RESUMO
Hepatitis E virus (HEV) is an enteric pathogen of humans and animals, and pigs have been considered an important reservoir of this virus. Recent evidence has indicated the cross-species transmission of hepatitis E virus (HEV) from pigs to humans, causing zoonosis, mostly via consumption of uncooked or undercooked animal meat/viscera. In this study, we have developed a one-step RT-LAMP assay for rapid detection of swine HEV. Specific primer sets targeting the ORF3 gene were designed. The sensitivity of the RT-LAMP assay was 10(1) copies/µl of RNA template, which was tenfold higher than that of RT-nPCR. The specificity of this assay was demonstrated by the lack of amplification of DNA/RNA from other swine viruses. Furthermore, a total of 41 bile samples were subjected to RT-LAMP and RT-nPCR. Eighteen positive samples were detected by RT-nPCR, while 36 positive samples were detected by RT-LAMP, indicating that the sensitivity of the RT-LAMP assay was higher than that of the conventional RT-nPCR assay. The RT-LAMP assay reported here may be used for diagnosis of swine HEV, not only in laboratories but also under field conditions.
Assuntos
Vírus da Hepatite E/isolamento & purificação , Hepatite E/veterinária , Técnicas de Amplificação de Ácido Nucleico/veterinária , Doenças dos Suínos/virologia , Animais , Bile/virologia , Regulação Viral da Expressão Gênica , Hepatite E/diagnóstico , Hepatite E/virologia , Vírus da Hepatite E/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Filogenia , RNA Viral/genética , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Sensibilidade e Especificidade , Suínos , Doenças dos Suínos/diagnósticoRESUMO
In China H3N2 and H1N1 swine influenza viruses have been circulating for many years. In January 2010, before swine were infected with foot and mouth disease in Guangdong, some pigs have shown flu-like symptoms: cough, sneeze, runny nose and fever. We collected the nasopharyngeal swab of all sick pigs as much as possible. One subtype H1N2 influenza viruses were isolated from the pig population. The complete genome of one isolate, designated A/swine/Guangdong/1/2010(H1N2), was sequenced and compared with sequences available in GenBank. The nucleotide sequences of all eight viral RNA segments were determined, and then phylogenetic analysis was performed using the neighbor-joining method. HA, NP, M and NS were shown to be closely to swine origin. PB2 and PA were close to avian origin, but NA and PB1were close to human origin. It is a result of a multiple reassortment event. In conclusion, our finding provides further evidence about the interspecies transmission of avian influenza viruses to pigs and emphasizes the importance of reinforcing swine influenza virus (SIV) surveillance, especially before the emergence of highly pathogenic FMDs in pigs in Guangdong.
Assuntos
Genoma Viral , Vírus da Influenza A Subtipo H1N2/classificação , Vírus da Influenza A Subtipo H1N2/genética , Infecções por Orthomyxoviridae/veterinária , RNA Viral/genética , Doenças dos Suínos/virologia , Animais , China , Análise por Conglomerados , Evolução Molecular , Genótipo , Vírus da Influenza A Subtipo H1N2/isolamento & purificação , Dados de Sequência Molecular , Nasofaringe/virologia , Infecções por Orthomyxoviridae/patologia , Infecções por Orthomyxoviridae/virologia , Filogenia , Vírus Reordenados/classificação , Vírus Reordenados/genética , Vírus Reordenados/isolamento & purificação , Análise de Sequência de DNA , Suínos , Doenças dos Suínos/patologia , Proteínas Virais/genéticaRESUMO
MicroRNAs (miRNAs) are a class of endogenous non-coding small RNAs of 18-22-nucleotides in length that regulate gene expression at the post-transcriptional level. The objective of this study was to examine the differences in the miRNA expression profiles of the lungs and trachea of beagle dogs infected with canine influenza virus (CIV). Total RNA was isolated from lung and trachea tissues of beagle dogs infected and non-infected with H3N2 CIV at 4 dpi. A total of 41,512,315 and 39,107,475 reads were obtained from the lung and trachea, respectively. Out of a total 288 dog miRNAs available in miRBase, 227 and 236 miRNAs were detected in the infected (Fg) and the non-infected lungs (Fc), respectively, whereas 242 miRNAs were detected in both the infected (Qg) and the non-infected trachea (Qc). From these, 34 and 45 miRNAs were differentially expressed in the lungs and trachea between the infected and non-infected dogs, respectively. More miRNAs were highly expressed in the non-infected tissues than in the infected tissues. miR-143 was the most abundantly expressed miRNA in the four samples, followed by let-7. In total, 252, 234, 196 and 235 novel miRNAs were identified in the Fc, Fg, Qc, and Qg groups, respectively. To our knowledge, this is the first study examining the miRNA gene expression in CIV infected dogs using the Solexa sequencing approach. We have revealed the existence of a large number miRNAs that are affected by CIV infection as well as identified some potentially new miRNAs. These findings will help us better understand the host-CIV interaction and its relationship to pathogenesis, as well as contribute to the prevention and control of CIV.