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Heumann 1962, 343AL, in the Genus International Journal of Systematic Bacteriology (1999), 49, 11 03-1 109 Printed in Great Britain Reclassification of Pseudomonas echinoides NOTE Heumann 1962, 343AL,in the genus I 1 Sphingomonas as Sphingomonas echinoides comb. nov. Ewald B. M. Denner,' Peter Kampfer,2 Hans-Jurgen Busse1n3 and Edward R. B. Moore4 Author for correspondence: Ewald B. M. Denner. Tel: +43 1 4277 54632. Fax: +43 1 4277 12876. e-mail : denner @ gem.univie.ac.at 1 lnstitut fur Mikrobiologie [Pseudomonas]echinoides DSM 1805T(= ATTC 14820T,DSM 5O40gT, ICBP 2835T, und Genetik, Universitdt NClB 94203 has been reinvestigated to clarify its taxonomic position. 16s rDNA Wien, A-1030 Wien, Austria sequence comparisons demonstrated that this species clusters phylogeneticallywith species of the genus Sphingomonas. Investigation of 2 lnstitut fur Angewandte Mikrobiologie, Justus- fatty acid patterns, polar lipid profiles, polyamine patterns and quinone Liebig-UniversitatGiessen, systems supported this delineation. Substrate utilization profiles and D-35390 Giessen, Germany biochemical characteristics displayed no distinct overall similarity to any 3 lnstitut fur Bakteriologie, validly described species of the genus Sphingomonas. Therefore, the Mykologie und Hygiene, reclassificationof [Pseudomonas]echinoides as Sphingomonas echinoides Vete r ind r med iz i n i sc he Universitat, Veterindrplatz comb. nov. is proposed, based upon the estimated phylogenetic position 1, A-1210 Wien, Austria derived from 165 rRNA gene sequence data, chemotaxonomic data and 4 Bereich Mikrobiologie, previously published genomic DNA G+C content data. Gesellschaft fur Biotechnologische Forschung, D-38124 Braunschweig, Germany Keywords: Sphingornonas echinoides comb. nov., phylogenetic position, chemotaxonomy, phenotypical characteristics, reclassification [Pseudomonas]echinoides was initially proposed as the classified as Sphingomonas trueperi; Kampfer et al., name for a yellow-pigmented bacterial strain, desig- 1997) and [P.] echinoides as the carotenoid nated B13 (Heumann, 1962a), which had been isolated nostoxanthin, so far not detected in other yellow- as a plate contaminant (Heumann, 1960). The species pigmented bacteria. A degree of genetic relationship description was based on morphological charac- between [P.]echinoides, S.paucimobilis, Sphingomonas teristics, i.e. a Gram-negative bacillus with polar capsulata (basonym [Flavobacterium]capsulatum) and flagellation, as well as several physiological traits. S. trueperi was observed and the reclassification of Further investigations concerned the genetics, ultra- these four species in a new genus has been proposed structure and carotenoid composition of this species by Owen & Jackman (1982) based on data from (Heumann, 1962b, 1972; Marx & Heumann, 1962; DNA-DNA hybridization studies. DNA-rRNA Heumann & Marx, 1964; Czygan & Heumann, 1967; hybridization studies have clearly distinguished [P.] Mayer et al., 1972). The species name has been echinoides from the genus Pseudomonas sensu strict0 registered in the Approved Lists of bacterial names and placed it close to [P.] paucimobilis and [F.] (Skerman et al., 1980). Previously, it was recognized capsulatum (De Vos et al., 1989). The G + C content of that [P.]echinoides is related genetically to Sphingo- the genomic DNA of [P.]echinoides was determined to monas paucimobilis (basonym Pseudomonas be 65-8 mol YO (Owen & Jackman, 1982). More re- paucimobilis) and related organisms (Segers et al., cently, using DNA-rRNA hybridization, Segers et al. 1994). (1994) observed that [P.] echinoides clusters with et al. species of the genus Sphingomonas (Yabuuchi et al., Jenkins (1979) identified the yellow pigment of 1990). [P.] paucimobilis, [P.] azotocolligans (recently re- ............................................. .......... ...... ............,...,.., ..... .................. ......... .... ................. .......... The genus Sphingomonas, as it is currently described, The EMBL accession number for the Sphingomonas echinoides 165 rDNA contains phylogenetically heterogeneous species and sequence is AJ012461. can be divided into several sub-groups (Van Bruggen 00955 0 1999 IUMS 1103 E. B. M. Denner and others 5. paucimobilis 5. aromaticivorans 0.05 suberifa ciens S. macrogoltabidus Brevundimonas diminuta Fig. 1. An unrooted dendrogram of the estimated phylogenetic relationships between Sphingornonas (Pseudomonas) echinoides DSM 1805T and other member species of the genus Sphingomonas, as derived by evolutionary distance calculations of pairwise comparisons of nearly complete 165 rRNA gene sequences, including the Jukes & Cantor (1969) correction factor. Nucleotide insertionddeletions were weighted at 0.5 x the value of nucleotide substitutions. The dendrogram