Forty isolates of Yersinia ruckeri from Italy (4), Turkey (24), the USA (10), and two type strains were compared by using biochemical test, pulsed-field gel electrophoresis (PFGE), and lipopolysaccharide (LPS) and outer membrane protein (OMP) profiles. Two biotype 1 and biotype 2 Y. ruckeri were determined based on motility and phospholipase activity. Twenty-two different pulse types were observed by cutting the DNA with SpeI restriction enzyme and running on PFGE. Four major clusters were generated using the UPGMA technique: cluster A contained Italian, Turkish and USA strains; cluster B included all the USA strains; and cluster C and cluster D contained all the Turkish strains. Based on the OMP profile, Y. ruckeri strains were divided into 4 clusters with 30 OMP types. A total of 30 different LPS types were observed. All the Italian, the USA, and most of the Turkish isolates were grouped together to form cluster A that consisted of 26 LPS types. It seems that all the four typing methods are highly discriminatory to distinguish even closely related strains. The overall similarities among the strains were 32.4 +/- 6.1%, 58.7 +/- 11.1%, and 79.5 +/- 3.9% in LPS, PFGE, and OMP profiles, respectively. The PFGE, biochemical, OMP, and LPS profiles of none of the strains were found to be similar. Hence, each typing method showed its own discriminatory characteristics to distinguish between the individual strains. It is apparent that there is sufficient genetic diversity to justify using PFGE for analysis of horizontal transmission of Y. ruckeri in trout rearing facilities. It is possible that diverse environmental conditions resulted in a relatively high degree of genetic diversity in Y. ruckeri.