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Phylogenetic Relationships of 38 Taxa of the Genus Staphylococcus Based

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International Journal of Systematic Bacteriology (1999), 49, 725-728 Printed in Great Britain 7 Phylogenetic relationships of 38 taxa of the 1 genus Staphylococcus based on 16s rRNA gene 1 sequence analysis Tatsufumi Takahashi, ltona Satoh and Naoya Kikuchi Author for correspondence: Tatsufumi Takahashi. Tel: +81 11 386 1112. Fax: +81 11 387 5890. e-mail : [email protected] Department of The nucleotide sequences of the 165 rRNA gene (rDNA) in 38 taxa of the genus Epizootiology, School of Staphylococcus were compared phylogenetically.Based on phylogenetic tree Veterinary Medicine, Rakuno Gakuen University, analysis, staphylococcal Species Were divided into 12 Cluster groups. These Ebetsu 069-8501, Japan cluster groups were in very good agreement with species groups determined by DNA-DNA reassociation studies. These genealogical classifications were consistent with the results of the production of coagulase or oxidase and with resistance to novobiocin.These suggest that the phylogenetic relationship of the genus Staphylococcus is accurately represented by the results obtained from the sequence analysis of 165 rDNA. Keywords: Staphylococcus, 16s rDNA, phylogeny The genus Staphylococcus comprised more than 30 However, phylogenetic relationships can be inferred taxa (species and subspecies) (Kloos et al., 1991). Five more precisely by the sequencing method because of its species and four subspecies of Staphylococcus have reliability in the calculation of DNA similarity (Olsen been added to the list of bacteria as newly identified & Woese, 1993). organisms from studies conducted in the 1990s & et al., Earlier studies on the taxonomy of species of the genus (Bannerman Kloos, 1991; Chesneau 1993; Staphylococcus, et al., et al., & based on DNA-DNA reassociation, Hiijek 1992; Igimi 1990; Kloos have indicated that genealogically distinct groups exist Wolfshohl, 199 1 ; Tanasupawat et al., 1992 ; Webster et al., 1994; Zakrzewska-Czerwinska et al., 1995). within the genus as species groups represented by the species of Staphylococcus epiderrnidis, Staphylococcus More recently, Gram-positive bacteria isolated from saprophyticus, Staphylococcus sirnulans, Staphylococ- European otters have been proposed as a new species, cus interrnedius, Staphylococcus hyicus, Staphylococcus Staphylococcus lutrae et al., (Foster 1997). Some of sciuri, Staphylococcus aureus and Staphylococcus these studies employed sequence analysis of the 16s caseolyticus & et al., rRNA or its gene (Hajek et al., 1992; Zakrzewska- (Kloos Schleifer, 1986; Kloos et al., 1991). Our previous study on 23 taxa of the genus Czerwinska 1995). This analysis has been Staphylococcus using 16s rDNA sequence analyses established as a method available for determining & has shown the existence of 10 cluster groups phylogenetic relationships (Olsen Woese, 1993). The (Takahashi et al., 1997). These classifications based on intermittent appearance of a highly conserved se- the whole genome and 16s rDNA analysis were nearly quence and integration of sequence data are useful for identical to each other ;however, the branching pattern correcting sequence alignment. in the phylogenetic tree may change at the nodes Ribotyping is a probe-based fragment analyses because of the limited number of species used for the targeting rRNA operons and the pattern of DNA analyses. Therefore, it is necessary to complete the 16s fragments hybridized with an rDNA probe can be rDNA sequence database for all of the type strains of highly discriminatory, and enable strain typing the undetermined taxa of the genus Staphylococcus. (Jordens, 1998). This method has been applied to the In order to have an overview of intrageneric taxonomy at the species and subspecies levels in the relationships of Staphylococcus, we examined the 16s genus