1887

Abstract

Multilocus sequence analysis (MLSA) was performed on representative species of the genus . Internal fragments of the genes selected, , , , , and 16S rRNA, were amplified by direct PCR and then sequenced from 38 strains representing 35 species. Neighbour-joining (NJ), maximum-likelihood (ML) and maximum-parsimony (MP) phylogenies of the individual genes were compared. The data confirm that the potential for discrimination of species is greater using MLSA of housekeeping genes than 16S rRNA genes. Among the housekeeping genes analysed, was the most informative, followed by . Analyses of concatenated sequences (4816 bp) of all six genes revealed robust phylogenetic relationships among different species when compared with the single-gene trees. The NJ, ML and MP trees were very similar, and almost fully resolved relationships of species were obtained, to our knowledge for the first time. In addition, analysis of a concatenation (2457 bp) of the , and genes produced essentially the same result. Ten distinct clades were recognized using the SplitsTree4 program. For the genus , we can define species as a group of strains that share at least 97.5 % gene sequence similarity based on the fragments of five protein-coding housekeeping genes and the 16S rRNA gene. This study demonstrates that MLSA of housekeeping genes is a valuable alternative technique for the identification and classification of species of the genus .

Funding
This study was supported by the:
  • Institute for Fermentation, Osaka (IFO, Osaka, Japan) (Award 2009-2011)
  • Japan Society for the Promotion of Science (Award 23580126)
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.052332-0
2011-12-01
2024-04-20
Loading full text...

Full text loading...

/deliver/fulltext/micro/157/12/3388.html?itemId=/content/journal/micro/10.1099/mic.0.052332-0&mimeType=html&fmt=ahah

References

  1. Ah-You N., Gagnevin L., Grimont P. A. D., Brisse S., Nesme X., Chiroleu F., Bui Thi Ngoc L., Jouen E., Lefeuvre P. et al. & other authors ( 2009). Polyphasic characterization of xanthomonads pathogenic to members of the Anacardiaceae and their relatedness to species of Xanthomonas . Int J Syst Evol Microbiol 59:306–318 [View Article][PubMed]
    [Google Scholar]
  2. Bakir M. A., Kitahara M., Sakamoto M., Matsumoto M., Benno Y. ( 2006a). Bacteroides intestinalis sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 56:151–154 [View Article][PubMed]
    [Google Scholar]
  3. Bakir M. A., Kitahara M., Sakamoto M., Matsumoto M., Benno Y. ( 2006b). Bacteroides finegoldii sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 56:931–935 [View Article][PubMed]
    [Google Scholar]
  4. Bakir M. A., Sakamoto M., Kitahara M., Matsumoto M., Benno Y. ( 2006c). Bacteroides dorei sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 56:1639–1643 [View Article][PubMed]
    [Google Scholar]
  5. Bilhère E., Lucas P. M., Claisse O., Lonvaud-Funel A. ( 2009). Multilocus sequence typing of Oenococcus oeni: detection of two subpopulations shaped by intergenic recombination. Appl Environ Microbiol 75:1291–1300 [View Article][PubMed]
    [Google Scholar]
  6. Bruen T. C., Philippe H., Bryant D. ( 2006). A simple and robust statistical test for detecting the presence of recombination. Genetics 172:2665–2681 [View Article][PubMed]
    [Google Scholar]
  7. Bryant D., Moulton V. ( 2004). Neighbor-Net: an agglomerative method for the construction of phylogenetic networks. Mol Biol Evol 21:255–265 [View Article][PubMed]
    [Google Scholar]
