Gordonia Amicalis Sp. Nov., a Novel Dibenzothiophene-Desulphurizing Actinomycete
Total Page：16
File Type：pdf, Size：1020Kb
International Journal of Systematic and Evolutionary Microbiology (2000), 50, 2031–2036 Printed in Great Britain Gordonia amicalis sp. nov., a novel dibenzothiophene-desulphurizing actinomycete Seung Bum Kim,1 Roselyn Brown,1 Christopher Oldfield,2 Steven C. Gilbert,2 Sergei Iliarionov3 and Michael Goodfellow1 Author for correspondence: Michael Goodfellow. Tel: j44 191 222 7706. Fax: j44 191 222 5228. e-mail: m.goodfellow!ncl.ac.uk 1 Department of The taxonomic position of a dibenzothiophene-desulphurizing soil Agricultural and actinomycete was established using a polyphasic taxonomic approach. The Environmental Science, T University of Newcastle, organism, strain IEGM , was shown to have chemical and morphological Newcastle upon Tyne properties typical of members of the genus Gordonia. The tested strain formed NE1 7RU, UK a distinct phyletic line within the evolutionary radiation occupied by the genus 2 Department of Biological Gordonia, with Gordonia alkanivorans DSM 44369T, Gordonia desulfuricans Sciences, Napier University, NCIMB 40816T and Gordonia rubropertincta DSM 43197T as the most closely Edinburgh EH10 5DT, UK related organisms. Strain IEGMT has a range of phenotypic properties that 3 Institute of Ecology and distinguish it from representatives of all of the validly described species of Genetics of Microorganisms, 13 Golev Gordonia. It was also sharply distinguished from the type strains of Gordonia Street, Perm 614081, Russia desulfuricans and Gordonia rubropertincta on the basis of DNA–DNA relatedness data. The combined genotypic and phenotypic data show that strain IEGMT merits recognition as a new species of Gordonia. The name proposed for the new species is Gordonia amicalis; the type strain is IEGMT (l DSM 44461T l KCTC 9899T). Keywords: Gordonia, polyphasic taxonomy, dibenzothiophene, desulphurization INTRODUCTION chemical, chemical and morphological features (Goodfellow et al., 1999). Gordonia Gordona The genus (formerly ) has had a The 12 validly described species of Gordonia, viz. short, but eventful, taxonomic history. It was proposed Gordonia aichiensis (Tsukamura, 1982) Klatte et al., by Tsukamura (1971), subsumed within the genus 1994, Gordonia alkanivorans Kummer et al., 1999, Rhodococcus (Tsukamura, 1974; Goodfellow & Gordonia amarae (Lechevalier and Lechevalier, 1974) et Alderson, 1977) and re-established by Stackebrandt Klatte et al., 1994, Gordonia bronchialis (Tsukamura, al Rhodococcus . (1988) for actinomycetes classified as 1971) Stackebrandt et al., 1988, Gordonia desulfuri- bronchialis Rhodococcus rubropertinctus Rhodo , and - cans Kim et al., 1999, Gordonia hirsuta Klatte et al., coccus terrae . The genus belongs to the mycolic acid 1996, Gordonia hydrophobica Bendinger et al., 1995, group of actinomycetes, that is, to the suborder Gordonia polyisoprenivorans Linos et al., 1999, Corynebacterineae et al (Stackebrandt ., 1997), which Gordonia rhizosphera Takeuchi and Hatano, 1998, form a distinct phyletic line that also encompasses the Gordonia rubropertincta (Hefferan, 1904) Stackebrandt Corynebacterium Dietzia Mycobacterium genera , , , et al., 1988, Gordonia sputi (Tsukamura, 1978) Riegel Nocardia Rhodococcus Skermania Tsukamurella , , , and et al., 1994 and Gordonia terrae (Tsukamura, 1971) Williamsia et al et al (Chun ., 1996; Goodfellow ., Stackebrandt et al., 1988, form a well-defined clade 1998a, b, 1999). Members of these taxa can be separ- within the evolutionary radiation occupied by the my- ated from one another by using a battery of bio- colic acid-containing actinomycetes (Kim et al., 1999; Kummer et al., 1999; Linos et al., 1999). Available ................................................................................................................................................. DNA–DNA relatedness and numerical phenetic data Abbreviation: meso-A2pm; meso-2,6-diaminopimelic acid. support the taxonomic integrity of these species The GenBank accession number for the 16S rDNA sequence of strain IEGMT (Mordarski et al., 1977, 1980; Zakrzewska-Czerwinska is AF101418. et al., 1988; Goodfellow et al., 1991, 1996; Klatte 01466 # 2000 IUMS 2031 S. B. Kim and others et al., 1994; Riegel et al., 1994; Takeuchi & Hatano, described previously (Kim et al., 1999). The package 1998; Kim et al., 1999, Kummer et al., 1999). Little is (Felsenstein, 1993) was used for all of the analyses. Bootstrap known about the biological properties of gordoniae. analyses (Felsenstein, 1985) of the neighbour-joining data However, they are a potentially rich source of meta- were carried out as described by Kim et al. (1999). bolic diversity, as exemplified by the isolation of strains DNA base composition and DNA relatedness. The DNA base