ABSTRACT Sixty-eight presumptive Xanthomonas translucens strains isolated from 15 small grains or grass species were studied by pathogenicity tests on barley, bread wheat, oat, and bromegrass species, and also by AFLP, analysis of fatty acid methyl esters (FAME), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of protein extracts. The X. translucens strains were divided into three pathogenicity types based on differences observed on barley and bread wheat. Two unspeciated strains producing atypical symptoms formed a fourth pathogenicity type. Pathogenicity on oat and bromegrass species varied within these types. Clusterings observed by AFLP analysis and, to a lesser extent, by FAME analysis were consistent with these pathogenicity groupings. The current results, as well as those of previous restriction fragment length polymorphism analyses of the same strains, support the recent reclassification of X. translucens pv. translucens and X. translucens pv. hordei as true synonyms. X. translucens pv. cerealis, X. translucens pv. translucens, and X. translucens pv. undulosa cluster in different groups by AFLP and FAME analyses. Even though distinction by simple biochemical tests is not clear-cut, the data indicate that the pathovars cerealis, translucens, and undulosa correspond to true biological entities.