  8. Bui Thi Ngoc L., Vernière C., Jouen E., Ah-You N., Lefeuvre P., Chiroleu F., Gagnevin L., Pruvost O. ( 2010). Amplified fragment length polymorphism and multilocus sequence analysis-based genotypic relatedness among pathogenic variants of Xanthomonas citri pv. citri and Xanthomonas campestris pv. bilvae . Int J Syst Evol Microbiol 60:515–525 [View Article][PubMed]
    [Google Scholar]
  9. Chassard C., Delmas E., Lawson P. A., Bernalier-Donadille A. ( 2008). Bacteroides xylanisolvens sp. nov., a xylan-degrading bacterium isolated from human faeces. Int J Syst Evol Microbiol 58:1008–1013 [View Article][PubMed]
    [Google Scholar]
  10. Enersen M., Olsen I., van Winkelhoff A. J., Caugant D. A. ( 2006). Multilocus sequence typing of Porphyromonas gingivalis strains from different geographic origins. J Clin Microbiol 44:35–41 [View Article][PubMed]
    [Google Scholar]
  11. Felsenstein J. ( 1985). Confidence limits of phylogenies: an approach using the bootstrap. Evolution 39:783–791 [View Article]
    [Google Scholar]
  12. Gevers D., Cohan F. M., Lawrence J. G., Spratt B. G., Coenye T., Feil E. J., Stackebrandt E., Van de Peer Y., Vandamme P. et al. & other authors ( 2005). Opinion: re-evaluating bacterial species. Nat Microbiol Rev 3:733–739 [View Article]
    [Google Scholar]
  13. Grosse-Herrenthey A., Maier T., Gessler F., Schaumann R., Böhnel H., Kostrzewa M., Krüger M. ( 2008). Challenging the problem of clostridial identification with matrix-assisted laser desorption and ionization-time-of-flight mass spectrometry (MALDI-TOF MS). Anaerobe 14:242–249 [View Article][PubMed]
    [Google Scholar]
  14. Guindon S., Gascuel O. ( 2003). A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol 52:696–704 [View Article][PubMed]
    [Google Scholar]
  15. Hanage W. P., Fraser C., Spratt B. G. ( 2006). Sequences, sequence clusters and bacterial species. Philos Trans R Soc Lond B Biol Sci 361:1917–1927 [View Article][PubMed]
    [Google Scholar]
  16. Hayashi H., Shibata K., Bakir M. A., Sakamoto M., Tomita S., Benno Y. ( 2007). Bacteroides coprophilus sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 57:1323–1326 [View Article][PubMed]
    [Google Scholar]
  17. Huson D. H., Bryant D. ( 2006). Application of phylogenetic networks in evolutionary studies. Mol Biol Evol 23:254–267 [View Article][PubMed]
    [Google Scholar]
  18. Jousimes-Somer H. R., Summanen P., Citron D. M., Baron E. J., Wexler H. M., Finegold S. M. ( 2002). Wadworth-KTL Anaerobic Bacteriology Manual, 6th edn. Belmont, CA: Star Publishing Co;
    [Google Scholar]
  19. Keymer D. P., Boehm A. B. ( 2011). Recombination shapes the structure of an environmental Vibrio cholerae population. Appl Environ Microbiol 77:537–544 [View Article][PubMed]
    [Google Scholar]
  20. Kim M.-S., Roh S. W., Bae J.-W. ( 2010). Bacteroides faecis sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 60:2572–2576 [View Article][PubMed]
    [Google Scholar]
  21. Kimura M. ( 1980). A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120 [View Article][PubMed]
    [Google Scholar]
  22. Kitahara M., Sakamoto M., Ike M., Sakata S., Benno Y. ( 2005). Bacteroides plebeius sp. nov. and Bacteroides coprocola sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 55:2143–2147 [View Article][PubMed]
    [Google Scholar]
  23. Kitahara M., Tsuchida S., Kawasumi K., Amao H., Sakamoto M., Benno Y., Ohkuma M. ( 2011). Bacteroides chinchillae sp. nov. and Bacteroides rodentium sp. nov., isolated from chinchilla (Chinchilla lanigera) faeces. Int J Syst Evol Microbiol 61:877–881 [View Article][PubMed]
    [Google Scholar]
