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.

Nucleotide sequence of the left-terminus of infectious laryngotracheitis virus (Gallid herpesvirus 1) SA-2 strain.

The nucleotide sequence of 10.6 kilobase pairs (kbp) at the leftterminus of infectious laryngotracheitis virus (ILTV) SA-2 vaccine strain was determined. Several features were elucidated, including, 102 base pair (bp) inverted repeats separated by 750 bp of unique sequence which contains an NF-1 binding site indicating that the terminal may be a site for an origin of replication. Other direct repeats were also found in this region. To the right of the inverted repeat region, a 2130 bp region was found to contain small open reading frames (ORFs) of less than 100 aa. Another potential ORF was found to the right of the region containing the small ORFs which consisted of two 184 bp direct repeats inserted into the reading frame, which would truncate the putative product. Only one copy of this repeat was found in the corresponding homologue of the wild type strain SA-0. Six other ORFs were found, which shared little or no identity to homologues of other alphaherpesviruses, suggesting that these putative genes are unique to ILTV.

Nucleotide sequence of canine herpesvirus homologues of herpes simplex virus type 1 US2, US3, glycoproteins I and E, US8.5 and US9 genes.

The partial nucleotide sequence of two BamHI fragments that span the unique short region (US), terminal repeat region (TR) and internal repeat region (IR) of canine herpesvirus (CHV) has been determined. Data obtained revealed several open reading frames (ORF's) identified as the US2, US3, gI, gE and US9 homologues of herpes simplex virus type 1 (HSV1). The CHV homologues also show significant identity in amino acid sequence with those encoded by feline herpesvirus type 1 (FHV1), bovine herpesvirus (BHV1) and equine herpesvirus (EHV1). Translation of another ORF showed little amino acid identity with the gene products of other alpha-herpesviruses. Its genomic position relative to the other CHV homologues would suggest it is the US8.5 gene of CHV.

Comparison of the genomic short regions of a vaccine strain (SA-2) and a virulent strain (CSW-1) of infectious laryngotracheitis virus (Gallid herpesvirus 1).

Restriction enzyme linkage maps were produced for the genomic short region of the virulent infectious laryngotracheitis virus (CSW-1 strain). After comparison with the equivalent restriction enzyme linkage maps for the infectious laryngotracheitis virus SA-2 strain (a vaccine strain), it was determined that the maps for the short regions of the two strains were identical, apart from a single section in each of the inverted terminal repeats. Each inverted terminal repeat of the SA-2 strain was discovered to contain 467 base pairs more DNA than the CSW-1 strain's inverted terminal repeats. This extra DNA was more precisely mapped entirely within the EcoRI fragments D and d of SA-2, which were found to form part of the SmaI fragments U and P of SA-2 and Q and b of SA-2 and to contain one SmaI restriction enzyme site.

Molecular evolution of infectious laryngotracheitis virus (ILTV; gallid herpesvirus 1): an ancient example of the Alphaherpesviridae?

An analysis of two essential genes of infectious laryngotracheitis virus (ILTV), glycoprotein D (gD) and the immediate early gene, herpes simplex virus homologue ICP27, was performed with the equivalent gene homologues from several alphaherpesviruses. Amino acid (aa) sequence analysis revealed that these ILTV genes shared limited homology to other alphaherpesvirus equivalents and were distinct from the two other avian herpesviruses, Marek's disease virus (MDV) and herpesvirus of turkeys (HVT). Simplex and varicella group viruses are clearly separate from the avian group. The amino acid sequences of these ILTV genes will be presented with comparisons to the homologues from other alphaherpes viruses, contributing further evidence of the evolution of this group of viruses from a common progenitor and that ILTV could be an ancient example of the Alphaherpesvirinae.

Nucleotide sequence of infectious laryngotracheitis virus (gallid herpesvirus 1) ICP4 gene.

The infectious laryngotracheitis virus (ILTV) gene encoding a homologue to the ICP4 protein of herpes simplex virus (HSV) has been mapped to the inverted repeat region. The complete nucleotide sequence of ILTV ICP4 has been determined. The ILTV ORF encoding ICP4 is 4386 nucleotides long, calculated from the first of four ATG codons, and has an overall G+C content of 59%. The ILTV ICP4 contains two domains of high homology which have been reported in other studies to be conserved in the ICP4 homologues of alphaherpesviruses, and to be functionally important. Several regulatory features were identified including a serine-rich domain in region one. A more extensive serine-rich domain was located in region five which is also found in varicella-zoster virus (VZV) and bovine herpesvirus 1. A 5.4 kb immediate early transcript was identified in infected primary kidney cells.

Sequence characteristics of a gene in infectious laryngotracheitis virus homologous to glycoprotein D of herpes simplex virus.

An infectious laryngotracheitis virus (ILTV, gallid herpesvirus 1) gene homologous to glycoprotein D of herpes simplex virus (HSV) was identified and characterized by its nucleotide and derived amino acid sequence. The ILTV gD gene is located in the unique short region (U(s)) and contains an open reading frame capable of specifying a polypeptide of 380 amino acids, including N- and C- terminal hydrophobic domains consistent with signal and anchor regions respectively, and no potential sites for N-glycosylation. Alignment of the amino acid sequence with those published for HSV gD, equine herpesvirus type 1 (EHV-1) gD, pseudorabies virus (PRV) gp50, Marek's disease virus (MDV) gD, herpesvirus of turkeys (HVT) gD and bovine herpesvirus type 1 (BHV-1) gD showed similarities over the N-terminal region, with the greatest differences occurring in the C-terminal. The identical positioning of 6 cysteine residues supports the hypothesis of common ancestry of herpesvirus family (McGeoch, 1990) and is consistent with the essential role of this glycoprotein.

ICP27 immediate early gene, glycoprotein K (gK) and DNA helicase homologues of infectious laryngotracheitis virus (gallid herpesvirus 1) SA-2 strain.

A 4.8 kilobase segment located at the left-terminal in the unique long (UL) region of infectious laryngotracheitis virus (ILTV) SA-2 strain contained three open reading frames (ORFs). The first of 421 amino acids (aa) was located at map units 0.065 to 0.07, and its predicted 48 kiloDaltons (kDa) protein product has significant homology to the immediate early regulatory protein ICP27 (UL54) of herpes simplex virus type-1 (HSV-1), to varicella-zoster virus (VZV) ORF4 and to equine herpesvirus 1 (EHV-1) ORF5. The zinc finger conserved in the C-terminal of the proteins from HSV-1, VZV and EHV-1, is poorly conserved in ILTV homologue. The second ORF of 336 aa, located at map units 0.075 to 0.08, has a predicted molecular weight (MW) of 38 kDa with significant homology to glycoprotein K (gK) of HSV-1 (UL53), ORF5 of VZV and ORF6 of EHV-1. ILTV gK has features characteristic of a membrane-bound glycoprotein. The 3' region of a third ORF was located at map units 0.08 to 0.095. Translation of the sequence revealed significant homology to the 3'-region of the DNA helicase-primase complex protein (UL52) of HSV-1, ORF6 of VZV and ORF 7 of EHV-1. Northern blot analyses were used to characterize the ILTV ICP27, gK and DNA helicase mRNAs. The data revealed that ILTV ICP27 is an immediate early gene that encodes a 1.6 kb mRNA, ILTV gK encodes a late transcript of 1.8 kb, while ILTV DNA helicase encodes a late transcript of 3.7 kb.