Immunosuppression in specific-pathogen-free broilers administered infectious bursal disease virus vaccines by in ovo route

The effect of two infectious bursal disease virus (IBDV) vaccines (IBDV-immune complex [Icx] and IBDV-2512), administered in ovo, on the cell-mediated immunity of specific-pathogen-free (SPF) broilers was examined. A decrease (P < 0.05) in the T-cell mitogenic response occurred in birds vaccinated with both vaccines on days 9 and 21 post in ovo vaccination (PIOV), but an increase (P < 0.05) occurred on day 15 PIOV. The T cells from birds given the IBDV-2512 were less responsive. There were no significant differences in proportions of lymphocytes expressing CT4+CT8 and CT8+CT4- except on day 21 PIOV, where an increase (P < 0.05) in IBDV-2512-vaccinated birds and a decrease (P < 0.05) in percentage of CT4+CT8- in IBDV-Icx-vaccinated birds was observed. There was an increase (P < 0.05) in percentage of CT8+CT4- T cells on day 21 PIOV in both vaccinated groups. A decrease (P < 0.05) in B-cell percentage was observed on day 21 PIOV in birds given both vaccines. Results indicated that although humoral immunosuppression is associated with destruction of B cells (bursal atrophy), cell-mediated immunosuppression induced by these two IBDV vaccines in SPF birds was not associated with altered helper (CT4+CT8-) or cytotoxic (CT8+CT4-) subpopulations of T lymphocytes.     

Prevention of avian lymphoid leukosis by induction of bursal atrophy with infectious bursal disease viruses.

Five groups of genetically susceptible chickens were inoculated at hatching with lymphoid leukosis virus; four of these were given infectious bursal viruses of varying virulence at 14 days of age and one group was not inoculated (control). All chickens in the control group developed evidence of lymphoid leukosis by 180 days. Two groups given relatively virulent bursal disease viruses, which destroyed bursal lymphoid cells, did not develop lymphoid leukosis. Treatment with avirulent vaccines had no visible effect on bursal morphology and did not significantly alter the incidence of lymphoid leukosis in two other groups, although the time of development was delayed. Results of our study show that viral-induced destruction of the bursa of Fabricius eliminates the development of lymphoid leukosis but that infection without bursal destruction has little effect on lymphoid leukosis.     

Functional restoration of the bursa of Fabricius following in ovo infectious bursal disease vaccination

The primary role of the avian bursa of Fabricius is to provide an essential microenvironment for B-lymphocytes to diversify their immunoglobulin genes by gene hyperconversion. Infectious bursal disease (IBD) vaccination using intermediate plus vaccine strains can temporarily deplete the bursal follicles and interrupt the normal B-cell development, which is generally followed by B-cell repopulation and histological regeneration. To find evidence that functional restoration of the bursa of Fabricius occurs in addition to the histological regeneration, we have analysed the chB1 gene expression, which indicates active bursal B-lymphocytes, and also the surface expression of a carbohydrate structure Lewis(x), a marker which identifies those bursal B-lymphocytes that are undergoing gene hyperconversion. In ovo vaccination with an immune complex vaccine (IBDV-BDA) caused transient bursal destruction in both the SPF and the maternally protected broiler groups with differences evident in the starting time, the severity and the duration of the effect. After the depletion phase, signs of histological regeneration appeared together with chB1- and Lewis(x) expression indicating that B-lymphocytes were functionally active and the bursa of Fabricius was serving again as an efficient primary lymphoid organ providing an appropriate microenvironment for B-cell development.     

Pathological and immunological study of an in ovo complex vaccine against infectious bursal disease

The appearance of very virulent strains of infectious bursal disease (IBD) virus at the end of the 1980s made it necessary to develop more effective immunization procedures. To facilitate this, the immunogenicity and the immunosuppressive effect of a mild (G-87), an intermediate (LIBD) and an intermediate-plus (IBDV 2512) IBDV strain were tested after the in ovo inoculation of 18-day-old SPF and broiler chicken embryos. It was established that no noteworthy difference existed between the immunized and the control embryos in hatching rate and hatching weight. The higher the virulence of the vaccine virus strain, the more severe damage it caused to the lymphocytes of the bursa of Fabricius. In SPF chickens, the haemagglutination inhibition (HI) titres induced by a Newcastle disease (ND) vaccine administered at day old decreased in inverse ratio to the virulence of the IBD vaccine strain, while in broiler chickens this was not observed. Despite the decrease of the HI titre, the level of protection did not decline, or did so only after the use of the 'hot' strain. SPF chickens immunized in ovo with a complex vaccine prepared from strain IBDV 2512 and IBD antibody showed the same protection against Newcastle disease as the broilers. In broiler chicken embryos immunized in ovo, only strain IBDV 2512 induced antibody production, and such chickens were protected against IBD at 3 weeks of age. The complex vaccine administered in ovo has been used successfully at farm hatcheries as well.     

