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Multilocus sequence typing

Data analysis in clinical microbiology and public health

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Abstract

Numerous computer-based statistical packages have been developed in recent years and it has become easier to analyze nucleotide sequence data and gather subsequent information that would not normally be available. Multilocus sequence typing (MLST) is used for characterizing isolates of bacterial and fungal species and uses nucleotide sequences of internal fragments of housekeeping genes. This method is finding a place in clinical microbiology and public health by providing data for epidemiological surveillance and development of vaccine policy. It adds greatly to our knowledge of the genetic variation that can occur within a species and has therefore been used for studies of population biology. Analysis requires the detailed interpretation of nucleotide sequence data obtained from housekeeping and nonhousekeeping genes. This is due to the amount of data generated from nucleotide sequencing and the information generated from an array of analytical tools improves our understanding of bacterial pathogens. This can benefit public health interventions and the development of enhanced therapies and vaccines. This review concentrates on the analytical tools used in MLST and their use in the clinical microbiology and public health fields.

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References

  1. Enright, M. C. and Spratt, B. G. (1998) A multilocus sequence typing scheme for Streptococcus pneumoniae: identification of clones associated with serious invasive disease. Microbiology 144 (Pt 11), 3049–3060.

    Article  PubMed  CAS  Google Scholar 

  2. Maiden, M. C., Bygraves, J. A., Feil, E., Morelli, G., Russell, J. E., Urwin, R., Zhang, Q., Zhou, J., Zurth, K., Caugant, D. A., Feavers, I. M., Achtman, M., and Spratt, B. G. (1998) Multilocus sequence typing: a portable approach to the identification of clones within populations of pathogenic microorganisms. Proc. Natl. Acad. Sci. USA 95, 3140–3145.

    Article  PubMed  CAS  Google Scholar 

  3. Enright, M. C. and Spratt, B. G. (1999) Multilocus sequence typing. Trends Microbiol. 7, 482–487.

    Article  PubMed  CAS  Google Scholar 

  4. Urwin, R. and Maiden, M. C. (2003) Multi-locus sequence typing: a tool for global epidemiology. Trends Microbiol. 11, 479–487.

    Article  PubMed  CAS  Google Scholar 

  5. van Leeuwen, W. B., Jay, C., Snijders, S., Durin, N., Lacroix, B., Verbrugh, H. A., Enright, M. C., Troesch, A., and van Belkum, A. (2003) Multilocus sequence typing of Staphylococcus aureus with DNA array technology. J. Clin. Microbiol. 41, 3323–3326.

    Article  PubMed  CAS  Google Scholar 

  6. Dingle, K. E., Colles, F. M., Wareing, D. R., Ure, R., Fox, A. J., Bolton, F. E., Bootsma, H. J., Willems, R. J., Urwin, R., and Maiden, M. C. (2001) Multilocus sequence typing system for Campylobacter jejuni. J. Clin. Microbiol. 39, 14–23.

    Article  PubMed  CAS  Google Scholar 

  7. Enright, M. C., Spratt, B. G., Kalia, A., Cross, J. H., and Bessen, D.E. (2001) Multilocus sequence typing of Streptococcus pyogenes and the relationships between emm type and clone. Infect. Immunol. 69, 2416–2427.

    Article  CAS  Google Scholar 

  8. Meats, E., Feil, E. J., Stringer, S., Cody, A. J., Goldstein, R., Kroll, J. S., Popovic, T., and Spratt, B. G. (2003) Characterization of encapsulated and noncapsulated Haemophilus influenzae and determination of phylogenetic relationships by multilocus sequence typing. J. Clin. Microbiol. 41, 1623–1636.

    Article  PubMed  CAS  Google Scholar 

  9. Robles, J. C., Koreen, L., Park, S., and Perlin, D. S. (2004) Multilocus sequence typing is a reliable alternative method to DNA fingerprinting for discriminating among strains of Candida albicans. J. Clin. Microbiol. 42, 2480–2488.

    Article  PubMed  CAS  Google Scholar 

  10. Bygraves, J. A., Urwin, R., Fox, A. J., Gray, S. J., Russell, J. E., Feavers, I. M., and Maiden, M. C. (1999) Population genetic and evolutionary approaches to analysis of Neisseria meningitidis isolates belonging to the ET-5 complex. J. Bacteriol. 181, 5551–5556.

