Analysis Of Genetic Variation Of VP3 Gene Of Goose Parvovirus And Study On Variation Of TAG13 Strain Passaged On Cell

Goose parvovirus(GPV)infection,also known as little Goose blast(Gosling plague),since the Chinese scholars party to set the equal inYangzhou since the first found in 1956,the virus has been widely popular among our Goose.Gosling blast event can cause massive death of cheeper,depressed disease goose show spirit,appetite waste,severe diarrhea,symptoms sometimes.The main pathological changes of intestinal inflammation,small intestinal mucosal necrotic fall off,and to form embolism,intestinal blockage.Its spread and popular raising goose industry in China has caused great economic losses,serious harm to raise the healthy development of goose industry in our country.Goose parvovirus VP3 protein structure has relatively higher conservative,at the same time,the body can be induced to produce neutralizing antibody,is GPV protective antigen,often used as GPV molecular biology diagnosis and analysis on the basis of genetic evolution.Therefore,study the biological characteristics,genetic evolution features of GPV and vaccine development become the disease focus.In parts of China from 2015 to 2017 in this study a suspected infection of GPV gaggle of collecting material,separation and 17 strains of the virus,using PCR amplification 17 strains of GPV isolates VP3 gene,the sequencing of splicing correction,17 strains of GPV virus VP3 gene nucleotide sequence,its open reading frame length are 1605 bp,encoding 535 amino acids.17 strains of isolates VP3 gene homology is higher,the nucleotide homology between 96% ~ 96%,and the classic strains SHFX1201 homology between 95.7% ~ 95.7%,the derivation of the amino acid homology was between 97.8% ~ 100%,and the classical strains SHFX1201 amino acid homology was between 98.7% ~ 100%.Nucleotide sequence alignment results showed that the nucleotide sequence of 17 strains,compared with 102 different bases,including 54 difference is bigger,the bases of the differences are mainly in the second half of the nucleotide sequence,in 723 ~ 525,789 ~ 789,1314 ~ 1605 bases vary considerably.Analysis of genetic evolution.,according to the results of GPV and MDPV distribution in two different branches of GPV from different areas and further evolved into three branches.Domestic mainly includes two branches,the main branch of Taiwan with the main isolates have close kinship,but domestic has gradually formed a new gene subtype.Most 17 strains of isolates with China before separation to the strains of close relatives,the gap with foreign isolates far away.The G2,G6,G13 and SHFX1201(KC478066chicago-brewed Goose China plant and 82-0308(AY382883 chicago-brewed Goose Taiwan)Taiwan plant close relatives,the G4 and G5,the G8,G11,G17 and Taiwan 82-0321-v plant relative higher.G9,G10,a G12 strains and other strains have a little gap,is located in a single branch.The above results show that the minimal VP3 gene variants,have higher conservative.Natural passive immunity is an effective method to prevent gosling plague.At present,gegenated attenuated goose vaccine or inactivated vaccine is mainly used.Because there is no SPF goose embryo in the country,there is a hidden danger of bio-safety goose quail vaccine and vaccine production is serious.It is limited by the cycle of goose embryos.Fibroblasts,as a vector for virus-increase,can be stored in large quantities or serially,so that cells with weakened virulence can be adapted to attenuated strains,thus providing favorable protection for the development of GPV attenuated vaccines.It is important for the prevention of gosling plagues.significance.In this study,a strain named TAG13 was selected from a strain of goose that was isolated.Animal regression experiments showed that this strain can cause a large number of deaths of goslings,resulting in obvious clinical symptoms,necropsy changes,and typical intestinal tract thrombosis.Observed tissue sections can also observe typical tissue lesions.The ELISA test of the diseased goslings infected with the virus proved that the strain produced higher antibodies.Because,it was determined that this strain is a virulent strain of goose parvovirus.They were alternately subcultured on GEF and DEF in two ways,aiming to obtain attenuated cells adapted to attenuated strains.Up to now,30 generations have passed.The experimental results show that as the number of passages increases,the TCID50 first increases and then decreases slightly and then tends to a stable trend.The lethality of different generations of cytotoxicity increases first and then decreases in the goslings,indicating that the strain The virus has started to weaken,but it still does not meet a safety standard.Different generations of cytotoxicity produced only minor differences between their whole genome sequence nucleotides and amino acids.In summary,the passage of the experimental cells can have a weak effect on the parvovirus.In order to obtain cells with weakened virulence to adapt to the attenuated strains,the passage should be continued.The attenuated strains of gooseparvovirus cells obtained in this study lay the foundation for the development of attenuated vaccines in the future,and the results of the sequence analysis of the parental and passaged viruses also provide a reference for studying the molecular mechanisms of virulence reduction of goose parvovirus.