Isolation and analysis of a very virulent Marek’s disease virus strain in China

Zhenhua Gong,Zhiliang Wang,Linlin Chen,Lijuan Zhang,Jianlin Wang,Hu Shan,Hongchao Ma

doi:10.1186/1743-422x-10-155

Zhenhua Gong, Zhiliang Wang + Show 5 more

Open Access

https://doi.org/10.1186/1743-422x-10-155

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Abstract

BackgroundA severe MD was broken out at a farm in Shandong, China, despite FC126 vaccination of the chickens at 1-day-old. The mortality of the flocks reached up to 38.3%. The infected chickens were found to have MD pathological changes, including enlargement of spleens, livers and kidneys, and tumors occured on organs later. Samples were collected from the chickens for diagnosis.MethodsThe collected samples were inoculated into primary duck embryo fibroblast (DEF) cells, and the MDV strain named SD2012-1 was isolated. In order to identify the isolate, amplification by PCR and sequencing of oncogenic Meq and vIL-8 gene were processed, the obtained sequences were compared with the sequences of reference strains, and SD2012-1 was used to challenge immunized SPF chickens.ResultsA very virulent MDV isolate strain, SD2012-1, was isolated from a chicken flock in Shandong Province, China, the isolate had the characteristics of very virulent MDV-1, nucleotide and deduced amino acid sequence comparisons of Meq and vIL-8 gene of SD2012-1 with those of reference strains showed SD2012-1 had high homology with MDV strains isolated from China, SD2012-1 could break through the protection provided by HVT vaccine and HVT + SB-1 vaccine immunization and caused the mortality of SPF chickens over 60%. The immune failure occured at the farm could be due to the improper selection of vaccines. SD2012-1 produced death later and the gross postmortem lesions of chickens died early and later were different.ConclusionsMDV strain SD2012-1 isolated from Shandong Province, China was found to have the characteristics of very virulent MDV-1, which could break through the protection provided by HVT vaccine and HVT + SB-1 vaccine, the virus seemed to have a long latent period, and cause different gross postmortem lesions of chickens between chickens died early and later. A better immunization way should be chosen to prevent infection of this MDV strain in field.

Highlights

A severe MD was broken out at a farm in Shandong, China, despite FC126 vaccination of the chickens at 1-day-old
Serotype 1 Marek’s Disease Virus (MDV) included all the oncogenic strains and their attenuated forms; serotype 2 MDVs were non-oncogenic viruses isolated in chickens; serotype 3 MDVs were nononcogenic viruses isolated in turkey, generally known as
PCR amplification for Meq and vIL-8 gene of SD2012-1 By PCR [11] of the extracted virus DNA with primers in Table 1, the specific presence of an approximately 1081 bp long DNA product for MDV Meq gene (Figure 1-A) and the specific presence of an approximately 886 bp long DNA product for MDV vIL-8 gene (Figure 1-B) were found in samples, including the duck embryo fibroblast (DEF) cultures infected with SD2012-1 isolate, samples collected from chickens infected with SD2012-1 at the farm and samples collected from challenged chickens with SD2012-1, the PCR products from all the samples were obvious, which verified the existence of infection with MDV to chickens and inoculated DEF cells

Summary

Methods

The collected samples were inoculated into primary duck embryo fibroblast (DEF) cells, and the MDV strain named SD2012-1 was isolated. Heparinised blood samples were collected from chickens with tumor lesions for diagnosis. This experimental research on animals were performed with the approval of Experimental Animal Administrative Center of Shandong Province. Virus isolation The collected samples were kept under refrigeration and transported, lymphocytes separated from the blood samples by lymphocyte separation medium were inoculated into primary duck embryo fibroblast (DEF) cells prepared from 11-day-old embryonated eggs, the inoculated DEF cells were incubated at 37°C with 5% CO2 for five days. The MDV strain named SD2012-1 was obtained by isolation, and the DEF cell cultures of SD2012-1 were collected and stored at-196°C in 10% dimethyl sulphoxide

Results

Discussion

Conclusion