Detection of infectious bursal disease vaccine viruses in lymphoid tissues after in ovo vaccination of specific-pathogen-free embryos.

Control of infectious bursal disease virus (IBDV) by vaccination is important for poultry production worldwide. Two vaccines, an IBDV immune complex (ICX) vaccine and an IBDV-2512 vaccine, were administered at 100 mean embryo infectious dose to specific-pathogen-free 18-day-old broiler embryos in ovo. At 3, 6, 9, 15, and 21 days post in ovo vaccination (PIOV), bursa, spleen, and thymus tissues were collected and analyzed for virus protein by antigen capture chemiluminescent enzyme-linked immunosorbent assay (ELISA). Chicks were bled and antibody titers were determined by the antibody ELISA. At 21 days PIOV, chickens were challenged with a 1:500 dilution of an antigenic standard IBDV strain. At 28 days PIOV, birds were euthanatized and bursa weight:body weight ratios were determined. Embryos vaccinated with either vaccine exhibited 92% hatchability; however, within 1 wk of hatch, birds vaccinated with IBDV-2512 showed 56% mortality, whereas those given IBDV-ICX had only 3.2% mortality. Both IBDV-ICX and IBDV-2512 vaccines were detected in bursa, spleen, and thymus at day 3 PIOV. A 5-day delay in virus replication was observed with IBDV-ICX vaccine. By day 15 PIOV, the IBDV-ICX was no longer detectable in the bursa and spleen but persisted in the thymus. The IBDV-2512 vaccine persisted in the spleen and thymus on day 15 PIOV. By day 21 PIOV, neither vaccine virus was detected in any lymphoid organ. This assay can be useful in the early detection of vaccine virus in the tissues of chickens vaccinated via the in ovo route. Both vaccines caused bursal atrophy at all times PIOV. The IBDV-2512 caused splenomegaly at day 6 PIOV, whereas splenomegaly was not seen in IBDV-ICX-vaccinated birds until day 9 PIOV. Thymus atrophy was observed in IBDV-2512-vaccinated chicks from day 3 PIOV, whereas this occurred on day 15 PIOV in IBDV-ICX-vaccinated birds. Bursa weight: body weight ratios in IBDV-ICX-vaccinated unchallenged and vaccinated challenged birds were not different (P < 0.05).     

Protective efficacy of intermediate and intermediate plus infectious bursal disease virus (IBDV) vaccines against very virulent IBDV in commercial broilers

The evolution of very virulent (vv) infectious bursal disease virus (IBDV) has led to significant economic losses in many poultry-producing areas. Despite vigorous vaccination strategies, IBDV has been difficult to control. The protective efficacy of IBDV vaccines is traditionally evaluated in specific pathogen-free (SPF) chickens. But under field conditions, residual maternal antibody (mAb) levels may interfere with vaccine efficacy. In this study, commercial broilers with various levels of maternally derived antibodies were vaccinated with IBDV vaccines of different virulence (vaccines 1-3, intermediate; vaccine 4, intermediate plus). At an average maternal virus-neutralizing antibody (mAb) level of log2 10.8 (range 7.6-11.6) at day of vaccination, only the intermediate plus vaccine induced IBDV antibodies after 18 days, while the other intermediate vaccines did not. At average mAb levels of log2 6.7 (range 5.6-8.6) at day of vaccination, all vaccines induced circulating antibodies, although the onset of antibody production differed significantly between strains (P < 0.05). While the intermediate plus vaccine induced enzyme-linked immunosorbent assay antibody levels already at 14 days postvaccination (PV), the intermediate vaccines induced significant antibody levels 28 (vaccines 1, 2) and 35 (vaccine 3) days PV. The time of IBDV antibody induction correlated with the onset of bursa lesions. The severity of lesions was comparable between vaccines 1, 3, and 4 (lesion score 4), while vaccine 2 induce only mild lesions of score 1 in 23% of the tested birds. Despite the induction of antibodies, none of the tested vaccines fully protected against challenge with vvIBDV. All challenged birds had either significantly higher bursal lesion scores or a higher IBDV antigen load in the bursa or sometimes both in comparison with nonchallenged birds (P < 0.05). Our study demonstrates that the evaluation of IBDV-vaccine efficacy is difficult in commercial broilers. For the first time, it was shown that the onset of bursa lesions and recovery of IBDV-vaccinated broilers is delayed in the presence of mAb in comparison with SPF chickens but not suppressed as previously assumed. At the time of challenge, vaccinated birds may still have significant bursa lesions and may lack target cells for IBDV-challenge virus. To be able to evaluate vaccine efficacy in commercial broilers, parameters such as intrabursal IBDV-antigen load should also be considered in conjunction with bursa lesion scores.     