Staphylococcus (Chesneau et al., 1993 ; Webster rDNA sequences of the type strain of each of 38 taxa et al., 1994; De Buyser et al., 1992; Thomson-Carter et by phylogenetic analysis and compared the results al., 1989). Probably, ribotyping is advantageous to 16s from DNA-DNA reassociation studies. rRNA sequence analysis in terms of detecting variations within and flanking the probe sequence. The Staphylococcus strains determined in this study ~ ~~~ 00887 0 1999 IUMS 725 T. Takahashi, I. Satoh and N. Kikuchi 0.03 0.00 I I I 1 Cluster group Species group Staphylococcus caseolyticus MAFF 91 13~7~ =S.caseolyticus 3S.caseolyticus Staphylococcus sciun’ ATCC 29062T 7s.sciuri Ssciuri Staphylococcus vitulinus ATCC 51 145T 1 Is.carnosus 7s*simu’ans1S.simulans I 1 S. hyicus- Sktermedius intermedius 3 S.hyicus =11 S.auricularis =S.auricularis bsp. urealyiicus ATCC 49330T x S.lugdunensis cus MAFF 91 1476T 1S.haemolyticus1 7S.warneri S.epidennidis midis ATCC 14990T 1 I ccus saccharolyticus ATCC 14953T s capitis subsp. urealyticus ATCC 49326T S.epidermidis cus caprae ATCC 3553BT coccus capitis subsp. capitis ATCC 27840T Staphylococcusaureus subsp. aureus ATCC 126OOT -Staphylococcus aureus subsp. anaerobius ATCC 35844T 7S‘aureus Fig. 1. Unrooted neighbour-joining tree based on 165 rRNA gene sequences showing the phylogenetic relationships among the species of the genus Staphylococcus. The numbers are the estimated confidence levels, expressed as percentages, for the positions of the branches, determined by bootstrap analysis (Felsenstein, 1985). The scale bar indicates the evolutionary distance value (K,J between sequences, determined by measuring the lengths of the horizontal lines connecting two organisms. ‘Cluster group’ was designated based on the branching pattern and was compared with the ‘species group‘ described by Kloos et a/. (1991). The accession numbers of the sequences of the organisms used in the analyses are as follows: 5. caseolyticus, D83359; 5. sciuri, S83569; 5. lentus, D83370; 5. pulvereri, AB009942; S. vitulinus, AB009946; 5. simulans, D83373; S. piscifermentans, AB009943; 5. carnosus, AB009934; S. muscae, 583566; 5. chromogenes, D83360; S. hyicus, D83368; S. lutrae, X84731; 5. felis, D83364; S. schleiferi subsp. coagulans, AB009945; S. schleiferi subsp. schleiferi, 583568; S. intermedius, D83369; S. delphini, AB009938; 5. auricularis, D83358; 5. kloosii, AB009940; 5. gallinarum, D83366; 5. arlettae, AB009933; 5. saprophyticus, D8337 1 ; 5. xylosus, D83374; 5. equorum, AB009939; 5. cohnii subsp. cohnii, D83361; S. cohnii subsp. urealyticus, AB009936; S. lugdunensis, AB009941; 5. haemolyticus, D83367; 5. hominis, X66101; 5. pasteuri, AB009944; 5. warneri, L37603; S. aureus subsp. aureus, D83357; 5. aureus subsp. anaerobius, D83355; 5. epidermidis, D83363; 5. saccharolyticus, L37602; 5. capitis subsp. urealyticus, AB009937; S. caprae, AB009935; and S. capitis subsp. capitis, L37599. The origin of strain codes, CCM, DSM and MAFF are the Czechoslovak Collection of Microorganisms, Deutsche Sammlung von Mikroorganismen und Zellkulturen and the Ministry of Agriculture, Forestry and Fisheries of Japan, respectively. were obtained from the American Type Culture construct a phylogenetic tree. The topology of the tree Collection (ATCC). The nucleotide sequence data was evaluated by a bootstrapping method (Felsenstein, reported in this paper appear in the GSDB, DDBJ, 1985). EMBL and GenBank nucleotide sequence databases with the accession numbers shown in the legend for The relatively close relationships among species of the Fig. 1. The bacterial culture, DNA extraction, gene genus Staphylococcus was confirmed by phylogenetic amplification and sequencing techniques were analyses based on the gene sequencing employed in described previously (Takahashi et al., 1997). Sequence this study. However, the phylogenetic tree showed that alignment was performed from nucleotide residues staphylococcal strains were divided into distinct sub- 28-1 