Compatibility of turkey herpesvirus-infectious bursal disease vector vaccine with Marek's disease rispens vaccine injected into day-old pullets

Mixtures of turkey herpesvirus (HVT) and Rispens poultry vaccines have been used worldwide for over 20 yr, mainly for vaccination of future layers and breeders. With increasing virulence of Marek's disease (MD) virus strains, vaccination strategies are evolving toward the use of vaccines combining HVT and Rispens. A single vaccination either in ovo or at 1 day of age with the HVT + infectious bursal disease (IBD) vector vaccine is efficient against IBD. However, with vaccination programs that include a hatchery administration of the HVT + IBD vaccine, additional protection against very virulent and very virulent-plus MD viruses is needed, especially for layers and breeders. This study looked at the combination of four commercially available Rispens vaccines with the HVT + IBD vector vaccine injected at 1 day of age. MD challenge tests that were superior to 90% in relative score in all the groups vaccinated with both vaccines showed that the mixture of HVT + IBD and Rispens vaccines had no effect on clinical protection against MD, and IBD challenge tests showed that the mixture of HVT + IBD and Rispens vaccines had no effect on clinical protection against IBD, which was equal to 100% protection in all the groups vaccinated with both vaccines.     

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.     

Field evaluations of safety and efficacy of an Australian Marek's disease vaccine

OBJECTIVE: To demonstrate the safety and efficacy of the Marek's Disease Virus-1 vaccine (strain BH 16) from field studies in comparison with the CVI 988 Rispens vaccine currently available in Australia. STUDY DESIGN: A small field trial was carried out on nine breeder flocks and a larger trial on 21 breeder flocks. All chickens were obtained from a commercial hatchery and each was vaccinated at hatch with cell-associated Herpes Virus of Turkeys vaccine. A group of chickens vaccinated with BH 16 vaccine was placed in one shed per property and the remainder were vaccinated with the Rispens vaccine and placed in the remaining sheds. At 25, 30, 35, and 40 weeks after hatch, the field veterinarian or farm manager examined all birds dying on two consecutive days in the designated placement sheds. RESULTS: In the small trial there was a significantly lower incidence of MD in birds vaccinated with the MDV-1 vaccine compared with the Rispens vaccine (P < 0.001). In a larger trial there was no difference in the incidence of MD between the treatment groups, due possibly to a lower rate of natural challenge. Egg production results and average weekly mortality results for both groups were similar. CONCLUSION: The present study describes an attenuated type 1 MD vaccine which is at least equivalent to a vaccine derived from the CVI 988 Rispens strain in terms of safety and efficacy when used in combination with HVT vaccine.     

Field trials to test the efficacy of polyvalent Marek's disease vaccines in broilers

The efficacies of trivalent (Md11/75C + SB-1 + HVT), bivalent (SB-1 + HVT), and turkey herpesvirus (HVT) vaccines against Marek's disease (MD) were compared in commercial broiler flocks in four trials involving 11 farm locations and 486,300 chickens. In all four trials, chickens receiving polyvalent vaccines had lower leukosis (MD) condemnation rates than chickens vaccinated with HVT alone; when data were summarized for each vaccine type in each trial, condemnation rates for the bivalent- or trivalent-vaccinated groups were 56-96% (mean 78%) lower than those for HVT-vaccinated chickens. Polyvalent vaccination was clearly mor efficacious than HVT in 8 of 11 individual farms, although it did not always reduce leukosis condemnations to acceptable levels. Body weights of chickens vaccinated with polyvalent vaccines did not differ consistently from those vaccinated with HVT. Chickens inoculated with the trivalent vaccine had slightly lower overall leukosis condemnation rates (0.24%) than those inoculated with the bivalent vaccine (0.45%) in trials 1-3, where direct comparisons were made. Bivalent vaccines containing either 1,500 or 200 plaque-forming units of SB-1 virus were equally effective; thus, HVT may need to be supplemented with only small amounts of SB-1 to obtain the benefits of protective synergism. SB-1 virus did not appear to carry over from polyvalent-vaccinated flocks to subsequent HVT-vaccinated flocks in the same houses, even when old litter was used.     