Safety and efficacy of in ovo administration of infectious bursal disease viral vaccines

In ovo vaccination against Marek's disease virus and infectious bursal disease virus (IBDV) in commercial broilers in the United States is common. Little information exists as to the safety and efficacy of intermediate IBDV vaccines given in ovo. Experiments were initiated to determine the safety and efficacy of three commercially available live intermediate IBDV vaccines by in ovo route. Commonly used vaccines were given at 18 days of embryonation to specific-pathogen-free (SPF) broiler embryos (first and second study) or to commercial broiler embryos (third study) that had maternal antibody against IBDV. When any of the antigenic standard vaccines was given at full dose to SPF embryos, embryonic and 3-wk posthatch mortality increased. Vaccines also caused significant microscopic lesions in the bursa of Fabricius at 1 and 3 wk posthatch. In contrast, there was no adverse effect on embryonic or posthatch mortality when vaccines were given at half dose to SPF or commercial broiler embryos. However, significant microscopic lesions were evident at 1 and 3 wk posthatch in the bursae of SPF embryos given the vaccines at half dose. When vaccines were given at half dose to commercial broiler embryos, lesions were evident at 1 but not 3 wk of age. In the third study, in ovo vaccinated chickens were challenged with either a virulent standard (APHIS) or antigenic variant (variant E) IBDV virus at 3 wk of age. All vaccines produced at least 87% protection against the standard and 60% protection against the variant challenge IBDV, as measured by bursal weight to body weight ratios. This study was the first to examine the safety and efficacy of the three commonly used intermediate IBDV vaccines given in ovo in protection against standard and antigenic variant IBDV challenge viruses.     

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.     

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.     

Comparative immunopathogenesis of mild,intermediate, and virulent strains of classic infectious bursal disease virus

Differences in the immunopathogenesis of several strains of infectious bursal disease virus (IBDV) were compared. The strains included a virulent virus (IBDV-IM) and three vaccine viruses that included an intermediate vaccine virus (IBDV-B2) and two mild vaccine viruses (IBDV-Lukert and IBDV-BVM). The most significant differences were found in the systemic effects of these strains. In comparison with other strains, IBDV-IM antigen was detectable for up to 8 days postinfection (PI) in lymphoid tissues that included spleen and cecal tonsils, whereas only a few IBDV-B2- and IBDV-Lukert- and no IBDV-BVM-inoculated birds had detectable IBDV antigen in these tissues. IBDV-IM induced systemic circulating nitrite levels in over 86% of the birds at days 2 and 3 PI. IBDV-IM suppressed most vigorously the splenic mitogenic response on days 3-8 PI. Among the three vaccine strains, IBDV-B2 was the most virulent of the three, inducing a significant suppression of the mitogenic response (P < 0.05) and the most vigorous lesions in the bursa of Fabricius with the highest possible lesion score of 4 at 3 days PI (P < 0.05). IBDV-BVM was the mildest strain, not inducing any detectable lesions in lymphoid tissue at the tested time points. Whereas all IBDV-BVM-inoculated and 67% and 33% of the IBDV-Lukert- and IBDV-B2-inoculated birds, respectively, had detectable IBDV antigen in the bursa at 4 days postchallenge, none of the IBDV-IM-inoculated birds was positive for IBDV by immunohistochemistry. IBDV-IM induced the highest enzyme-linked immunosorbent assay (ELISA) antibody levels detected at days 8-29 PI (P < 0.05) and the best protection against challenge virus replication in comparison with IBDV-B2 and IBDV-Lukert. Only one of five IBDV-BVM-inoculated birds developed anti-IBDV ELISA antibodies at 29 days PI, and none of the birds was protected against IBDV challenge. We speculate that better protection with more virulent strains was due to more systemic antigenic stimulation on the basis of higher replication of IBDV in extrabursal lymphoid tissues. Interestingly, IBDV-IM did not differ from IBDV-B2 and IBDV-Lukert in its ability to induce T cell accumulation in the bursa at 8 days PI and local interferon-gamma induction from days 2 to 5 PI. These results suggested that the local T cell events in the bursa alone may not be indicative of a rapid and protective immune response.     

