Comparative analysis of Marek's disease virus (MDV) glycoprotein-, lytic antigen pp38- and transformation antigen Meq-encoding genes: association of meq mutations with MDVs of high virulence

Marek's disease (MD) is a highly contagious lymphoproliferative and demyelinating disorder of chickens. MD is caused by Marek's disease virus (MDV), a cell-associated, acute-transforming alphaherpesvirus. For three decades, losses to the poultry industry due to MD have been greatly limited through the use of live vaccines. MDV vaccine strains are comprised of antigenically related, apathogenic MDVs originally isolated from chickens (MDV-2), turkeys (herpesvirus of turkeys, HVT) or attenuated-oncogenic strains of MDV-1 (CVI-988). Since the inception of high-density poultry production and MD vaccination, there have been two discernible increases in the virulence of MDV field strains. Our objectives were to determine if common mutations in the major glycoprotein genes, a major lytic antigen phosphoprotein 38 (pp38) or a major latency/transformation antigen Meq (Marek's EcoRI-Q-encoded protein) were associated with enhanced MDV virulence. To address this, we cloned and sequenced the major surface glycoprotein genes (gB, gC, gD, gE, gH, gI, and gL) of five MDV strains that were representative of the virulent (v), very virulent (vv) and very virulent plus (vv+) pathotypes of MDV. We found no consistent mutations in these genes that correlated strictly with virulence level. The glycoprotein genes most similar among MDV-1, MDV-2 and HVT (gB and gC, approximately 81 and 75%, respectively) were among the most conserved across pathotype. We found mutations mapping to the putative signal cleavage site in the gL genes in four out of eleven vv+MDVs, but this mutation was also identified in one vvMDV (643P) indicating that it did not correlate with enhanced virulence. In further analysis of an additional 12 MDV strains, we found no gross polymorphism in any of the glycoprotein genes. Likewise, by PCR and RFLP analysis, we found no polymorphism at the locus encoding the pp38 gene, an early lytic-phase gene associated with MDV replication. In contrast, we found distinct mutations in the latency and transformation-associated Marek's EcoRI-Q-encoded protein, Meq. In examination of the DNA and deduced amino acid sequence of meq genes from 26 MDV strains (9 m/vMDV, 5 vvMDV and 12 vv+MDVs), we found distinct polymorphism and point mutations that appeared to correlate with virulence. Although a complex trait like MDV virulence is likely to be multigenic, these data describe the first sets of mutations that appear to correlate with MDV virulence. Our conclusion is that since Meq is expressed primarily in the latent/transforming phase of MDV infection, and is not encoded by MDV-2 or HVT vaccine viruses, the evolution of MDV virulence may be due to selection on MDV-host cell interactions during latency and may not be mediated by the immune selection against virus lytic antigens such as the surface glycoproteins.     

A comparative evaluation of the protective efficacy of rMd5deltaMeq and CVI988/ Rispens against a vv+ strain of Marek's disease virus infection in a series of recombinant congenic strains of White Leghorn chickens

Marek's disease (MD) is a lymphoproliferative disease of domestic chickens caused by a highly infectious, oncogenic alpha-herpesvirus known as Marek's disease virus (MDV). MD is presently controlled by vaccination. Current MD vaccines include attenuated serotype 1 strains (e.g., CVI988/Rispens), avirulent serotype 2 (SB-1), and serotype 3 (HVT) MDV strains. In addition, recombinant MDV strains have been developed as potential new and more efficient vaccines to sustain the success of MD control in poultry. One of the candidate recombinant MDV strains, named rMd5deltaMeq, was derived from Md5, a very virulent strain of MDV lacking the MDV oncogene Meq. Our earlier reports suggest that rMd5deltaMeq provided protection equally well or better than commonly used MD vaccines in experimental and commercial lines of chickens challenged with very virulent plus (vv+) strains of MDV. In this study, maternal antibody-positive (trial 1) and negative (trial 2) chickens from a series of relatively MD resistant lines were either vaccinated with the rMd5deltaMeq or CVI988/Rispens followed by infection of a vv+ strain of MDV, 648A, passage 10. This report presents experimental evidence that the rMd5deltaMeq protected significantly better than the CVI988/Rispens (P < 0.01) in the relatively resistant experimental lines of chickens challenged with the vv+ strain of MDV. Together with early reports, the rMd5deltaMeq appeared to provide better protection, comparing with the most efficacious commercially available vaccine, CVI988/Rispens, for control of MD in lines of chickens regardless of their genetic background.     

