Technical Abstract: Flavobacterium columnare is a Gram-negative pathogen of many species of wild and cultured fish, especially channel catfish (Ictalurus punctatus). This motile microbe is responsible for severe economic losses to the catfish industry. Flavobacterium columnare isolates from diseased channel catfish belong to either genomovar I or II. Genomovar II isolates were found to be more virulent than genomovar I isolates. The objective of the present study determine if differences exist in the chemotactic response of these genomovars to mucus obtained from the skin, gills and intestines of healthy channel catfish using the capillary chemotaxis assay. To accomplish this objective, F. columnare motility without mucus was determined. Next, a genomovar II isolate was used to determine the chemotactic response to mucus from skin, gill and intestine. The same genomovar was used to determine the chemotactic response to skin mucus from 15 individual healthy catfish. Mucus from skin and gill induced a greater chemotactic response by F. columnare than mucus from the intestine. Sixty percent of mucus from the skin of individual catfish yielded a positive chemotactic response from F. columnare. Finally, skin mucus induced a greater chemotactic response by genomovar II F. columnare than for genomovar I F. columnare isolates. The data indicates that mucus from channel catfish results in a chemotactic response by F. columnare. This positive chemotactic response may be an important first step for F. columnare colonization of channel catfish skin or gills. Although the role that chemotaxis plays in virulence of F. columnare is not fully defined, the chemotactic response of genomovar ll isolates suggest that chemotaxis is associated with virulence.