  24. Ko K. S., Kuwahara T., Haehwa L., Yoon Y.-J., Kim B.-J., Lee K.-H., Ohnishi Y., Kook Y.-H. ( 2007). RNA polymerase β-subunit gene (rpoB) sequence analysis for the identification of Bacteroides spp. Clin Microbiol Infect 13:48–54 [View Article][PubMed]
    [Google Scholar]
  25. Lan P. T. N., Sakamoto M., Sakata S., Benno Y. ( 2006). Bacteroides barnesiae sp. nov., Bacteroides salanitronis sp. nov. and Bacteroides gallinarum sp. nov., isolated from chicken caecum. Int J Syst Evol Microbiol 56:2853–2859 [View Article][PubMed]
    [Google Scholar]
  26. Larkin M. A., Blackshields G., Brown N. P., Chenna R., McGettigan P. A., McWilliam H., Valentin F., Wallace I. M., Wilm A. et al. & other authors ( 2007). clustal w and clustal_x version 2.0. Bioinformatics 23:2947–2948 [View Article][PubMed]
    [Google Scholar]
  27. Lee C. S., Lee J. ( 2010). Evaluation of new gyrB-based real-time PCR system for the detection of B. fragilis as an indicator of human-specific fecal contamination. J Microbiol Methods 82:311–318 [View Article][PubMed]
    [Google Scholar]
  28. Leon A., Pronost S., Fortier G., Andre-Fontaine G., Leclercq R. ( 2010). Multilocus sequence analysis for typing Leptospira interrogans and Leptospira kirschneri . J Clin Microbiol 48:581–585 [View Article][PubMed]
    [Google Scholar]
  29. Librado P., Rozas J. ( 2009). DnaSP v5: a software for comprehensive analysis of DNA polymorphism data. Bioinformatics 25:1451–1452 [View Article][PubMed]
    [Google Scholar]
  30. Liu C., Song Y., McTeague M., Vu A. W., Wexler H., Finegold S. M. ( 2003). Rapid identification of the species of the Bacteroides fragilis group by multiplex PCR assays using group- and species-specific primers. FEMS Microbiol Lett 222:9–16 [View Article][PubMed]
    [Google Scholar]
  31. Maiden M. C., Bygraves J. A., Feil E., Morelli G., Russell J. E., Urwin R., Zhang Q., Zhou J., Zurth K. et al. & other authors ( 1998). Multilocus sequence typing: a portable approach to the identification of clones within populations of pathogenic microorganisms. Proc Natl Acad Sci U S A 95:3140–3145 [View Article][PubMed]
    [Google Scholar]
  32. Margos G., Vollmer S. A., Cornet M., Garnier M., Fingerle V., Wilske B., Bormane A., Vitorino L., Collares-Pereira M. et al. & other authors ( 2009). A new Borrelia species defined by multilocus sequence analysis of housekeeping genes. Appl Environ Microbiol 75:5410–5416 [View Article][PubMed]
    [Google Scholar]
  33. Martens M., Delaere M., Coopman R., De Vos P., Gillis M., Willems A. ( 2007). Multilocus sequence analysis of Ensifer and related taxa. Int J Syst Evol Microbiol 57:489–503 [View Article][PubMed]
    [Google Scholar]
  34. Martens M., Dawyndt P., Coopman R., Gillis M., De Vos P., Willems A. ( 2008). Advantages of multilocus sequence analysis for taxonomic studies: a case study using 10 housekeeping genes in the genus Ensifer (including former Sinorhizobium). Int J Syst Evol Microbiol 58:200–214 [View Article][PubMed]
    [Google Scholar]
  35. Mellmann A., Cloud J., Maier T., Keckevoet U., Ramminger I., Iwen P., Dunn J., Hall G., Wilson D. et al. & other authors ( 2008). Evaluation of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry in comparison to 16S rRNA gene sequencing for species identification of nonfermenting bacteria. J Clin Microbiol 46:1946–1954 [View Article][PubMed]
    [Google Scholar]
  36. Nagy E., Maier T., Urban E., Terhes G., Kostrzewa M. ESCMID Study Group on Antimicrobial Resistance in Anaerobic Bacteria ( 2009). Species identification of clinical isolates of Bacteroides by matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry. Clin Microbiol Infect 15:796–802 [View Article][PubMed]
    [Google Scholar]
