assuntos
HIV-1/isolamento & purificação , Parvovirus Suíno/isolamento & purificação , Vírion/isolamento & purificação , Animais , Linhagem Celular , HIV-1/química , Humanos , Parvovirus Suíno/química , Suínos , Vírion/química
4.
Arch Virol ; 166(8): 2249-2254, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33999261

RESUMO

Porcine parvovirus 1 (PPV1) is a major cause of reproductive failure in pigs. To date, six additional porcine parvoviruses (PPV2-PPV7) have been identified. In this study, we detected 11 PPV1 strains, five PPV3 strains, three PPV4 strains, six PPV5 strains, five PPV6 strains, and one PPV7 strain in Korean wild boars. PPV1, -3, and -5, and PPV6 from Korean wild boars harbor conserved motifs within the Ca2+ binding loop and the catalytic center of the PLA1 motif. Intra-species recombination among PPV7 strains was also identified. Genetic characterization revealed that PPV1 from Korean wild boars may be similar to virulent PPV strains.


Assuntos
DNA Viral/genética , Infecções por Parvoviridae/virologia , Parvovirus Suíno/classificação , Sus scrofa/virologia , Substituição de Aminoácidos , Animais , Feminino , Técnicas de Genotipagem , Masculino , Parvovirus Suíno/genética , Parvovirus Suíno/isolamento & purificação , Filogenia , República da Coreia , Suínos
5.
Microb Pathog ; 141: 104027, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32007620

RESUMO

The clinical implications of recently discovered porcine circovirus 3 (PCV3) infections are still unknown. The potential role of this emerging virus in reproductive loss in swine has been described. Herein, we report a high prevalence of PCV3 in mummified fetuses from sows maintained in modern farms in Rio Grande do Sul, Santa Catarina, Paraná, Goiás, and Mato Grosso do Sul states, Brazil. For this analysis, 276 mummified fetuses from 11 commercial swine farms were included. The presence of PCV3 DNA was confirmed using PCR, and the complete sequence of five different viral strains was obtained. Sequences of PCV3 genomes available on GenBank were then used for phylogenetic tree construction. Of the 276 mummified fetuses examined, 270 (nearly 97%) were positive for PCV3. In 93.1% of the fetuses, co-infections with at least one of the following agents were identified: porcine parvovirus (PPV), porcine circovirus 2 (PCV2) and Leptospira spp. Twelve fetuses were positive for PCV3 alone. The amino acid sequence of the capsid gene for the five viral strains shared 98-100% homology among them. Analysis of the DNA sequence indicates that the viruses identified in this study belong to the PCV3a1 subgroup. In summary, PCV3 DNA was detected in mummified fetuses at a surprisingly high rate. The role of PCV3 in porcine circovirus-associated disease (PCVAD) is still uncertain. However, considering that PCV3 has been detected in a variety of conditions, even in healthy animals, the present results confirm the need to investigate PCV3 as a causative agent of fetal mummification in swine.


Assuntos
Circovirus/genética , Feto/virologia , Genoma Viral , Animais , Brasil/epidemiologia , Proteínas do Capsídeo/genética , Infecções por Circoviridae/epidemiologia , Infecções por Circoviridae/veterinária , Circovirus/classificação , Circovirus/patogenicidade , Coinfecção/epidemiologia , Coinfecção/veterinária , Fazendas , Leptospira/isolamento & purificação , Leptospirose/veterinária , Infecções por Parvoviridae/veterinária , Parvovirus Suíno/isolamento & purificação , Filogenia , Prevalência , Suínos , Doenças dos Suínos/virologia
6.
Microbiol Immunol ; 64(5): 366-376, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32096557

