J. COMP.
PATH.
1975.
VOL.
597
85.
COMPARISON
OF VACCINES AGAINST BURSAL DISEASE
DENISE
and M. PATTISON*
H. THORNTON Central
Veterinary
INFECTIOUS
Laboratory,
Wqbridge,
Surrey
INTRODUCTION
Infectious bursal disease (IBD), or Gumboro disease, is a viral disease affecting chicks causing a high rate of morbidity and an average mortality of 5 per cent. (Faragher, 1972). In Britain, the disease appears as early as 10 days of age in contrast to the findings in Europe and U.S.A. where it does not appear before 3 weeks of age (Rinaldi, Cessi, Cervio and Lodetti, 1972; Hitchner, 1971). The infectious bursal agent (IBA) multiplies in the bursa of Fabricius which is the maturation site for immunocompetent cells (B cells) and the agent has been shown to be immunosuppressiveafter inoculation at one day of age (Faragher, Allan and Cullen, 1972). Damage in the form of lymphocyte necrosis occurs in the bursa and this can be detected by histological examination. The economic importance of the diseasemay therefore be due not only to mortality, but to impaired weight gain in broilers (Rinaldi et al., 1972) and to unsatisfactory response to vaccination against other diseases(Allan, Faragher and Cullen, 1972). The use of hygienic measures to eradicate the diseaseis normally unsuccessful (Benton, Cover, Rosenberger and Lake, 1967). Attempts to control the diseaseby vaccination have been reported. Planned infection with unmodified strains of IBA was used by Dorn, Kronthaler and Schindler (1968) and Edgar and Cho (1973). The use of vaccine partly attenuated by egg passagewas reported by Snedeker, Wills and Moulthrop (1967), while Winterfield (1969) and Rinaldi et al. (1972) used further attenuated vaccines. The serological response to a vaccine prepared by attenuation in baby mice was described by Bengelsdorff and Bernhardt (1971), but no evidence of protective capacity was presented. Vaccines currently available commercially in Europe and U.S.A., are live vaccines intended for administration to chicks during the first few weeks of life. At the time of writing, no preparation is available for general use in Britain. This report is a comparative account of the properties of various vaccinal strains of IBA which were studied for their suitability for use in Britain. It was considered that vaccines should be capable of administration during the first few days of life and should confer rapid protection because of the early age at which the disease occurs in this country. It was, therefore, necessary to ensure that, in addition to being effective, the vaccines were safe for administration at this age and were not significantly immunosuppressive. MATERIALS
AND
METHODS
Vaccines. Final container samples of vaccines were obtained from seven sources
and designated A to G. Samples of 2 batches were obtained direct from 2 of the manufacturers : these were designated El and E2, and Fl and F2. It was not possible to test all the vaccines simultaneously and not all the tests described below were *Present
address:
Ross Poultry
Ltd.,
Newbridge,
Midlothian.
598
D.
H.
THORNTON
AND
M.
