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There is considerable evidence that infection by avian lymphoid leukosis viruses can lead to tumor development in the target organ of the host. The mechanism by which virus-induced oncogenic transformation occurs, however, is not clearly understood. As a first step toward deciphering this process, we have characterized the proviruses ofthe lymphoid leukosis viruses in DNAs extracted from the leukotic and metastatic tumors by using restriction enzyme digestion and filter hybridization analysis with radioactive probes specific for the infecting genome. Our results indicate (i) that lymphoid leukosis tumors are clonal in origin; (ii) that there are multiple sites in the cellular genome of the target tissue where the virus DNA can integrate and that, in the majority of the tumors, at least one such site of each tumor is adjacent to a cellular sequence related to the oncogene of MC-29 virus; and (iii) that deletions and other structural alterations in the proviral DNA may facilitate tumorigenesis.The oncogenic retroviruses can be separated into at least two classes that appear to induce neoplasms by different molecular mechanisms. The more extensively characterized group includes viruses that induce rapid neoplasms, encode genes for cell transformation (probably of host origin), and are often defective, requiring a helper virus for infectivity or replication (1,2). The second group induces neoplasms that have long latent periods, have no known genes coding directly for cell transformation, and are not defective in replication. Among these, some appear to have the potential for inducing several types of neoplasms (1, 2). The first class ofviruses, although ofbasic interest in studies ofin vitro cell transformation, are probably laboratory products, while the second class of viruses is likely to be responsible for the majority of naturally occurring retrovirus-induced neoplasms. Viral induction of avian lymphoid leukosis (LL) is an excellent model of neoplasm by a virus of the second group. The steps leading to mortality with LL include the infection of the target cell in the bursa of Fabricius, the transformation of the target cells not earlier than 3 to 4 weeks of age, the development of the grossly visible bursal tumor at 10-16 weeks of age, and the metastasis to visceral organs leading to massive lymphoid tumors and death, usually after 16 weeks of age (3).The present studies are aimed at characterizing the newly integrated exogenous proviruses in LL tumor cell DNA to provide insight into the molecular events that lead to the development of LL. MATERIALS AND METHODSCell Culture, Viruses, and Biochemicals. A RAV-1 virus stock, purified by three cycles of propagation at limiting dilutions, was used. Infection ofchicken embryo fibroblast cultures was carried out at a multiplicity of 0.1, and the infected cells were passaged at least four times before DNA extraction. The media of such cultures contained a high level of reverse-transcriptase activity (4). For the synthesis of cDNA probes, concentrated Prague C ...

Several subgroup J-like avian leukosis viruses (ALV-Js) were isolated from broiler breeder (BB) and commercial broiler flocks experiencing myeloid leukosis (ML) at 4 wk of age or older. In all cases, diagnosis of ML was based on the presence of typical gross and microscopic lesions in affected tissues. The isolates were classified as ALV-J by 1) their ability to propagate in chicken embryo fibroblasts (CEF) that are resistant to avian leukosis virus (ALV) subgroups A and E (C/AE) and 2) positive reaction in a polymerase chain reaction with primers specific for ALV-J. The prototype strain of these isolates, an isolate termed ADOL-Hc1, was obtained from an adult BB flock that had a history of ML. The ADOL-Hc1 was isolated and propagated on C/AE CEF and was distinct antigenically from ALV of subgroups A, B, C, D, and E, as determined by virus neutralization tests. Antibody to ADOL-Hc1 neutralized strain HPRS-103, the prototype of ALV-J isolated from meat-type chickens in the United Kingdom, but antibody to HPRS-103 did not neutralize strain ADOL-Hc1. On the basis of both viremia and antibody, prevalence of ALV-J infection in affected flocks was as high as 87%. Viremia in day-old chicks of three different hatches from a BB flock naturally infected with ALV-J varied from 4% to 25%; in two of the three hatches, 100% of chicks that tested negative for virus at hatch had evidence of viremia by 8 wk of age. The data document the isolation of ALV-J from meat-type chickens experiencing ML as young as 4 wk of age. The data also suggest that strain ADOL-Hc1 is antigenically related, but not identical, to strain HPRS-103 and that contact transmission of ALV-J is efficient and can lead to tolerant infection.

We injected chick syncytial strain of reticuloendotheliosis virus (CS-REV) and wild type and recombinant avian leukosis virus (ALV) near the blastoderm of unincubated fertilized embryos and CS-REV intra-abdominally at day of hatch, and we progeny tested the surviving ALV viremic males and REV viremic males and females for transmitted viral genetic material. A number of positive progeny were identified and their deoxyribonucleic acid (DNA) analyzed for restriction enzyme fragments that hybridized with viral genetic material. Most of the progeny had simple restriction enzyme patterns unlike the viremic parents or congenitally infected progeny. This is suggestive evidence that retroviral genetic information has been inserted into the germ line of chickens.