Comparison of in ovo and post-hatch vaccination with particular reference to infectious bursal disease. A review

In ovo vaccination is an alternative approach to post-hatch vaccination of chickens, particularly in broilers. Vaccination at embryonation day 18 helps to 'close the window' of susceptibility i.e. the time between vaccination and early exposure to infectious agents compared with post-hatch vaccination. Attempts on embryonal vaccination as a mode of vaccine delivery were approached from the observation that chickens already develop certain immunologic functions before hatching. The immune system in birds begins to develop early during embryogenesis and various immune reactions have been induced in the late stage chicken embryos. Compared with post-hatch vaccination, in ovo vaccination stimulates both the innate and adaptive immune responses with the advantage that because of the prenatal immunization, in ovo vaccinated chicks have developed an appreciable degree of protection by the time of hatch. Effects of maternal antibodies on vaccines to be used for in ovo vaccination can be prevented by developing vaccines that are insensitive to maternal antibodies. It has been described that vaccination of chicken embryos at embryonation day 18 did not significantly affect the immune competence of hatched chickens. The apparent absence of tolerance in chicks hatched from embryos exposed to an antigen at the late stage of embryonation implies the feasibility of in ovo vaccination. Investigations on in ovo vaccination to produce safe and efficient vaccines are still in progress. Currently a large number of vaccines are under investigation for viral, bacterial and protozoal diseases.     

Bilateral effects of vaccination against infectious bursal disease and Newcastle disease in specific-pathogen-free layers and commercial broiler chickens

Different infectious bursal disease virus (IBDV) live vaccines (intermediate, intermediate plus) were compared for their immunosuppressive abilities in specific-pathogen-free (SPF) layer-type chickens or commercial broilers. The Newcastle disease virus (NDV) vaccination model was applied to determine not only IBDV-induced immunosuppression but also bilateral effects between IBDV and NDV. None of the IBDV vaccines abrogated NDV vaccine-induced protection. All NDV-vaccinated SPF layers and broilers were protected against NDV challenge independent of circulating NDV antibody levels. Sustained suppression of NDV antibody development was observed in SPF layers, which had received the intermediate plus IBDV vaccine. We observed a temporary suppression of NDV antibody development in broilers vaccinated with one of the intermediate, as well as the intermediate plus, IBDV vaccines. Different genetic backgrounds, ages, and residual maternal antibodies might have influenced the pathogenesis of IBDV in the different types of chickens. Temporary suppression of NDV antibody response in broilers was only seen if the NDV vaccine was administered before and not, as it was speculated previously, at the time the peak of IBDV-induced bursa lesions was detected. For the first time, we have demonstrated that the NDV vaccine had an interfering effect with the pathogenesis of the intermediate as well as the intermediate plus IBDV vaccine. NDV vaccination enhanced the incidence of IBDV bursa lesions and IBDV antibody development. This observation indicates that this bilateral effect of an IBDV and NDV vaccination should be considered in the field and could have consequences for the performance of broiler flocks.     

Immunogenicity of infectious bursal disease viruses in chickens.

Cross-protective properties of infectious bursal disease viruses (IBDVs) were studied. Viruses represented different subtypes of serotype 1, including recently isolated viruses (variants), and a serotype 2 virus. Chickens were vaccinated at 3 weeks of age with inactivated vaccines containing 10(5), 10(6), 10(7), or 10(8) mean tissue-culture infectious dose of a given virus and challenged 2 weeks later using either 10(2) or 10(3.5) mean embryo infectious dose (EID50) of either a standard virus or a variant serotype 1 virus. Protection was evaluated at 5 and 10 days post-challenge, based on gross and microscopic lesions, body weight, and bursa/body-weight ratios. The serotype 2 virus did not confer protection on birds challenged with the serotype 1 viruses. Vaccines made of variant viruses at the low doses protected chickens challenged with the high or low doses of either the standard or the variant viruses. Vaccines made of the standard or variant strains at low doses protected against high or low challenge doses of the standard strain. Vaccines made of the high dose of any of the standard strains protected chickens against the variant virus when the low challenge dose (10(2) EID50) was used, but not when the high challenge dose (10(3.5) EID50) was used. The lowest dose of the standard viruses vaccines required to confer protection against the variant virus varied depending on the strain. Results indicated that protection depended on the strain and dose of both the vaccine and challenge viruses and that the variant strains and standard strains share a common protective antigen(s).     