possessing hydrocarbon-oxidizing and aromatic de- composition of strain IEGMT was estimated by following an sulphurizing pathways (Gilbert et al., 1998; Rhee et al., established procedure (Kim et al., 1999). DNA–DNA values 1998; Finkel’shtein et al., 1999; Kummer et al., 1999; between this organism and the type strains of Gordonia Linos et al., 1999). desulfuricans and Gordonia rubropertincta were carried out by the identification service at the Deutsche Sammlung von The aim of the present study was to determine the Mikroorganismen und Zellkulturen (Braunschweig, Ger- taxonomic relationships of a putatively novel member many), as described by Kim et al. (1999). of the genus Gordonia, strain IEGMT, which uses dibenzothiophene as a sole source of sulphur. Geno- RESULTS AND DISCUSSION typic and phenotypic data show that the strain should be recognized as a new species of Gordonia, for which The chemotaxonomic and morphological properties the name Gordonia amicalis sp. nov. is proposed. of strain IEGMT are consistent with its assignment to the genus Gordonia (Stackebrandt et al., 1988; METHODS Goodfellow et al., 1998a, 1999). The organism is T aerobic, Gram-positive, slightly acid–alcohol-fast, Isolation, maintenance and cultivation. Strain IEGM was amycelial and forms red colonies on various complex isolated by enrichment culture from garden soil collected in media, including modified Bennett’s, ISP medium 2 Perm, Russia. Ten grams of soil was shaken at 27 mC for 7 d and peptone–glucose–yeast-extract agars. It contains in a flask containing 100 ml mineral medium (Pfennig, 1965). Inocula from the enrichment culture were spread over meso-A#pm, arabinose and galactose in whole-organ- the surface of corresponding mineral agar plates, which were ism hydrolysates (wall chemotype IV sensu Lechevalier incubated at 27 mC for 10 d, at which point the characteristic & Lechevalier, 1970), predominant amounts of dihy- pink colonies were seen. A representative colony was drogenated menaquinones with nine isoprene units streaked onto a beef-extract agar plate (Atlas, 1993), which [MK-9 (H#)], mycolic acids with 50–58 carbon atoms was incubated at 27 mC for 4 d. The purified strain was and has DNA with a GjC value of 64n2mol%. maintained on modified Bennett’s-agar slopes (Jones, 1949) at 30 mC and as a suspension in 20% (v\v) glycerol at Comparison of the nearly complete 16S rDNA se- V20 mC. Biomass for the chemotaxonomic and molecular quence (1448 nucleotides) of the tested strain with the systematic investigations was prepared by growing the strain corresponding sequences of representatives of the in shake flasks of modified Bennett’s broth for 7 d at 30 mC. suborder Corynebacterineae shows that it forms a Cells for the chemical studies were washed in distilled water, monophyletic clade, in all three analyses, with the type whereas those used for the molecular systematic studies were strains of G. alkanivorans, G. desulfuricans and G. washed in NaCl\EDTA buffer (0n1 M EDTA, pH 8n0, 0n1M rubropertincta (Fig. 1). These relationships are also NaCl). supported by the high nucleotide-similarity values T Phenotypic characterization. The strain was examined for a between strain IEGM and G. alkanivorans DSM T range of phenotypic characteristics (notably those shown in 44369 (98n4% similarity, which corresponds to 23 Table 1) and for the presence of menaquinones, mycolic nucleotide differences), G. desulfuricans NCIMB T acids, whole-organism sugars and the isomers of 2,6- 40816 (98n5% similarity, which corresponds to 21 diaminopimelic acid (A#pm), as described previously (Kim nucleotide differences) and G. rubropertincta DSM et al., 1999). Mass spectra of the purified mycolic esters were T obtained with an Autospec M instrument (Micromass) 43197 (99n0% similarity, which corresponds to 14 operating in electron-impact mode with an ionizing voltage nucleotide differences). The separation of the latter two strains is also supported by DNA–DNA related- of 55 eV and a probe temperature of 150–300 mC. The type T strains of Gordonia alkanivorans and Gordonia polyisopreno- ness data (Kim et al., 1999); strain IEGM showed vorans were also examined for the phenotypic characteristics DNA relatedness values of 31n8 and 37n3% with the shown in Table 1. type strains of G. desulfuricans and G. rubropertincta, Small-subunit rDNA sequencing. Isolation of chromosomal respectively. The type strains of G. alkanivorans and G. DNA, PCR amplification and direct sequencing of the rubropertincta, which share a 99n1% 16S rDNA purified PCR products were carried out using the procedure similarity value (which corresponds to 13 nucleotide of Kim et al. (1998). The resultant 16S rDNA sequence was differences), show a DNA–DNA relatedness value of aligned manually with the corresponding sequences of the 52% (Kummer et al., 1999). All of these