Discovery of an expanded set of avian leukosis subgroup E proviruses in chickens using Vermillion, a novel sequence capture and analysis pipeline [corrected].

Transposable elements (TEs), such as endogenous retroviruses (ERVs), are common in the genomes of vertebrates. ERVs result from retroviral infections of germ-line cells, and once integrated into host DNA they become part of the host's heritable genetic material. ERVs have been ascribed positive effects on host physiology such as the generation of novel, adaptive genetic variation and resistance to infection, as well as negative effects as agents of tumorigenesis and disease. The avian leukosis virus subgroup E family (ALVE) of endogenous viruses of chickens has been used as a model system for studying the effects of ERVs on host physiology, and approximately 30 distinct ALVE proviruses have been described in the Gallus gallus genome. In this report we describe the development of a software tool, which we call Vermillion, and the use of this tool in combination with targeted next-generation sequencing (NGS) to increase the number of known proviruses belonging to the ALVE family of ERVs in the chicken genome by 4-fold, including expanding the number of known ALVE elements on chromosome 1 (Gga1) from the current 9 to a total of 40. Although we focused on the discovery of ALVE elements in chickens, with appropriate selection of target sequences Vermillion can be used to develop profiles of other families of ERVs and TEs in chickens as well as in species other than the chicken.

Genetic variation at the tumour virus B locus in commercial and laboratory chicken populations assessed by a medium-throughput or a high-throughput assay.

The tumour virus B (TVB) locus encodes cellular receptors mediating infection by three subgroups of avian leukosis virus (B, D, and E). Three major alleles, TVB*S1, TVB*S3, and TVB*R, have been described. TVB*S1 encodes a cellular receptor mediating infection of subgroups B, D, and E. TVB*S3 encodes the receptor for two subgroups, B and D, and TVB*R encodes a dysfunctional receptor that does not permit infection by any of the subgroups, B, D, or E. Genetic diversity at the TVB locus of chickens was investigated in both layer and broiler commercial pure lines and laboratory lines. Genotyping assays were developed for both medium-throughput and high-throughput analysis. Of the 36 broiler lines sampled, 14 were fixed for the susceptible allele TVB*S1. Across all broiler lines, 83% of chickens were typed as TVB*S1/*S1, 3% as TVB*R/*R, and 14% as TVB*S1/*R. In the egg-layer lines, five of the 16 tested were fixed for TVB*S1/*S1. About 44% of egg-layers were typed as TVB*S1/*S1, 15% as TVB*R/*R, with the rest segregating for two or three of the alleles. In the laboratory chickens, 60% were fixed for TVB*S1/*S1, 6% for TVB*S3/*S3, 14% for TVB*R/*R, and the rest were heterozygotes (TVB*S1/*S3 or TVB*S1/*R). All commercial pure lines examined in this study carry the TVB*S1 allele that sustains the susceptibility to avian leukosis viruses B, D, and E. More importantly, the TVB*R allele was identified in multiple populations, thus upholding the opportunities for genetic improvement through selection.

Acutely transforming avian leukosis virus subgroup J strain 966: defective genome encodes a 72-kilodalton Gag-Myc fusion protein.

Avian leukosis virus subgroup J (ALV-J), the most recent member of the avian retroviruses, is predominantly associated with myeloid leukosis in meat-type chickens. We have previously demonstrated that the acutely transforming virus strain 966, isolated from an ALV-J-induced tumor, transformed peripheral blood monocyte and bone marrow cells in vitro and induced rapid-onset tumors, suggesting transduction of oncogenes (L. N. Payne, A. M. Gillespie, and K. Howes, Avian Dis. 37:438-450, 1993). In order to understand the molecular basis for the rapid transformation and tumor induction, we have determined the complete genomic structure of the provirus of the 966 strain. The sequence of the 966 provirus clone revealed that its genome is closely related to that of HPRS-103 but is defective, with the entire pol and parts of the gag and env genes replaced by a 1,491-bp sequence representing exons 2 and 3 of the c-myc gene. LSTC-IAH30, a stable cell line derived from turkey monocyte cultures transformed by the 966 strain of ALV-J, expressed a 72-kDa Gag-Myc fusion protein. The identification of the myc gene in 966 virus as well as in several other ALV-J-induced tumors suggested that the induction of myeloid tumors by this new subgroup of ALV occurs through mechanisms involving the activation of the c-myc oncogene.

Development and application of polymerase chain reaction (PCR) tests for the detection of subgroup J avian leukosis virus.

Subgroup J avian leukosis virus (ALV) is a recently identified avian retrovirus associated with myeloid leukosis in meat-type chickens. The env gene of the HPRS-103 strain of ALV, the prototype of this subgroup, differs considerably from that of other subgroups, but shows close homology to the env-like sequences of members of the EAV family of endogenous retroviruses. Polymerase chain reaction (PCR) tests using two sets of primers were developed for the specific detection of the members of this new subgroup along with another pair of primers for detecting other subgroup viruses. The specificity and sensitivity of this detection system was compared with the conventional detection methods in experimentally and naturally infected samples. The use of PCR was found to be rapid, specific and more sensitive than the conventional diagnostic tests for the detection of ALV. Moreover, the two subgroup J ALV-specific PCR tests were found to be capable of differentiating between 'prototype-like' viruses and more recent isolates which show extensive antigenic and sequence variations. The use of this test as a rapid and sensitive method of detection of viruses in epidemiological studies and eradication programs is discussed.

Variation in plasma cholesterol concentration over time in the domestic fowl.

1. Variations in the concentration of plasma cholesterol available to the developing oocyte over a three week period in Gallus domesticus are described. 2. There are small changes in concentration between weeks for individual birds, but no changes between consecutive days within weeks or times within days. 3. It is recommended that future attempts to assess the relationship between variation in blood and yolk cholesterol concentrations estimate blood cholesterol concentration from two samples taken a week apart.

The relationship between yolk cholesterol and total lipid concentration throughout the first year of egg production in the domestic fowl.

1. Variation of egg yolk cholesterol and total lipid concentration (mg/g wet yolk) throughout the first year of egg production for the domestic fowl is described. 2. Yolk cholesterol concentration decreases between 20 and 30 weeks of age, and then remains constant until 70 weeks of age. Yolk total lipid concentration increases to a maximum at 40 weeks of age before decreasing to its original value at 60 and 70 weeks of age. 3. Bird means for yolk cholesterol and total lipid concentration are negatively related at 20 weeks of age, and essentially independent for the remainder of the first year of egg production. 4. Variation for yolk cholesterol concentration between 30 and 70 weeks of age appears to be a consequence of differences in the proportion of cholesterol to other lipid components. 5. Uptake of plasma lipoproteins characteristic of the immature bird is likely to be responsible for the high yolk cholesterol concentration at 20 weeks of age.

Variation in egg yolk cholesterol concentration between and within breeds of the domestic fowl.

1. The pattern of variation for egg yolk cholesterol concentration between 5 commercial egg layer lines and a cross of Gallus domesticus is described. 2. Yolk cholesterol concentration in the cross was lower than in the lines, and 6.7% lower than the midparent value. 3. It is proposed that the reduced yolk cholesterol concentration of the cross may be a consequence of heterosis, although sex-linkage and/or maternal effects cannot be discounted. 4. The difference between the cross and parental lines is consistent with a physiological relationship between yolk cholesterol concentration and rate of egg production, but not between yolk cholesterol concentration and yolk weight.