    PubMed  CAS  Google Scholar 

  11. Feavers, I. M., Gray, S. J., Urwin, R., Russell, J. E., Bygraves, J. A., Kaczmarski, E. B., and Maiden, M. C. (1999) Multilocus sequence typing and antigen gene sequencing in the investigation of a meningococcal disease outbreak. J. Clin. Microbiol. 37, 3883–3887.

    PubMed  CAS  Google Scholar 

  12. Clarke, S. C., Diggle, M. A., and Edwards, G. F. (2002) Multilocus sequence typing and porA gene sequencing differentiates strains of Neisseria meningitidis during case clusters. Br. J. Biomed. Sci. 59, 160–162.

    PubMed  CAS  Google Scholar 

  13. Clarke, S. C., Diggle, M. A., Molling, P., Unemo, M., and Olcen, P. (2003) Analysis of PorA variable region 3 in meningococci: implications for vaccine policy? Vaccine 21, 2468–2473.

    Article  PubMed  CAS  Google Scholar 

  14. Wisplinghoff, H., Rosato, A. E., Enright, M. C., Noto, M., Craig, W., and Archer, G. L. (2003) Related clones containing SCCmec type IV predominate among clinically significant Staphylococcus epidermidis isolates. Antimicrob. Agents Chemother. 47, 3574–3579.

    Article  PubMed  CAS  Google Scholar 

  15. Feil, E. J. (2004) Small change: keeping pace with microevolution. Nat. Rev. Microbiol. 2, 483–495.

    Article  PubMed  CAS  Google Scholar 

  16. Chan, M. S., Maiden, M. C., and Spratt, B. G. (2001) Database-driven multi locus sequence typing (MLST) of bacterial pathogens. Bioinformatics 17, 1077–1083.

    Article  PubMed  CAS  Google Scholar 

  17. Jolley, K. A., Chan, M. S., and Maiden, M. C. (2004) mlstdbNet—distributed multi-locus sequence typing (MLST) databases. BMC Bioinformatics 5, 86.

    Article  PubMed  CAS  Google Scholar 

  18. Jolley, K. A., Feil, E. J., Chan, M. S., and Maiden, M. C. (2001) Sequence type analysis and recombinational tests (START). Bioinformatics 17, 1230–1231.

    Article  PubMed  CAS  Google Scholar 

  19. Sreevatsan, S., Pan, X., Stockbauer, K. E., Connell, N. D., Kreiswirth, B. N., Whittam, T. S., and Musser, J. M. (1997) Restricted structural gene polymorphism in the Mycobacterium tuberculosis complex indicates evolutionarily recent global dissemination. Proc. Natl. Acad. Sci. USA 94, 9869–9874.

    Article  PubMed  CAS  Google Scholar 

  20. Achtman, M., Zurth, K., Morelli, G., Torrea, G., Guiyoule, A., and Carniel, E. (1999) Yersinia pestis, the cause of plague, is a recently emerged clone of Yersinia pseudotuberculosis. Proc. Natl. Acad. Sci. USA 96, 14043–14048.

    Article  PubMed  CAS  Google Scholar 

  21. Clarke, S. C., Diggle, M. A., and Edwards, G. F. (2001) Semiautomation of multilocus sequence typing for the characterization of clinical isolates of Neisseria meningitidis. J. Clin. Microbiol. 39, 3066–3071.

    Article  PubMed  CAS  Google Scholar 

  22. Clarke, S. C. (2002) Nucleotide sequence-based typing of bacteria and the impact of automation. Bioessays 24, 858–862.

    Article  PubMed  CAS  Google Scholar 

  23. Jefferies, J., Clarke, S. C., Diggle, M. A., Smith, A., Dowson, C., and Mitchell, T. (2003) Automated pneumococcal MLST using liquid-handling robotics and a capillary DNA sequencer. Mol. Biotechnol. 24, 303–308.