CAV与REV共感染SPF鸡对疫苗免疫反应的抑制作用

用1日龄SPF鸡人工感染鸡贫血病毒(CAV)和禽网状内皮增生病病毒(REV),探讨病毒感染对鸡体疫苗免疫反应的影响。结果表明,在用禽流感病毒(AIV,H5和H9)疫苗免疫后,CAV与REV单独感染均显著抑制了鸡体对H5和H9亚型禽流感病毒灭活疫苗的HI抗体反应,在CAV与REV共感染后,这种抑制作用更为明显。CAV单独感染后鸡体对新城疫病毒(NDV)和传染性法氏囊病病毒(IBDV)疫苗的免疫反应受到抑制,但与对照组在统计学上的差异不显著,然而,CAV可以显著加重REV感染对鸡体在NDV和IBDV疫苗免疫后抗体反应的抑制作用。从而证实CAV与REV共感染在疫苗免疫抑制上有协同作用。     

Infectious bursal disease virus variant from commercial Leghorn pullets

An infectious bursal disease virus (IBDV) was isolated from 39-to-43-day-old commercial leghorn pullets suspected of having infectious bursal disease (IBD). These chickens had been vaccinated with a commercial live IBDV vaccine at 28 and 35 days of age. An isolate designated IN was recovered using specific-pathogen-free (SPF) chickens and the BGM-70 established cell line. Experimental studies using SPF chickens vaccinated with either inactivated vaccines made from the vaccine strain used in the problem flock or a standard-type vaccine indicated no protection against the IN isolate. However, two variants and another standard-type vaccine induced protection against the IN isolate. Cross-neutralization tests indicated that the IN isolate differed antigenically from commercial vaccine strains and was related to the variant IBDV strains recently isolated from broilers. To our knowledge, this is the first report of a variant IBDV recovered from commercial layer chickens in the United States.     

The effects of age, route of exposure, and coinfection with infectious bursal disease virus on the pathogenicity and transmissibility of chicken anemia agent (CAA)

Specific-pathogen-free (SPF) chickens were inoculated with several different concentrations of chicken anemia agent (CAA) by the intra-abdominal, intratracheal, or oral routes. Based on lowered hematocrit values, the birds were most susceptible to CAA introduced by the intra-abdominal route. When SPF chickens were infected with infectious bursal disease virus (IBDV) at 1 day of age, they remained susceptible to CAA up to at least 21 days, whereas birds inoculated with CAA alone were susceptible only at 1 day of age. Infectious bursal disease virus introduced at 1 day of age also increased the susceptibility of birds to contact infection with CAA and resulted in increased mortality rates in CAA inoculates. The response of SPF birds to CAA infection varied following exposure at 1 day of age to two different strains of IBDV (STC and Variant-E). Chicken anemia agent contacts and inoculates infected with the Variant-E strain were affected 1 week earlier by CAA than by STC inoculates, as evidenced by depressed hematocrits. However, the total number of birds affected was similar for both the Variant-E and STC-inoculated chickens. Commercial broiler chickens inoculated at 1, 7, 10, and 14 days of age by non-parenteral routes with CAA or a combination of CAA and IBDV had mean hematocrits that were lower than controls. Several CAA-inoculated birds were considered anemic, with hematocrit values of 25 or less, while uninoculated birds remained within normal ranges.     

Pathogenicity of fowl adenovirus isolated from gizzard erosions to immuno-suppressed chickens

Pathogenicity of a fowl adenovirus (FAV), JM1/1 strain of serotype 1 derived from gizzard erosions of a broiler chicken, was examined to specific pathogen-free (SPF) chickens pre-treated with infectious bursal disease viruses (IBDVs) or cyclophosphamide (CY). Virulent IBDVs, classical type, were inoculated orally at 3 days of age of SPF chickens. CY was treated subcutaneously for 3 days after hatch. FAV was given orally at 30 days of age. At 40 days of age, all chickens were bled and autopsied for serology and gross observation. Gizzard lesions were ranked by the scores depending on their severities. IBDV- or CY-treated chickens showed significantly higher gizzard lesion scores than non treated birds. There were no gross lesions in any other organs except for bursal atrophy. Serologically, antibody production against FAV was highly suppressed by IBDV infection or CY treatment.     

Humoral and cell-mediated immune responses in chickens with infectious bursal disease

Primary and secondary immune responses to Newcastle disease virus (NDV) was evaluated in chickens infected with infectious bursal disease virus (IBDV) at one and 28 days of age. The geometric mean primary hemagglutination-inhibition antibody titers (GMT) of chickens infected with IBDV at one day of age was significantly lower (P less than or equal to 0.01) than those infected at 28 days of age. Infection with IBDV had no influence on secondary immune response to NDV. The effect of IBDV infection at one day of age on the cell-mediated immunity of chickens was evaluated by skin allograft acceptance or survival time. There was no significant difference between the percentage of grafts accepted in IBDV infected and noninfected control chickens. However, the mean graft survival time in the IBDV infected chickens was significantly longer (P less than or equal to 0.05) than those in the control group. This suggested a suppression of cell-mediated immunity due to IBDV infection.     