  37. Nishiyama T., Ueki A., Kaku N., Watanabe K., Ueki K. ( 2009). Bacteroides graminisolvens sp. nov., a xylanolytic anaerobe isolated from a methanogenic reactor treating cattle waste. Int J Syst Evol Microbiol 59:1901–1907 [View Article][PubMed]
    [Google Scholar]
  38. Pascual J., Macián M. C., Arahal D. R., Garay E., Pujalte M. J. ( 2010). Multilocus sequence analysis of the central clade of the genus Vibrio by using the 16S rRNA, recA, pyrH, rpoD, gyrB, rctB and toxR genes. Int J Syst Evol Microbiol 60:154–165 [View Article][PubMed]
    [Google Scholar]
  39. Posada D. ( 2008). jModelTest: phylogenetic model averaging. Mol Biol Evol 25:1253–1256 [View Article][PubMed]
    [Google Scholar]
  40. R Development Core Team( 2009). R: a language and environment for statistical computing. http://www.R-project.org
  41. Redondo M. C., Arbo M. D., Grindlinger J., Snydman D. R. ( 1995). Attributable mortality of bacteremia associated with the Bacteroides fragilis group. Clin Infect Dis 20:1492–1496 [View Article][PubMed]
    [Google Scholar]
  42. Rivas R., Martens M., de Lajudie P., Willems A. ( 2009). Multilocus sequence analysis of the genus Bradyrhizobium . Syst Appl Microbiol 32:101–110 [View Article][PubMed]
    [Google Scholar]
  43. Robert C., Chassard C., Lawson P. A., Bernalier-Donadille A. ( 2007). Bacteroides cellulosilyticus sp. nov., a cellulolytic bacterium from the human gut microbial community. Int J Syst Evol Microbiol 57:1516–1520 [View Article][PubMed]
    [Google Scholar]
  44. Saitou N., Nei M. ( 1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425[PubMed]
    [Google Scholar]
  45. Sakamoto M., Benno Y. ( 2006). Reclassification of Bacteroides distasonis, Bacteroides goldsteinii and Bacteroides merdae as Parabacteroides distasonis gen. nov., comb. nov., Parabacteroides goldsteinii comb. nov. and Parabacteroides merdae comb. nov. Int J Syst Evol Microbiol 56:1599–1605 [View Article][PubMed]
    [Google Scholar]
  46. Sakamoto M., Ohkuma M. ( 2010). Usefulness of the hsp60 gene for the identification and classification of Gram-negative anaerobic rods. J Med Microbiol 59:1293–1302 [View Article][PubMed]
    [Google Scholar]
  47. Sakamoto M., Suzuki M., Umeda M., Ishikawa I., Benno Y. ( 2002). Reclassification of Bacteroides forsythus (Tanner et al. 1986) as Tannerella forsythensis corrig., gen. nov., comb. nov. Int J Syst Evol Microbiol 52:841–849 [View Article][PubMed]
    [Google Scholar]
  48. Sakamoto M., Kitahara M., Benno Y. ( 2007). Parabacteroides johnsonii sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 57:293–296 [View Article][PubMed]
    [Google Scholar]
  49. Sakamoto M., Suzuki N., Benno Y. ( 2010). hsp60 and 16S rRNA gene sequence relationships among species of the genus Bacteroides with the finding that Bacteroides suis and Bacteroides tectus are heterotypic synonyms of Bacteroides pyogenes . Int J Syst Evol Microbiol 60:2984–2990 [View Article][PubMed]
    [Google Scholar]
  50. Sawabe T., Kita-Tsukamoto K., Thompson F. L. ( 2007). Inferring the evolutionary history of vibrios by means of multilocus sequence analysis. J Bacteriol 189:7932–7936 [View Article][PubMed]
    [Google Scholar]
  51. Schmidt H. A., Strimmer K., Vingron M., von Haeseler A. ( 2002). TREE-PUZZLE: maximum likelihood phylogenetic analysis using quartets and parallel computing. Bioinformatics 18:502–504 [View Article][PubMed]
    [Google Scholar]
  52. Sears C. L. ( 2009). Enterotoxigenic Bacteroides fragilis: a rogue among symbiotes. Clin Microbiol Rev 22:349–369 [View Article][PubMed]
    [Google Scholar]