was generated using the least-squares distance algorithm of the FITCH program in PHYLIP (Phylogeny Inference Package, version 3.5~).Sequences were extracted from the EMBL database and their accession numbers are as follows: 5. adhaesiva JCM 7370T, X72720; S. aromaticivorans SMCC F19gT, U20765; S. asaccharolytica IF0 1549gT, Y09639; 5. capsulata IF0 12533T, D16147; S. chlorophenolica ATCC 33790T, X87161; S. mali IF0 15500T, Y09638; 5. rnacrogoltabidus IF0 1 5033T, D13723; 5. parapaucimobilis JCM 7510T, X72721; 5. paucimobilis DSM 1098T, X72722; 5. pruni IF0 15498T, Y09637; 5. rosa IF0 15208l, D13945; 5. sanguinis IF0 13937T, D13726; 5. stygia SMCC B0712T, U20775; 5. subarctica HAMBI 21 loT, X94102; 5. subterranea SMCC B0478T, U20773; 5. terrae IF0 1 5098T, D13727; 5. trueperi AlTC 12417T, X97776; 5. yanoikuyae JCM 7371T, X72725; Rhizomonas suberifaciens IF0 1521 lT,D13737; Agrobacteriurn tumefaciens DSM 30150, M11223; Brevundimonas diminuta ATCC 11 568T, M59064; Erythrobacter longus JCM 61 70T, D12699; Porphyrobacter neustonensis ACM 2844T, LO1 785. The bar represents the scale of estimated evolutionary distance (i.e. a mean of 5 substitutions per 101 nt) from the point of divergence of the 165 rRNA gene sequences. et al., 1993; Takeuchi et al., 1994; Balkwill et al., major compound (Busse & Auling, 1988; Seger et al., 1997). Members of the genus Sphingomonas are charac- 1994; Hamana & Matsuzaki, 1991). terized by the presence of the respiratory isoprenoid quinone, ubiquinone- 10 (Q- lo), a unique sphingo- To our knowledge only one strain of [P.]echinoides has glycolipid, the presence of 2-hydroxymyristic acid been described and is available in culture collections. (14: 0 2-OH), the absence of 3-hydroxy fatty acids and Considering the recent description of the genus spermidine or sym-homospermidine as the major Sphingomonas, we have reinvestigated the taxonomic polyamine (Yabuuchi et al., 1990; Takeuchi et al., position of [P.]echinoides DSM 1805T (= ATTC 1993,1995; Nohynek et al., 1996; Balkwill et al., 1997; 14820T,DSM 50409T, ICBP 2835T, NCIB 9420T)with Kampfer et al., 1997). Therefore, the presence of respect to all validly described Sphingomonas species. ubiquinone Q-lO,2-hydroxymyristic acid (14: 0 2-OH) and the lack of 3-hydroxy fatty acids are characteristic [P.]echinoides has been characterized by determination for all species of the genus Sphingomonas analysed so of polyamine content, using HPLC (Busse et al., 1997), far. Improved resolution for discrimination within the the respiratory isoprenoid quinone system genus Sphingomonas can be obtained by the deter- (Kroppenstedt, 1982) and the polar lipid pattern, using mination of the polyamine content. Species of the TLC (Tindall, 1990), and by analysis of fatty acid genus Sphingomonas sensu stricto, as suggested by composition, using GLC (Kampfer et al., 1992), Takeuchi et al. (1999, including S. paucimobilis, physiological and biochemical characterizations Sphingomonas parapaucimobilis, Sphingomonas (Kampfer & Altwegg, 1992 ; Kampfer et al., 199 I), and sanguin is, Sph ingomonas asaccharoly t ica, Sphingo- 16s rRNA gene sequence determination, analysis monas adhaesiva, Sphingomonas mali, Sphingomonas (Moore et al., 1993) and comparison with related pruni and S. trueperi (Kampfer et al., 1997), are located member species of the genus Sphingomonas. The nearly on a separate branch of the genus (Fig. 1). These complete 16s rRNA gene sequence (positions 28-1 524, species are characterized by the presence of sym- E. coli 16s rRNA gene sequence numbering) of [P.] homospermidine as the predominant polyamine. echinoides (1442 nt) was determined by direct Other member species of the genus Sphingomonas, sequencing of 16s rDNA after PCR amplification. which are located on different branches, have a Comparative analysis and estimation of phylogenetic polyamine pattern that contains spermidine as the relationships demonstrated that [P.]echinoides clusters 1104 International Journal of Systematic Bacteriology 49 Sphingornonas echinoides comb. nov. Table 1. Sequence similarities between the 165 rRNA gene of Sphingomonas (Pseudomonas)echinoides DSM 1805Tand member species of the genus Sphingomonas as well as species of the a-subclass of the froteobacteria Species Strain* Sequence similarity (%) with S. echinoides S. paucimobilis DSM 109gT 94.3 S. parapaucimobilis JCM 7510T 94-2 S. trueperi LMG 2 142T 93-8 S. adhaesiva JCM 7370T 94.6 S. sanguinis IF0 13937T 93.9 S. mali IF0 15500T 96.4 S. pruni IF0 1549gT 96.2 S. asaccharolytica IF0 15499T 95-9 S. yanoikuyae JCM 7370T 91.9 S. chlorophenolica ATCC 33790T 91.3 S. aromaticivorans SMCC F199T 92.4 ............... ........................................ ............................................. ............................................
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