Effect of Vaccine, Route, and Schedule on Antibody Response of Rabbits to Pasteurella tularensis

John E. Nutter
1969 Applied microbiology  
The response of the rabbit to viable or killed whole-cell Pasteurella tularensis vaccines was studied. The most practical preparation for the production of anti-P. tularensis antibodies was viable organisms of the live vaccine strain (LVS). The intravenous route of administration proved superior to either the subcutaneous or intradermal routes, and incorporation of LVS into Freund's adjuvants did not result in increased levels of antibody. Short-term hyperimmunization, three injections at
more » ... injections at weekly intervals, constituted the most efficient method for increasing levels of the antibodies. The rabbit is commonly used for the production of antisera against a wide variety of antigens (3). There is, however, little information concerning the antibody response of this animal to Pasteurella tularensis (2, 4, 6). The objectives of this study were to determine optimal procedures for antibody production and to study the response of the rabbit to viable or killed whole-cell P. tularensis vaccines. MATERIALS AND METHODS Rabbits. New Zealand white rabbits, weighing between 1.8 and 2.5 kg, were used throughout the investigation. Except where noted, all experimental groups contained five animals. Vaccines and vaccination techniques. P. tularensis was grown in a modified casein hydrolysate medium (MCPH) similar to that described by Mills et al. (Bacteriol. Proc., p. 37, 1949). Numbers of viable cells were estimated by plating appropriate dilutions on glucose-cysteine-blood-agar (6). The vaccine strain (LVS) described by Eigelsbach and Downs (5) was used to produce viable vaccine. The usual preparation was a saline dilution of an MCPH culture containing 108 or 109 viable organisms per ml. In one experiment, equal portions of LVS in saline were mixed with either Freund's complete or incomplete adjuvant (BBL) to give a final concentration of 109 viable organisms per ml. Strain SCHU S4, a fully virulent North American strain, was employed as a killed (phenol-Merthiolate) or viable vaccine; the killed preparation contained 109 organisms per ml. To infect rabbits intravenously (iv), 1,000 viable cells were used; after clinical symptoms appeared (3 to 4 days), the animals were treated with streptomycin (6). Serological techniques. Trial bleedings were made from the marginal ear vein, and animals were exsanguinated by cardiac puncture. Three serological tests were performed on sera. Bacterial agglutination tests were performed with formalinized SCHU S4 cells as antigen (1). Passive hemagglutination tests were done with polysaccharide-treated sheep erythrocytes (9). Precipitins were assayed by double diffusion in agar by use of the technique and strain SCHU S4 antigen (sonic lysate) described by Tulis and Eigelsbach (Bacteriol. Proc., p. 138, 1961). Precipitates were allowed to develop for 21 days at 37 C in a moist atmosphere. RESULTS Primary and secondary antibody responses to three vaccines. The agglutinin responses of three groups of 10 rabbits, each inoculated iv with 109 killed strain SCHU S4, 109 viable strain LVS, or 103 viable strain SCHU S4, respectively, are presented in Table 1 . During the primary response the highest peak titer was obtained with viable strain SCHU S4; killed strain SCHU S4 induced the poorest response. The peak titers were compared by use of Student's t test, and the difference between titers produced by viable SCHU S4 and killed strain SCHU S4 was the only significant one (P = 0.05). Rabbits receiving viable strain SCHU S4 maintained significantly higher titers on the 21st and 28th days than those given LVS; at these times, animals inoculated with killed SCHU S4 exhibited the lowest titers. Twenty-eight days after vaccination, the killed SCHU S4 and LVS groups were revaccinated with 109 cells of the appropriate vaccine by the iv route. The animals in the viable SCHU S4 group were not revaccinated, because only three animals survived the infection despite streptomycin therapy. Both revaccinated groups of rabbits had elevated 355 on December 10, 2020 by guest http://aem.asm.org/ Downloaded from
doi:10.1128/am.17.3.355-359.1969 fatcat:ktjnuyzbona7bfztfjetbjzkji