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Actinokineospora Spheciospongiae Sp. Nov., Isolated from the Marine Sponge Spheciospongia Vagabunda

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Actinokineospora Spheciospongiae Sp. Nov., Isolated from the Marine Sponge Spheciospongia Vagabunda View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by OceanRep International Journal of Systematic and Evolutionary Microbiology (2015), 65, 879–884 DOI 10.1099/ijs.0.000031 Actinokineospora spheciospongiae sp. nov., isolated from the marine sponge Spheciospongia vagabunda Peter Ka¨mpfer,1 Stefanie P. Glaeser,1 Hans-Ju¨rgen Busse,2 Usama Ramadan Abdelmohsen,33 Safwat Ahmed4 and Ute Hentschel3 Correspondence 1Institut fu¨r Angewandte Mikrobiologie, Justus-Liebig-Universita¨t Giessen, D-35392 Giessen, Peter Ka¨mpfer Germany [email protected] 2Institut fu¨r Bakteriologie, Mykologie und Hygiene, Veterina¨rmedizinische Universita¨t, giessen.de A-1210 Wien, Austria 3Department of Botany II, Julius-von-Sachs-Institute for Biological Sciences, University of Wuerzburg, D-97082 Wuerzburg, Germany 4Department of Pharmacognosy, Faculty of Pharmacy, Suez Canal University, Ismailia 41522, Egypt A Gram-staining-positive, aerobic organism, isolated from the Red Sea sponge Spheciospongia vagabunda was investigated to determine its taxonomic position. On the basis of results of 16S rRNA gene sequence analysis strain EG49T was most closely related to Actinokineospora cibodasensis and Actinokineospora baliensis (both 97.3 % similarity) and Actinokineospora diospyrosa and Actinokineospora auranticolor (both 97.0 % similarity). The 16S rRNA gene sequence similarity to all other species of the genus Actinokineospora was ,97.0 %. The quinone T system of strain EG49 contained the menaquinones MK-9(H4) (47 %), MK-9(H6) (27 %) and MK-9(H2) (15 %) in major amounts. Minor amounts of MK-7(H4) (2 %), MK-9(H0) (1 %), MK- 9(H8) (3 %) and MK-10(H4) (3 %) were detected as well in addition to MK-8(H4), MK-8(H6), MK- 10(H2) and MK-10(H6) (all ,1 %). The diagnostic diamino acid of the peptidoglycan was meso- diaminopimelic acid. In the polar lipid profile, diphosphatidylglycerol, phosphatidylethanolamine and hydroxyphosphatidylethanolamine were predominant. Phosphatidylinositol-mannoside, two unidentified phospholipids and two glycoglipids as well as one aminoglycolipid, one aminolipid and one unidentified lipid were found in addition. The fatty acid profile was composed of mainly iso-branched fatty acids: iso-C16 : 0, iso-C14 : 0, iso-C15 : 0 and iso-C16 : 1H. All these findings clearly supported the classification of the strain as representing a member of the genus Actinokineospora. In addition, the results of physiological and biochemical tests also allowed phenotypic differentiation of strain EG49T from the most closely related species of the genus Actinokineospora. Strain EG49T represents a novel species of the genus Actinokineospora, for which we propose the name Actinokineospora spheciospongiae sp. nov., with strain EG49T (5DSM 45935T5CCM 8480T5LMG 27700T) as the type strain. The genus Actinokineospora was proposed by Hasegawa fastidiosa. The genus contains, at the time of writing, 13 (1988), initially for motile, arthrospore-producing organ- species: Actinokineospora riparia (the type species), Actino- isms of the class Actinomycetes. Labeda et al. (2010) kineospora inagensis, Actinokineospora globicatena, Actinokineo- recently emended the description of the genus, which now spora terrae, Actinokineospora diospyrosa, Actinokineospora harbours also species for which the production of motile auranticolor, Actinokineospora enzanensis, Actinokineospora fasti- spores was not observed, and proposed the transfer of diosa, Actinokineospora baliensis, Actinokineospora bangkokensis, Amycolatopsis fastidiosa to this genus as Actinokineospora Actinokineospora cianjurensis, Actinokineospora cibodasensis and Actinokineospora soli (Hasegawa, 1988; Tamura et al., 1995; 3Permanent address: Department of Pharmacognosy, Faculty of Otoguro et al., 2001; Labeda et al., 2010, Lisdiyanti et al., 2010; Pharmacy, Minia University, Minia 61519, Egypt Tang et al., 2012; Intra et al., 2013). These actinomycetes The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene are characterized by having meso-diaminopimelic acid as a T sequence of strain EG49 is GU318361. cell-wall diamino acid, MK-9(H4) as the predominant 000031 G 2015 IUMS Printed in Great Britain 879 P. Ka¨mpfer and others menaquinone, phospholipid type II, iso-C16 : 0 fatty acid as the the SILVA Incremental Aligner (SINA; v1.2.11; Pruesse et al., predominant fatty acid and DNA G+C contents of 69– 2012) according to the SILVA seed alignment [http://www. 74 mol%. arb-silva.de; Pruesse et al. (2007)]. The aligned sequence was implemented into the LTP database and added to the Strain EG49T was isolated by Abdelmohsen et al. (2010) database tree using the ARB