Pasteurella multocida possesses a characteristically gram-negative ultrastructure, yet its inability to grow in the presence of hydrophobic compounds and the general penicillin susceptibility of genera making up the family Pasteurellaceae suggest a cell envelope having atypical permeability properties. The cell surface hydrophobicity properties of strains representing 15 of the 16 somatic serotypes of P. multocida and three strains of Actinobacillus lignieresii were assessed with hydrocarbon adherence and hydrophobic interaction chromatographic assays. These methods revealed surface hydrophobicity to vary dramatically among strains in both species. No direct correlation was observed with species, growth rate, or susceptibility to the antibiotics oxytetracycline (polar), polymyxin B (amphiphilic), or novobiocin (nonpolar) as measured with MIC determinations. All strains were susceptible to the antibiotics, although A. lignieresii was significantly less susceptible than P. multocida to novobiocin. These data suggest that cell surface hydrophobicity in P. multocida may be influenced by the type of lipopolysaccharide present but is not directly related to permeability of the antibiotics examined. The wide diversity of hydrophobic properties exhibited by strains of both P. multocida and A. lignieresii precludes the use of this parameter as a taxonomic aid.