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Title: The role of humoral and cellular defences of the pig against Actinobacillus pleuropneumoniae
Author: Cullen, Janice Mitchell
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 1993
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Abstract:
The main objective of this research was to assess the defensive roles of both the humoral and cellular defences of the pig against Actinobacillus pleuropneumoniae (A, pleuropneumoniae) in vitro. The aims were firstly to establish the effects of complement on the survival of A. pleuropneumoniae and secondly, to study the ability of alveolar macrophages to phagocytose A. pleuropneumoniae. In Chapter 3 the effect of complement on the survival of A. pleuropneumoniae was examined. All strains of A, pleuropneumoniae tested were found to be resistant to pig complement. The strains were also found to be resistant to both immune pig serum and hyperimmune rabbit serum containing specific antibodies against A, pleuropneumoniae. Three of the four strains were found however to have a delayed sensitivity to normal human serum. The mode of complement resistance in A, pleuropneumoniae was compared to a known serum resistant Escherichia coli strain using the outer membrane disorganising chemical polymyxin B. All A, pleuropneumoniae strains tested could not be sensitised to complement bactericidal activity by this method. This was in contrast to the serum resistant E, coli which was sensitised to the effects of complement following treatment with polymyxin B. In Chapter 4 the method by which A, pleuropneumoniae avoided complement-mediated damage was investigated. A, pleuropneumoniae was found to consume large amounts of complement activity via the classical and alternative pathways from both pig and human serum. Activation of the complement component C3 also occurred in both pig and human serum. Activation of factor B, which occurs via the alternative pathway only, was also demonstrated in human serum during incubation with the bacteria. In Chapter 5 non-viable A. pleuropneumoniae were found not to consume complement activity suggesting the ability of A. pleuropneumoniae to consume complement was dependant on bacterial viability. The extracellular secreted toxins of A. pleuropneumoniae were found not to affect consumption of complement activity and therefore some other function of viable A. pleuropneumoniae appeared to be responsible. In Chapter 6 the ability of pig alveolar macrophages to phagocytose A. pleuropneumoniae was investigated. Both haemolytic and cytotoxic activities are known to be associated with 109 (haemolysin II, Hly II) and 120 (pleurotoxin, Ptx) kDa proteins respectively. Both HK 361 (Hly II+, Ptx+) and mutant e (Hly 11-, Ptx+) were found to rapidly destroy all macrophages when incubated with normal pig serum and any phagocytosis could not be determined. Mutant h (Hly II-, Ptx-) was found initially not to cause damage to alveolar macrophages. However, after prolonged incubation, a level of damage was observed, although not to the same extent as seen with HK 361 and mutant e. A few phagocytosed mutant h were observed initially, however the numbers did not increase with time. No reduction of toxicity was seen for macrophages incubated with HK 361 and mutant e in the presence of immune serum. The reduced toxicity of mutant h for alveolar macrophages was diminished following the addition of immune pig serum and the numbers of phagocytosed mutant h greatly increased. The nature of the reduced toxicity of mutant h was compared to killed mutant h. No toxicity of killed mutant h (by heat or formaldehyde) was observed in the presence of normal pig serum, and similar levels of phagocytosed bacteria were seen. This suggested the toxicity associated with mutant h was both heat labile and neutralised by immune pig serum. In Chapter 7 the immunological relationships of the haemolytic and cytotoxic proteins (109 and 120 kDa respectively) of A, pleuropneumoniae serotype 2 strain HK 361 were investigated. Monoclonal antibodies (Mabs) were raised against both the 109 or 120 kDa protein secreted from HK 361. Mabs raised against each protein cross-reacted with other proteins on Western blots suggesting they were immunologically related. Cross-reactive proteins were found in all the serotype reference strains of A. pleuropneumoniae, including some of the related pathogens. The presence of similar proteins in all serotypes of A. pleuropneumoniae suggested a similar virulence factor of A, pleuropneumoniae* The crossreaction seen with related pathogens was in agreement with others who suggested the similarity of the A, pleuropneumoniae toxins to the RTX (repeat in structural toxin) toxin-producing family.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.796860  DOI: Not available
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