Li, F.

Turner, S.L.

Young, J.P.W.

Zhang, X.

English

White Rose Research Online

This is a repository copy of What does a bacterial genome sequence represent? Mis-assignment of MAFF 303099 to the genospecies Mesorhizobium loti.White Rose Research Online URL for this paper:http://eprints.whiterose.ac.uk/292/Article:Zhang, X., Li, F., Young, J.P.W. orcid.org/0000-0001-5259-4830 et al. (1 more author) (2002) What does a bacterial genome sequence represent? Mis-assignment of MAFF 303099 to the genospecies Mesorhizobium loti. Microbiology. pp. 3330-3331. ISSN 1465-2080 eprints@whiterose.ac.ukhttps://eprints.whiterose.ac.uk/Reuse Items deposited in White Rose Research Online are protected by copyright, with all rights reserved unless indicated otherwise. They may be downloaded and/or printed for private study, or other acts as permitted by national copyright laws. The publisher or other rights holders may allow further reproduction and re-use of the full text version. This is indicated by the licence information on the White Rose Research Online record for the item. Takedown If you consider content in White Rose Research Online to be in breach of UK law, please notify us by emailing eprints@whiterose.ac.uk including the URL of the record and the reason for the withdrawal request. Microbiology Comment..................................................................................................................................................................................................................Fig. 1. Phylogenetic trees showing evolutionary relationships among Mesorhizobium strains for fivepartial gene sequences : 16S rrn (606–1420 bp, gaps ignored in pair-wise comparisons), glnA (936 bp),glnII (828 bp), recA (440 bp) and nodA (470 bp). Sequences labelled A1–4 (strains HN8A1, ZJ15B7,JX2B5 and JS5A15) and B1–3 (HB5A4, HN14A16 and HN15B23) are from isolates that nodulateAstragalus sinicus (7), MAFF 303099 indicates sequences taken from the published genome sequence.These sequences were amplified and sequenced according to the methods described in and compared tothe corresponding sequences of Mesorhizobium type strains described in: ref. 2 for 16S rrn and recA ;ref. 5 for glnA and glnII and ref. 7 for nodA. All trees were calculated using PAUP* (version 4.0b10;http:}}paup.csit.fsu.edu}) using the neighbour-joining algorithm with the HKY model. Trees wererooted using the corresponding Sinorhizobium meliloti sequence (not shown on trees). Bootstrappercentages shown on each branch were estimated from 1000 replicates. New sequences for the A.sinicus isolates have been deposited in the databases under accession numbers AJ459585–AJ459605.sMMO genes are expressed in heterologoushosts. We have recently identified a groELgene located 5« of the mmoX gene cluster inM. trichosporium OB3b which, when muta-ted, results in a mutant with an sMMO-minusphenotype (Murrell et al., unpublished obser-vations). This groEL, which is not present onthe sMMO gene cluster constructs used inexperiments by Wood and colleagues (1, 2),may be essential for the correct assembly ofthe sMMO or sMMO regulatory proteins,which could account for the high level expres-sion of sMMO in the homologous host sincethis groEL is present both on the chromo-some of M. trichosporium OB3b and on theexpression plasmid used (3).J. Colin MurrellDepartment of Biological Sciences,University of Warwick, Coventry, CV4 7AL,UKTel: ­44 24 76523553. Fax: ­44 2476523568. e-mail :cmurrell!bio.warwick.ac.uk1. Jahng, D. & Wood, T. K. (1994). Trichloroethyleneand chloroform degradation by a recombinantpseudomonad expressing soluble methanemonooxygenase from Methylosinus trichosporium OB3b.Appl Environ Microbiol 60, 2473–2482.2. Jahng, D., Kim, C. S., Hanson, R. S. & Wood, T. K.(1996). Optimization of trichloroethylene degradationusing soluble methane monooxygenase of Methylosinustrichosporium OB3b expressed in recombinant bacteria.Biotechnol Bioeng 51, 349–359.3. Lloyd, J. S., Finch, R., Dalton, H. & Murrell, J. C.