Susceptibility of adult chickens, with and without prior vaccination, to challenge with Marek's disease virus

Marek's disease (MD) outbreaks can occur in previously healthy adult layer or breeder flocks. However, it is not clear whether such outbreaks are caused by recent challenge with highly virulent (vv and vv+) strains of MD virus (MDV; i. e., new infection hypothesis) or by exacerbation of an earlier MDV infection (i. e., old infection hypothesis). To discriminate between these hypotheses, adult White Leghorn chickens of laboratory strains or commercial crosses with or without prior vaccination or MDV exposure were challenged at 18-102 wk of age with highly virulent MDVs, and lesion responses were measured. Horizontal transmission was studied in one trial. Challenge of adult chickens, which were free from prior MDV vaccination or exposure, with highly virulent MDV strains induced transient paralysis or tumors in 60%-100% of 29 groups (mean = 91%), and horizontal spread of virus was detected. The magnitude of the response was similar to that induced by challenge at 3 wk of age. In contrast, comparable challenge of adult chickens, which had been vaccinated or exposed to MDV early in life, induced transient paralysis or tumors in 0%-6% of 12 groups (mean = 0. 5%), although some birds showed limited virologic evidence of infection and transmission of the virus to contacts. The MD responses were influenced by the virulence of the challenge virus strain, and to a lesser extent by virus dose and route of exposure. Strong inflammatory lesions were induced in the brain and nerves of adult specific pathogen-free (SPF) chickens at 9-15 days after infection. The low susceptibility of previously vaccinated and exposed groups to challenge at > or =18 wk of age suggests that late outbreaks of MD in commercial flocks are not likely a result of recent challenge alone and that additional factors could be involved.     

The efficacy of recombinant fowlpox vaccine protection against Marek's disease: its dependence on chicken line and B haplotype

Earlier studies have shown that the B haplotype has a significant influence on the protective efficacy of vaccines against Marek's disease (MD) and that the level of protection varies dependent on the serotype of MD virus (MDV) used in the vaccine. To determine if the protective glycoprotein gene gB is a basis for this association, we compared recombinant fowlpox virus (rFPV) containing a single gB gene from three serotypes of MDV. The rFPV were used to vaccinate 15.B congenic lines. Nonvaccinated chickens from all three haplotypes had 84%-97% MD after challenge. The rFPV containing gB1 provides better protection than rFPV containing gB2 or gB3 in all three B genotypes. Moreover, the gB proteins were critical, since the B*21/*21 chickens had better protection than chickens with B*13/*13 or B*5/*5 using rFPV with gB1, gB2, or gB3. A newly described combined rFPV/gB1gEgIUL32 + HVT vaccine was analyzed in chickens of lines 15 x 7 (B*2/*15) and N (B*21/*21) challenged with two vv+ strains of MDV. There were line differences in protection by the vaccines and line N had better protection with the rFPV/gB1gEgIUL32 + HVT vaccines (92%-100%) following either MDV challenge, but protection was significantly lower in 15 X 7 chickens (35%) when compared with the vaccine CVI988/Rispens (94%) and 301B1 + HVT (65%). Another experiment used four lines of chickens receiving the new rFPV + HVT vaccine or CVI988/Rispens and challenge with 648A MDV. The CVI 988/Rispens generally provided better protection in lines P and 15 X 7 and in one replicate with line TK. The combined rFPV/gB1gEgIUL32 + HVT vaccines protected line N chickens (90%) better than did CVI988/Rispens (73%). These data indicate that rFPV + HVT vaccines may provide protection against MD that is equivalent to or superior to CVI988/ Rispens in some chicken strains. It is not clear whether the rFPV/gB1gEgIUL32 + HVT vaccine will offer high levels of protection to commercial strains, but this vaccine, when used in line N chickens, may be a useful model to study interactions between vaccines and chicken genotypes and may thereby improve future MD vaccines.     