RESUMO

Worldwide, many emerging porcine parvoviruses (PPVs) have been linked to porcine circovirus-2 (PCV2) associated disease (PCVAD), which includes post-weaning multi-systemic wasting syndrome (PMWS), PCV2-related reproductive failure (PCV2-RF), as well as other syndromes. To determine the DNA prevalence of PPVs and their relationship with PMWS and PCV2-RF in Mexico, 170 formalin-fixed paraffin-embedded tissues were selected from archival collections to detect PPVs using a nested polymerase chain reaction. The tissues were composed of 50 PMWS cases, 20 age-matched tissues from healthy pigs, 56 PCV2-related reproductive failure (PCV2+ -RF) cases, and 44 PCV2- -RF cases. Overall, PPV2 and PPV6 were the most prevalent species (90.0% and 74.7%, respectively). In 8-11 week old pigs, the highest prevalence was for PPV6 and PPV3. Concerning reproductive failure, the PCV2-affected farms had a significantly higher prevalence for PPV6 (61.6%) and PPV5 (36.4%) than the PCV2-unaffected farms (35.0% and 5.0%, respectively). The concurrent infection rate was high, being significant for PPV2/PPV4 and PPV1/PPV5 within the PMWS cases and for PPV6/PPV5 among the PCV2+ -RF tissues. PPV5 showed a significant relationship with PMWS, whereas PPV5 and PPV6 were significant for PCVAD. The prevalence and coinfection rate of PPVs in Mexico were markedly higher than that described in other countries, denoting that PPV5 and PPV6 might have a potential role in PCVAD in Mexico. It is concluded that it is likely that the density population of pigs in Mexico is contributing to high PPV inter-species and PCV2 coinfections which might lead to a different pathogenic outcome.


Assuntos
Infecções por Circoviridae/veterinária , Circovirus/isolamento & purificação , Coinfecção , Infecções por Parvoviridae/veterinária , Parvovirus Suíno/isolamento & purificação , Doenças dos Suínos/virologia , Animais , Infecções por Circoviridae/epidemiologia , Infecções por Circoviridae/virologia , Circovirus/genética , Coinfecção/veterinária , Coinfecção/virologia , DNA Viral/isolamento & purificação , México , Infecções por Parvoviridae/epidemiologia , Infecções por Parvoviridae/virologia , Parvovirus Suíno/genética , Prevalência , Estudos Retrospectivos , Suínos/virologia , Doenças dos Suínos/epidemiologia
7.
BMC Vet Res ; 16(1): 57, 2020 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-32059673

RESUMO

BACKGROUND: PPV is one of the most important pathogens causing porcine reproductive disorder. It has been shown in clinical cases to be a commonly mixed infection with other important swine diseases which can aggravate the severity of the disease and bring serious economic losses to the pig industry. Serological methods, such as hemagglutination inhibition assays (HAI), serum neutralization (SN), and the modified direct complement-fixation (MDCF) test were utilized earlier, whereas the enzyme-linked immunosorbent assay (ELISA) is the most frequently applied assay to detect PPV-specific antibodies. RESULTS: We establish the visible protein chip and the cyanine dye 3 (Cy3)-labeled protein chip to detect the clinical serum from pigs. In this study, the recombinant protein VP2 of PPV was expressed in E.coli, purified with nickel magnetic beads, and then printed onto epoxy-coated glass slides for preparation of the protein chip. After a series of experiments, the conditions of antigen protein concentration, incubation time of primary antibody or secondary antibody, and optimal serum dilution fold were optimized, resulting in a successful visible protein chip and Cy3-labeled protein chip. The results showed that the positive serum, diluted up to 6000-fold, can be detected by the visible protein chip, and the positive serum, diluted up to 12,800-fold, can be detected by the Cy3-labeled protein chip, suggesting the high sensitivity of these protein chips. Moreover, the positive detection ratio, sensitivity, and specificity of these two kinds of protein chips were higher than those of commercial ELISA antibody detection kits. CONCLUSION: Overall, these two protein chips can be used to rapidly diagnose clinical samples with high throughput.


