The phenotypic characteristics as well as the prevalence and quantity (colony forming units/ml/sample) of 39 bacterial isolates of agar pitting Gram-negative rods, from two diseased and one healthy subgingival site from each of 16 pet dogs with naturally occurring periodontitis, were investigated. Phenotypic features were determined with use of standard biochemical methods, by enzymatic profiling with the API ZYM system, and by cellular fatty acid profiling. The organisms detected were motile, catalase-negative Campylobacter sp., present in 69% of the dogs and in 29% of the subgingival samples (69/29), motile, catalase-positive Campylobacter sp. (63/29), Eikenella corrodens (25/10), organisms closely resembling E. corrodens but nitrate-negative and unable to grow in air, designated E. corrodens-like (19/8), Bilophila wadsworthia (6/2), and non-motile Campylobacter sp. (6/2). The most frequent organisms were the motile Campylobacter isolates constituting 72% of all isolates. No statistically significant differences were detected between the diseased and healthy subgingival sites, with regard to the prevalence of any of these groups of organisms. Furthermore, the bacterial isolates were detected in almost equal numbers in the diseased and healthy sample sites. Hence, no association between dog periodontitis and the agar pitting Gram-negative rods was established. The phenotypic data also suggest, that the organisms that were only presumptively identified in the present study (Campylobacter sp., E. corrodens-like), may not have been previously described. (C) 2000 Academic Press.