Deoxyribonucleic acid (DNA) base composition, intergenic transformation efficiency, and DNA hybridization were used to determine the relatedness of a variety of established or proposed species of Neisseria and Branhamella. These studies indicated that these bacteria form three genetic groupings. Group I, comprised of N . meningitidis, N . gonorrhoeae, N . subflava, N . flava, N . perflava, N . sicca, N . mucosa, N . cinerea, N . flavescens, N . lactamica, N . elongata, N . canis, and N . denitrificans, was characterized by DNA base compositions ranging between 49.3 and 55.6 mol% guanine plus cytosine. Group 11, comprised of N . cuniculi, N . caviae, and N . ovis, was characterized by DNA base compositions ranging between 45.3 and 47.3 mol% guanine plus cytosine. Group 111, comprised of one species, B . catarrhalis, was characterized by DNA base compositions between 41 and 42 mol% guanine plus cytosine. Transformation and DNA hybridization results revealed that members of each group, with few exceptions, exhibited high DNA homology with other members of the same group but most often distinctly lower levels of homology with members of a different group. These data suggest that N . ovis, N . caviae, and N . cuniculi may be significantly different from other neisseriae and from branhamellae to warrant their separation in a distinct genus. The results of nucleic acid hybridizations performed by Kingsbury (17) revealed that the genus Neisseria was a heterogeneous group comprised of at least three distinct subgroups and questioned the proper classification of N . catarrhalis and N . caviae as members of the genus Neisseria. On the basis of nucleic acid hybridization studies, Bdvre reported that N . ovis, N . caviae, and B. catarrhalis showed distinct degrees of relatedness to several Moraxella species (3, 4). This supported the inclusion of Moraxella in the family Neisseriaceae.The purpose of this study was to use DNA base composition determinations, transformation, and DNA hybridization studies to clarify the taxonomic positions of recognized or proposed species of Neisseria and Branhamella. Special attention was focused on evaluating the potential of transformation as a toxonomic tool for determining the relatedness of members of these genera.
MATERIALS AND METHODSThe bacterial strains examined are listed in Table 1. Stock cultures were preserved by freeze-drying. Working cultures were maintained at -70°C in Trypticase soy broth (BBL Microbiology Systems) supplemented with 6% lactose. Cultures were routinely passaged on GC agar consisting of GC medium base (Difco Laboratories) and 1 % (vol/vol) chemically defined supplements (36) and were incubated at 36°C for 18 h (5% C02, humidity).Preparation of DNA for base composition determinations. A modification of the procedure originally described by Marmur (25) was followed for the extrac-
57