Assuntos
Anticorpos Antivirais/sangue , Dispositivos Lab-On-A-Chip/veterinária , Infecções por Parvoviridae/veterinária , Parvovirus Suíno/isolamento & purificação , Doenças dos Suínos/virologia , Animais , Dispositivos Lab-On-A-Chip/virologia , Infecções por Parvoviridae/diagnóstico , Infecções por Parvoviridae/virologia , Sensibilidade e Especificidade , Suínos , Doenças dos Suínos/sangue , Doenças dos Suínos/diagnóstico
8.
BMC Vet Res ; 16(1): 113, 2020 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-32295585

RESUMO

BACKGROUND: with the advantage of sequencing technology, many novel porcine parvoviruses (PPV) rather than PPV1 has been reported. This study ultilized specific PCR- based method and gene- based analysis to study the presence and genetic diversity of porcine parvoviruses in South Korea in 2018. RESULTS: The present study was conducted in 2018 and found PPV1 and PPV7 in nine out of 151 field samples (organs and semen) by the PCR method. Among these, the complete genome sequences of five strains (N2, N91, N108, N133, and N141) were recovered. Phylogenic analysis revealed that the strains N2, N91, and N108 belong to the PPV1 genotype, while N133 and N141 belong to PPV7 genotype. The PPV7 strains collected in this study had deletion mutations in the VP2 gene but differed from that of PPV7 strains collected in 2017. Among the PPV1 strains, the amino acid variations in the B cell epitopes of the VP2 protein were observed between three Korean PPV1 field strains (N2, N91, and N108) and the reference PPV1 strains. Those substitutions resulted in six out of 12 predicted epitopes having significant differences in antigenic index compared to the other PPV1 strains. CONCLUSIONS: This study confirmed the presence of different genotypes of porcine parvoviruses in South Korea. The PPVs circulating in South Korea were phylogenetically classified as PPV1 and PPV7 genotypes. Three Korean PPV1 strains collected in 2018 were predicted to have antigenic alteration in VP2 compared to several reference strains of PPV1.


Assuntos
Infecções por Parvoviridae/veterinária , Parvovirus Suíno/genética , Parvovirus Suíno/isolamento & purificação , Doenças dos Suínos/virologia , Animais , Epitopos de Linfócito B , Variação Genética , Genótipo , Infecções por Parvoviridae/epidemiologia , Infecções por Parvoviridae/virologia , Parvovirus Suíno/classificação , Filogenia , Reação em Cadeia da Polimerase/veterinária , República da Coreia/epidemiologia , Suínos , Doenças dos Suínos/epidemiologia
9.
J Nanobiotechnology ; 18(1): 7, 2020 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-31910833

RESUMO

BACKGROUND: Antibodies are an important reagent to determine the specificity and accuracy of diagnostic immunoassays for various diseases. However, traditional antibodies have several shortcomings due to their limited abundance, difficulty in permanent storage, and required use of a secondary antibody. Nanobodies, which are derived from single-chain camelid antibodies, can circumvent many of these limitations and, thus, appear to be a promising substitute. In the presented study, a sandwich ELISA-like immunoassay and direct fluorescent assay with high sensitivity, good specificity, and easy operation were the first time to develop for detecting porcine parvovirus (PPV). After screening PPV viral particles 2 (VP2) specific nanobodies, horseradish peroxidase (HRP) and enhanced green fluorescent protein (EGFP) fusions were derived from the nanobodies by recombinant technology. Finally, using the nanobody-HRP and -EGFP fusions as probes, the developed immunoassays demonstrate specific, sensitive, and rapid detection of PPV. RESULTS: In the study, five PPV-VP2 specific nanobodies screened from an immunised Bactrian camel were successfully expressed with the bacterial system and purified with a Ni-NTA column. Based on the reporter-nanobody platform, HRP and EGFP fusions were separately produced by transfection of HEK293T cells. A sandwich ELISA-like assay for detecting PPV in the samples was firstly developed using PPV-VP2-Nb19 as the capture antibody and PPV-VP2-Nb56-HRP fusions as the detection antibody. The assay showed 92.1% agreement with real-time PCR and can be universally used to surveil PPV infection in the pig flock. In addition, a direct fluorescent assay using PPV-VP2-Nb12-EGFP fusion as a probe was developed to detect PPV in ST cells. The assay showed 81.5% agreement with real-time PCR and can be used in laboratory tests. CONCLUSIONS: For the first time, five PPV-VP2 specific nanobody-HRP and -EGFP fusions were produced as reagents for developing immunoassays. A sandwich ELISA-like immunoassay using PPV-VP2-Nb19 as the capture antibody and PPV-VP2-Nb56-HRP fusion as the detection antibody was the first time to develop for detecting PPV in different samples. Results showed that the immunoassay can be universally used to surveil PPV infection in pig flock. A direct fluorescent assay using PPV-VP2-Nb12-EGFP as a probe was also developed to detect PPV in ST cells. The two developed immunoassays eliminate the use of commercial secondary antibodies and shorten detection time. Meanwhile, both assays display great developmental prospect for further commercial production and application.