Partial protection against Marek's disease in chickens immunized with glycoproteins gB purified from turkey-herpesvirus-infected cells by affinity chromatography coupled with monoclonal antibodies

Glycoproteins gB of Marek's disease virus (MDV) and herpesvirus of turkeys (HVT) related to virus neutralization were purified from HVT-infected cells by affinity chromatography. Immunization of chickens with purified glycoproteins gB resulted in partial protection against MD. Neutralizing antibodies were detected in chickens immunized with HVT-gB.     

Coinfection of specific-pathogen-free chickens with Marek's disease virus (MDV) and chicken infectious anemia virus: effect of MDV pathotype

Both Marek's disease virus (MDV) and chicken infectious anemia virus (CIAV) infections are prevalent in chickens throughout the world. In the past decade, MDV strains with increased virulence (very virulent plus MDV pathotype [vv+MDV]) have been isolated. The purpose of this experiment was to determine the effects of coinfection of chickens with CIAV and a vv+MDV isolate. Specific-pathogen-free chickens were inoculated at 1 day posthatch with RB1B (very virulent MDV pathotype [vvMDV]) only, 584A (vv+MDV) only, CIAV only, RB1B + CIAV, 584A + CIAV, or nothing. Samples of spleen, thymus, and bursa of Fabricius were collected at 4, 7, 10, and 13 days postinoculation (DPI). Thymic and bursal atrophy at 13 DPI and final mortality at 30 DPI were significantly greater in chickens inoculated with 584A with or without added CIAV, or with RB1B plus CIAV, compared with birds inoculated with RB1B alone. Both amounts of virus reisolated and levels of virus detected by quantitative-competitive polymerase chain reaction were greater at 4 DPI in 584A inoculates compared with RB1B inoculates. To monitor the early cytolytic infection, northern analysis was done with a probe for the MDV immediate early gene ICP4 (infected cell protein 4). In the absence of CIAV, ICP4 expression was more apparent in chickens inoculated with 584A than in those inoculated with RB1B. CIAV coinfection increased ICP4 expression in the spleens of chickens infected with RB1B. These results indicated that inoculation of chickens with the 584A isolate caused a more robust early cytolytic infection compared with inoculation with RB1B alone and support the classification of 584A as a vv+MDV strain. Coinfection with CIAV exacerbated vvMDV strain RB1B infection. The extent of this exacerbation was less evident when birds were coinfected with 584A and CIAV.     

Protective efficacy of Marek's disease virus (MDV) CVI-988 CEF65 clone C against challenge infection with three very virulent MDV strains

Comparative 50% protective dose (PD50) assays were performed using a plaque-purified preparation of Marek's disease virus (MDV) strain CVI-988 at the 65th chicken embryo fibroblast (CEF) passage level (MDV CVI-988 CEF65 clone C) and three commercial MD vaccines: herpesvirus of turkeys (HVT) FC126, MDV CVI-988 CEF35, and a bivalent vaccine composed of HVT FC126 and MDV SB-1. In addition, comparative PD50 assays were performed in groups of chickens with maternal antibody to each of the three vaccines. Three representatives of the newly emerged biovariant very virulent (vv) MDV strains-RB/1B, Tun, and Md5-were employed as challenge virus. The experiments made feasible the differentiation between virulent MDV and vvMDV strains, within serotype 1. Vaccination with CVI-988 clone C vaccine resulted in PD50 estimates of about 5 plaque-forming units (PFUs) against challenge infection with each of the three vvMDV strains. The PD50 estimate of CVI-988 clone C vaccine was 12-fold below the PD50 of HVT FC126. The protective synergism of bivalent vaccine, composed of HVT and SB-1, was confirmed by groups given the lowest vaccine doses. The bivalent vaccine, however, resulted in incomplete protection in groups given the highest vaccine doses. Homologous maternal antibodies to serotype 1 caused a fivefold increase in the PD50 estimate of CVI-988 clone C. Heterologous maternal antibodies against HVT did not interfere with efficacy of CVI-988 clone C vaccination. However, the combination of maternal antibodies against both HVT and SB-1 (serotypes 2 and 3) showed a strong adverse effect on CVI-988 clone C vaccine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of diluting Marek's disease vaccines on the outcomes of Marek's disease virus infection when challenged with highly virulent Marek's disease viruses