PATTISON
applied to each product if the results of other tests indicated that the sample was unsuitable. All but one of the vaccines were recommended for drinking water administration, but for the current work it was necessary to ensure that each chick received a full dose of vaccine; the products were therefore reconstituted in distilled water to contain one field dose in O-025 ml. and given by eye-drop. One vaccine was recommended for injection intramuscularly (i/m) and so was reconstituted in the diluent provided and given i/m. After inoculation of the birds in each experiment, the inoculum was checked for viability by the injection of groups of 10 g-day chick embryos, onto the chorioallantoic membrane (CAM). All inocula were found to be viable on each occasion. Birds. In addition to normal SPF test requirements (Thornton, 1974) the parent flocks were certified free from IBA infection. Chicks from SPF hatching eggs* had no antibody to IBA detectable by the agar gel precipitin (AGP) test. Each group was separately housed in virus-proof isolators, except that after challenge with virulent virus the relevant groups were transferred to a single room to eliminate possible effects of individual environments on the response to challenge. Antibody response. Serum samples were obtained from birds 14 and 24 days after inoculation. AGP tests were done using IBA antigen prepared from the Cheville strain? of IBA (Faragher, 1971). This preparation was free from detectable antigen to reovirus. Test for sufe9. Observations were made of the effect of vaccination on groups of 30 birds, each bird being vaccinated with one field dose at 7 days of age. Vaccines B, Es and G were given to further groups of 20 chicks at 1 day of age. In each case, a comparison was made with birds given a known virulent strain, 52170. Sickness and death were recorded over a period of 24 days. Depression of weight gain was assessed using groups of 10 birds weighed just prior to inoculation at 7 days of age. Comparison was made with uninoculated control birds by weighing each bird 4 days after the inoculation. Approximately 10 birds from each group were killed at 4, 14 and 24 days after vaccination at 7 days of age, bursa and body weights were obtained and histological examination was made of 2 bursae from each group at these times. Observations of the effect of vaccination were also made in other experiments described below. Test for virus content. Titration was performed by inoculation of serial ten-fold dilutions of vaccine reconstituted in peptone broth onto the CAM of SPF eggs, 9 days of age, using 7 eggs per level. Embryos were judged to be infected when characteristic lesions (Hitchner, 1970) were found in embryos that died and in those that survived the 7 day observation period. The median embryo infective dose (EID,,) was calculated by the method of Spearman and Karber (Finney, 1964). Test for potency. Vaccination was carried out at 7 days of age using groups of 10 or 20 birds. Ten days later, the birds together with unvaccinated controls were challenged by the eye-drop route. The challenge viruses, designated 52/70 and 10/69, had been isolated at the Central Veterinary Laboratory from outbreaks of disease in Britain and given 2 or 3 passages in chickens. Bursal homogenate was used to challenge the groups of experimental chicks. The birds were observed for 14 days, the incidence of sickness and death being recorded daily. In some experiments, the birds and bursae were weighed at the end of the observation period and histological examination was made of at least 2 bursae from each group. Test for immunosufipression. A selection of vaccines was tested for immunosuppressive properties excluding those which were considered to confer inadequate protection and those causing the most immediate and serious damage to the bursa. IBD vaccine was given at 1, 7 or 12 days of age to groups of 15 to 30 chicks. Newcastle disease (ND) vaccine Bl strain was given by the intranasal installation of one field dose of commercial vaccine at 21 days of age. The source of the B, vaccine varied from experiment to experiment. Thirteen days later, the haemagglutination inhibition *SAN Eggs Inc, Salisbury, toriginally obtained from
Wiltshire. Dr N. F. Cheville,
Ames,
Iowa.
COMPARISON
OF
VACCINES
AGAINST
INFECTIOUS
BURSAL
599
DISEASE
(HI) antibody response to ND vaccine was determined and the following day, all birds including a group given B,, but no IBD vaccine, were challenged i/m with virulent ND virus, Herts 33/56 strain. Deaths were recorded over the following week. In one experiment, antibody response to sheep red blood cells given at 14 days of age was also determined in order to assess the degree of immunosuppression using a non-replicating antigen. The agglutination titre was measured 7 days after the i/m inoculation of 107 red blood cells. In the first experiment, the immunosuppressive effect of virulent IBA strain 52/70 was determined as a positive control. Histological technique. Tissues were fixed in 10 per cent buffered neutral formalin and processedin a routine manner. Sections were cut at 5 urn. and stained with HE. RESULTS
Antibody Response The virulent IBA strain 52/70 and most of the vaccines induced the formation of antibody detectable by the AGP test. Antibody was detected in a proportion of birds at 14 days and in all birds at 24 days. No antibody was detected in uninoculated controls or in any bird given vaccines El or Fl. Safeety The virulent strain 52170 caused a 90 per cent. morbidity and 35 per cent. to 50 per cent. mortality in chicks when given at 1 or 7 days of age. No vaccine given at 7 days of age produced sickness or death in any chicks in any of the experiments. Vaccines B, E2 and G were also given to further groups of 20 chicks one day of age, when vaccines E2 and G caused no symptoms, but vaccine B caused a 40 per cent. morbidity and 2 birds died. TABLE1 WEIGHT
(g.)