Field efficacy of different vaccines against infectious bursal disease in broiler flocks

A field study was performed to determine the efficacy of three commercially available vaccines against infectious bursal disease (IBD) in commercial broilers raised in a high IBD virus (IBDV) risk area. Live attenuated intermediate and intermediate plus vaccines were used in four flocks. Birds were vaccinated orally at the estimated vaccination time. Three broiler flocks were vaccinated subcutaneously with a turkey herpesvirus (HVT)-IBD vector vaccine at one day old. Evaluation of the efficacy of different vaccines was focused on humoral immune response, bursa/body weight (B/Bw) ratio, molecular detection of IBDV in ileocaecal tonsils and bursa of Fabricius, and production parameters. The serological results showed that although the uptake of all three vaccine strains was confirmed in the lymphoid organs, no significant antibody response to vaccination was detected in flocks vaccinated with intermediate and intermediate plus vaccines. A significant increase in antibody titres detected in flocks vaccinated with the vector vaccine indicated its ability to induce an immune response in birds with a high level of maternally derived antibodies. Observations obtained in this field trial did not confirm the expected reduction of the B/Bw ratio in flocks vaccinated with less attenuated vaccines. No significant differences were observed between birds vaccinated with the vector vaccine and those immunised with the intermediate plus vaccine. Very virulent IBDV was confirmed in the flock vaccinated with the intermediate vaccine. The infection induced reduced B/Bw and moderate mortality but did not affect the production parameters. Field infection was not detected in broilers vaccinated with the intermediate plus vaccine and the vector vaccine.     

Effect of in ovo administered reovirus vaccines on immune responses of specific-pathogen-free chickens

We investigated the effect of in ovo administered reovirus vaccines on the immune responses of specific-pathogen-free chickens. T-cell mitogenic responses to concanavalin A were numerically lower at 9 and 12 days of age and significantly lower at 6 days of age in birds vaccinated with a commercial reovirus vaccine compared with unvaccinated birds or birds vaccinated with an experimental reovirus-antibody complex vaccine. There were no significant differences in proportions of subpopulations of helper (CD4+CD8-) or cytotoxic (CD4-CD8+) T cells except at 12 days of age, when the percentages of CD4-CD8+ cells in the two vaccinated groups were statistically higher than in the nonvaccinated group. B-cell populations were not different among vaccine groups except at 9 days of age, when the vaccinated groups had the highest level of B cells. This commercial reovirus vaccine should not be given in ovo to embryos having little or no maternal antibody, otherwise immunosuppression may occur in the chicks. The addition of the antibody complex to the vaccine prevented this T-cell immunosuppression.     

Field trial in commercial broilers with a multivalent in ovo vaccine comprising a mixture of live viral vaccines against Marek's disease,infectious bursal disease,Newcastle disease,and fowl pox

A multivalent in ovo vaccine (MIV) was tested for safety and efficacy in a commercial broiler complex. The MIV comprised five replicating live viruses including serotypes 1, 2, and 3 of Marek's disease virus (MDV), an intermediate infectious bursal disease virus (IBDV) and a recombinant fowl poxvirus (FPV) vector vaccine containing HN and F genes of Newcastle disease virus (NDV). The performance of MIV-vaccinated broilers was compared with that of hatchmates that received turkey herpesvirus (HVT) alone (routinely used in ovo vaccine in the broiler complex). The chickens that hatched from the MIV-injected and HVT-injected eggs were raised under commercial conditions in six barns. Barn 1 housed 17,853 MIV-vaccinated chickens and each of the barns 2-6 housed 18,472-22,798 HVT-vaccinated chickens. The HVT-vaccinated chickens were given infectious bronchitis virus (IBV) and NDV vaccines at hatch and at 2 wk of age. The MIV-vaccinated chickens received IBV vaccine at hatch and IBV + NDV at 2 wk of age. The relative values of hatchability of eggs, livability and weight gain of chickens, and condemnation rates at processing were comparable between the MIV and the HVT groups (P > 0.05). Chickens from the MIV- and the HVT-vaccinated groups were challenged with virulent viruses under laboratory conditions. The resistance of vaccinated chickens against Marek's disease could not be assessed because of high natural resistance of unvaccinated commercial broilers to virulent MDV. The relative resistances of the MIV- and the HVT-vaccinated groups, respectively, against other virulent viruses were as follows: IBDV, 100% for both groups; NDV, 81% vs. 19%; FPV, 86% vs. 0%. The successful use of MIV under field conditions expands the usefulness of the in ovo technology for poultry.