Dilution of Marek's disease (MD) vaccines is a common practice in the field to reduce the cost associated with vaccination. In this study we have evaluated the effect of diluting MD vaccines on the protection against MD, vaccine and challenge MD virus (MDV) kinetics, and body weight when challenged with strains Md5 (very virulent MDV) and 648A (very virulent plus MDV) by contact at day of age. The following four vaccination protocols were evaluated in meat-type chickens: turkey herpesvirus (HVT) at manufacturer-recommended full dose; HVT diluted 1:10; HVT + SB-1 at the manufacturer-recommended full dose; and HVT + SB-1 diluted 1:10 for HVT and 1:5 for SB-1. Vaccine was administered at hatch subcutaneously. One-day-old chickens were placed in floor pens and housed together with ten 15-day-old chickens that had been previously inoculated with 500 PFU of either Md5 or 648A MDV strains. Chickens were individually identified with wing bands, and for each chicken samples of feather pulp and blood were collected at 1, 3, and 8 wk posthatch. Body weights were recorded at 8 wk for every chicken. Viral DNA load of wild-type MDV, SB-1, and HVT were evaluated by real time-PCR. Our results showed that dilution of MD vaccines can lead to reduced MD protection, reduced relative body weights, reduced vaccine DNA during the first 3 wk, and increased MDV DNA load. The detrimental effect of vaccine dilution was more evident in females than in males and was more evident when the challenge virus was 648A. However, lower relative body weights and higher MDV DNA load could be detected in chickens challenged with strain Md5, even in the absence of obvious differences in protection.     

鸡马立克氏病活疫苗免疫效力比较试验

用ＨＶＴ冻干苗、ＨＶＴ细胞结合苗、ＣＶＩ９８８细胞结合苗、ＳＢ１＋ＦＣ１２６双价活疫苗、３０１Ｂ／１＋ＦＣ１２６双价活疫苗和Ｚ４＋ＦＣ１２６双价活疫苗等６种鸡马立克氏病（ＭＤ）疫苗免疫ＳＰＦ白来航鸡或普通伊莎鸡，用鸡马立克氏病病毒（ＭＤＶ）强毒ＧＡ株、京－１血毒以及鸡马立克氏病超强毒ｖｖＭＤＶ－Ｍｄ５毒株分别攻击进行免疫效力比较试验。试验表明，ＭＤ单价苗的免疫效力强弱顺序依次是ＣＶＩ９８８、ＨＶＴ细胞结合苗和ＨＶＴ冻干苗，这３种ＭＤ单价苗均能给免疫鸡群提供有效的免疫保护力。ＳＢ１＋ＦＣ１２６、Ｚ４＋ＦＣ１２６和３０１Ｂ／１＋ＦＣ１２６等３种ＭＤ双价苗免疫效力显著高于ＭＤ单价苗，均能给免疫鸡群提供较强的免疫保护力，并能有效地抵抗ｖｖＭＤＶ－Ｍｄ５毒株的致瘤作用。Ｚ４＋ＦＣ１２６和３０１Ｂ／１＋ＦＣ１２６ＭＤ双价苗免疫效力无显著差异     

Identification of the cross-reactive antigens between Marek's disease virus and pseudorabies virus by monoclonal antibodies

Among the 33 monoclonal antibodies (MAbs) against pseudorabies virus (PRV) examined, three MAbs (24-17, 74-26, and 8) were found to react with cells infected with Marek's disease virus (MDV)-related viruses by immunofluorescence test. Two of the MAbs (24-17 and 74-26) reacted with the nuclei of cells infected with MDV serotype 1 (MDV1), MDV serotype 2 (MDV2), and herpesvirus of turkeys (HVT), whereas MAb 8 reacted with the cytoplasm of MDV2- and HVT-infected cells. However, none of the MAbs against MDV1, MDV2, and HVT that were examined reacted with PRV-infected cells. None of these three MAbs against PRV reactive with MDV-related viruses cross-reacted with the cells infected with other herpesviruses, such as herpes simplex virus type 1, herpes simplex virus type 2, varicella zoster virus, Epstein-Barr virus, or human herpesvirus 6. Southern-blot hybridization under stringent or less-stringent conditions showed that no significant DNA homology was detected between PRV DNA and MDV DNA.     