Antibody response of chickens to serotype 1, 2, or 3 Marek's disease vaccines based on ELISA with infected cells as antigen

An enzyme-linked immunosorbent assay (ELISA) was applied to evaluate the antibody response of commercial White Leghorn chickens to vaccination against Marek's disease (MD) at hatch (day 0) with serotype-1 (Rispens), -2 (SB-1), or -3 (turkey herpesvirus, HVT) vaccine virus and to challenge on day 21 with MD virus. Antigens for the test were whole chicken embryo fibroblast cells infected with Rispens, SB-1, or HVT. The chickens were progeny of stock that had been vaccinated with HVT, and on day 21 the nonvaccinated group had higher levels of maternal antibodies to HVT than to other antigens (P < 0.05). Only SB-1 vaccine had induced antibodies by day 21, and this was detected only against homologous antigens. On day 49, all three vaccines had induced higher levels of antibodies to homologous than to heterologous antigens. Marek's Disease virus (MDV) induced antibodies to all three antigens, but challenging vaccinated chicks did not significantly increase levels of antibodies on day 81 to any of the three antigens. It was concluded that an ELISA using whole cells as antigens would have potential value for monitoring the antibody response induced by MD vaccines and virulent MDV.     

Isolation of very virulent Marek''s disease viruses from vaccinated chickens in Australia

Very virulent Marek's disease viruses (vvMDV), defined as isolates against which the herpesvirus of turkey (HVT) vaccine provide poor protection, have been isolated from poultry flocks in both the United States and Europe. Twenty-one samples from vaccinated Australian flocks, experiencing problems with excessive Marek's disease (MD), were tested for the presence of transmissible MD viruses (MDV). Of the 16 samples which contained a transmissible agent, 14 were pathogenic in chickens, based on the development of MD lesions or depression of the bursa/body weight ratio. Of the pathogenic isolates which have been successfully typed 10 were serotype 1, and one was serotype 2 MDV. Pathogenicity of isolates varied. Several isolates caused tumours in 20-30% of both vaccinated and unvaccinated chickens. Two isolates, MPF6 and MPF23, caused tumours in more than 50% of chickens. When MPF6 and MPF23 were tested in vaccine trials bivalent vaccine gave no better protection against development of MD lesions than a monovalent vaccine. Isolate MPF23 was so pathogenic that lesions were produced in all chickens, regardless of the vaccine protocol used. Therefore vvMDV have been isolated in Australia, and unlike the vaccines tested overseas, bivalent Australian vaccines do not appear to provide greater protection against these vvMDV.     

Derivation, safety and efficacy of a Marek's disease vaccine developed from an Australian isolate of very virulent Marek's disease virus

Objective To develop a serotype 1 Marek's disease (MD) vaccine from a very virulent MDV (vvMDV) pathotype and demonstrate safety and efficacy against early challenge with very virulent field strains in the presence of maternal antibody.
Study design Strain BH 16 was isolated and attenuated by serial cell culture passage. One of two cloned passages was selected for vaccine development following early laboratory-scale protection trials in commercial birds. Comparative protection trials were carried out on the BH 16 vaccine and on a CVI 988 Rispens vaccine using commercial and SPF chickens. Challenge viruses used were either a low passage strain BH 16 virus, the Woodlands No. 1 strain or MPF 57 strain of MDV. The BH 16 vaccine was back-passaged in SPF chickens six times and virus recovered from the final passage and the original vaccine virus were tested for safety. The immunosuppressive potential of the BH 16 and Rispens vaccines was also assessed in parallel.
Results The BH 16 and Rispens vaccines induced comparable levels of protection when used as monovalent or multi-valent vaccines, although protection achieved with the mono-valent vaccines was lower. No gross tumour formation was evident in any birds receiving the BH 16 vaccine or bird-passaged virus, although microscopic lesions were present in 2/12 birds that received the bird-passaged virus. In tests for immunosuppression, there was no histological evidence of damage to either the bursa of Fabricius or the thymus.
Conclusion The BH 16 vaccine was shown to be safe and at least as protective as the Rispens vaccine against three highly virulent MD challenge viruses.     

