Complete Genome Sequence of a J Subgroup Avian Leukosis Virus Isolated from Local Commercial Broilers

Abstract

Subgroup J avian leukosis virus (ALV-J) isolate GDKP1202 was isolated from a 50-day-old local yellow commercial broiler in the Guangdong province of China in 2012. Here we report the complete genomic sequence of the GDKP1202 isolate, which caused high mortality, serious growth suppression, thymic atrophy, and liver enlargement in commercial broilers. A novel potential binding site (5′-GGCACCTCC-3′) for c-myb was identified in the GDKP1202 genome. These findings will provide additional insights into the molecular characteristics in the genomes and pathogenicity of ALV-J.

GENOME ANNOUNCEMENT

Subgroup J avian leukosis virus (ALV-J) belongs to the genus Alpharetrovirus of the Retroviridae family and was first detected from meat-type chickens in 1999 in China (10). ALV-J primarily induces myeloid leukosis (ML) in meat-type chickens and causes enormous economic losses for the poultry industry worldwide (2, 8). A new isolate, designated GDKP1202, was isolated from 50-day-old broilers from a local commercial flock that suffered from high mortality, serious growth suppression, thymic atrophy, and liver enlargement with diffuse gray-white nodules in the Guangdong province of China in 2012. Histological examination indicated that the tumor cells proliferated in tissues were myelocytes with eosinophilic granules in cytoplasm. The virus was propagated in a chicken embryo fibroblast cell line (DF-1 cells). The whole genome of GDKP1202 was amplified by PCR and cloned into a pMD19-T vector (TaKaRa Bio Inc., Japan), sequenced three times, and assembled using DNAStar (version 7). Multiple-sequence alignment was performed with Clustal X (BioEdit version 7). The transcriptional regulatory elements in noncoding regions of the genome were analyzed with SoftBerry (Softberry, Inc., Mt. Kisco, NY).

Comparative analyses showed that env, gag, and pol of the GDKP1202 isolate genome shared the most homology—95.0%, 96.5%, and 97.3%, respectively—with the reported hemangioma ALV-J sequences in GenBank. The long terminal repeat (LTR) and E elements shared more identity with ML isolates (1, 3). The GDKP1202 did not harbor the 19-bp insertion (5′CGGTTGCTCTGCGTGATTC3′) in leader sequence which usually appears in hemangioma strains (7). Almost all of the putative transcription regulatory elements identified in the U3 region of the LTR of the GDKP1202 isolate were conserved and homologous to those of ALV-J broiler isolates, indicating the importance of these regulatory elements during viral replication. However, the second PRE box in GDKP1202 rarely shared a single nucleotide substitution (G to A) with the sequence GATGG, even though the first box was reportedly variable (9). Almost all of the redundant transmembrane region (rTM) of the GDKP1202 genome was deleted by comparing it with other ML isolates, such as HRPS103, NX0101, JS-nt, and SD07LK1, suggesting that this region is not necessary for viral replication and tumor type determination (3). The binding site (5′TAACTG3′) for the Myb was identified in the E element of GDKP1202 genome, as for other ALV-J isolates (4–6). However, a single nucleotide substitution (T27C) of the E element resulted in the identification of a novel potential binding site (5′-GGCACCTCC-3′) for c-myb by using the SoftBerry program. In addition, an analysis of the E-element sequence revealed unique nucleotide substitutions (G66A, T98C, G123C, and C124T) and a deletion of three consecutive nucleotides (GAA) (bases 10 to 12) in this region of the GDKP1202 genome in comparison to other E-element sequences. Although an intact E element was not a critical requirement for the ALV-J induction of tumors, the presence of the E element could be associated with a higher frequency of tumors in susceptible chickens (3). These present findings will help us to understand the molecular characteristics of genomes and pathogenicity of ALV-J.

Nucleotide sequence accession number.

The complete genome sequence of the GDKP1202 isolate was submitted to GenBank, and the assigned accession number is JX453210.