表达马立克氏病病毒CVI988／Rispens株糖蛋白B基因的重组鸡痘病毒疫苗的免疫保护研究

用鸡痘病毒载体表达马立克氏病病毒（MDV）糖蛋白B（gB）基因构建成重组鸡痘病毒（rFPV）。以MDV　GA或Md5和RBIB混合攻毒，在不同品种的鸡中评价rFPV－gB／R单价苗和与火鸡疱疹病毒（HVT）组成的二价苗的免疫保护效力。试验结果表明，MDV疫苗和FPV母源抗体阴性的SPF鸡和母源抗体阳性的商品鸡中，rFDV－gB／R均能提供免疫保护作用，且其中一种商品蛋鸡中rFPV－gB／R与HVT液氮苗或HVT冻干苗结合使用，均有显著的免疫协同保护作用。免疫学研究指出，rFPV－gB／R与常规疫苗一样，可显著地降低疫苗免疫攻毒鸡的病毒血症和羽囊排毒。     

Pathogenicity studies with plaque-purified preparations of Marek's disease virus strain CVI-988

The pathogenicity of Marek's disease (MD) strain CVI-988 vaccine, eight plaque-purified preparations originating from this strain, and the vaccine HVT FC126 (based on herpesvirus of turkeys) was determined by intramuscular administration of high virus doses to day-old specific-pathogen-free Rhode Island Red (RIR) chickens, which are extremely MD-susceptible. Paralysis and neuritis were observed in 88% of RIR chickens inoculated with MDV CVI-988 at the cell-passage level of the commercial vaccine. HVT FC126 caused paralysis in two of 39 RIR chickens tested, of which one had an endoneural lymphoma, and another three had endoneural inflammation. Five plaque-purified MDV CVI-988 virus preparations at various cell-culture-passage levels caused no lesions. Of another three clones, two caused inflammatory B-type lesions in the nerves of 1/10 chickens, and the third clone caused inflammatory nonneoplastic MD lesions in the liver of 1/11 chickens.     

Attenuated revertant serotype 1 Marek's disease viruses: safety and protective efficacy.

In earlier studies, a revertant serotype 1 Marek's disease virus (MDV), clone Md11/75C/R2, was found to be a highly protective vaccine virus but was mildly pathogenic for susceptible chickens. The term "revertant" indicates that the virus, after attenuation, gained virulence following backpassage in chickens. The present study is an attempt to develop a more attenuated but still protective vaccine virus from Md11/75C/R2. Forty-two derivative viruses or clones from Md11/75C/R2 were evaluated. Two of these, designated clones R2/23 and R2/29, induced viremia but little or no pathology in preliminary trials and were selected for further study. In a series of nine trials, both clones provided protection against challenge with very virulent MDV strains that was superior to that induced by turkey herpesvirus (HVT) and was not significantly different (P greater than 0.05) from that induced by a bivalent (HVT + SB-1) vaccine. Both clones appeared fully attenuated based on pathogenicity tests in susceptible antibody-negative chickens. Both clones gained virulence on backpassage in chickens, but this seemed of little concern because neither virus spread by contact to other chickens. Although the two clones were very similar, clone R2/23 appeared to have a slightly lower pathogenic potential following backpassage and thus best meets the combined criteria of safety and efficacy.     