Assuntos
Proteínas de Fluorescência Verde/metabolismo , Peroxidase do Rábano Silvestre/metabolismo , Imunoensaio , Parvovirus Suíno/isolamento & purificação , Proteínas Recombinantes de Fusão/metabolismo , Anticorpos de Domínio Único/metabolismo , Sequência de Aminoácidos , Escherichia coli/metabolismo , Células HEK293 , Humanos , Cadeias Pesadas de Imunoglobulinas/química , Cadeias Pesadas de Imunoglobulinas/metabolismo , Região Variável de Imunoglobulina/química , Região Variável de Imunoglobulina/metabolismo , Indicadores e Reagentes , Anticorpos de Domínio Único/química , Proteínas Virais/imunologia
10.
Analyst ; 144(18): 5486-5496, 2019 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-31386701

RESUMO

Traditional virus detection methods require ligands that bind to either viral capsid proteins or viral nucleic acids. Ligands are typically antibodies or oligonucleotides and they are expensive, have limited chemical stability, and can only detect one specific type of virus at a time. Here, the biochemical surface properties of viruses are exploited for ligand-free, nonspecific virus detection. It has been found that the osmolyte mannitol can preferentially aggregate virus, while leaving proteins in solution. This led to the development of a ligand-free detection of virus using gold nanoparticle (AuNP) aggregation. Porcine parvovirus (PPV) was incubated with AuNPs and aggregation of the PPV-AuNP complex with mannitol was detected by dynamic light scattering (DLS). The lowest detectable concentration of PPV was estimated to be 106 MTT50 per mL, which is lower than standard antibody assays. PPV was also detected when swabbed from a dry surface and in the presence of a protein solution matrix. The enveloped bovine viral diarrhea virus (BVDV) was also detected using mannitol-induced aggregation of BVDV-coated AuNPs. The lowest detectable concentration of BVDV was estimated to be 104 MTT50 per mL. This demonstrates that gold nanoparticle aggregation can detect virus without the use of specific ligands.


Assuntos
Ouro/química , Manitol/química , Nanopartículas Metálicas/química , Vírion/isolamento & purificação , Vírus da Diarreia Viral Bovina/isolamento & purificação , Limite de Detecção , Parvovirus Suíno/isolamento & purificação , Propriedades de Superfície
11.
Arch Virol ; 163(1): 209-213, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29022179

RESUMO

Porcine parvovirus (PPV) are small, non-enveloped and single-stranded DNA viruses, taxonomically classifiable within the family Parvoviridae. Seven PPV genotypes (PPV1 to PPV7) have been identified to date. PPV7, the most recently discovered PPV genotype, was first reported in US pigs in 2016. To explore PPV7 status in Chinese pig populations a total of 64 serum samples collected from two commercial farms in Guangdong province in 2014 were analyzed. PPV7 DNA was detected in 32.8% (21/64) of tested samples. On the porcine circovirus type 2 (PCV2) positive farm, the prevalence rate of PPV7 was 65.5% (19/29) which was significantly higher than that on the PCV2 negative farm (2/35, 5.7%), indicating a possible association between PCV2 and PPV7 infections. The sequences of three PPV7 strains were determined. Phylogenetic analysis revealed that the identified PPV7 strains circulating in China shared 98.7%-99.7% nucleotide homology with the US strain. Further sequence comparison analysis indicated that GD-2014-2 and GD-2014-3 possess a consecutive 9-nt deletion in the VP gene. This is the first report of the existence of PPV7 in China and this finding will strengthen understanding of the epidemiology of porcine parvovirus in Chinese pigs.


