| Marek's disease (MD), reticuloendotheliosis (RE), and avian leukosis (AL) are the most common neoplastic diseases or tumors in chicken. The gross lesions and histological findings overlap making conventional field diagnosis problematical. The infections of the oncogenic viruses may cause damages of immune cells and immune organs other than the forming of tumors, and the infected birds experienced immunosuppression and were susceptible of bacteria, viruses, or parasites afterward, which is responsible for increasing morbidity, mortality and great economic losses. Subclinical infection and co-infections of pathogenic agents make worse to diagnose and differentiate avian tumors. A new technique featuring rapidity, sensitivity, and specificity should be employed to identify the causative viruses and/or their genomes in the differential diagnosis of avian tumors.A multiplex PCR protocol was successfully developed for the detection of avian tumors. The optimal ratio of the primers for the oncogenic viruses, reaction components and anealing temperature were evaluated, while the innovation of the extraction of the sample DNAs was made, the whole procedure of diagnosis, from sampling to the final conclusion, was finished within 5 hours in our protocol.1,458 samples, from 457 chickens in 125 flocks, were diagnosed for avian tumors by using the developed PCR protocol in the recent 2 years. The epidemiological survey showed that MD was the most common tumor in Guangxi (accounted 95.56% of the total tumors), and followed by RE and AL (accounted 17.55% and 11.59% of the total tumors, respectively). Infected birds without gross tumor lesions were also oncogenic viruses positive in the survey (accounted 44.4%, 9.88%, 8.64%, respectively). The analysis of epidemiological data showed that the birds suffered from tumors experienced retardation in growth, depression ofimmune responses and co-infection with other pathogens, as well as increasing mortality.One-day-old chickens were inoculated with vaccine strain CVI988/Rispens and detected for the distribution and duration of viruses in different visceral organs, feather follicles and blood post-innoculation(PI). The results showed that the total positive rates of viral DNAs in feather follicle (FF), peripheral blood lymphocyte (PBL), spleen, liver and thymus, kidney and proventriculus were 20% (2/10), 30%(6/20), 20%(4/20), 5%(1/20) and 15%(3/20), 0 and 0, respectively within 4 weeks PI, but almost negative in all the visceral organs, FF and PBL in the birds from 5 weeks PI.A dot-hybridization technique, with MDV-1 specific pp38-based DNA probe for MD and e/iv-based DNA probe for RE, was developed and the comparison with the PCR-based protocol was also conducted. Only 65.1%, 35.2% of the PCR positive samples respectively for MD and RE were detected by the dot-hybridization assay. The results demonstrated that the developed PCR-based protocol has advantages in the aspects of sensitivity, rapidity, labor and expense compared with the dot-hybridization technique for the diagnosis of MD and RE.The results of the study demonstrated that the PCR-based protocol for differential diagnosis of avian tumors was characteristic of rapidity, sensitivity, and specificity, and valuable for clinical diagnosis, epidemiological survey, veterinary quarantine and the detection of the contamination of REV and ALV in vaccine. |