(1999). Homologous expression of soluble methanemonooxygenase genes in Methylosinus trichosporiumOB3b. Microbiology 145, 461–470.What does a bacterialgenome sequencerepresent? Mis-assignment of MAFF303099 to the genospeciesMesorhizobium lotiDoolittle (1) recently discussed the potentialand limitations of bacterial genome sequen-cing, and emphasized the importance oflateral gene transfers in bacterial adaptation,e.g. to become pathogens. The transferablegenes, or ‘accessory genome’, should com-prise genes that are advantageous intermit-tently and are not uniformly distributedamong individuals of a species, though theymay be shared between species. The import-ant phenotypes that they encode are often ofeconomic interest to medicine, e.g. patho-genicity islands and antibiotic resistance, andagriculture, e.g. symbiotic nitrogen fixationby rhizobia in association with plant roots.Such traits are sometimes used to identifybacteria, but this can lead to an ‘ identitycrisis ’. This is illustrated by the largestgenome that Doolittle (1) discusses, that of astrain of rhizobia (MAFF 303099) isolatedfrom a Lotus corniculatus root nodule. Thestrain was identified as Mesorhizobium lotibut we suggest, based on phylogeny, that itbelongs to a different species : Mesorhizobiumhuakuii biovar loti. Host-range variants inrhizobia are generally called ‘biovars ’, inwhich the chromosome type (basic genome)defines the species and the symbiosis type(accessory genome) the biovar. MAFF 303099differs considerably from the M. loti typestrain at four chromosomal loci (rrn, glnA,glnII, recA ; Fig. 1) and more resembles thetype strain of M. huakuii (every bacterialspecies is defined by similarity to a singlerepresentative, the ‘ type strain ’).The affiliation with M. huakuii is par-ticularly striking for glutamine synthetase II(glnII), as MAFF 303099 shares a distinctivesequence thought to have been acquired bythe ancestor of the M. huakuii lineage throughlateral gene transfer from a Rhizobium-likespecies (5). We found this same glnII signaturein four strains representing the dominant(" 96%) chromosomal type (6) among 204isolates from Astragalus sinicus, the typicalhost of M. huakuii (type A1–4 in Fig. 1).Three strains representing the remaininggenotypic diversity belong to a different lin-eage (type B1–3 in Fig. 1). We conclude thatCCBAU 2609T is truly representative of M.huakuii, and that MAFF 303099 belongs tothis species.In contrast, MAFF 303099 has symbiosisgenes (represented here by nodA) that aremost similar to those of otherLotus symbionts(Fig. 1). Symbiosis genes are mobile, beingusually found on plasmids or transmissiblegenetic islands. All A. sinicus symbiontsscreened to date have identical nodA se-quences (7), indicating relatively recent (inevolutionary time) transfer of their symbiosisgenes between species. More directly, Sullivanet al. (4) detected transfer of the genes fornodulation of Lotus from an inoculant straininto several different chromosomal back-grounds within four years.Lateral gene transfer thus shapes the bac-Microbiology 148, November 20023330Microbiology Comment(b)(c)(a)..................................................................................................................................................................................................................Fig. 1. (a) Structure of lysylphosphatidylglycerol (L-PG). The -lysine is shown in grey. (b) L-PG-mediated resistance against CAMPs is based on electrostatic repulsion from the bacterial membrane.Modified with permission from (4). (c) Production of L-PG by Bacillus subtilis (lane 1) Staphylococcusaureus (lane 3) and an S. aureus mprF mutant (lane 2). Polar lipids were prepared, separated by TLCand stained with ninhydrin as described recently (5).terial genome on two different time scales.The transfer of glnII from Rhizobium toMesorhizobium was apparently a single, rareevent, since other basic ‘housekeeping’ genesgenerally share a consistent phylogeny (2, 5).By contrast, the accessory genome, here repre-sented by symbiosis genes, undergoes detect-able transfers within and between species.Accessory DNA makes up 10–25% of theDNA in the three rhizobial genomes se-quenced so far [M. loti (NCj002678), Sino-rhizobium meliloti (NCj003047) and Agro-bacterium tumefaciens (NCj003305)].Sequencing an individual bacterial genomewill provide a clear picture of the basicgenome of the species, but only an arbitrary‘snapshot ’ of that part of the accessory genepool that happens to be in the chosen isolate.When genome sequences are available frommore than one strain of a species, e.g.Escherichia coli, we see clearly that they differin their complement of accessory genes (3), sothat a single strain does not adequatelyrepresent the whole species. Characterizingand sequencing more than one member of aspecies is therefore important, since this willdefine the common core of genes whichdefines the species. It will also allow identi-fication of the associated accessory gene poolthat provides a species with its adaptations todifferent niches and determines the variety ofkey properties such as symbioses or diseaseswith which the species is associated (1).Sarah L. Turner,1† Xue-Xian Zhang,2† Fu-DiLi2 and J. Peter W. Young11Department of Biology, University of York,PO Box 373, York YO10 5YW, UK.2Department of Microbiology, HuazhongAgricultural University, China.†Present address : CEH Oxford, MansfieldRoad, Oxford OX1 3SR, UK.Author for correspondence: Sarah L. Turner.Tel : ­44 1865 281 630. Fax: ­44 1865 281696. e-mail : sltu!ceh.ac.uk1. Doolittle, R. F. (2002). Microbial genomes multiply.Nature 416, 697–700.2. Gaunt, M. J., Turner, S. L., Rigottier-Gois, L., Lloyd-Macgilp, S. A. & Young, J. P. W. (2001). Phylogenies ofatpD and recA support the SSU rRNA-basedclassification of rhizobia. Int J Syst Evol Microbiol 51,2037–2048.3. Perna, N. T., Plunkett, G., Burland, V. & 25 otherauthors (2001). Genome sequence of enterohaemorrhagicEscherichia coli O157:H7. Nature 409, 529–533.4. Sullivan, J. T., Eardly, D. B., van Berkum, P. &Ronson, C. W. (1996). Four unnamed species ofnonsymbiotic rhizobia isolated from the rhizosphere ofLotus corniculatus. Appl Environ Microbiol 62,2818–2825.5. Turner, S. L. & Young, J. P. W. (2000). The glutaminesynthetases of rhizobia: phylogenetics and evolutionaryimplications. Mol Biol Evol 17, 309–319.6. Zhang, X. X., Guo, X. W., Terefework, Z., Cao,Y. Z., Hu, F. R, Lindstrom, K & Li, F. D. (1999). Geneticdiversity among rhizobial isolates from field grownAstragalus sinicus of Southern China. Syst ApplMicrobiol 22, 312–320.7. Zhang, X. X., Turner, S. L., Guo, X. W., Yang, H. J.,Debelle! , F., Yang, G. P., De! narie! , J., Young, J. P. W. &Li, F. D. (2000). The common nodulation genes ofAstragalus sinicus rhizobia are conserved despitechromosomal diversity. Appl Environ Microbiol 66,2988–2995.MprF-mediated lysinylationof phospholipids in Bacillussubtilis – protectionagainst bacteriocins interrestrial habitats?A common strategy for organisms to inhibitbacteria is the production of antimicrobialpeptides that damage bacterial membranesand that usually have cationic properties toenable efficient interactions with the anionicpolymers in bacterial cell envelopes. Suchcationic antimicrobial peptides (CAMPs) areproduced by the innate immune systems ofhumans, animals and plants (e.g. defensins)(7) and many of the bacterial bacteriocinsbelong to the same class of molecules (3).The recently described bacterial mechan-isms protecting against a wide range ofCAMPs may be beneficial in many kinds ofenvironments. Most of these mechanismsinvolve modulations of the net charge in thebacterial cell envelope to reduce accumulationof cationic peptides (4). Modification ofteichoic acids with positively charged -Microbiology 148, November 2002 3331

What does a bacterial genome sequence represent? Mis-assignment of MAFF 303099 to the genospecies Mesorhizobium loti

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