Journal of Applied Bacteriology zyxwvutsrq 1996, 80, zyxwvutsrqpo 105-1 13 zyxwvutsr Comparison of extracellular proteases produced by zy Aeromonas salmonicida strains, isolated from various fish species B.K. Gudmundsdottir Institute for Experimental Pathology, University of Iceland, Keldur, Reykjavik, Iceland 5347/05/95 : received 18 May 1995 and accepted 24 July 1995 B.K. GUDMUNDSD~TTIR. 1996. zyxwvutsrq Extracellular products (ECPs) of five typical and 25 atypical zyx Arromonas salmonicida isolates from various fish species and geographical locations were analysed by substrate specificity, inhibition of proteolytic activity and substrate SDS-PAGE. The type strains of Aer. salmonicidu subsp. salmonicida and Aer. salmonicida subsp. achrumugenes were included for comparison. The results indicated that the strains formed six protease groups. T h e proteases produced by the two type strains were of a different nature. All the typical strains belonged to one group and showed proteolytic activities comparable to P1 and P2 proteases. Three atypical (oxidase- negative) strains secreted a protease comparable to P1. With the exception of these three, all strains produced metallo-gelatinases. A metallo-caseinase (AsaP1) was detected in the ECP of subsp. achromugenes type strain and 10 of the atypical strains. A number of proteolytic components with different apparent molecular weights (AMWs) were identified. These include caseinases with AMWs of > 100, 80, 60 and 30 kDa and gelatinolytic components with different AMWs, including some with AMW higher than P1 and lower than P2. The protease production of the isolates was not found to be host specific. INTRODUCTION The species Arromunris salmonirirlii is according to Austin et a/. (1989) and Bergty's zyxwvutsrqpo Mirnuirl of'Determinutice Bacteriology (1994) divided into four subspecies : subsp. snlmonicida, subsp. achromogenes, subsp. tnasouciiiu and subsp. smithia. Some authors have suggested different subgrouping of the species (McCarthy and Roberts 1980; Belland and Trust 1988) and new isolates that do not fit into the existing classi- fication are frequently reported (reviewed in : Austin and Austin 1993 ; Wiklund eta/. 1994; Hirvela-Koski et al. 1994). Aeromonus salmonicido subsp. salmonicida, the aetiological agent of furunculosis of salmonids and often referred to as the typical strain, has been described as a homogenous taxon, with respect to biochemical and genotypic characteristics (reviewed in : Austin and Austin 1993). The other Aer. sal- moniridu subspecies, often termed atypical strains, cause ulcerative diseases in a wide variety of both marine and fresh Correspondrnr.r to : Bjcirnliridiir K Guc/?nurrr/sdij,/ir.. 1~1stIt~4~c~j~r Exprrinrmrcrl fci/holo~ty, Unii~twitl~ n/'lct.lurrr~', Kddur, zyxwvutsrq D/ G>srrirltindswg. IS-112. Rtyk~iitik, Icrluncl. 0 1996 The Society for Applied Bacteriology water fish (reviewed in: Austin and Austin 1993; Nak- atsugawa 1994). Infections due to atypical Aer. salmonicidu strains are of augmented economic importance in aquaculture worldwide as an increasing number of fish species are being cultivated. Furthermore, infections of wild fish stocks have been reported (Wiklund and Bylund 1993). Subsp. sul- monicida has been phenotypically easily distinguishable from the other subspecies, but recently aberrant strains have been described (Wiklund eta/. 1993). Available information on extracellular virulence factors of Aer. sdmonicidu strains indicates a difference between the exotoxin production of typical and atypical strains (Gud- mundsdottir et al. 1990 ; Lee and Ellis 1990). Extracellular proteases are considered to play a major role in the virulence and pathogenicity of the bacterium (Tajima et al. 1983 ; Shieh 1985 ; Lee and Ellis 1989 ; Gudmundsdottir et a/. 1990). Two extracellular proteases (P1 and P2) of Aer. salmonicidn subsp. snlmonicidfi and one isolated from a strain of subsp. rich- romugenes (AsaP1) have been described. The PI protease is a caseinolytic, gelatinolytic and collagenolytic serine protease with a molecular weight of 70 kDa and is described as one of