OF CHICKS
Expt. Inoculum at 7 days of age
Vaccine .4 Vaccine B Vaccine C Vaccine D Vaccine El Vaccine Fl Virulent strain 52170 Uninoculated controls
Weight at 7 days of age
4
DAYS
AFTER
INOCULATIOS
I
Expt. Weight at II days of age
42.1 39.0
72.6 67.6
Weight at 7 days of age
2 Weight at I I days of age
39.4 39.8 37.4
74.4 7943 82.8
42.7 40.3 42.1 43.4 42.7 43.2
39.2
56.0
43.5
57.2
37.8
82.3
44.1
78.8
Not
done
73.6 66.8 68.9 69.9 76.0 74.2
The effect of 6 of the vaccines on weight gain was assessedusing groups of 10 chicks which were weighed 4 days after inoculation. Significant reduction in weight gain occurred within 4 days with the virulent strain 52/70 (P < 0.001) and with vaccine B (P < 0.01). The results of 2 experiments are shown in Table 1. The effect of the IBA preparations on the weight of the bursa and on bursa: body weight ratio is shown in Figs 1 and 2. The virulent strain 52/70 significantly H
600
D.
H.
THORNTON
AND
M.
PATTISON
reduced the size of the bursa within 4 days (P < O-001) and, although these birds were lighter than the controls, the bursa : body weight ratio was still significantly less than that of the controls (P < 0.001). Vaccine B reduced the size of the bursa and the bursa body weight ratio (P < O-05), but no other vaccine had such a pronounced effect at 4 days. In the case of the virulent strain and vaccines A, B and C, the bursae and bursa : body weight ratios were 52/70 100 I-
EC l-
s E .$ 3
A
B
-.--1
C
D
E2
FI
F2
G,
ooo
r
X
2 Xf
go i i
6C l-
:: 5 m 4c )-
J
X
\
1
‘f: “i,
2c )-
0
X
4 .I424
yl424
41424
41424
p
1 I
(I41424 Days after
Fig.
El
.% 41424
41424
414#
41424
414:
J! %I
lnOCdatiOn
1. Effect of IBA preparations on the bursa at intervals after inoculation. The bursa weight is expressed as a percentage of the values obtained for uninoculated control birds. The bursa weights for the control birds were 0.28 g. at 4 days, 0.49 g. at 14 days and I.00 g. at 24 days. (x-x) Bursa weight; (O---O) Histological damage; (---) Values for bursal weight below the lines are sigmficantly different from the values for uninoculated controls. (P < 0.01) ; (- - -) Values for bursal weight above the lines are significantly different from the values obtained with the virulent strain 52/70 (P < 0.01). Histological damage: 0, No effect. 1, Isolated follicles only show mild necrosis. 2, Mild lymphoid depletion with slight reticuloendothelial cell hyperplasia and connective tissue formation in plicae. 3, Acute change with loss ofmature follicles, epithelial corrugation and cyst formation. 4, Very severe change with complete loss of follicular architecture. 5, Reticulum cell hyperplasia and fibroplasia causing plicae to be shrunken with corrugated epithelium.
much reduced by 14 days and a similar picture was seen at 24 days. Vaccine D produced a less rapid decrease in the size of the bursa. Vaccine F2 caused a significant reduction in size by 14 days, but with some recovery by 24 days. Vaccines El, E2 and Fl produced a transient decrease and vaccine G had no significant effect on bursa weight or bursa : body weight ratio. Histological
Observations on Bursae of Inoculated Birds
The results are summarized in Fig. 1. Vaccine A. Four days after vaccination isolated follicles showed necrosis in the medulla. By 14 days, the damage had progressed
lymphocyte and mature
COMPARISON
OF
VACCINES
AGAINST
INFECTIOUS
BURSAL
DISEASE
601
follicles had disappeared. The remaining follicles were atrophic, the epithelium had become cystic and the plicae contained an increased amount of connective tissue. At 24 days, one bursa was severely damaged, but the other showed evidence of regeneration.