    Article  PubMed  CAS  Google Scholar 

  24. Diggle, M. A. and Clarke, S. C. (2002) What a load of old sequence! J. Clin. Microbiol. 40, 2707.

    Article  PubMed  Google Scholar 

  25. Nicolas, P., Decousset, L., Riglet, V., Castelli, P., Stor, R., and Blanchet, G. (2001) Clonal expansion of sequence type (ST-)5 and emergence of ST-7 in serogroup A meningococci, Africa. Emerg. Infect. Dis. 7, 849–854.

    Article  PubMed  CAS  Google Scholar 

  26. Eldridge, J., Sutcliffe, E. M., Abbott, J. D., and Jones, D. M. (1978) Serological grouping of meningococci and detection of antigen in cerebrospinal fluid by coagglutination. Med. Lab. Sci. 35, 63–66.

    PubMed  CAS  Google Scholar 

  27. Frasch, C. E., Zollinger, W. D., and Poolman, J. T. (1985) Serotype antigens of Neisseria meningitidis and a proposed scheme for designation of serotypes. Rev. Infect. Dis. 7, 504–510.

    PubMed  CAS  Google Scholar 

  28. Smart, L. E. (1986) Serotyping of Streptococcus pneumoniae strains by coagglutination. J. Clin Pathol. 39, 328–331.

    PubMed  CAS  Google Scholar 

  29. Ni, H., Knight, A. I., Cartwright, K., Palmer, W. H., and McFadden, J. (1992) Polymerase chain reaction for diagnosis of meningococcal meningitis. Lancet 340, 1432–1434.

    Article  PubMed  CAS  Google Scholar 

  30. Kristiansen, B.E., Ask, E., Jenkins, A., Fermer, C., Radstrom, P., and Skold, O. (1991) Rapid diagnosis of meningococcal meningitis by polymerase chain reaction. Lancet 337, 1568–1569.

    Article  PubMed  CAS  Google Scholar 

  31. Maiden, M.C. and Stuart, J.M. (2002) Carriage of serogroup C meningococci 1 year after meningococcal C conjugate polysaccharide vaccination. Lancet 359, 1829–1831.

    Article  PubMed  Google Scholar 

  32. Zhou, J., Enright, M. C., and Spratt, B. G. (2000) Identification of the major Spanish clones of penicillin-resistant pneumococci via the Internet using multilocus sequence typing. J. Clin. Microbiol. 38, 977–986.

    PubMed  CAS  Google Scholar 

  33. Martin, I. M., Ison, C. A., Aanensen, D. M., Fenton, K. A., and Spratt, B. G. (2004) Rapid sequence-based identification of gonococcal transmission clusters in a large metropolitan area. J. Infect. Dis. 189, 1497–1505.

    Article  PubMed  CAS  Google Scholar 

  34. Bougnoux, M.E., Morand, S., and d’Enfert, C. (2002) Usefulness of multilocus sequence typing for characterization of clinical isolates of Candida albicans. J. Clin. Microbiol. 40, 1290–1297.

    Article  PubMed  CAS  Google Scholar 

  35. Bougnoux, M. E., Tavanti, A., Bouchier, C., Gow, N. A., Magnier, A., Davidson, A. D., Maiden, M. C., D’Enfert, C., and Odds, F. C. (2003) Collaborative consensus for optimized multilocus sequence typing of Candida albicans. J. Clin. Microbiol. 41, 5265–5266.

    Article  PubMed  CAS  Google Scholar 

  36. Tavanti, A., Gow, N. A., Senesi, S., Maiden, M. C., and Odds, F. C. (2003) Optimization and validation of multilocus sequence typing for Candida albicans. J. Clin. Microbiol. 41, 3765–3776.

    Article  PubMed  CAS  Google Scholar 

  37. Staden, R. (1996) The Staden sequence analysis package. Mol. Biotechnol. 5, 233–241.

    Article  PubMed  CAS  Google Scholar 

  38. Staden, R., Beal, K. F., and Bonfield, J. K. (2000) The Staden package, 1998. Methods Mol. Biol. 132, 115–130.

    PubMed  CAS  Google Scholar 

  39. Diggle, M. A. and Clarke, S. C. (2002) Rapid assignment of nucleotide sequence data to allele types for multi-locus sequence analysis (MLSA) of bacteria using an adapted database and modified alignment program. J. Mol. Microbiol. Biotechnol. 4, 515–517.

