Pyocines and the treatment of pseudomonas aueruginosa infections
Williams, Rosamund J.
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The aim of this investigation was to study the interaction of different kinds of pyocine with a single sensitive indicator strain of Pseudomonas aeruginosa, and to examine the possible therapeutic applications of these pyocines to infections caused by the indicator strain. After surveying the literature concerning Is. aeruginosa. with particular emphasis on the pathogenicity of the organism, the past and present therapy of Ps. aeruginosa infections was discussed. The literature on the pyooines was reviewed and it wa3 clear that two train groups of pyocines were distinguishable; the high and low molecular weight groups, and within the high molecular weight pyooines, two structural types were defined; contraotile and filamentous. Members of the low molecular weight group were referred to as small pyocines. Ninety-four strains of Ps. aeruginosa were examined for pyocine production against two selected indicator strains, Ps. aeruginosa 6a and P14. Thirteen strains which produced pyocines with properties characteristic of each of the three types of pyocine were selected and examined in greater detail. The inducibility of the strains was examined and the effects of heat, trypsin and ultracentrifugation on the extraoted pyocinea were investigated. Eleotronmicroscopy was used to oonfirm the classifioation of the pyocines. 6. Three pyocinogenic strains were selected for further investigation! one (5893) which produced contractile pyocines, another (5882) which produced filamentous pyocines, and a third (H108) whioh produced small pyocines. These pyocinogenic strains were defined with respect to their pyocine activity against indioator strain P14 only and this indicator strain alone was used throughout the remainder of the study. The pyooines were numbered according to their producer strain, e.g. pyocine 5893 was the pyocine produced by strain 5893 and inhibitory to indicator strain P14. 7. Various methods for purifying and concentrating the pyocine preparations were examined and precipitation of the proteins with ammonium sulphate proved the most satisfactory. 8. Mouse toxicity testing of pyocine preparations at various degrees of purity revealed that in some forma, certain pyocine preparations caused adverse effects. Uninduced, low activity preparations of the pyocine were more lethal, suggesting that factors other than pyocine were responsible for toxicity, but examination of fractions of an induced preparation in vivo favoured the pyocine itself as the toxic agent. In view of the toxicity problems, another pyocinogenic strain (1577)# which produced contractile pyocines inhibitory to indicator strain HL4, replaced the tacie pyocine (5893). 10, An extended pyocine purification scheme was devised that included ion-exchange chromatography. The method gave satisfactory results with the high molecular weight pyocines but the small pyocine proved more problematic. 11, The interactions between the pyooines and cultures of strain P14 were examined in vitro and the effects of dosage and incubation time on the survival of P14 cells were studied. 12, The pyocines were examined in mic^ alone, and in the presence of infections caused by strain P14. When the pyocines were injected into mice by various routes, inhibitory activity against strain P14 could be identified in tie serum for several hours. 13, High molecular weight pyooines given intraperitoneally in the presence of a lethal dose of strain P14 administered by the same route, were unable to prevent the fatal outcone of the infection unless they were given before or simultaneously with the bacteria. The 3mall pyocine preparation had no protective effect. 14, Infection of experimentally-induced burns with strain P14 was examined; although colonisation of the burn was easily established, lethal infections were more difficult to achieve. The effects of topical application of the high moleoular weight pyocines on infected burns were examined and it was eonoluded that the pyocines did not improve the ohanoes of survival of burned, infected aioe, 15, Despite the problems of pyocine resistance of strain P14 recognised in vitro, this did not appear to be the cause of failure of pyocine therapy in vivo. 16, A single, intraperitoneal dose of pyocine was sufficient to cause the production of pyocine-neutralising antibody, but topically-applied pyooine did not elicit an antibody response. 17, The future of pyocine therapy was discussed.