Derivation,safety and efficacy of a Marek's disease vaccine developed from an Australian isolate of very virulent Marek's disease virus

OBJECTIVE: To develop a serotype 1 Marek's disease (MD) vaccine from a very virulent MDV (vvMDV) pathotype and demonstrate safety and efficacy against early challenge with very virulent field strains in the presence of maternal antibody. STUDY DESIGN: Strain BH 16 was isolated and attenuated by serial cell culture passage. One of two cloned passages was selected for vaccine development following early laboratory-scale protection trials in commercial birds. Comparative protection trials were carded out on the BH 16 vaccine and on a CVI 988 Rispens vaccine using commercial and SPF chickens. Challenge viruses used were either a low passage strain BH 16 virus, the Woodlands No. 1 strain or MPF 57 strain of MDV. The BH 16 vaccine was back-passaged in SPF chickens six times and virus recovered from the final passage and the original vaccine virus were tested for safety. The immunosuppressive potential of the BH 16 and Rispens vaccines was also assessed in parallel. RESULTS: The BH 16 and Rispens vaccines induced comparable levels of protection when used as monovalent or multivalent vaccines, although protection achieved with the monovalent vaccines was lower. No gross tumour formation was evident in any birds receiving the BH 16 vaccine or bird-passaged virus, although microscopic lesions were present in 2/12 birds that received the bird-passaged virus. In tests for immunosuppression, there was no histological evidence of damage to either the bursa of Fabricius or the thymus. CONCLUSION: The BH 16 vaccine was shown to be safe and at least as protective as the Rispens vaccine against three highly virulent MD challenge viruses.     

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.     

Embryo vaccination against Marek's disease with serotypes 1, 2 and 3 vaccines administered singly or in combination

Marek's disease virus (MDV) vaccines of serotypes 1 and 2 administered in 18-day-old embryonated eggs induced better protection against post-hatch challenge at 3 days with virulent MDV than vaccines given at hatch. Embryonal vaccination with a polyvalent vaccine containing equal quantities of serotypes 1 and 2 of MDV and serotype 3 virus (turkey herpesvirus, HVT) was also significantly more effective than post-hatch vaccination. These and earlier results indicate that protective efficacy of single or combined Marek's disease vaccine serotypes against post-hatch challenge at 3 days can be substantially improved if the vaccines are injected into 18-day embryos rather than at hatch. Injection of vaccines of serotypes 1 or 2 into embryonated eggs or hatched chicks did not cause detectable gross or microscopic lesions in chickens. Vaccine viruses of serotypes 1 and 2 could be isolated from spleen cells of chickens 1 week post-vaccination, and the titer of recoverable viruses was higher in chickens that received the vaccines at the 18th day of embryonation than in chickens vaccinated at hatch. Although embryo vaccination with HVT usually provided better protection than post-hatch vaccination against early post-hatch challenge with variant pathotypes of MDV, the protection was poor regardless of vaccination protocol. If challenge with variant pathotypes of MDV was delayed until embryonally or post-hatch HVT-vaccinated chickens were 21 days of age, protection of chickens by HVT was not enhanced. Thus, resistance induced by embryonal vaccination with HVT was qualitatively similar to that induced by post-hatch vaccination with this virus.     