Assuntos
Infecções por Parvoviridae/veterinária , Parvovirus Suíno/classificação , Parvovirus Suíno/isolamento & purificação , Doenças dos Suínos/virologia , Animais , Sequência de Bases , China/epidemiologia , Regulação Viral da Expressão Gênica , Infecções por Parvoviridae/epidemiologia , Infecções por Parvoviridae/virologia , Parvovirus Suíno/genética , Filogenia , Suínos , Doenças dos Suínos/epidemiologia , Proteínas Virais/genética , Proteínas Virais/metabolismo
12.
J Gen Virol ; 97(6): 1408-1413, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-26939976

RESUMO

Inactivated whole-virus vaccines against porcine parvovirus (PPV) can prevent disease but not infection and virus shedding after heterologous virus challenge. Here, we showed that the same is true for a homologous challenge. Pregnant sows were vaccinated with an experimental inactivated vaccine based on PPV strain 27a. They were challenged on day 40 of gestation with the virulent porcine parvovirus PPV-27a from which the vaccine was prepared (homologous challenge). On day 90 of gestation, the fetuses from vaccinated sows were protected against disease, while the fetuses of the non-vaccinated sows (control group) exhibited signs of parvovirus disease. All gilts, whether vaccinated or not vaccinated, showed a boost of PPV-specific antibodies indicative of virus infection and replication. Low DNA copy numbers, but not infectious virus, could be demonstrated in nasal or rectal swabs of immunized sows, but high copy numbers of challenge virus DNA as well as infectious virus could both be demonstrated in non-vaccinated sows.


Assuntos
Infecções por Parvoviridae/veterinária , Parvovirus Suíno/imunologia , Doenças dos Suínos/prevenção & controle , Vacinas Virais/imunologia , Eliminação de Partículas Virais , Animais , Mucosa Nasal/virologia , Infecções por Parvoviridae/prevenção & controle , Parvovirus Suíno/isolamento & purificação , Reto/virologia , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/virologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/administração & dosagem
13.
Mol Cell Probes ; 30(5): 300-305, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27593155

RESUMO

Porcine parvovirus (PPV) is a major cause of swine reproductive failure and reported in many countries worldwide. Recombinase polymerase amplification (RPA) assays using a real-time fluorescent detection (PPV real-time RPA assay) and a lateral flow dipstick (PPV RPA LFD assay) were developed targeting PPV NS1 gene. The detection limit of PPV real-time RPA assay was 300 copies per reaction within 9 min at 38 °C, while the RPA LFD assay has a detection limit of 400 copies per reaction in less than 20 min at 38 °C. In both assays, there were no cross-reactions with porcine circovirus type 2, pseudorabies virus, porcine reproductive and respiratory syndrome virus, classical swine fever virus, and foot-and-mouth disease virus. Based on a total of 128 clinical samples examined, the sensitivity and the specificity of the developed RPA assays for identification of PPV was 94.4% and 100%, respectively, when compared to real-time (qPCR) assay. Therefore, the RPA assay provides a rapid, sensitive and specific alternative for PPV detection.


Assuntos
Parvovirus Suíno/genética , Parvovirus Suíno/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Recombinases/metabolismo , Temperatura , Animais , Sensibilidade e Especificidade , Suínos
14.
Arch Virol ; 161(4): 981-4, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26711454

RESUMO

Porcine hokovirus (PHoV), also referred to as porcine parvovirus 4 (P-PARV4), a recently discovered parvovirus of swine that is closely related to human parvovirus 4/5 (H-PARV4/5), was first described in Hong Kong. To evaluate the occurrence of P-PARV4 in Portuguese wild boars in the hunting season of 2011/2012, liver and serum samples were tested. P-PARV4 was detected in 24 % of the wild boars analyzed. Phylogenetic analysis showed a close relationship between the P-PARV4 isolates and other P-PARV4 reference strains. This virus appears to be emerging, with yet unknown implications for public health.