    PubMed  CAS  Google Scholar 

  40. Clarke, S. C., Lawrie, D. I., and Diggle, M. A. (2004) Genetic relatedness of antibiotic-resistant pneumococci isolated during case clusters. J. Med. Microbiol. 53, 1097–1099.

    Article  PubMed  CAS  Google Scholar 

  41. Caugant, D. A., Mocca, L. F., Frasch, C. E., Froholm, L. O., Zollinger, W. D., and Selander, R. K. (1987) Genetic structure of Neisseria meningitidis populations in relation to serogroup, serotype, and outer membrane protein pattern. J. Bacteriol. 169, 2781–2792.

    PubMed  CAS  Google Scholar 

  42. Feil, E. J., Cooper, J. E., Grundmann, H., Robinson, D. A., Enright, M. C., Berendt, T., Peacock, S. J., Smith, J. M., Murphy, M., Spratt, B. G., Moore, C. E., and Day, N. P. (2003) How clonal is Staphylococcus aureus? J. Bacteriol. 185, 3307–3316.

    Article  PubMed  CAS  Google Scholar 

  43. Feil, E. J., Holmes, E. C., Bessen, D. E., Chan, M. S., Day, N. P., Enright, M. C., Goldstein, R., Hood, D. W., Kalia, A., Moore, C. E., Zhou, J., and Spratt, B. G. (2001) Recombination within natural populations of pathogenic bacteria: short-term empirical estimates and long-term phylogenetic consequences. Proc. Natl. Acad. Sci. USA 98, 182–187.

    Article  PubMed  CAS  Google Scholar 

  44. Feil, E. J., Li, B. C., Aanensen, D. M., Hanage, W. P., and Spratt, B. G. (2004) eBURST: inferring patterns of evolutionary descent among clusters of related bacterial genotypes from multilocus sequence typing data. J. Bacteriol. 186, 1518–1530.

    Article  PubMed  CAS  Google Scholar 

  45. Mokrousov, I., Narvskaya, O., Limeschenko, E., Vyazovaya, A., Otten, T., and Vyshnevskiy, B. (2004) Analysis of the allelic diversity of the mycobacterial interspersed repetitive units in Mycobacterium tuberculosis strains of the Beijing family: practical implications and evolutionary considerations. J. Clin. Microbiol. 42, 2438–2444.

    Article  PubMed  CAS  Google Scholar 

  46. Sugita, T., Ichikawa, T., Matsukura, M., Sueda, M., Takashima, M., Ikeda, R., Nishikawa, A., and Shinoda, T. (2001) Genetic diversity and biochemical characteristics of Trichosporon asahii isolated from clinical specimens, houses of patients with summer-type-hypersensitivity pneumonitis, and environmental materials. J. Clin. Microbiol. 39, 2405–2411.

    Article  PubMed  CAS  Google Scholar 

  47. Barsotti, O., Decoret, D., and Renaud, F. N. (2002) Identification of Streptococcus mitis group species by RFLP of the PCR-amplified 16S–23S rDNA intergenic spacer. Res. Microbiol. 153, 687–691.

    Article  PubMed  CAS  Google Scholar 

  48. Enright, M. C. and Spratt, B. G. (1999) Extensive variation in the ddl gene of penicillin-resistant Streptococcus pneumoniae results from a hitchhiking effect driven by the penicillin-binding protein 2b gene. Mol. Biol. Evol. 16, 1687–1695.

    PubMed  CAS  Google Scholar 

  49. Drouin, G., Prat, F., Ell, M., and Clarke, G. D. (1999) Detecting and characterizing gene conversions between multigene family members. Mol. Biol. Evol. 16, 1369–1390.

    PubMed  CAS  Google Scholar 

  50. Sawyer, S. (1989) Statistical tests for detecting gene conversion. Mol. Biol. Evol. 6, 526–538.

    PubMed  CAS  Google Scholar 

  51. Maynard Smith, J. and Smith, N. H. (1998) Detecting recombination from gene trees. Mol. Biol. Evol. 15, 590–599.

    PubMed  CAS  Google Scholar 

  52. Maynard-Smith, J., Smith, N. H., O’Rourke, M., and Spratt, B. G. (1993) How clonal are bacteria? Proc. Natl. Acad. Sci. USA 90, 4384–4388.

