TWENTY-FIFTH ANNUAL EASTERN FISH HEALTH WORKSHOP

MARCH 10-13, 2000

Analysis Of Chondroitin AC Lyase Activity And Virulence Of Flavobacterium columnare

Kelly Stringer, Laurie Caslake, and Wayne Yunghans

Biology Department, State University of New York at Fredonia, 14063

Flavobacterium columnare (formally Flexibacter columnaris) is a Gram negative, rod-shaped, aquatic bacterium that causes columnaris disease in warm- and cold-water species of fish. The bacterium is a detrimental pathogen, particularly in game fish raised in hatcheries. Flavobacterium columnare strains are classified as either high or low virulence and are distinguished by the temperature at which each is capable of causing infection. Strains with high virulence can cause infection at lower water temperatures than strains with low virulence. The bacterium produces an extracellular enzyme, chondroitin AC lyase, which degrades polysaccharides in cartilaginous tissue. Virulence differences of strains may be related to chondroitinase activity. We grew warm-water and cold-water isolates in liquid culture to assess changes in culture absorbance, cell culture protein, supernatant protein, and cell numbers over a growth period of 10 days. Absorbance of the cultures and amount of supernatant protein present correlated to the number of cells present. The results verified the need to be cognizant of the variable growth patterns of individual strains while analyzing cell culture supernatants for chondroitinase activity. To assess and compare the ability of ten isolates of F. columnare to degrade chondroitin sulfate A, a technique was adapted that used acidified albumin and chondroitin sulfate A to produce turbidity. Cold-water isolates degraded more chondroitin sulfate A than warm-water isolates.