OL
4 14 24 4
1424
4 14 24 Days
on Fig. 2. Effect of IB.4 preparations are significantly less than the Values for uninoculated control and 0.45 per cent at 24 days. those obtained for the virulent
after
inoculation
the bursa: body weight ratio. (- - - -) Values below the lines value s obtained for uninoculated control birds (P < 0.01). birds were 0.34 per cent at 4 days, 0.40 per cent at 14 days ( -----) Values above the lines are significantly greater than strain 52/70 iP < 0.01).
VaccineB. At 4 days there was acute inflammation with oedema, heterophil infiltration and lymphocyte depletion. Signs of regeneration were seen at 14 days, but this had progressed no further by 24 days. Vaccine C. At 4 days, there was acute inflammation with oedema and lymphocyte depletion and necrosis. By 14 days the plicae were corrugated and contained atrophic follicles, but there was evidence of regeneration which continued to some extent in one bursa, but not in the other, at 24 days. Vaccine D. At 4 days no change was observed in either bursa, but severe damage was seen at 14 days, with the follicular architecture obliterated by reticula-endothelial hyperplasia. Regeneration was seen in one bursa at 24 days but not in the other. Vaccine El. No change was seen at any stage. Vaccine E2. At 4 days there was slight lymphoid depletion without inflammation. By 14 days lymphoid depletion was more evident and epithelial damage was present, but by 24 days the appearance was normal.
602
D.
H.
THORNTON
AND
M.
PATTISON
Vaccine Fl. No change was seen at any stage. Vaccine FL?. At 4 days necrosis was seen in about half the follicles, and by 14 days damage had progressed, but regeneration was taking place and this was more complete by 24 days. Vaccine G. No change was seen at any stage. Virulent strain 52170. At 4 days there was necrosis and atrophy of follicles with reticula-endothelial hyperplasia and oedema and corrugation of the epithelium. Damage was still severe at 14 days when the plicae were shrunken TABLE 2 PROTECTION
1
A B C:
19 ii
11
20 20 20 20 10 10 10 10
Controls E2 2 F2 G
Controls
AGAIMT
CHALLENGE*
Sick and dead birds
Dead
0 1 3 6 12 13 13 2 0
0 1 1 3 8 6 7 1
NS
0 0
P < 0.05 P < 0.05
0
9
P
PATH.
1975.
VOL.
597
85.
COMPARISON
OF VACCINES AGAINST BURSAL DISEASE
DENISE
and M. PATTISON*
H. THORNTON Central
Veterinary
INFECTIOUS
Laboratory,
Wqbridge,
Surrey
INTRODUCTION
Infectious bursal disease (IBD), or Gumboro disease, is a viral disease affecting chicks causing a high rate of morbidity and an average mortality of 5 per cent. (Faragher, 1972). In Britain, the disease appears as early as 10 days of age in contrast to the findings in Europe and U.S.A. where it does not appear before 3 weeks of age (Rinaldi, Cessi, Cervio and Lodetti, 1972; Hitchner, 1971). The infectious bursal agent (IBA) multiplies in the bursa of Fabricius which is the maturation site for immunocompetent cells (B cells) and the agent has been shown to be immunosuppressiveafter inoculation at one day of age (Faragher, Allan and Cullen, 1972). Damage in the form of lymphocyte necrosis occurs in the bursa and this can be detected by histological examination. The economic importance of the diseasemay therefore be due not only to mortality, but to impaired weight gain in broilers (Rinaldi et al., 1972) and to unsatisfactory response to vaccination against other diseases(Allan, Faragher and Cullen, 1972). The use of hygienic measures to eradicate the diseaseis normally unsuccessful (Benton, Cover, Rosenberger and Lake, 1967). Attempts to control the diseaseby vaccination have been reported. Planned infection with unmodified strains of IBA was used by Dorn, Kronthaler and Schindler (1968) and Edgar and Cho (1973). The use of vaccine partly attenuated by egg passagewas reported by Snedeker, Wills and Moulthrop (1967), while Winterfield (1969) and Rinaldi et al. (1972) used further attenuated vaccines. The serological response to a vaccine prepared by attenuation in baby mice was described by Bengelsdorff and Bernhardt (1971), but no evidence of protective capacity was presented. Vaccines currently available commercially in Europe and U.S.A., are live vaccines intended for administration to chicks during the first few weeks of life. At the time of writing, no preparation is available for general use in Britain. This report is a comparative account of the properties of various vaccinal strains of IBA which were studied for their suitability for use in Britain. It was considered that vaccines should be capable of administration during the first few days of life and should confer rapid protection because of the early age at which the disease occurs in this country. It was, therefore, necessary to ensure that, in addition to being effective, the vaccines were safe for administration at this age and were not significantly immunosuppressive. MATERIALS
AND
METHODS
Vaccines. Final container samples of vaccines were obtained from seven sources
and designated A to G. Samples of 2 batches were obtained direct from 2 of the manufacturers : these were designated El and E2, and Fl and F2. It was not possible to test all the vaccines simultaneously and not all the tests described below were *Present
address:
Ross Poultry
Ltd.,
Newbridge,
Midlothian.
