Deletion of the non-essential UL0 gene of infectious laryngotracheitis (ILT) virus leads to attenuation in chickens, and UL0 mutants expressing influenza virus haemagglutinin (H7) protect against ILT and fowl plague

Abstract

Infectious laryngotracheitis virus (ILTV), a member of the Alphaherpesvirinae, possesses several unique genes. One of them, UL0, encodes an abundantly expressed protein that accumulates in the nuclei of ILTV-infected cells. This study demonstrates that this protein is dispensable for in vitro virus replication and that UL0 deletion mutants exhibit only minor growth defects in cultured cells. The UL0 gene locus of ILTV was also used for insertion of foreign DNA sequences encoding enhanced GFP or haemagglutinin (HA), subtype H7, of a highly pathogenic avian influenza virus under the control of the human cytomegalovirus immediate-early gene promoter. Expression of foreign proteins was shown by (immuno)fluorescence tests and Western blot analyses. After experimental infection of chickens, UL0 deletion mutants proved to be attenuated when compared to both parental wild-type ILTV and an UL0 rescue mutant. Nevertheless, all animals immunized with UL0-negative ILTV were protected from clinical disease after subsequent infection with virulent ILTV. Furthermore, all animals immunized with HA-expressing ILTV survived a lethal challenge with H7 subtype avian influenza virus with minimal clinical signs. Thus, an UL0-negative and HA-expressing ILTV recombinant may be used as a bivalent live virus vaccine against ILT and fowl plague. Unlike inactivated influenza virus vaccines, HA-expressing ILTV recombinants should be suitable for mass application and would also permit serological discrimination between vaccinated and virus-infected animals in the field.

Epidemiology of Avian Infectious Laryngotracheitis with Special Focus to South America: an update

“…Some viruses with deleted genes may retain their ability to induce immune response while not producing clinical signs or latency. ILT viruses with deletions of the genes gC (Pavlova et al, 2010), gG (Devlin et al, 2006), gJ (Fuchs et al, 2005), TK (Han et al, 2002), UL0 (Veits et al, 2003), and UL 47 (Helferich et al, 2007) showed attenuation and may be used for vaccine production. A gG-deficient ILTV strain administered to 3-week-old SPF chickens via drinking water and eye drop induced adequate immunity against challenge with a wild strain and may potentially be used for large-scale vaccination; however, further studies are needed before it is applied to commercial poultry flocks (Devlin et al, 2008).…”

Attenuation and protection efficacy of ORF C gene-deleted recombinant of infectious laryngotracheitis virus

“…Growth kinetics analysis of these gene deletion recombinants indicated that they are non-essential for virus replication in cell culture (Fuchs et al, 2007). In chickens the DU L 50 (dUTPase) (Fuchs et al, 2000) and DU L 44 (gC) (Pavlova et al, 2010) gene-deleted recombinants still exhibited some degree of virulence, while the DU S 4 (gG) (Devlin et al, 2006), DU L 47 (tegument protein) (Helferich et al, 2007), DU L 23 (thymidine kinase) (Han et al, 2002), DU L 0 (nuclear protein) (Veits et al, 2003b), and DU S 5 (gJ) recombinants exhibited moderate to marked degrees of attenuation.…”

“…Among the 20 gene-deleted ILTV strains generated, only seven have been evaluated in vivo for attenuation and protection efficacy. Individually, these mutants contained deletions in the genes encoding a nuclear protein of unknown function (U L 0 gene), (Veits et al, 2003b), thymidine kinase (U L 23) (Han et al, 2002;Schnitzlein et al, 1995), dUTPase (U L 50) (Fuchs et al, 2000), glycoprotein G (U S 4) (Devlin et al, 2006), glycoprotein J (U S 5) Mundt et al, 2011), glycoprotein C (U L 44) (Pavlova et al, 2010), and the tegument protein (U L 47) (Helferich et al, 2007). Growth kinetics analysis of these gene deletion recombinants indicated that they are non-essential for virus replication in cell culture (Fuchs et al, 2007).…”

Molecular biology of avian infectious laryngotracheitis virus

“…Thus, these genes apparently do not represent crucial adaptations to avian host cells, although more important functions during infection of chickens cannot be excluded. Deletion of the ILTV-specific UL0 gene also caused only minor in vitro growth defects, whereas isolation of ILTV mutants lacking the adjacent UL(-1) gene was not possible [92]. This finding indicates that UL0 and UL(-1), although structurally related, are not functionally equivalent.…”