490, based on the Escherichia coli numbering groups that were recovered in high percentages of the system (Brosius et al., 1978). Evolutionary distance bootstrap trees. Based on criteria of the branching values (Knuc)were calculated by Kimura’s two-par- patterns and the bootstrap values (80% or more), 12 ameter method (Kimura, 1980) with the BioResearch cluster groups were designated, as shown in Fig. 1. SINCA program package. The neighbour-j oining They are S. caseolyticus, S. sciuri, S. carnosus, S. method of Saitou & Nei (1987) was employed to simulans, S. hyicus-intermedius, S. auricularis, S. 726 International Journal of Systematic Bacteriology 49 Phylogeny of Staphylococcus spp. saprophyticus, S. lugdunensis, S. haemolyticus, S. ococcus haemolyticus, Staphylococcus hominis, warneri, S. epidermidis and S. aureus. Staphylococcus lugdunensis, Staphylococcus saccharo- lyticus and Staphylococcus warneri were somewhat The phylogenetic position of S. caseolyticus is the most Staphy- closely related with each other and with the recently distant from any other taxa of the genus added species Staphylococcus pasteuri. However, lococcus. A root for Bacillus subtilis as an outgroup was branched off from a line between S. caseolyticus specific association among these species was not and the others (data not shown). The staphylococcal exhibited in our treeing analysis. Based on the above- described criteria, we divided these species into five species groups determined by DNA-DNA re- cluster groups : S. lugdunensis, S. haemolyticus, S. association studies (Kloos & Schleifer, 1986; Kloos et S. S. al., warneri, epidermidis and aureus (Fig. 1). The 199 1) have been supported by phenotypic character branching order of S. lugdunensis, S. haemolyticus and analysis. In
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  • The Genome Sequence of a Type ST239 Methicillin-Resistant Staphylococcus Aureus Isolate from a Malaysian Hospital

    The Genome Sequence of a Type ST239 Methicillin-Resistant Staphylococcus Aureus Isolate from a Malaysian Hospital

    Standards in Genomic Sciences (2014) 9: 933-939 DOI:10.4056/sigs.3887716 The Genome Sequence of a Type ST239 Methicillin-Resistant Staphylococcus aureus Isolate from a Malaysian Hospital LS Lee1,2, LK Teh1, ZF Zainuddin2 and MZ Salleh1* 1Integrative Pharmacogenomics Centre, Faculty of Pharmacy, Universiti Teknologi MARA Malaysia, 42300 Bandar Puncak Alam, Selangor, Malaysia. 2School of Health Sciences, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan, Ma- laysia * Corresponding author: MZ Salleh ([email protected]) Keywords: Staphylococcus aureus, MRSA, Malaysia, Genomics We report the genome sequence of a healthcare-associated MRSA type ST239 clone isolated from a patient with septicemia in Malaysia. This clone typifies the characteristics of ST239 lineage, including resistance to multiple antibiotics and antiseptics. Introduction cin, ciprofloxacin and co-trimoxazole. Antibiotic resistance in S. aureus is a major con- Genome sequencing information cern, as an increasing number of infections are caused by methicillin-resistant S. aureus (MRSA). Genome project history Figure 1 shows the phylogenetic position of S. This organism was selected for sequencing as a aureus in relation to other staphylococci. In Ma- representative of MRSA infection in a local Malay- laysia, the incidence of MRSA-related infections is sian hospital. The genome sequences of this or- a cause of concern in hospitals country-wide. ganism were deposited in GenBank (WGS data- Health-associated MRSA (HA-MRSA) has been base). Sequencing, finishing and annotation were dominated by a few lineages in Southeast Asia, performed at the Pharmacogenomics Centre particularly ST239. Sequence type 239 is an inter- (PROMISE), UiTM. Table 2 presents the project in- national healthcare-associated (HA) MRSA lineage formation and its association with MIGS version prevalent in Asia, South America and Eastern Eu- 2.0 compliance [14].