  53. Shah H. N., Olsen I., Bernard K., Finegold S. M., Gharbia S., Gupta R. S. ( 2009). Approaches to the study of the systematics of anaerobic, Gram-negative, non-sporeforming rods: current status and perspectives. Anaerobe 15:179–194 [View Article][PubMed]
    [Google Scholar]
  54. Shimodaira H. ( 2002). An approximately unbiased test of phylogenetic tree selection. Syst Biol 51:492–508 [View Article][PubMed]
    [Google Scholar]
  55. Shimodaira H., Hasegawa M. ( 1999). Multiple comparisons of log-likelihoods with applications to phylogenetic inference. Mol Biol Evol 16:1114–1116 [CrossRef]
    [Google Scholar]
  56. Shimodaira H., Hasegawa M. ( 2001). CONSEL: for assessing the confidence of phylogenetic tree selection. Bioinformatics 17:1246–1247 [View Article][PubMed]
    [Google Scholar]
  57. Snydman D. R., Jacobus N. V., McDermott L. A., Ruthazer R., Golan Y., Goldstein E. J. C., Finegold S. M., Harrell L. J., Hecht D. W. et al. & other authors ( 2007). National survey on the susceptibility of Bacteroides fragilis group: report and analysis of trends in the United States from 1997 to 2004. Antimicrob Agents Chemother 51:1649–1655 [View Article][PubMed]
    [Google Scholar]
  58. Song Y. L., Liu C. X., McTeague M., Finegold S. M. ( 2004). “Bacteroides nordii” sp. nov. and “Bacteroides salyersae” sp. nov. isolated from clinical specimens of human intestinal origin. J Clin Microbiol 42:5565–5570 [View Article][PubMed]
    [Google Scholar]
  59. Song Y., Liu C., Bolanos M., Lee J., McTeague M., Finegold S. M. ( 2005). Evaluation of 16S rRNA sequencing and reevaluation of a short biochemical scheme for identification of clinically significant Bacteroides species. J Clin Microbiol 43:1531–1537 [View Article][PubMed]
    [Google Scholar]
  60. Stackebrandt E., Frederiksen W., Garrity G. M., Grimont P. A. D., Kämpfer P., Maiden M. C. J., Nesme X., Rosselló-Mora R., Swings J. et al. & other authors ( 2002). Report of the ad hoc committee for the re-evaluation of the species definition in bacteriology. Int J Syst Evol Microbiol 52:1043–1047 [View Article][PubMed]
    [Google Scholar]
  61. Tamura K., Dudley J., Nei M., Kumar S. ( 2007). MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599 [View Article][PubMed]
    [Google Scholar]
  62. Tanigawa K., Watanabe K. ( 2011). Multilocus sequence typing reveals a novel subspeciation of Lactobacillus delbrueckii . Microbiology 157:727–738 [View Article][PubMed]
    [Google Scholar]
  63. Thompson F. L., Gevers D., Thompson C. C., Dawyndt P., Naser S., Hoste B., Munn C. B., Swings J. ( 2005). Phylogeny and molecular identification of vibrios on the basis of multilocus sequence analysis. Appl Environ Microbiol 71:5107–5115 [View Article][PubMed]
    [Google Scholar]
  64. Thompson F. L., Gomez-Gil B., Vasconcelos A. T. R., Sawabe T. ( 2007). Multilocus sequence analysis reveals that Vibrio harveyi and V. campbellii are distinct species. Appl Environ Microbiol 73:4279–4285 [View Article][PubMed]
    [Google Scholar]
  65. Whitehead T. R., Cotta M. A., Collins M. D., Falsen E., Lawson P. A. ( 2005). Bacteroides coprosuis sp. nov., isolated from swine-manure storage pits. Int J Syst Evol Microbiol 55:2515–2518 [View Article][PubMed]
    [Google Scholar]
  66. Yamamoto S., Harayama S. ( 1995). PCR amplification and direct sequencing of gyrB genes with universal primers and their application to the detection and taxonomic analysis of Pseudomonas putida strains. Appl Environ Microbiol 61:1104–1109[PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.052332-0
Loading
/content/journal/micro/10.1099/mic.0.052332-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error