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Title: Studies on immunity to Pasteurella haemolytica
Author: Sutherland, A. D.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1990
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This thesis investigated the interaction of humoral mechanisms of immunity in the sheep with antigens of Pasteurella haemolytica. An assay was developed to detect toxic activity in crude preparations of P. haemolytica cytotoxin (CC). A cytotoxin neutralisation (CN) assay showed that a serotype A2 CC vaccine could raise CN antibodies in a rabbit and that immune sera and lung washings from lambs 'convalescent' from serotype A2 infection had CN titres. Thus CN titres correlated with protection in lambs, indicating A2 CC had potential as a vaccine component. Analysis of the CC showed it to contain outer membrane proteins, lipopolysaccharide (LPS), serotype specific capsular antigens and proteases as well as cytotoxin antigens. Microtitre plate assays were developed to detect antibodies involved in bactericidal activity and opsonophagocytosis. Convalescent lamb sera and lung washings, in combination with complement, were bactericidal for P. haemolytica A2 organisms, and adsorption studies indicated LPS to be a target for antibodies involved in bactericidal activity. In contrast, P. haemolytica serotype T10 organisms were resistant to bactericidal activity, this resistance being associated with the 'smooth' type LPS found in all T serotype organisms. All A serotype organisms had 'rough' type LPS, except for serotypes A5 and all which were intermediate in LPS profile. Both complement and antibodies in convalescent serum opsonised serotype A2 organisms causing their increased uptake of phagocytes. Adsorption studies were not conclusive, but indicated that LPS was not a target for opsonic antibodies, whereas opsonins were removed by adsorption with a sodium salicylate extract (SSE) of serotype A2 cells which contained capsular polysaccharide antigens and outer membrane proteins as well as LPS. Serotype A2 organisms grown in vivo were susceptible to bactericidal and opsonophagocytic mechanisms of immunity. Also, passive protection studies in lambs indicated that sera, which variously stimulated CN, bactericidal and opsonophagocytic mechanisms, were highly protective. These combined findings suggested that humoral mechanisms of immunity were active against organisms grown both in vivo and in vitro, and that humoral immunity alone was highly protective against serotype A2 infection. As a vaccine, the serotype A2 CC gave 86% protection against experimental homologous challenge. Protection correlated (P < 0.001) with serum CN titres and bactericidal capacity, but opsonophagocytosis was not stimulated. CN and bactericidal activity appeared to be stimulated by cytotoxin and LPS antigens in the CC. This study, therefore, identified in vitro correlates of immunity to P. haemolytica infection in lambs, and demonstrated the ability of a cytotoxin based vaccine to stimulate some of these mechanisms of immunity and engender a high degree of protection against infection.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available