鸡马立克氏病CVI988／Rispens疫苗免疫效力试验

应用荷兰农业部提供的鸡马立克氏病（ＭＤ）ＣＶＩ９８８／Ｒｉｓｐｅｎｓ Ⅰ型致弱种毒, 在农业部批准的符合ＧＭＰ 要求的生产车间研制出鸡马立克氏病ＣＶＩ９８８／Ｒｉｓｐｅｎｓ 疫苗。将按国际标准检验合格的三批疫苗及进口商品ＣＶＩ９８８／Ｒｉｓｐｅｎｓ 疫苗接种１ 日龄ＳＰＦ 雏鸡, 于７ 日龄经腹腔攻击鸡马立克氏病强毒（北京－ １ 株） 血毒, 全部鸡只隔离饲养观察至６０ 日龄并作全群剖检。经测定： 非免疫攻毒组１００％ 发病,健康对照组全部阴性, 三批国产ＣＶＩ９８８／Ｒｉｓｐｅｎｓ 疫苗保护指数分别为９０－０, ９０－０, ９３－３ , 进口商品苗保护率为９３－３ 。结果表明国产和进口ＣＶＩ９８８／Ｒｉｓｐｅｎｓ疫苗均能提供对ＭＤ 较高的免疫保护力, 国产疫苗的保护效果达到了国际同类产品的先进水平。     

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。     

Induction of strong protection by vaccination with partially attenuated serotype 1 Marek's disease viruses

Studies were conducted to better understand the relationship among Marcek's disease (MD) vaccine strains between induction of protective immunity and the degree of attenuation (or virulence). To obtain viruses at different stages of attenuation, very virulent plus MD strains 584A and 648A and selected clones of these strains were serially passaged in chicken and duck cells. These viruses were considered fully attenuated after passage for 70-100 times in chicken embryo cell cultures until they no longer induced gross lesions in susceptible, maternal antibody-negative (ab-) chickens. Lower passages of the same strains were considered partially attenuated, provided their virulence was less than that of the parent strain. Four of five partially attenuated preparations derived from MD virus strains 584A and 648A or the previously attenuated Md11 strain induced 28%-62% higher levels of protection in maternal antibody-positive (ab+) chickens against virulent MD challenge than the fully attenuated counterpart viruses. The partially attenuated 584A/d2/3 strain replicated in chickens but was totally nonprotective. Data from two subsequent trials in ab+ chickens confirmed that protection induced by the partly attenuated (passage 80) preparations was 79% and 118% higher, respectively, than that induced by the fully attenuated (passage 100) preparations of strain 648A. However, in one trial with ab- chickens, no difference in protection between partially and fully attenuated virus was observed. Strong protection (up to 85%) against highly virulent challenge also was provided by preparations of 648A at passages 40-60, which were moderately oncogenic when used alone. Partially attenuated strains tended to replicate to higher titers in both ab+ and ab- chickens compared with fully attenuated vaccines. Also, ab+ and ab- chickens vaccinated with partially attenuated strains developed three- to nine fold more extensive microscopic lesions in peripheral nerves at 14 and 22 days after virulent challenge than chickens vaccinated with fully attenuated strains. When measured in ab+ chickens, loss of lesion induction by 648A was achieved 30 passages earlier (at passage 70) than when measured in ab- chickens. Thus, maternal antibodies appeared to abrogate the pathogenicity of some partially attenuated strains. These studies establish for MD the principle that at least some partially attenuated MD viruses may replicate better and induce stronger immunity against virulent challenge than fully attenuated preparations of the same strain, at least when tested in ab+ chickens. Moreover, depending on passage level, partially attenuated vaccine strains may be relatively innocuous for ab+ chickens, causing few or no lesions.     

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.     

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     

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CVI988/Rispens疫苗免疫效力试验

本试验用北京市农林科学院畜牧兽医研究所制备的ＣＶＩ９８８／Ｒｉｓｐｅｎｓ疫苗和进口的ＣＶＩ９８８／Ｒｉｓｐｅｎｓ疫苗免疫１日龄来航鸡,７日龄以ＭＤＶ北京－１株血毒进行攻击,６０日龄全群剖检。经免疫效力试验两次测定,３批北京所制备的ＣＶＩ９８Ｒｉｓｐｅｎｓ疫苗产品保护指数分别为试验的９０．０、９０．０、９３．３和试验（２）的１００．０、１００．０、９４．５与进口商品ＣＶＩ／９８８Ｒｉｓｐｅｎｓ苗的保     

Protection of chickens from Newcastle and Marek's diseases with a recombinant herpesvirus of turkeys vaccine expressing the Newcastle disease virus fusion protein.

