Effects of Intranasal Immunization and Mucosal Adjuvants on the Immunogenicity of Pasteurella multocida in Rats.
Effects of Intranasal Immunization and Mucosal Adjuvants on the Immunogenicity of Pasteurella multocida in Rats.
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Date
2015-04-12
Authors
Geffer, Enass
Journal Title
Journal ISSN
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Publisher
UOFK
Abstract
This study was carried out to study the effects of intranasal
immunization, mucosal adjuvants and vaccine-adjuvant ratio on the
immunogenicity of Pasteurell multocida in rats. Forty-eight rats were kept in
a conventional animal house and allowed to adapt to the environment, food
and water for one week. They were assigned to one of five groups (G1 to
G5). G1, G2 and G3 were further divided into subgroups (a) and (b) each
composing 6 rats. Groups 4 and 5 were control groups of 6 rats each. The
rats were immunized intranasally with P. multocida broth bactrin mixed with
mucosal adjuvants at day one and day 15. Mucosal adjuvants used were
olive oil for group 1, sesame oil for group 2 and vitamin D3 for group 3. The
vaccine adjuvant ratio for all a subgroups was 1:1 and 2:1 for all subgroups
b. Rats in group 4 received plain P. multocida broth bacterin and rats in
group 5 were sham-vaccined with phosphate-buffered saline (PBS). All rats
were sacrificed on day 21 post initial immunization. Lung and trachea from
each rat were carefully removed and the trachea and bronchial tree were
washed thoroughly with 5 ml PBS. The tracheobronchial (TB) lavage was
collected, centrifuged and the supernatant was kept at -20 ºC for the
detection of P. multocida specific mucosal Ab using gel diffusion
precipitation test (GDPT) and indirect haemagglutination (IHA) assay. P.
multocida whole cell lysate (WCL) was used as antigen for GDPT and for
coating formalized tanned RBCs. In GDPT, precipitation lines were obtained
using TB lavage from all Pasteurella multocida intranasally immunized rats,
and relatively sharp precipitation lines were obtained in TB lavage of rats of
G1b, G2a and G3a, indicating the presence of P. multocida specific Ab in
their TB lavage. The IHA was performed using two-fold dilution of TB
lavage in v-shaped microtitre plates, starting with 1/200 dilution. Significant
iv
(P<0.05) titre were obtained in TB lavage of G1b, G2a and G3a. The IHA
titre was expressed as the reciprocal of the highest dilution of TB lavage
showing a diffuse matt of erythrocytes as compared to negative control
showing a clearly delineated spot of erythrocyte within the vertex of the
well. Rats of G1b that received P. multocida broth bactrin in olive oil in a
ratio of 2:1 gave high Ab activity in TB lavage as was determined by IHA
assay compared to ratio of group 4 ( Adjuvant control) that received plain
broth bactrin vaccine. This difference is highly significant (P=0.0082). Rats
in this group G1b showed also higher Ab activity in their TB lavage than
rats in group 2b that received P. multocida broth bactrin in sesame oil in a
ratio of 2:1 (P=0.9991) as well as to rats in G3b (P=0.9661). On the other
hand, rats in group 2a that received P. multocida broth bactrin in sesame oil
in a 1:1 ratio showed high Ab activity in their TB lavage as determined by
IHA assay. The Ab titre of this group 2a was significantly higher (P=0.0227)
than in rats of G2b, G1a and G3b. Similarly, rats in G3a that received P.
multocida broth bactrin in vitamin D3 in the ratio of 1:1 had significant Ab
titres in their TB lavage, in comparison to G4 with P values equal to 0.0272.
In addition, sesame oil and vitamin D3 in 1:1 ratio with P. multocida broth
bactrin had induced high Ab activity at the respiratory tract, whereas olive
oil vaccine-adjuvant ratio of 2:1 induced Ab activity of the same magnitude.
It seems that the booster dose was necessary to ensure a prolonged and
strong local immune response for P. multocida broth bactrin. Since the
effectiveness of this broth bactrin vaccine was expected to be much lower
than live vaccine, mucosal adjuvants as potent delivery systems for antigens
were incorporated to enhance the uptake of the vaccine-antigen and
ameliorate the induction of local Ab response at the respiratory tract as
measured by IHA and GDPT assay.
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Keywords
Intranasal,Immunization, Mucosal,Adjuvants,Pasteurella, multocida,Rats.