Doctor of Philosophy
Department of Biology
Dai, Yung, Potential role of pseudomonas aeruginosa in sheep fleece rot and the associated serological response, Doctor of Philosophy thesis, Department of Biology, University of Wollongong, 1997. http://ro.uow.edu.au/theses/1073
Fleece rot is a bacterial infection of the fleece and skin predominantly caused by the bacterium Pseudomonas aeruginosa (P. aeruginosa). The bacterial infection can attract the blowfly, Lucilia cuprina (L. cuprina), which lays its eggs at the fleece rot site. The hatching larvae cause a problem known as fly strike or body strike, a debilitating disease which can cause sudden death in affected sheep.
Nine bacterial species have been isolated and identified in the fleece and on the skin of sheep. They remain unimportant until there is an environmental change such as rain and heat. Wetting by rain results in the proliferation of all these bacterial species in the first 24 hours but after this time P. aeruginosa predominates in the fleece. The inhibitory activity of P. aeruginosa on other bacteria was shown by co-culture of each bacterial species with a P. aeruginosa lawn. Pyocyanin, which is produced by P. aeruginosa, inhibited the growth of other fleece bacteria in vitro. Partial purification of pyocyanin from P. aeruginosa culture supernatants revealed that this inhibitory activity was most effective against gram positive bacteria such as Bacillus cereus, Bacillus coagulans and Staphylococcus epidermidis.
Apart from pyocyanin, P. aeruginosa produces many extracellular enzymes which may facilitate the invasion of fleece rot. Culture supernatant from the growth media of P. aeruginosa contained two proteolytic enzymes. Protease I had a molecular mass of approximately 50 kDa and contains subunits of 25 - 30 kDa. Protease II had a molecular mass of 150 kDa and contains a 50 kDa subunit. Protease I was similar to elastase and protease II appeared to be an alkaline proteinase as previously described in the literature. The production of these proteases from P. aeruginosa culture was not only affected by basic media ingredients but also by the amount of sheep wool added to the cultural media. Skin biopsies indicated that proteases could cause an acute inflammatory reaction, as assessed by the appearance of haemorrhages and neutrophil infiltration with a mild hyperaemia and oedema after only 2 hours. Skin thickness was found to increase within 4 hours of exposure to these proteases when compared with the controls. This pathological change occurred only when a mixture of both proteases was present but not with either protease alone.
Studies on the sheep serological response to surface membrane proteins of P. aeruginosa showed that the outer membrane proteins were more antigenic to sheep when they were subjected to intradermal injection. Evidence is also presented to suggest that the so-called genetically fleece rot "resistant" and "susceptible" sheep have a qualitative and quantitative difference in their serological responses.
Experiments were conducted to introduce P. aeruginosa infection with and without L. cuprina infestation on the skin of sheep. In all sheep, the serological specificity was higher against the outer membrane proteins purified from P. aeruginosa than the inner membrane proteins or the whole cell proteins regardless of the presence or absence of fly strike. These data demonstrated differences in sero-reactivity to bacterial and larval antigen and there was no correlation between antibody levels and intensity of fly strike.
The results in this research contribute further to the understanding of the potential role of P. aeruginosa in sheep fleece rot and could assist in the development of strategies and recommendations for the control of fleece rot which in turn will minimise the development of fly strike.