ISOLATION AND PARTIAL CHARACTERIZATION OF EXTRACELLULAR PROTEASES PRODUCED BY ISOLATES OF FLAVOBACTERIUM COLUMNARE DERIVED FROM CHANNEL CATFISH

Proteases of 23 isolates of Flavobacterium columnare derived primarily from channel catfish Ictalurus punctatus raised in the southeastern U nited States were isolated and partially characterized. The bacterial isolates were divided into two groups according to the apparent molecu lar masses of proteases after zymographic resolution by nonreducing, n ondenaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with gelatin as the protease substrate. The 15 isolates in group 1 had two proteases with apparent molecular masses of 58 and 53 .5 kilodaltons (kDa). Eight group-2 isolates produced three proteases with apparent molecular masses of 59.5, 48, and 44.5 kDa. Culture medi um had an effect on the amount of protease produced by F. columnare LA 88-173. More protease was produced in a medium with low nutrients and salt (Ordal's medium) than in media with higher concentrations of nut rients or salts (TYES, Hsu-Shotts, modified Shieh's media). No differe nces were observed in the apparent molecular masses of the two proteas es of F. columnare LA 88-173 produced in the various media or with dif ferent incubation times. Two proteases with apparent molecular masses of 58 and 53.5 kDa were seen as early as 1 d after inoculation, and th ese molecular masses did no: change during the 7-d experiment. A sharp increase in protease production occurred during the first 24 h of inc ubation with minimal increase during the remaining 7 d of the experime nt. All 23 isolates of F. columnare degraded the gelatin and casein in corporated into TYES agar medium but only 7 of the 23 isolates degrade d elastin.