    Article  Google Scholar 

  53. Diggle, M. A. and Clarke, S. C. (2003) Detection and genotyping of meningococci using a nested PCR approach. J. Med. Microbiol. 52, 51–57.

    Article  PubMed  CAS  Google Scholar 

  54. Lewis, C., Diggle, M. A., and Clarke, S. C. (2003) Nucleotide sequence analysis of the sialyl-transferase gene of meningococcal serogroups B, C, y and w135. J. Mol. Microbiol. Biotechnol. 5, 82–86.

    Article  PubMed  CAS  Google Scholar 

  55. Urwin, R., Kaczmarski, E. B., Guiver, M., Fox, A. J., and Maiden, M. C. (1998) Amplification of the meningococcal porB gene for non-culture serotype characterization. Epidemiol. Infect. 120, 257–262.

    Article  PubMed  CAS  Google Scholar 

  56. Iannelli, F., Oggioni, M. R., and Pozzi, G. (2002) Allelic variation in the highly polymorphic locus pspC of Streptococcus pneumoniae. Gene 284, 63–71.

    Article  PubMed  CAS  Google Scholar 

  57. Enright, M. C., Day, N. P., Davies, C. E., Peacock, S. J., and Spratt, B. G. (2000) Multilocus sequence typing for characterization of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus. J. Clin. Microbiol. 38, 1008–1015.

    PubMed  CAS  Google Scholar 

  58. Frenay, H. M., Bunschoten, A. E., Schouls, L. M., van Leeuwen, W. J., Vandenbroucke-Grauls, C. M., Verhoef, J., and Mooi, F. R. (1996) Molecular typing of methicillin-resistant Staphylococcus aureus on the basis of protein A gene polymorphism. Eur. J. Clin. Microbiol. Infect. Dis. 15, 60–64.

    Article  PubMed  CAS  Google Scholar 

  59. Montesinos, I., Salido, E., Delgado, T., Cuervo, M., and Sierra, A. (2002) Epidemiologic genotyping of methicillin-resistant Staphylococcus aureus by pulsed-field gel electrophoresis at a university hospital and comparison with antibiotyping and protein A and coagulase gene polymorphisms. J. Clin. Microbiol. 40, 2119–2125.

    Article  PubMed  CAS  Google Scholar 

  60. Oliveira, D. C., Crisostomo, I., Santos-Sanches, I., Major, P., Alves, C. R., Aires-de-Sousa, M., Thege, M. K., and de Lencastre, H. (2001) Comparison of DNA sequencing of the protein A gene polymorphic region with other molecular typing techniques for typing two epidemiologically diverse collections of methicillin-resistant Staphylococcus aureus. J. Clin. Microbiol. 39, 574–580.

    Article  PubMed  CAS  Google Scholar 

  61. Luneberg, E., Glenn-Calvo, E., Hartmann, M., Bar, W., and Frosch, M. (1998) The central, surface-exposed region of the flagellar hook protein FlgE of Campylobacter jejuni shows hypervariability among strains. J. Bacteriol. 180, 3711–3714.

    PubMed  CAS  Google Scholar 

  62. Vitali, L. A., Zampaloni, C., Prenna, M., and Ripa, S. (2002) PCR m typing: a new method for rapid typing of group a streptococci. J. Clin. Microbiol. 40, 679–681.

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Scottish Meningococcus and Pneumococcus Reference Laboratory North Glasgow University Hospital NHS Trust, Glasgow, U.K.

    Christopher B. Sullivan, Matthew A. Diggle & Stuart C. Clarke

  2. Faculty of Biomedical and Life Sciences, University of Glasgow, U.K.

    Stuart C. Clarke

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  1. Christopher B. Sullivan
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  2. Matthew A. Diggle
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  3. Stuart C. Clarke
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Correspondence to Stuart C. Clarke.

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Sullivan, C.B., Diggle, M.A. & Clarke, S.C. Multilocus sequence typing. Mol Biotechnol 29, 245–254 (2005). https://doi.org/10.1385/MB:29:3:245

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