In situ production of interferon in tissues of chickens exposed as embryos to turkey herpesvirus and Marek's disease virus

Chicken eggs at embryonation day (ED) 18 or newly hatched chicks were inoculated with turkey herpesvirus (HVT), Marek's disease virus (MDV), or virus-free diluent and, at intervals after inoculation, tissue homogenates of virus-exposed and virus-free chickens or chicken embryos were examined for interferon (IFN) activity. Homogenates of lung, thymus and spleen specimens from chickens given HVT at ED 18 had IFN activity. Activity of IFN in the lungs was studied further. Homogenates of lung specimens from chickens exposed to HVT at hatching also had IFN activity, although the concentration of IFN was lower than that in chickens given HVT at ED 18. The pathogenic isolates of MDV (JM-MDV), but not the attenuated (Md11/75C-MDV) or nonpathogenic (SB1-MDV) isolates, inoculated at ED 18 also induced high lung IFN activity. Exposure to a combination of HVT and SB1-MDV induced IFN activity comparable with that in chickens given HVT alone. The IFN activity in homogenates of lung specimens from virus-exposed chickens was species specific and heat and pH stable, but was destroyed by trypsin treatment. Occasionally, low IFN activity also was detected in homogenates of tissue specimens from virus-free chickens or chicken embryos. This IFN activity could have been produced constitutively or may have been induced by substances (inducers) in the environment.     

马立克氏病毒单克隆抗体杂交瘤细胞株的建立与鉴定

以马立克氏病毒814株(MDV-814)作为免疫用抗原,建立了2个MDV特异性单克隆抗体(McAb)杂交瘤细胞株—4A6、3D7。鉴定结果表明;两株细胞所分泌的McAb均为IgM类免疫球蛋白,具有MDVⅠ型病毒特异性,无中和反应特性和沉淀反应特性。利用两种McAb对国内标准强毒MDV-京1株进行鉴定,肯定其为MDV-Ⅰ型毒株。     

Effects of vaccine dose, virus challenge dose and interval from vaccination to challenge on protection of broiler chickens against Marek's disease virus challenge

OBJECTIVES: To examine the effects of varying the doses of turkey herpesvirus (HVT) vaccine and Marek's disease virus (MDV) challenge at two intervals after vaccination on the protection of chickens against challenge with MDV. DESIGN AND PROCEDURE: Experiment 1, a dose response study, consisted of 11 doses of HVT vaccine administered at hatch followed by challenge with 100 plaque forming units (pfu) of MDV 5 days post vaccination. Experiment 2, a 2 x 6 x 2 factorial design, included two HVT vaccine types, six different doses of HVT vaccine and 50 pfu and 200 pfu of MDV challenge 2 days post vaccination. All chickens were reared up to day 56 post challenge when all survivors were killed humanely. Dead and killed chickens were examined for gross MD tumours. RESULTS: Experiment 1 showed a significant positive linear relationship between dose of HVT vaccine and protective index in chickens challenged 5 days post vaccination. However the range of protective index observed was limited. In Experiment 2 neither HVT vaccine provided significant protection at any dose. There was no significant effect of vaccine type or MDV challenge dose on overall protection against challenge. Chickens challenged with 200 pfu of MDV had significantly higher mortality and MD incidence than those with 50 pfu. CONCLUSIONS: HVT vaccine dose had a significant impact on protective index, but vaccination to challenge interval appeared to have greater impact on the protective efficacy of vaccination. A fourfold increase in challenge dose increased mortality rate and incidence of MD.     

Marek's disease virus infection in the brain: virus replication, cellular infiltration, and major histocompatibility complex antigen expression

Marek's disease virus (MDV) infection in the brain was studied chronologically after inoculating 3-week-old chickens of two genetic lines with two strains of serotype I MDV representing two pathotypes (v and vv+). Viral replication in the brain was strongly associated with the development of lesions. Three viral antigens (pp38, gB, and meq) were detected in the brain of infected chickens. Marked differences between v and vv+ pathotypes of MDV were identified for level of virus replication, time course of brain lesions, and expression of major histocompatibility complex (MHC) antigens. Two pathologic phenomena (inflammatory and proliferative) were detected in the brain of chickens inoculated with vv+MDV, but only inflammatory lesions were observed in those inoculated with vMDV. Inflammatory lesions, mainly composed of macrophages, CD4+ T cells, and CD8+ T cells, started at 6-10 days postinoculation (dpi) and were transient. Proliferative lesions, characterized by severe infiltrates of CD4+CD8- T cells (blasts), started at 19-26 dpi and persisted. Expression of MHC antigens in endothelial cells and infiltrating cells within the brain was influenced by MDV infection. Upregulation of MHC class II antigen occurred in all treatment groups, although it was more severe in those inoculated with vv+MDV. MHC class I antigen was downregulated only in those groups inoculated with vv+MDV. These results enhance our understanding of the nature and pattern of MDV infection in the brain and help to explain the neurovirulence associated with highly virulent MDV.     

