Studies on ecology and lipolytic activity of Corynebacterium bovis

The ecology and lipolytic activity of C. bovis was studied to evaluate its physiological role in bovine mastitis and its affect upon milk quality. Ecological studies revealed that C. bovis was present in 47.3 percent of 1447 quarter samples from the University of Missouri dairy herd. California Mastitis Tests indicated that the degree of inflamation with quarters infected with C. bovis was more than twice as high as the inflamation from quarters which contained no microorganisms. However, the degree of inflamation exhibited by C. bovis infections was 28 percent less than that produced by hemolytic microorganisms. An unsuccessful effort was made to improve C. bovis isolation by development of a selective medium. Several antibiotics, potassium tellurite, and sodium azide inhibited C. bovis and were unsuitable for use in a selective medium. Oleic acid and sodium oleate inhibited growth of C. bovis; yet, Tween 80 stimulated growth. Microbiological analyses of samples taken from various external and internal locations of the udder strongly suggested that C. bovis was located primarily in the upper portion of the teat canal. Lipolytic studies revealed that C. bovis possessed a very versatile lipolytic enzyme system. Esters of fatty acids and triglycerides were hydrolyzed. Lipolytic activity was associated primarily with the cells. Attempts to concentrate and isolate the enzyme by conventional methods were not successful. Isolation was hampered by relatively low activity and the requirement for long incubation periods to obtain measurable activity. Studies utilizing washed cells as the enzyme source, revealed that C. bovis hydrolyzed -naphthyl esters of fatty acids and triglycerides. Washed cells hydrolyzed -naphthyl laurate g and -naphthyl myristate. A cell concentration of 1 x 10 per milliliter exhibited lipolytic activity comparable to that of normal human blood serum. A study of the effects of added salts revealed that the addition of magnesium increased the hydrolysis of -naphthyl laurate while added calcium decreased hydrolytic activity towards -naphthyl myristate. The optimum pH for the hydrolysis of -naphthyl laurate and -naphthyl myristate ranged from 7.7 to 8.6. Triglyceride hydrolysis by washed cells was detected by incubating the cells with the substrate and titrating the liberated fatty acids. Several triglycerides. Tween 80, and milk fat were hydrolyzed by C. bovis. Hydrolysis of -naphthyl esters and the non-specific activity towards triglycerides, suggested the possibility of two enzyme systems or more likely, one enzyme with a broad spectrum of activity. The slow rate of hydrolytic activity exhibited by C. bovis, especially at lower temperatures, indicated that this microorganism would not effect the quality of milk.