Characterization of a recombinant fowlpox virus expressing the native hexon of hemorrhagic enteritis virus.

The structure of the icosahedral adenovirus capsid is highly conserved among Adenoviridae. In its native form, the hexon is the major capsid protein. The nascent hexon requires the 100 kDa folding protein to fold into its native, trimeric form.

Protection and synergism by recombinant fowl pox vaccines expressing genes from Marek's disease virus.

Recombinant fowl poxviruses (rFPV) were constructed to express genes from serotype 1 Marek's disease virus (MDV) coding for glycoproteins B (gB1), C (gC), and D (gD) and tegument proteins UL47 and UL48, as well as genes from serotypes 2 and 3 MDV coding for glycoprotein B (gB2 and gB3). These rFPVs, alone and in various combinations, including combinations of fowl poxvirus (FPV)/gBs with turkey herpesvirus (HVT), were evaluated for ability to protect maternal antibody-positive (ab+) and -negative (ab-) chickens against challenge with highly virulent MDV isolates. The protective efficacy was also compared with that of prototype Marek's disease (MD) vaccines.

A recombinant fowlpox virus expressing the envelope antigen of subgroup A avian leukosis/sarcoma virus.

A recombinant fowlpox virus (FPV) was constructed by inserting cloned sequences from Schmidt-Ruppin subgroup A avian sarcoma virus coding for the viral envelope (env) antigen into a nonessential region of FPV DNA downstream from a synthetic promoter. Sera from chickens hyperimmunized with the recombinant FPV neutralized the infectivity of the homologous subgroup A virus (RCASBP/AP) but only weakly neutralized the infectivity of Rous sarcoma virus, another subgroup A avian leukosis virus. Similarly, vaccination of 1-day-old chicks with this recombinant FPV protected against infection with RCASBP/AP virus but not against infection with another subgroup A Rous-associated virus (RAV-1).

Mutational analysis of the proteolytic cleavage site of glycoprotein B (gB) of Marek's disease virus.

The Marek's disease virus (MDV) glycoprotein B (gB) precursor, gp100, is proteolytically cleaved into two disulfide-linked subunits, gp60 and gp49. In the gB homologs of most other herpesviruses, a tetrapeptide, Arg-Xaa-Arg-Arg, is immediately upstream from the predicted cleavage site. We have investigated the specificity of the proteolytic cleavage in gp100 by introducing mutations within its predicted cleavage site (Arg-Leu-Arg-Arg) and expressed these mutants in recombinant fowlpox virus (FPV).

Identification and characterization of a Marek's disease virus gene homologous to glycoprotein L of herpes simplex virus.

We have identified three Marek's disease virus (MDV) open reading frames (ORFs) within the BamHI D fragment of MDV genome. The predicted polypeptides are homologous to UL1 (glycoprotein L, gL), UL2 (uracil-DNA glycosylase), and UL3 (nuclear localizing phosphoprotein) of herpes simplex virus type 1 (HSV-1). Comparison of the deduced amino acid sequences of these three ORFs with HSV-1 counterparts revealed overall identities of 18, 43, and 49%, respectively.

The glycoprotein B genes of Marek's disease virus serotypes 2 and 3: identification and expression by recombinant fowlpox viruses.

The nucleotide sequences of the glycoprotein B (gB) genes of Marek's disease virus (MDV) serotypes 2 and 3 were determined (gB-2 and gB-3, respectively). The genomic locations of these genes coincide with that of the gB gene of serotype 1 MDV (gB-1). Alignment with gB-1 (Ross et al.

An immunoperoxidase plaque assay for reticuloendotheliosis virus and its application to a sensitive serum neutralization assay.

A rapid assay for the enumeration of reticuloendotheliosis virus (REV) is described. Chicken embryo fibroblast monolayer cultures were infected with REV and incubated 6 days under an agar overlay. After removal of the overlay, cells were fixed with acetone/ethanol.

Nucleotide and predicted amino acid sequences of Marek's disease virus homologues of herpes simplex virus major tegument proteins.