Assuntos
Infecções por Parvoviridae/veterinária , Parvovirus Suíno/isolamento & purificação , Sus scrofa/virologia , Doenças dos Suínos/virologia , Animais , Feminino , Masculino , Infecções por Parvoviridae/virologia , Parvovirus Suíno/genética , Filogenia , Portugal/epidemiologia , Suínos , Doenças dos Suínos/epidemiologia
15.
Virol J ; 12: 170, 2015 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-26475593

RESUMO

BACKGROUND: Currently, eight species in four genera of parvovirus have been described that infect swine. These include ungulate protoparvovirus 1 (classical porcine parvovirus, PPV), ungulate tetraparvovirus 2 (PPV3), ungulate tetraparvovirus 3 (which includes PPV2, porcine hokovirus, porcine partetravirus and porcine PARV4), ungulate copiparvovirus 2 (which includes PPV4 and PPV5), ungulate bocaparvovirus 2 (which includes porcine bocavirus 1, 2 and 6), ungulate bocaparvovirus 3 (porcine bocavirus 5), ungulate bocaparvovirus 4 (porcine bocavirus 7) and ungulate bocaparvovirus 5 (porcine bocavirus 3, 4-1 and 4-2). PPV6, the most recently described porcine parvovirus, was first identified in China in late 2014 in aborted pig fetuses. Prevalence of PPV6 in China was found to be similar in finishing age pigs from farms with and without evidence of swine reproductive failure. METHODS: Porcine parvovirus 6 (PPV6) was detected by sequence-independent single primer amplification (SISPA) and confirmed by overlapping and real-time PCR in the serum of porcine reproductive and respiratory virus (PRRSv) positive samples. RESULTS: Seven nearly complete genomes of PPV6 were identified in PRRSv genotype 2 positive serum samples submitted to state veterinary diagnostic laboratories in 2014. Further testing using overlapping and real-time PCR determined PPV6 to be present in 13.2 % of the serums tested. Additionally, PPV6 was present in samples from all of the geographic locations sampled encompassing nine states in the United States and one state in Mexico. The presence of PPV6 in serum indicates that the PPV6 infection is disseminated and not localized to a specific tissue type. Alignments of the near full length genomes, NS1, and capsid genes identified one of the five PPV6 isolates from China (98.6-99.5 % identity with the North American strains) to be the North American strains nearest relative. CONCLUSIONS: These results are the first to report the presence of PPV6 in North America and demonstrate that the virus is found in multiple geographic areas in the United States and in Mexico. The overall prevalence of PPV6 in PRRSv viremic animals is relatively low. Further, all of the PPV6 genomes found in North America are most closely related to a PPV6 strain first identified in 2014 in healthy pigs from the Tianjin province of China.


Assuntos
Infecções por Parvoviridae/veterinária , Parvovirus Suíno/isolamento & purificação , Síndrome Respiratória e Reprodutiva Suína , Soro/virologia , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/virologia , Animais , Análise por Conglomerados , Metagenômica , México/epidemiologia , Dados de Sequência Molecular , Infecções por Parvoviridae/epidemiologia , Infecções por Parvoviridae/virologia , Parvovirus Suíno/classificação , Parvovirus Suíno/genética , Filogenia , Reação em Cadeia da Polimerase , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , Análise de Sequência de DNA , Homologia de Sequência , Suínos , Estados Unidos/epidemiologia
16.
Arch Virol ; 160(5): 1339-44, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25742931

RESUMO

A total of 450 samples from domestic pigs in China were tested for porcine parvoviruses (PPVs) and co-infections with porcine circovirus type 2 (PCV2), and their complete capsid genes were sequenced. The prevalence of PPV1, PPV2, PPV3, PPV4, and PCV2 was 5.56 %, 39.56 %, 45.11 %, 21.56 %, and 47.33 %, respectively, and co-infection with PCV2 occurred in 4 % (PPV1), 22.44 % (PPV2), 24 % (PPV3), and 12 % (PPV4) of the samples. Phylogenetic analysis revealed two main lineages for each virus, and residues that differentiated these viruses were identified. The co-infections of emerging PPVs and PCV2 were prevalent, indicating their cooperative roles in porcine circovirus-associated diseases.