Parsimony (Quick add marked) from the marine sponge Spheciospongia vagabunda col- tool. The alignment, including all members of the family lected from the Red Sea (Ras Mohamed, Sinai, Egypt; GPS Pseudonocardiaceae and some outgroup species of the u 9 u 9 coordinates 27 47.655 N34 12.904 W). The metabolo- family Micromonosporaceae, was checked manually before mic and the genomic analyses of the strain showed its richness reconstruction of phylogenetic trees. A maximum-like- with diverse bioactive natural products. (Abdelmohsen et al., lihood tree was reconstructed using RAxML v7.04 2014; Harjes et al., 2014; Grkovic et al., 2014). (Stamatakis, 2006) with GTR-GAMMA and rapid boot- The strain showed the presence of aerial mycelium with strap analysis. The tree was based on 16S rRNA gene spore chains. The spores were rod-shaped and were formed sequence termini 73–1443 (Escherichia coli numbering; by fragmentation of the hyphae (arthrospores). Cultural Brosius et al., 1978). The phylogenetic tree showed a clear T characteristics were recorded after 14 days of incubation at allocation of strain EG49 , clustering among species of the 28 uC on International Streptomyces Project (ISP) 2 medium, genus Actinokineospora. Based on these results, further tryptone soy agar (TSA; Oxoid)) and nutrient agar (Oxoid). phylogenetic analyses were performed including all species Light yellow to brown colonies were produced on these of the genus Actinokineospora and species of the genus media. The isolate exhibited good growth on all of the media. Kutzneria as the outgroup. Again, a maximum-likelihood tree, a neighbour-joining tree using ARB neighbour-joining Cells displayed Gram-positive staining (analysed as and the Jukes–Cantor correction (Jukes & Cantor, 1969) described by Gerhardt et al., 1994) and were negative for and a maximum-parsimony tree using DNAPARS v 3.6 cytochrome oxidase, determined by using an oxidase test (Felsenstein, 2005) were generated. The phylogenetic (Merck). Endospores could not be detected. Temperature- trees were reconstructed with 100 resamplings (bootstrap dependent growth was determined on ISP2 agar at 4, 15, analysis; Felsenstein, 1985) and based on 16S rRNA gene 25, 28, 32, 37 and 42 uC. Salinity- and pH-dependent sequence termini 62–1459 (E. coli numbering, Brosius et al., growth were analysed in ISP2 broth either supplemented 1978). Pairwise sequence similarities among the type strains with 1–10 % (w/v) NaCl or adjusted to pH values between of species of the genus Actinokineospora were calculated pH 4 and 12 (at intervals of 0.5 pH units, adjusted by the using the ARB neighbour-joining tool without the use of an addition of HCl or NaOH) and cultured at 28 uC. evolutionary substitution model. Detailed phylogenetic analysis based on 16S rRNA gene The sequenced 16S rRNA gene of strain EG49T represents sequences was performed in ARB release 5.2 (Ludwig et al., a continuous stretch of 1481 nt spanning E. coli positions 2004) using the ‘All-Species Living Tree’ Project (LTP) 9–1514 (E. coli numbering; Brosius et al., 1978). Strain (Yarza et al., 2008) database LTPs115 (March, 2014). The EG49T shared highest 16S rRNA gene sequence similarity 16S rRNA gene sequence of strain EG49T was aligned with with the type strains of Actinokineospora cibodasensis and 95 Actinokineospora enzanensis IFO 16517T (AB058395) ** Actinokineospora bangkokensis 44EHWT (JQ922512) Actinokineospora riparia NRRL B–16432T (AF114802) 100 Actinokineospora soli YIM 75948T (JN005785) ** Actinokineospora fastidiosa NRRL B–16697T (GQ200601) Actinokineospora cibodasensis NBRC 104212T (AB447489) * Actinokineospora spheciospongiae EG49T (GU318361) Actinokineospora diospyrosa NRRL B–24047T (AF114797) Actinokineospora auranticolor IFO 16518T (AB058396) Actinokineospora baliensis NBRC 104211T (AB447488) Actinokineospora cianjurensis BTCC B–558T (AB473945) Actinokineospora globicatena NRRL B-24048T (AF114798) 100 Actinokineospora terrae IFO 15668T (AB058394) Actinokineospora inagensis NRRL B–24050T (AF114799) 100 3 Kutzneria 0.10 Fig. 1. Maximum-likelihood tree showing the phylogenetic position of strain EG49T among type strains of species of the genus Actinokineospora. The tree was generated in ARB using RAxML (GTR-GAMMA, Rapid Bootstrap analysis) and based on nucleotide sequences at 16S rRNA gene sequence positions 62–1459 (E. coli numbering). Type strains of species of the genus Kutzneria were used as the outgroup. Bootstrap values ¢70 % are given in the tree. Nodes marked with asterisks also occurred in the maximum-parsimony and neighbour-joining trees; those nodes marked with two asterisks also showed bootstrap values .70 %. Bar, 0.1 substitutions per nucleotide position. 880 International Journal of Systematic and Evolutionary Microbiology 65 Actinokineospora spheciospongiae sp. nov. Actinokineospora baliensis (both 97.3 %) and Actinokineospora
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