No abstract

The cytotoxic action of leukocidin from Pseudomonas aeruginosa was studied in vitro by following the release of various intracellular markers from polymorphonuclear leukocytes from cattle (PMLC). Low-molecular markers (K+, 86Rb+, glucose) were lost from PMLC within 1 to 2 min after the addition of leukocytes. The release of high-molecular-weight indicators (51Cr, bound to intracellular protein; lactate dehydrogenase) occurred only after swelling of the cells, leading to an increased permeability of the plasma membrane. Calcium ions stimulated the leakage of granule enzymes but retarded or inhibited the release of cytoplasmic markers. At 4 C, leukocytes were unaffected by the toxin. Leukocidin, bound at 4 C to leukocytes and treated with antiserum against leukocidin, did not damage the cells upon increasing the incubation temperature to 37 C.

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S U M M A R YLeucocidin from Pseudomonas aeruginosa strain I 58 was released from bacteria by autolysis and purified Ig-fold by ammonium sulphate precipitation (20 % saturation) and combined 'tandem' gel filtration on Sephadex G-IOO superfine and Bio Gel P-100. The product gave a single band (mol. wt. 27000) after polyacrylamide gel electrophoresis with sodium dodecyl sulphate (SDS). However, it was separated into two active peaks (PI 5.0 and 5.2) by isoelectric focusing, and into five bands by disc electrophoresis without SDS. All bands contained leucocidic activity of about the same specific activity and retained their homogeneity.The purified toxin was thermolabile and was inactivated by pronase, but not by several other proteases. The ultraviolet light absorbancy was typical of proteins. Antibodies directed against leucocidin were detected by passive haemagglutination and by toxin-neutralization. These antibodies inhibited the cytotoxic action of leucocidin bound to granuloyctes. The toxin damaged all tested leucocytes (granulocytes of various animal species and lymphocytes of humans) and a number of tissue cultures, but was ineffective against erythrocytes, thrombocytes and isolated granules from polymorphonuclear leucocytes. The intravenous lethal dose for mice was about I pg.

SummarySurvival has been determined for Pa ste ure lla pne um o tro pic a on various surfaces found in an anim al room at 23 1 C and 50 10% relati ve humidity. Longest survival (120 min) was found on mouse hair, shortest ( < 30 min) on laborat ory coat fabric . Transmission experiments were performed using sentinel animals in order to evaluate the efficiency of their use for the detection of P. pne um o tro pi ca in quaran tined mice. In sentinels exposed to infected mice by close contact, P. pne um o tro pic a was detected by culture 2 weeks post-exposure and seroconversion 3 weeks after contact . Transfer of soiled bedding from Pa ste ure lla -infected mice did not infect sentinels within a period of 12 weeks as tested by cultivation or serum antibodies.