Serotype 1 viruses modified by backpassage or insertional mutagenesis: approaching the threshold of vaccine efficacy in Marek's disease

Improved vaccines to control Marek's disease (MD) in chickens are desired by the poultry industry but have been difficult to develop. Studies were conducted to evaluate strategies for deriving MD vaccines of high protective efficacy, irrespective of virulence. Candidate viruses from parent strains representing v and vv+ pathotypes were modified by cell culture passage, backpassage in chickens, or insertional mutagenesis following cocultivation with retroviruses. Ten strains considered most likely to exhibit high protective efficacy were selected for further study. The ability of these modified viruses to protect commercial or maternal antibody-positive (ab+) chickens against virulent MD virus (MDV) challenge was compared with that of strain CVI988, the standard commercial MD vaccine. Modified strains were also evaluated for the ability to induce lymphomas or other pathologic changes in ab+ and antibody-negative (ab-) chickens. Two of the 10 modified viruses, strains RM1 and CVI988/BP5, provided high levels of protection against highly virulent MDV challenge. The magnitude of protection was greater than that of one laboratory and two commercial preparations of CV1988, but was approximately equal to that of two other commercial preparations of CVI988 in laboratory and field tests. Three of the strains, including RMI and CVI988/BP5, induced lymphoid organ atrophy in ab-chicks but not in ab+ commercial chicks, a property designated here as L phenotype. Seven strains, including two L+ strains, were mildly oncogenic for ab- chicks, a property designated here as O phenotype. Five of these strains caused no tumors in ab+ chickens. The two fully attenuated strains induced neither lymphomas nor lymphoid organ atrophy. The L and O phenotypes appeared not to be linked, and both (especially the L phenotype) appeared associated with high levels of protection. These studies also illustrated differences in the protective efficacy of different preparations of CVI988 vaccine, indicating the need to choose carefully the most protective strains as controls for efficacy studies. A new vv+ strain, designated as 686, is described and appears useful as a challenge virus; it is the most virulent of the 48 field isolates of MDV thus far pathotyped in this laboratory. These findings support the conclusion that new virus strains with high levels of protective immunity comparable to that of CVI988 can be developed. However, the question of whether strains can be developed that exceed the efficacy of current CVI988-based vaccines remains unanswered. After more than 30 years of unsuccessful endeavor by many laboratories toward this goal, it now may be useful to consider whether the efficacy of MD vaccines is limited by some type of biologic threshold.     

鸡传染性喉气管炎病毒WG株ICP4基因的鉴定及其在潜伏位点的表达

根据传染性喉气管炎病毒(Infectious laryngotracheitis virus,ILTV)SA-2株ICP4基因序列设计并合成3对引物,以ILTV中国王岗株(WG)DNA为模板扩增ICP4基因,并对其进行了序列测定。将ILTV WG株的ICP4基因及其推导的氨基酸序列,分别与ILTV SA-2株、BHV-1、EHV-1、EHV-4、MDV-1、MDV-2、HVT、PRV、VZV、HSV-1和HSV-2的ICP4基因及其推导的氨基酸序列比较后发现,ILTV毒株之间ICP4基因相对保守,核苷酸和氨基酸水平的同源性分别为99.7%和99.1%,但与其它α-疱疹病毒的ICP4基因的同源性则较低,低于3.0%。对潜伏感染鸡三叉神经节中病毒基因的检测显示,在人工感染ILTV WG株后第10 d～60 d内均能检测到ICP4特异RNA,而gB、gC、TK则未能检出。鉴于目前国内外对α-疱疹病毒潜伏感染相关基因以及ICP4基因序列和结构功能的研究,ILTV WG株ICP4基因的克隆和序列测定,以及病毒基因在潜伏感染鸡三叉神经节中的差异表达,为进一步研究ICP4基因的功能及确定潜伏感染相关基因奠定了基础。