598
D.
H.
THORNTON
AND
M.
PATTISON
applied to each product if the results of other tests indicated that the sample was unsuitable. All but one of the vaccines were recommended for drinking water administration, but for the current work it was necessary to ensure that each chick received a full dose of vaccine; the products were therefore reconstituted in distilled water to contain one field dose in O-025 ml. and given by eye-drop. One vaccine was recommended for injection intramuscularly (i/m) and so was reconstituted in the diluent provided and given i/m. After inoculation of the birds in each experiment, the inoculum was checked for viability by the injection of groups of 10 g-day chick embryos, onto the chorioallantoic membrane (CAM). All inocula were found to be viable on each occasion. Birds. In addition to normal SPF test requirements (Thornton, 1974) the parent flocks were certified free from IBA infection. Chicks from SPF hatching eggs* had no antibody to IBA detectable by the agar gel precipitin (AGP) test. Each group was separately housed in virus-proof isolators, except that after challenge with virulent virus the relevant groups were transferred to a single room to eliminate possible effects of individual environments on the response to challenge. Antibody response. Serum samples were obtained from birds 14 and 24 days after inoculation. AGP tests were done using IBA antigen prepared from the Cheville strain? of IBA (Faragher, 1971). This preparation was free from detectable antigen to reovirus. Test for sufe9. Observations were made of the effect of vaccination on groups of 30 birds, each bird being vaccinated with one field dose at 7 days of age. Vaccines B, Es and G were given to further groups of 20 chicks at 1 day of age. In each case, a comparison was made with birds given a known virulent strain, 52170. Sickness and death were recorded over a period of 24 days. Depression of weight gain was assessed using groups of 10 birds weighed just prior to inoculation at 7 days of age. Comparison was made with uninoculated control birds by weighing each bird 4 days after the inoculation. Approximately 10 birds from each group were killed at 4, 14 and 24 days after vaccination at 7 days of age, bursa and body weights were obtained and histological examination was made of 2 bursae from each group at these times. Observations of the effect of vaccination were also made in other experiments described below. Test for virus content. Titration was performed by inoculation of serial ten-fold dilutions of vaccine reconstituted in peptone broth onto the CAM of SPF eggs, 9 days of age, using 7 eggs per level. Embryos were judged to be infected when characteristic lesions (Hitchner, 1970) were found in embryos that died and in those that survived the 7 day observation period. The median embryo infective dose (EID,,) was calculated by the method of Spearman and Karber (Finney, 1964). Test for potency. Vaccination was carried out at 7 days of age using groups of 10 or 20 birds. Ten days later, the birds together with unvaccinated controls were challenged by the eye-drop route. The challenge viruses, designated 52/70 and 10/69, had been isolated at the Central Veterinary Laboratory from outbreaks of disease in Britain and given 2 or 3 passages in chickens. Bursal homogenate was used to challenge the groups of experimental chicks. The birds were observed for 14 days, the incidence of sickness and death being recorded daily. In some experiments, the birds and bursae were weighed at the end of the observation period and histological examination was made of at least 2 bursae from each group. Test for immunosufipression. A selection of vaccines was tested for immunosuppressive properties excluding those which were considered to confer inadequate protection and those causing the most immediate and serious damage to the bursa. IBD vaccine was given at 1, 7 or 12 days of age to groups of 15 to 30 chicks. Newcastle disease (ND) vaccine Bl strain was given by the intranasal installation of one field dose of commercial vaccine at 21 days of age. The source of the B, vaccine varied from experiment to experiment. Thirteen days later, the haemagglutination inhibition *SAN Eggs Inc, Salisbury, toriginally obtained from
Wiltshire. Dr N. F. Cheville,
Ames,
Iowa.