Embryo vaccination of specific-pathogen-free chickens with infectious bursal disease virus: tissue distribution of the vaccine virus and protection of hatched chickens against disease

Vaccination of specific-pathogen-free chickens as 18-day embryos with the BVM isolate of infectious bursal disease virus (IBDV) resulted in extensive replication of the vaccine virus in the embryonic tissues. The virus was recovered from lung, thymus, proventriculus, liver, kidney, and spleen of embryos 1 day postvaccination, and recoverable virus persisted for at least 7 days. Replication and spread of the vaccine virus in chickens vaccinated as 18-day embryos was compared with that in chickens vaccinated at hatch. Distribution of the virus in tissues was more extensive, virus levels in tissues were generally higher, and detectable virus persisted longer in chickens vaccinated as 18-day embryos than in those vaccinated at hatch. Effective vaccine response could be initiated with 6.2 median embryo lethal doses, the lowest dose tested. Chickens immunized as embryos developed neutralizing antibody against IBDV and resisted challenge with pathogenic IBDV at 4, 6, 8, and 10 weeks of age.     

Detection of infectious bursal disease viruses using in situ hybridization and non-radioactive probes.

The in situ hybridization assay was developed for the detection of infectious bursal disease virus (IBDV) infections in chickens. Bursal tissue samples were harvested 4 days following infection with the ST-C, MD, E, IN, or SAL IBDV strain. The cDNA clones STC-243, located on genome segment A, and STC-119, located on genome segment B, were used to prepare non-radioactive probes. Probes were labeled with digoxigenin and detected the homologous ST-C virus and also heterologous viruses in bursal tissue sections. No positive cells were observed in tissue sections from uninfected control chickens.     

Experimental cryptosporidiosis and infectious bursal disease virus infection of specific-pathogen-free chickens

Specific-pathogen-free chickens orally inoculated at 4 days of age with a moderately pathogenic vaccine strain of infectious bursal disease virus (IBDV) and/or at 5 days of age with Cryptosporidium baileyi oocysts remained free of overt clinical signs throughout a 16-day period postinoculation (PI). The prepatency period for C. baileyi oocyst shedding was shorter in chickens receiving higher numbers of oocysts, but once shedding was detected, there were no obvious differences in shedding patterns among groups receiving 10(3) through 10(6) oocysts. On days 8 and 16 PI, cryptosporidia were located primarily in the bursae of Fabricius. IBDV exposure was associated with bursal follicle atrophy, whereas C. baileyi infection resulted in bursal epithelial hypertrophy and hyperplasia, mild follicle atrophy, and heterophil infiltration of the bursal mucosa. Examination of experimental groups of 30 birds each indicated that concurrent infection with both agents resulted in more severe bursal lesions, more infected birds, and greater numbers of cryptosporidia in infected tissues. At the termination of the trial, 16 days PI, Cryptosporidium infection was associated with a 6% decrease in mean body weight compared with controls.     

Antigenic diversity of infectious bursal disease viruses

Statistically significant antigenic differences were detected among serotype I infectious bursal disease viruses (IBDV) using the virus-neutralization test. Eight serotype I commercial vaccine strains, five serotype I field strains, and two serotype II field strains were tested. Hyperimmune guinea pig antisera against heterologous and homologous IBDV strains were used in cross-neutralization tests. Relatedness values were calculated from geometric mean antibody titers based on a minimum of three tests. Six subtypes were distinguished among the 13 serotype I strains tested.     

Embryo vaccination with infectious bursal disease virus alone or in combination with Marek's disease vaccine

Studies with specific-pathogen-free chickens revealed that chicks hatching from eggs inoculated at the 18th day of embryonation with infectious bursal disease (IBD) vaccine viruses of low virulence (isolates TC-IBDV and BVM-IBDV) developed antibody against IBD virus (IBDV) and resisted challenge with virulent IBDV at 3 weeks of age or older. Embryo vaccination did not adversely affect hatchability of chicks or survival of hatched chicks. Chicks embryonally vaccinated with TC-IBDV had transient histologic lesions in the bursa of Fabricius at hatch. Similar but milder lesions were also noted in chickens that received TC-IBDV at hatch. The level of protection following embryo vaccination with TC-IBDV and BVM-IBDV was similar to that following vaccination with the same vaccines at hatch. Vaccine viruses of moderate virulence (isolates BV-IBDV and 2512-IBDV) were not suitable as vaccines in embryos lacking maternal antibody to IBDV, because the vaccinated chicks developed acute IBD after hatch. Isolate 2512-IBDV was not pathogenic for embryos bearing maternal antibody to IBDV. Maternal antibody against IBDV interfered with efficacy of embryo vaccination with BVM-IBDV but not with 2512-IBDV. Embryo vaccination with a mixture of vaccines against IBD and Marek's disease resulted in protection of hatched chicks against challenge with virulent IBDV and Marek's disease virus.