Identification of non-essential loci within the Meleagrid herpesvirus 1 genome

Abstract

BackgroundMeleagrid herpesvirus 1 (MeHV-1) infectious bacterial artificial chromosomes (iBACs) are ideal vectors for the development of recombinant vaccines for the poultry industry. However, the full potential of iBACS as vectors can only be realised after thorough genetic characterisation, including identification of those genetic locations that are non-essential for virus replication. Generally, transposition has proven to be a highly effective strategy for rapid and efficient mutagenesis of iBAC clones. The current study describes the characterisation of 34 MeHV-1 mutants containing transposon insertions within the pMeHV1-C18 iBAC genome.MethodsTn5 and MuA transposition methods were used to generate a library of 76 MeHV-1 insertion mutants. The capacity of each mutant to facilitate the recovery of infectious MeHV-1 was determined by the transfection of clone DNA into chicken embryo fibroblasts.ResultsAttempts to recover infectious virus from the modified clones identified 14 genetic locations that were essential for MeHV-1 replication in cell culture. Infectious MeHV-1 was recovered from the remaining 14 intragenic insertion mutants and six intergenic insertion mutants, suggesting that the respective insertion locations are non-essential for MeHV-1 replication in cell culture.ConclusionsThe essential and non-essential designations for those MeHV-1 genes characterised in this study were generally in agreement with previous reports for other herpesviruses homologues. However, the requirement for the mardivirus-specific genes LORF4A and LORF5 are reported for the first time. These findings will help direct future work on the development of recombinant poultry vaccines using MeHV-1 as a vector by identifying potential transgene insertion sites within the viral genome.Electronic supplementary materialThe online version of this article (doi10.1186/s12985-015-0362-9) contains supplementary material, which is available to authorized users.

Unraveling the evolutionary origin of the complex Nuclear Receptor Element (cNRE), a cis-regulatory module required for preferential expression in the atrial chamber

“…What could be the source of the cNRE? Galliform birds have been locked into a long co-evolutionary battle with viruses of the Alphaherpesviridae family, such as Marek's disease virus (MDV/GaHV2), a DNA virus capable of integrating its sequences into the genome of the host germline (Nair, 2005;Waidner et al, 2009;Morissette & Flamand, 2010;Hall et al, 2015). We found at least one cNRE-like signature in GaHV1 and six cNRE-like sequences in HSV1, suggesting that cNREs do reside in the sequence of Alphaherpesvirus.…”

“…To test this hypothesis, we searched for cNRE-like sequences in Alphaherpesviruses that infect galliform birds, such as GaHV1/ILTV, the virus responsible for avian infectious laryngotracheitis in chicken, GaHV2/MDV1, GaHV3/MDV2, a non-pathogenic virus, and MeHV1/HTV, a non-pathogenic Meleagrid herpesvirus 1. The human herpes simplex virus 1 (HSV1) was included as an outgroup (Waidner et al, 2009;Hall et al, 2015).…”

Herpesvirus of Turkeys as a Vaccine Vector in Viral Diseases: Pros and Cons

“…The disruption of intergenic regions usually does not alter the virulence of herpesviruses, whereas virulenceassociated gene indels do; therefore, the latter is considered to reduce virulence and insert an expression cassette simultaneously. At least eleven non-essential genes have been recognized in the HVT genome so far (Hall et al, 2015) and US2, US7 or UL40 genes; US3/ US4, UL54/UL55, UL45/46 or HVT065/HVT066 intergenic regions have been used for foreign gene insertion. Foremost examples of the recombinant HVT vaccines are Newcastle disease (ND) vaccines.…”