Assuntos
Infecções por Circoviridae/epidemiologia , Circovirus/isolamento & purificação , Coinfecção/epidemiologia , Infecções por Parvoviridae/epidemiologia , Parvovirus Suíno/isolamento & purificação , Sus scrofa/virologia , Animais , Proteínas do Capsídeo/genética , China/epidemiologia , Infecções por Circoviridae/virologia , Análise por Conglomerados , Coinfecção/virologia , DNA Viral/química , DNA Viral/genética , Dados de Sequência Molecular , Infecções por Parvoviridae/virologia , Filogenia , Prevalência , Análise de Sequência de DNA , Homologia de Sequência
17.
Microbiol Immunol ; 59(2): 82-8, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25431024

RESUMO

Porcine parvovirus (PPV) causes reproductive failure in sows and has spread worldwide. Several new types of porcine parvoviruses have recently been identified in pig herds. The prevalence of five porcine parvoviruses in the Chiangmai area of Thailand was studied. The prevalence in 80 pigs was 53% for PPV (PPV-Kr or -NADL2 being the new abbreviations), 83% for PPV2 (CnP-PARV4), 73% for PPV3 (P-PARV4), 44% for PPV4 (PPV4), and 18% for PBo-likeV (PBoV7). Over 60% of the pigs carried more than three of the five porcine parvoviruses and occurrence together of the two pairs of viral genes, PPV1/PPV3 and PPV2/PBo-likeV were observed. Phylogenetic analyses for PPV2 and PPV3 indicated the existence of only two major clades of PPV2 and one major clade of PPV3.


Assuntos
Infecções por Parvoviridae/veterinária , Parvovirus Suíno/classificação , Parvovirus Suíno/genética , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/virologia , Animais , Análise por Conglomerados , Coinfecção/epidemiologia , Coinfecção/veterinária , Coinfecção/virologia , DNA Viral/química , DNA Viral/genética , Dados de Sequência Molecular , Infecções por Parvoviridae/epidemiologia , Infecções por Parvoviridae/virologia , Parvovirus Suíno/isolamento & purificação , Filogenia , Prevalência , Análise de Sequência de DNA , Suínos , Tailândia/epidemiologia
18.
Appl Microbiol Biotechnol ; 99(2): 919-28, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25557628

RESUMO

A novel multiplex detection array based on Luminex xMAP technology was developed and validated for simultaneous detection of five major viruses causing swine reproductive diseases. By combining one-step asymmetric multiplex reverse transcription polymerase chain reaction (RT-PCR) with xMAP bead-based hybridization and flow cytometry analysis, the resulting multiplex assay was capable of detecting single and mixed infections of PRRSV, PCV-2, PRV, CSFV, and PPV in a single reaction. The assay accurately detected and differentiated 23 viral strains used in this study. The low detection limit was determined as 2.2-22 copies/µL (corresponding to 0.5-6.8 fg/µL DNA template) on plasmid constructs containing viral fragments. The intra-assay and inter-assay variances (CV%) were low that ranged from 2.5 to 5.4 % and 4.1 to 7.6 %, respectively. The assay was applied to test field samples and detected single and mixed viral infections. The detection rate was higher than that of uniplex conventional PCR and RT-PCR methods. The detection of PRRSV by the bead-based multiplex assay was comparable with a commercially available real time RT-PCR kit. The test procedure on purified DNA or RNA samples could be completed within 2 h. In conclusion, the bead-based suspension array presented here proved to be a high-throughput practical tool that provided highly specific and sensitive identification of single and multiple infections of five major viruses in pigs and boar semen.