Recombinant strains of herpesvirus of turkeys (HVT) were constructed that contain either the fusion protein gene or the hemagglutinin-neuraminidase gene of Newcastle disease virus (NDV) inserted into a nonessential gene of HVT. Expression of the NDV antigens was regulated from a strong promoter element derived from the Rous sarcoma virus long terminal repeat. Recombinant HVT strains were stable and fully infectious in cell culture and in chickens. Chickens receiving a single intra-abdominal inoculation at 1 day of age with recombinant HVT expressing the NDV fusion protein had an immunological response and were protected (> 90%) against lethal intramuscular challenge at 28 days of age with the neurotropic velogenic NDV strain Texas GB. Recombinant HVT expressing the NDV hemagglutinin-neuraminidase provided partial protection (47%) against the same challenge. Chickens vaccinated with recombinant HVT vaccines had low levels of protection against NDV replication in the trachea when challenged ocularly. Recombinant HVT vaccines and the parent HVT strain provided similar levels of protection to chickens challenged with the very virulent RB1B strain of Marek's disease virus, indicating that insertion of foreign sequences into the HVT genome did not compromise the ability of HVT to protect against Marek's disease.     

Load of challenge Marek's disease virus DNA in blood as a criterion for early diagnosis of Marek's disease tumors

Outbreaks of Marek's disease (MD) in vaccinated flocks still occur sporadically and lead to economic losses. Unfortunately, adequate methods to predict MD outbreaks are lacking. In the present study, we have evaluated whether high load of challenge MD virus (MDV) DNA in peripheral blood could aid in the early diagnosis of MD and in monitoring efficacy of vaccines against MD. One experiment was conducted to simulate field conditions by combining various vaccines (turkey herpesvirus [HVT] and HVT + MDV serotype 2 [SB1]) and challenge viruses (GA, Md5, and 648A). Vaccine efficacy among our experimental groups ranged from 13.3% to 94.2%. Each chicken was sampled three times during the length of the experiment (3, 5, and 15 wk postchallenge [wpc]), and gross lesions were evaluated in chickens that died and at termination of the experiment. DNA was extracted from whole blood and buffy coats from each sample, and the load of challenge MDV DNA and HVT DNA were quantified by real-time polymerase chain reaction. Chickens that developed MD by the end of the experiment had higher load of challenge MDV DNA (threshold cycle [Ct] glyceraldehyde-3-phosphate dehydrogenase [GAPDH]/Ct glycoprotein B [gB] ratios of 1.0, 1.04, and 1.05 at 3, 5, and 15 wpc, respectively) than those that did not develop MD (Ct GAPDH/Ct gB ratios of 0.7, 0.69, and 0.46 at 3, 5, and 15 wpc, respectively). However, load of HVT DNA in blood was not correlated with the development of tumors (Ct GAPDH/Ct HVT ratios from 0.04 to 0.10 in both groups). Vaccinated groups with >75% protection had statistically significant less challenge DNA virus (Ct GAPDH/Ct gB ratios of 0.76, 0.70, and 0.45 at 3, 5, and 15 wpc, respectively) than less protected groups (Ct GAPDH/Ct gB ratios of 0.92, 0.97, and 0.85 at 3, 5, and 15 wpc, respectively). No differences in the load of HVT DNA could be found between protected and nonprotected groups at any time point of the study (Ct GAPDH/Ct HVT from 0.05 to 0.09 in both groups). Our results showed that load of challenge MDV DNA but not load of HVT DNA in blood can be used as criterion for early diagnosis of MD.     

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)