| Hydropericardium syndrome(HPS)is a highly infectious disease caused by fowl adenovirus serotype 4(FAV-4)affecting poultry especially broiler chickens,which has been report in many countries and region in the world.Occasionally reported in parts of china,however,Henan province began to outbreak in summer and autumn in 2015,and then spread to the whole province,which brought huge economic losses to the poultry production in our province and even the whole country.Currently,no effective and safe vaccine is available for the control of HHS virus(HHSV)infection in chicken.Therefore,it is of practical significance to study the biological characteristics of the pathogen and the innate immune mechanism.Avian beta defensins are small,cationic,and amphipathic cystein-rich antibiotic petides with broad-spectrum antimicrobial activity,they has been widely studied as a new type of antibacterial and vaccine adjuvant,which acts on microbial membrane or host cells,resulting in very low microbial resistance.In September 2015,the poultry farm of Henan outreak case which has a characterisitic of pericardial effusion with a highly mortality rate.According to the analysis of clinical anatomy,it is suspected to be HHS,the pathogen may be Avian adenovirus.Take the pericardial effusion of liver and heart and other tissues inoculated with SPF chicken embryo to isolate virus,while using aviadenovirus group?specific primers by PCR detection and sequence analysis of embryo allantoic fluid and tissues.The results of PCR amplification show that size about 900 bp fragments were amplified in the tissues and chicken embryo allantoic fluid,and the isolate was named HNXZ.The results of homology analysis based on hexon gene nucleotide sequence show that the isolate had the highest homology with adenovirus subgroup C in 96.3%-100% similarity and in 97.9%-100% similarity with serotype 4.Phylogenetic analysis showed that the nearest genetic distance with group? avian adenovirus serotype 4.Restriction endonuclease Sty I and Mlu I enzyme digestion analysis showed that the isolate was 4.According to the results of above,it is proved that the isolate belongs to group?avian Avian adenovirus.In order to further understand the biological characterisitics of the isolated strain,it was studied by observing the characteristics of cell culture,animal regression test and histopathological examination.The results showed that the isolated strain could produce a typical avian adenovirus lesion in the chicken embryo liver(CEL)cell,which showed that the cells were round and the refraction was enhanced.The results of animal regression test showed that the clinical symptoms and pathological changes were consistent with that of naturally occurring chickens.The histopathological changes were characterized by the inclusion of basic inclusions and large necrosis of liver cells in the nucleus of hepatocytes.The above studies showed that the isolated virus was highly pathogenic.This study provides a reference for the prevention and control of avian adenovirus.RT-PCR was used to detect the expression distribution of 14 kinds of Av BDs in different tissues.The results showed that Av BD1,2 and 10 were widely distributed,and the expression of Av BD14 in brain tissue was first detected.At the same time,the establishment of Av BDs,IL-1 and IL-6 SYBR Green I Real-time f quantitative polymerase chain method(q PCR),the method is simple,economic and fast,for the further study of Av BDs,IL-1?and IL-6 m RNA expression research laid the technical foundation?Using the established q PCR method,we detected the expression profile of Av BDs,IL-1 and IL-6 in HNXZ infected chicken liver,and also analyzed the relationship between them in vivo.The results showed that the expression level of Av BD4,8,10,13 and IL-1 was significantly up regulated(P<0.05)and Av BD5,11 were significantly down regulated(P>0.05)in chickens infected with chicken.Av BD2,Av BD3,Av BD6,IL-6 m RNA expression did not change significantly compared with the control group(P>0.05).Av BD9 expression was significantly decreased 4 day after infection(P<0.05).These results indicated that FAd V-4 could induce the expression of Av BD8,Av BD4,Av BD10,Av BD13 and IL-1?in chicken liver,but had no effect on the expression of IL-6.These results suggest that Av BDs may be an important antiviral molecule involved in innate immune regulation after fowl adenovirus infection.This study lays the foundation for further understanding the immune regulation mechanism of Av BD,IL-1?and IL-6 in the liver of adenovirus infection. |
PDF Full Text Request