COMPARISON
OF
VACCINES
AGAINST
INFECTIOUS
BURSAL
599
DISEASE
(HI) antibody response to ND vaccine was determined and the following day, all birds including a group given B,, but no IBD vaccine, were challenged i/m with virulent ND virus, Herts 33/56 strain. Deaths were recorded over the following week. In one experiment, antibody response to sheep red blood cells given at 14 days of age was also determined in order to assess the degree of immunosuppression using a non-replicating antigen. The agglutination titre was measured 7 days after the i/m inoculation of 107 red blood cells. In the first experiment, the immunosuppressive effect of virulent IBA strain 52/70 was determined as a positive control. Histological technique. Tissues were fixed in 10 per cent buffered neutral formalin and processedin a routine manner. Sections were cut at 5 urn. and stained with HE. RESULTS
Antibody Response The virulent IBA strain 52/70 and most of the vaccines induced the formation of antibody detectable by the AGP test. Antibody was detected in a proportion of birds at 14 days and in all birds at 24 days. No antibody was detected in uninoculated controls or in any bird given vaccines El or Fl. Safeety The virulent strain 52170 caused a 90 per cent. morbidity and 35 per cent. to 50 per cent. mortality in chicks when given at 1 or 7 days of age. No vaccine given at 7 days of age produced sickness or death in any chicks in any of the experiments. Vaccines B, E2 and G were also given to further groups of 20 chicks one day of age, when vaccines E2 and G caused no symptoms, but vaccine B caused a 40 per cent. morbidity and 2 birds died. TABLE1 WEIGHT
(g.)
OF CHICKS
Expt. Inoculum at 7 days of age
Vaccine .4 Vaccine B Vaccine C Vaccine D Vaccine El Vaccine Fl Virulent strain 52170 Uninoculated controls
Weight at 7 days of age
4
DAYS
AFTER
INOCULATIOS
I
Expt. Weight at II days of age
42.1 39.0
72.6 67.6
Weight at 7 days of age
2 Weight at I I days of age
39.4 39.8 37.4
74.4 7943 82.8
42.7 40.3 42.1 43.4 42.7 43.2
39.2
56.0
43.5
57.2
37.8
82.3
44.1
78.8
Not
done
73.6 66.8 68.9 69.9 76.0 74.2
The effect of 6 of the vaccines on weight gain was assessedusing groups of 10 chicks which were weighed 4 days after inoculation. Significant reduction in weight gain occurred within 4 days with the virulent strain 52/70 (P < 0.001) and with vaccine B (P < 0.01). The results of 2 experiments are shown in Table 1. The effect of the IBA preparations on the weight of the bursa and on bursa: body weight ratio is shown in Figs 1 and 2. The virulent strain 52/70 significantly H
600
D.
H.
THORNTON
AND
M.
PATTISON
reduced the size of the bursa within 4 days (P < O-001) and, although these birds were lighter than the controls, the bursa : body weight ratio was still significantly less than that of the controls (P < 0.001). Vaccine B reduced the size of the bursa and the bursa body weight ratio (P < O-05), but no other vaccine had such a pronounced effect at 4 days. In the case of the virulent strain and vaccines A, B and C, the bursae and bursa : body weight ratios were 52/70 100 I-
EC l-
s E .$ 3
A
B
-.--1
C
D
E2
FI
F2
G,
ooo
r
X
2 Xf
go i i
6C l-
:: 5 m 4c )-
J
X
\
1
‘f: “i,
2c )-
0
X
4 .I424
yl424
41424
41424
p
1 I
(I41424 Days after
Fig.