Assuntos
Circovirus/isolamento & purificação , Vírus da Febre Suína Clássica/isolamento & purificação , DNA Viral/isolamento & purificação , Parvovirus Suíno/isolamento & purificação , Vírus da Síndrome Respiratória e Reprodutiva Suína/isolamento & purificação , RNA Viral/isolamento & purificação , Animais , Circovirus/classificação , Peste Suína Clássica/diagnóstico , Peste Suína Clássica/virologia , Primers do DNA , DNA Viral/genética , Reação em Cadeia da Polimerase Multiplex , RNA Viral/genética , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Manejo de Espécimes , Suínos/virologia , Doenças dos Suínos/diagnóstico , Doenças dos Suínos/virologia
19.
Virol J ; 11: 203, 2014 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-25442288

RESUMO

BACKGROUND: Parvoviruses are classified into two subfamilies based on their host range: the Parvovirinae, which infect vertebrates, and the Densovirinae, which mainly infect insects and other arthropods. In recent years, a number of novel parvoviruses belonging to the subfamily Parvovirinae have been identified from various animal species and humans, including human parvovirus 4 (PARV4), porcine hokovirus, ovine partetravirus, porcine parvovirus 4 (PPV4), and porcine parvovirus 5 (PPV5). METHODS: Using sequence-independent single primer amplification (SISPA), a novel parvovirus within the subfamily Parvovirinae that was distinct from any known parvoviruses was identified and five full-length genome sequences were determined and analyzed. RESULTS: A novel porcine parvovirus, provisionally named PPV6, was initially identified from aborted pig fetuses in China. Retrospective studies revealed the prevalence of PPV6 in aborted pig fetuses and piglets(50% and 75%, respectively) was apparently higher than that in finishing pigs and sows (15.6% and 3.8% respectively). Furthermore, the prevalence of PPV6 in finishing pig was similar in affected and unaffected farms (i.e. 16.7% vs. 13.6%-21.7%). This finding indicates that animal age, perhaps due to increased innate immune resistance, strongly influences the level of PPV6 viremia. Complete genome sequencing and multiple alignments have shown that the nearly full-length genome sequences were approximately 6,100 nucleotides in length and shared 20.5%-42.6% DNA sequence identity with other members of the Parvovirinae subfamily. Phylogenetic analysis showed that PPV6 was significantly distinct from other known parvoviruses and was most closely related to PPV4. CONCLUSION: Our findings and review of published parvovirus sequences suggested that a novel porcine parvovirus is currently circulating in China and might be classified into the novel genus Copiparvovirus within the subfamily Parvovirinae. However, the clinical manifestations of PPV6 are still unknown in that the prevalence of PPV6 was similar between healthy pigs and sick pigs in a retrospective epidemiological study. The identification of PPV6 within the subfamily Parvovirinae provides further insight into the viral and genetic diversity of parvoviruses.


Assuntos
DNA Viral/genética , Genoma Viral , Parvovirus Suíno/genética , Parvovirus Suíno/isolamento & purificação , Animais , China , Análise por Conglomerados , Dados de Sequência Molecular , Infecções por Parvoviridae/veterinária , Infecções por Parvoviridae/virologia , Parvovirus Suíno/classificação , Filogenia , Análise de Sequência de DNA , Homologia de Sequência , Suínos , Doenças dos Suínos/virologia
20.
Arch Virol ; 159(6): 1533-6, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24327097

RESUMO

To investigate porcine parvovirus 5 (PPV5) infections in swine herds in China, clinical specimens of piglet lungs were examined for the presence of PPV5 using a polymerase chain reaction method. A strain of PPV5 was detected, and its genome was sequenced and analyzed. In the sequence alignment and phylogenetic analysis, the Chinese PPV5 strain clustered into a distinct clade with the reference PPV5 strains. These results provide direct evidence that PPV5 is present in pigs in China. Extensive epidemiological studies are warranted to determine the geographic distribution of PPV5 in China.


Assuntos
DNA Viral/química , DNA Viral/genética , Genoma Viral , Parvovirus Suíno/genética , Animais , China , Análise por Conglomerados , Dados de Sequência Molecular , Parvovirus Suíno/isolamento & purificação , Filogenia , Análise de Sequência de DNA , Homologia de Sequência , Suínos
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