Journal of General Microbiology (1992), 138, 2283-2286. Printed in Great Britain 2283 Taxonomic studies of the genera Acidomonas, Acetobacter and Gluconobacter by 5s ribosomal RNA sequencing EUGENIA s. BULYGINA,’ *f’ OLGA M. GULIKOVA,2 ELLA M. DIKANSKAYA,3 ALEXANDER I. NETRUSOV,~ TATYANA P. TOUR OVA^ and KONSTANTIN M. CHUMAKOV~ $ Institute of Microbiology, Russian Academy of Sciences, Prop. 60-let Oktyabrya 7, I1 781 I Moscow, Russia Department of Microbiology, Moscow State University, 119899 Moscow, Russia Institute for Protein Biosynthesis, 109004 Moscow, Russia (Received 14 May 1992; revised 28 July 1992; accepted 31 July 1992) Ribosomal SSRNA (rRNA) was isolated from 12 strains belonging to the genera Acidomonas, Acetobacter and Gluconobacter and sequenced. A dendrogram constructed from the data indicated that methylotrophic and non- methylotrophic strains of the genus Acetobacter formed two separate clusters. The non-methylotrophicmembersof the genus Acetobacter were phylogenetically closer to Gluconobacter than to the methylotrophic strains of Acetobacter. The methylotrophic strains of Acetobacter were recovered as a clade with the type strain of Acidomonas methanolica. These data support an earlier proposal which reclassified methylotrophic strains of Acetobacter into the genus Acidomonas. Introduction The genus Acetobacter is composed of aerobic, Gram- negative, non-sporeforming, acidophilic rod-shaped bac- teria, which utilize a wide variety of organic compounds (De Ley et al., 1984). Some strains of this genus are able to use one-carbon compounds. The validity of placing methylotrophic and non-methylotrophic species in a single genus has been questioned for several genera (Green & Bousfield, 1982; Doronina & Govorukhina, 1987). For example, pink-pigmented facultative methy- lotrophic bacteria, previously assigned to the genus Pseudomonas have been given a separate generic status mainly as a result of their ability to grow on methanol and methylamines (Patt et al., 1976; Hood et al., 1987). The same feature was used for distinguishing the genera Hyphomicrobium and Hyphomonas (Gebers et al., 1986). * Author for correspondence. Tel. (612)4717741; fax (612) 471 9070; email gfO1020@vx.acs.umn.edu. t Present address : Gray Freshwater Biological Institute, University of Minnesota, PO Box 100, County Roads 15 & 19, Navarre, MN 55392, USA. 1 Present address: FDA Center for Biologics Evaluation and Research, 55 16 Nicholson Lane, Bldg. 2-B, Kensington, MD 20895, USA. The nucleotide sequence data reported in this paper have been submitted to GenBank and have been assigned the accession numbers M76569 to M76580. Recently, Urakami et al. (1989) proposed that the methylotrophic species of Acetobacter should be trans- ferred to a new genus Acidomonas on the basis of chemo- and genotaxonomic data. To address this question, we have performed a phylogenetic study of the genus Acidomonas and related genera by comparing 5s rRNA sequences. Met hods Strains and growth conditions. The strains used in this study and their sources are listed in Table 1. Methylotrophic strains of Acetobacter and Acidomonas methanolica were grown at 30°C, with shaking, in a mineral medium (pH4.0; Loginova et al., 1981) containing 0.5% methanol. Non-methylotrophic strains of Acetobacter and Glucono- bucter were grown at 30 “C in a medium containing 0.3% peptone, 0.5% yeast extract and 1.5% (w/v) glucose; this medium was adjusted to pH 4-5 with HCl. Isolation and sequencing of 5s rRNA, 5s rRNAs were isolated and their nucleotide sequences were determined as described previously (Bulygina et al., 1990). [5’-32P] cytidine 3’,5’-diphosphate (pCp, Isotope, USSR) and T, phage RNA-ligase (Ferment, USSR) were used for 3’-end-labelling. Phylogenetic analysis. The 5s rRNA nucleotide sequences were aligned as previously described (Wolters & Erdmann, 1988)and used to calculate the mutation distance matrix (Md). The distance between two sequences was expressed as a proportion of differing positions. A dendrogram was constructed using a UPGMA method (Sneath & Sokal, 1973). A tentative unrooted phylogenetic tree was constructed using the ‘maximum topological simiiarity’ (MTS) method (Chumakov 0001-7581 0 1992 SGM