The DNA sequence of an 8.4 kbp BamHI-EcoRI fragment of Marek's disease virus (MDV) strain GA was determined. Three of the predicted polypeptides are homologous to UL47, UL48 and UL49 encoding the major tegument proteins of herpes simplex virus type 1 (HSV-1), and four are homologous to HSV-1 UL45, UL46, UL49.

Fowlpox virus recombinants expressing the envelope glycoprotein of an avian reticuloendotheliosis retrovirus induce neutralizing antibodies and reduce viremia in chickens.

Eight stable fowlpox virus (FPV) recombinants which express the envelope glycoprotein of the spleen necrosis virus (SNV) strain of reticuloendotheliosis virus (REV), an avian retrovirus, were constructed. These recombinants differ in the genomic location of the inserted genes, in the orientation of the insert relative to flanking viral sequences, and in the promoter used to drive expression of the env gene. Of these variables, promoter strength seems to be the most crucial.

Insertional inactivation of a fowlpox virus homologue of the vaccinia virus F12L gene inhibits the release of enveloped virions.

Insertion of the Escherichia coli lacZ gene into a ClaI restriction enzyme site of a 5.7 kb HindIII fragment of the fowlpox virus (FPV) genome resulted in the generation of stable recombinants. These recombinants produced plaques that were significantly smaller than those produced by parental FPV or by FPV recombinants containing the lacZ gene at other non-essential sites.

Identification and functional analysis of the fowlpox virus homolog of the vaccinia virus p37K major envelope antigen gene.

A fowlpox virus (FPV) gene with homology to the vaccinia virus p37K major envelope antigen gene was identified and sequenced. The predicted product has a molecular weight of 43,018 Da (p43K). The FPV p43K gene has 37.

Protection against Marek's disease by a fowlpox virus recombinant expressing the glycoprotein B of Marek's disease virus.

Fowlpox virus (FPV) recombinants expressing the glycoprotein B and the phosphorylated protein (pp38) of the GA strain of Marek's disease virus (MDV) were assayed for their ability to protect chickens against challenge with virulent MDV. The recombinant FPV expressing the glycoprotein B gene elicited neutralizing antibodies against MDV, significantly reduced the level of cell-associated viremia, and, similar to the conventional herpesvirus of turkeys, protected chickens against challenge with the GA strain and the highly virulent RB1B and Md5 strains of MDV. The recombinant FPV expressing the pp38 gene failed to either elicit neutralizing antibodies against MDV or protect the vaccinated chickens against challenge with MDV.

Recombinant fowlpox viruses expressing the glycoprotein B homolog and the pp38 gene of Marek's disease virus.

Two Marek's disease virus (MDV) genes, one homologous to the glycoprotein B gene of herpes simplex virus and encoding the B antigen complex and the other encoding a 38-kDa phosphorylated protein (pp38), were inserted into the fowlpox virus (FPV) genome under the control of poxvirus promoters. Randomly selected nonessential regions of FPV were used for insertion, and the vaccinia virus 7.5 kDa polypeptide gene promoter or a poxvirus synthetic promoter was used for expression of MDV genes.

Structural analysis of unstable intermediate and stable forms of recombinant fowlpox virus.

The stability and structure of the products of recombination in a fowlpox virus (FPV) system using the thymidine kinase (TK) gene as the insertion site were examined. A 4.6 kb chimeric DNA fragment from the pUV1 expression vector, containing the bacterial lacZ gene and the vaccinia virus P7.

Structural polypeptides of type II avian adenoviruses analyzed by monoclonal and polyclonal antibodies.

Polypeptides of hemorrhagic enteritis virus (HEV) of turkeys and marble spleen disease virus (MSDV) of pheasants were analyzed by immune precipitation and immunoblot assays. A total of 11 polypeptides ranging in molecular weight from 14,000 to 97,000 were detected in lysates of HEV-infected turkey cells analyzed by immunoblot assay using a polyclonal antibody against HEV. Identical patterns were observed with preparations of MSDV.

