ISOLATION OF INFECTIOUS BURSAL DISEASE VIRUS (IBDV), ITS COMPARISON WITH VACCINE STRAINS AND EFFICACY OF DIFFERENT VACCINATION REGIMENS IN PASSIVELY IMMUNE CHICKS
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Date
1995
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AAU, Anand
Abstract
Infectious bursal disease (IBD) which was characterised by relatively less mortality but more immunosuppression was earlier prevalent in India. During a past couple of years, there is country-wide change in pattern of disease with high mortality rates. Some of the farms investigated in the present study have also experienced high mortality among both, layers (six to 14 weeks) and broilers (four to six weeks). Present study was undertaken on isolation of IBDV from field samples, study their pathogenicity and antigenicity in comparison with existing vaccine strains; and formulation of vaccination regimen effective enough to give protection to chicks in presence of maternal antibody. Before taking up virus isolation, tissue homogenates from clinical cases were examined by agar gel diffusion test (AGDT) against reference antiserum and their identity as IBDV was confirmed. Overall AGDT positive samples ranged between 40.0 and 70.5 per cent. Positive field samples were grouped under five groups (UNO, KAM, AND, HAL and NAV), according to different geographical regions. An attempt was made for isolation of IBDV from all the five representative samples in different indicator systems. Among experimentally inoculated chicks, the clinical disease was reproduced in case of UND, KAM and AND isolate infected group. No clinical symptoms were observed in NAL and NAV isolates infected groups. The birds inoculated with each samples and sacrified 48 hours PI, had gross and histopathologic changes suggestive of IBD. Specific precipitation lines were demonstrated in bursal homogenates. Out of five field isolates, three isolates (UND, KAM and AND) were adapted to growth in chick embryos by chorioallantoic membrane (CAM) route. Predominant changes were observed in the embryos which included slow and stunted growth, cutaneous edema and hemorrhages in subcutis. UND isolate was serially passed onto chicken embryo fibroblast (CEF) cell culture for its adaptation to growth. Characteristic cytopathic effect (CPE) was observed earliest, at 30 hours PI, and maximum CPE showing degeneration of about 80 per cent cell monolayer, at 48 hours PI. Syncytia formation due to aggregation of large number of nuclei of dead cells was observed in later stages. Other field samples failed to induce CPE in cell culture upto five serial passages. Comparative pathogenicity studies on five field isolates, one reference isolate (BAN) and vaccine strain of intermediate (Vac-I) and mild (Vac-M) virulence each, were carried in chicks, chick enbryos and CEF cell culture. Birds infected with UHD and BAM isolates showed severe clinical synptons and also hemorrhagic lesions in the bursa of Fabricius (BF), skeletal muscles and proventriculus-gizzard junction. Concurrent histopathologic lesions in the BF, at 48 hours PI, includes marked edema, hyperemia, necrosis of lymphoid cells, and formation of vacuoles or cystic follicles. In case of birds inoculated with Vac-I strain, the enlargement and edema was more marked which persisted longer. Vaccine strain induced histopathological lesions in BF which were of mild degree and comparable to those found in birds infected with NAL and NAV isolates. As regard to the pathogenicity of field isolates in chick embryos inoculated by CAM route, UND and BAN isolates caused almost 80 per cent mortality. Among other isolates, AND and KAM showed relatively low mortality rates followed by HAL and NAV. Unlike any of the recent field isolates tested in the present study, the UND isolate grew well in CEF cells exhibiting CPE in infected monolayers. Thus, only the UND and BAN isolates, were able to maintain consistent pathogenic potential in every laboratory system tested. Slight variation as regard to period required for initiation of CPE was observed in UND and BAN isolates when compared with that of vaccine strains. Highest virus infectivity titre was recorded in Vac-I strain, while in Vac-M strain the titre was lowest. The virus titres in both field isolates (UND and BAN) were sane. Neutralization and cross neutralization tests were carried out to assess the antigenic relatedness of a field isolate, reference isolate and vaccine strains using homologus and heterologus innune sera. The results indicated close antigenic relationship of the isolates with vaccine strains. In the present study, efficacy of three vaccination regiuens was evaluated in the progeny obtained from breeder hens having high levels of maternal antibody (MA). The levels of precipitating antibodies of progeny chicks at hatch indicated, on an average, 50 to 55 per cent passive transfer of MA to progeny. After hatch, higher levels of MA sustained for one week and afterwards declined steadily at the half-life of approximately six to eight days. MA attained low levels between 21 and 28 days of age. While after 28 days of age, precipitating antibodies were undetected by quantitative AGDT (QAGDT), still these antibodies were detected by enzyme linked immunosorbent assay (ELISA). Live vaccine (mild) was used at day-old age for 'priming' of all the birds and subsequently, combination of live (intermediate) and/or inactivated vaccines was tested in three groups. One group served as control. The response of birds, vaccinated once or twice with live intermediate vaccine was not satisfactory as indicated by low levels of antibodies even after vaccination. On challenge, some birds showed clinical symptoms and bursal damage. The group in which birds received inactivated vaccine, at seven days of age and live vaccine at 14 and 28 days of age, showed significantly (P <0.05) higher levels of antibody. On challenge, the birds did not show clinical symptons and mortality, but still these birds were not prevented from infection because the bursal danage was apparent indicating inadequate response against virulent IBDV. Overall studies indicated the presence of highly virulent IBDV in sone pockets of Gujarat State. Therefore, it is important to locate such pockets at an earliest to prevent possible spread of highly virulent IBDV and carry out effective vaccination coupled with strict biosecurity measures, where immunization has failed probably because of increased virulence of IBDV. Serological monitoring of progeny flock is necessary to determine when MA in chicks decline to levels that the vaccine can overcome. Without this information, it is very difficult to control highly virulent IBDV.
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VETERINARY MICROBIOLOGY, A STUDY