El
.% 41424
41424
414#
41424
414:
J! %I
lnOCdatiOn
1. Effect of IBA preparations on the bursa at intervals after inoculation. The bursa weight is expressed as a percentage of the values obtained for uninoculated control birds. The bursa weights for the control birds were 0.28 g. at 4 days, 0.49 g. at 14 days and I.00 g. at 24 days. (x-x) Bursa weight; (O---O) Histological damage; (---) Values for bursal weight below the lines are sigmficantly different from the values for uninoculated controls. (P < 0.01) ; (- - -) Values for bursal weight above the lines are significantly different from the values obtained with the virulent strain 52/70 (P < 0.01). Histological damage: 0, No effect. 1, Isolated follicles only show mild necrosis. 2, Mild lymphoid depletion with slight reticuloendothelial cell hyperplasia and connective tissue formation in plicae. 3, Acute change with loss ofmature follicles, epithelial corrugation and cyst formation. 4, Very severe change with complete loss of follicular architecture. 5, Reticulum cell hyperplasia and fibroplasia causing plicae to be shrunken with corrugated epithelium.
much reduced by 14 days and a similar picture was seen at 24 days. Vaccine D produced a less rapid decrease in the size of the bursa. Vaccine F2 caused a significant reduction in size by 14 days, but with some recovery by 24 days. Vaccines El, E2 and Fl produced a transient decrease and vaccine G had no significant effect on bursa weight or bursa : body weight ratio. Histological
Observations on Bursae of Inoculated Birds
The results are summarized in Fig. 1. Vaccine A. Four days after vaccination isolated follicles showed necrosis in the medulla. By 14 days, the damage had progressed
lymphocyte and mature
COMPARISON
OF
VACCINES
AGAINST
INFECTIOUS
BURSAL
DISEASE
601
follicles had disappeared. The remaining follicles were atrophic, the epithelium had become cystic and the plicae contained an increased amount of connective tissue. At 24 days, one bursa was severely damaged, but the other showed evidence of regeneration.
OL
4 14 24 4
1424
4 14 24 Days
on Fig. 2. Effect of IB.4 preparations are significantly less than the Values for uninoculated control and 0.45 per cent at 24 days. those obtained for the virulent
after
inoculation
the bursa: body weight ratio. (- - - -) Values below the lines value s obtained for uninoculated control birds (P < 0.01). birds were 0.34 per cent at 4 days, 0.40 per cent at 14 days ( -----) Values above the lines are significantly greater than strain 52/70 iP < 0.01).
VaccineB. At 4 days there was acute inflammation with oedema, heterophil infiltration and lymphocyte depletion. Signs of regeneration were seen at 14 days, but this had progressed no further by 24 days. Vaccine C. At 4 days, there was acute inflammation with oedema and lymphocyte depletion and necrosis. By 14 days the plicae were corrugated and contained atrophic follicles, but there was evidence of regeneration which continued to some extent in one bursa, but not in the other, at 24 days. Vaccine D. At 4 days no change was observed in either bursa, but severe damage was seen at 14 days, with the follicular architecture obliterated by reticula-endothelial hyperplasia. Regeneration was seen in one bursa at 24 days but not in the other. Vaccine El. No change was seen at any stage. Vaccine E2. At 4 days there was slight lymphoid depletion without inflammation. By 14 days lymphoid depletion was more evident and epithelial damage was present, but by 24 days the appearance was normal.
602
D.
H.
THORNTON
AND
M.
PATTISON
Vaccine Fl. No change was seen at any stage. Vaccine FL?. At 4 days necrosis was seen in about half the follicles, and by 14 days damage had progressed, but regeneration was taking place and this was more complete by 24 days. Vaccine G. No change was seen at any stage. Virulent strain 52170. At 4 days there was necrosis and atrophy of follicles with reticula-endothelial hyperplasia and oedema and corrugation of the epithelium. Damage was still severe at 14 days when the plicae were shrunken TABLE 2 PROTECTION
1
A B C:
19 ii
11
20 20 20 20 10 10 10 10
Controls E2 2 F2 G
Controls
AGAIMT
CHALLENGE*
Sick and dead birds
Dead
0 1 3 6 12 13 13 2 0
0 1 1 3 8 6 7 1
NS
0 0
P < 0.05 P < 0.05
0
9
P
