Hemolytic Activity of Leptospira interrogans Serovar canicola Cultured in Protein-Free Medium

Hemolytic activity of the culture supernatant of Leptospira interrogans serovar canicola strain Moulton grown in protein-free medium was demonstrated. The activity began to appear in the late logarithmic phase of growth of the organism and reached a plateau after 2 weeks of cultivation. It was inactivated by the addition of dipotassium ethylenediaminetetraacetate, but was effectively restored by Mg²⁺ . Hemolysis by the culture supernatant was stimulated by "hot-cold" incubation. Sheep erythrocytes treated with the culture supernatant of the organism were transformed into spherocytes, in which invagination was observed. A hemolysin inhibitor in rabbit serum was found to be in the chloroform-methanol soluble fraction of the serum. The hemolysin of Leptospira may be phospholipase C.