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Title: Factors influencing the attenuation of serotype 1 Marek's disease virus by serial cell culture passage, and evaluation of attenuated strains for protection and replication

Author
item DUDNIKOVA, EKATERINA - NARVAC - MOSCOW RUSSIA
item VLASOV, ANATOLY - NARVAC - MOSCOW RUSSIA
item NORKINA, SVETLANA - NARVAC - MOSCOW RUSSIA
item KIREEV, DMITRY - NARVAC - MOSCOW RUSSIA
item Witter, Richard

Submitted to: Avian Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/10/2008
Publication Date: 3/31/2009
Citation: Dudnikova, E., Vlasov, A., Norkina, S., Kireev, D., Witter, R.L. 2009. Factors Influencing the Attenuation of Serotype 1 Marek's Disease Virus by Serial Cell Culture Passage and Evaluation of Attenuated Strains for Protection and Replication. Avian Diseases. 53(1):63-72.

Interpretive Summary: Marek’s disease (MD), a virus-induced cancer-like disease of chickens, is considered as a major disease problem in commercial poultry. Vaccination has dramatically reduced the incidence of the disease, but efforts to develop improved vaccines have met with limited success in the past. The objective of this research was to determine the most efficient way to develop a weakened vaccine strain, starting with a fully virulent MD virus. We have determined that the number of passages in cell culture required to derive a weakened strain is strongly influenced by the strength of the original strain. We also found cell culture passage techniques that would speed the weakening process. There was considerable variation in results depending on the specific virus strain used. This important information will help scientists in academia and industry understand the prospects for deriving new candidate vaccine strains which will eventually lead to better control of the disease.

Technical Abstract: This study was to better understand factors that influenced the process of attenuation of Marek’s disease (MD) virus by serial passage in cell cultures. Three virulent (v) pathotype and 3 very virulent plus (vv+) pathotype strains were passed by 3 techniques up to 131 times and the passage level at attenuation was determined. The 18 attenuated or partially attenuated viruses were evaluated for protection against challenge with virulent MD virus, and the virus load (latent infection) in blood lymphocytes at 14-21 days post vaccination was determined. Viral pathotype strongly influenced the rate of attenuation. The mean passage level at attenuation for v and vv+ strains was 74 and >109, respectively. Full attenuation was achieved for 9 of 9 passage series with v pathotype strains but for only 4 of 9 passage series with vv+ pathotype strains. Time to attenuation was not significantly influenced by multiplicity of infection at passage or by cell type although a possible advantage of alternate high and low multiplicity passage was noted. Protection was not significantly influenced by pathotype or time to attenuation. Protection varied from 50-95% for the 18 passaged virus preparations; 6 attenuated viruses provided high protection that did not differ from that of the prototype Rispens strain. Virus load was not influenced by pathotype or by passage strategy and showed no positive correlation with protection. In several cases the most protective vaccines had the least virus load. This finding differs from previous reports and warrants further study. Variation among different strains within the same pathotype was documented for attenuation rate, protection and virus load. Also, variation was evident when the same strain was passaged by different strategies, probably reflecting random changes during serial passage. Strain 596A (v pathotype) was the first to become attenuated, provided the best protection and had one of the lowest virus loads. In contrast, strain 617A (v pathotype) provided the least protection and had one of the highest virus loads. Such strains provide fertile opportunities for further study.