A double-antibody enzyme-linked immunosorbent assay for the detection of turkey hemorrhagic enteritis virus antibody and antigen.

A highly sensitive and specific double-antibody enzyme-linked immunosorbent assay (ELISA) is described for the detection of antigen and antibody of turkey hemorrhagic enteritis virus (HEV). The assay utilizes a virus-neutralizing monoclonal antibody (MAb) to capture the antigen and turkey antiserum against HEV as the second antibody. Microtiter plates were first coated with a dilution of 1:3000 of the MAb (300 ng immunoglobulin/well) and are used for detection of both antigen and antibody.

Transient expression assay for qualitative assessment of gene expression by fowlpox virus.

A transient expression assay for fowlpox virus (FPV) was developed to assess the feasibility of using heterologous promoters in FPV and to qualitatively determine relative promoter strength. A transient expression system for FPV has not been reported, and various methods used for transient expression in vaccinia-virus-infected cells produced negative results when used with FPV. Here a successful method for transient expression of E.

Hemorrhagic enteritis of turkeys: influence of maternal antibody and age at exposure.

The effect of maternal antibody (MAB) to hemorrhagic enteritis (HE) on the response of turkeys to infection with virulent and avirulent strains of HE virus (HEV) was examined. The influence of age at exposure and treatment with HEV antibody on development of clinical HE also was studied. MAB protected poults from clinical HE for up to 6 weeks of age.

Structural proteins of two different plaque-size phenotypes of fowlpox virus.

Structural polypeptides of two plaque-purified variant isolates of fowlpox virus differing in plaque morphology and size were examined by Coomassie blue-staining and immunoblot analysis of purified virions. A total of 30 structural polypeptides were observed, ranging in molecular weight from 14,100 to 122,600. A late polypeptide of 36,400 molecular weight was quite prominent in the small-plaque clone but absent in the large-plaque clone.

Some observations on the response of ring-necked pheasants to inoculation with various strains of cell-culture-propagated type II avian adenovirus.

The response of ring-necked pheasants to inoculation with three strains of cell-culture-propagated type II avian adenovirus was examined. Marble spleen disease (MSD) virus of pheasants and both avirulent and virulent strains of hemorrhagic enteritis virus (HEV) of turkeys all induced typical gross and microscopic splenic lesions of MSD; neither MSD-associated lung lesions nor mortality were noted in inoculated pheasants, regardless of strain of virus used. Pheasants inoculated with a cell-culture-propagated avirulent strain of HEV were properly immunized against challenge with virulent HEV, as indicated by seroconversion and by protection against virus-induced splenic lesions.

Properties of producer and non-producer clones of a Marek's disease turkey lymphoblastoid cell line.

The MDTC-RP30 lymphoblastoid cell line established from Marek's disease (MD) tumors in turkeys consisted of a heterogeneous population of cells 10 to 25 micron in diameter. Large-cell fractions obtained from a bovine fetal serum gradient had a higher titer of cell-associated MD virus (MDV) than the small-cell fractions. Seven single-cell clones were established from MDTC-RP30 cell line: two consisted of large cells, and the other clones consisted of small cells.

Further studies on in vitro and in vivo assays of hemorrhagic enteritis virus (HEV).

Isolation of hemorrhagic enteritis virus (HEV) from spleens of infected turkeys in the MDTC-RP19 lymphoblastoid cell line was compared with detection of HEV antigen in the same spleens using the agar gel precipitation (AGP) test. A concordance of 80% was found between the two assays. Virus isolation had a sensitivity of 84% and a specificity of 88% compared with the AGP test.

Field vaccination against hemorrhagic enteritis of turkeys by a cell-culture live-virus vaccine.

A cell-culture-propagated (CC) live-virus hemorrhagic enteritis (HE) vaccine was evaluated for efficacy and safety in two field trials conducted in North Carolina (NC) and Minnesota (MN). At 4 or 5 1/2 weeks of age, 9,839 poults in NC and 15,857 poults in MN were vaccinated with a CC HE vaccine administered via the drinking water. A comparable number of poults were maintained as unvaccinated controls.