Chapter 43 : Molecular Tools for Epidemiological Investigations into Infections

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This chapter describes the theoretical framework within which typing networks can be established. The definitions used for identifying bacterial strains and clones will be provided, and the clinical value of typing will be illustrated as well. The chapter discusses the current tools available for molecular typing of and their reproducibility. Some of the current developments in the field of bacterial genetic identification are pinpointed, and ongoing and future developments in the field of establishing typing networks are summarized. The most common overall methodological distinction is the one between phenotyping and genotyping procedures. Genes can be specifically amplified and analyzed for sequence variability on the basis of restriction site polymorphism or variable length of the PCR products. Random amplification of polymorphic DNA (RAPD) analysis and arbitrary primed PCR are methods that are essentially based on random priming of PCR primers, often using quite relaxed reaction conditions. Amplification fragment length polymorphism (AFLP) analysis is another recent PCR-mediated procedure with which subsets of DNA restriction fragments can be successfully amplified. DNA sequencing technology has revolutionized microbiology in general because it enabled whole genome sequencing for a variety of microorganisms. Multilocus sequence typing (MLST) is one example: the sequence polymorphism detected in a number of slowly evolving genes allows for the categorization of strains on the basis of allelic diversity. Future technology improvement may increase the self-sustained development of exchangeable data subdirectories in individual laboratories, but we still have to go along way before we get there.

Citation: van Belkum A. 2002. Molecular Tools for Epidemiological Investigations into Infections, p 227-236. In Marre R, Abu Kwaik Y, Bartlett C, Cianciotto N, Fields B, Frosch M, Hacker J, Lück P (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555817985.ch43
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Image of FIGURE 1

RAPD analysis of reference strains and isolates obtained during water contamination or nosocomial outbreaks of infection. A combination of two primers (ERIC2 and BG2) was applied during amplification. Lanes: A, NCTC12008-OLDA; B, NCTC11404-Bellingham; C, Hospital B, clinical isolate; D, water isolate hospital A; E through H, patient isolates obtained in hospital A. Patients F, G, and ? were hospitalized simultaneously. The identity of the RAPD fingerprints suggests an ongoing outbreak of infections involving patients F, G, and H. Note, however, that not all patients who are epidemiologically linked are infected by the same strain (lane E).

Citation: van Belkum A. 2002. Molecular Tools for Epidemiological Investigations into Infections, p 227-236. In Marre R, Abu Kwaik Y, Bartlett C, Cianciotto N, Fields B, Frosch M, Hacker J, Lück P (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555817985.ch43
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Image of FIGURE 2

Principle of AFLP (Keygene NV, Wageningen, The Netherlands) and example of its experimental output. Panel A delineates the various stages in the typing protocol. Initially, DNA is digested with the help of two different restriction enzymes. Specific linkers are coupled to the respective sticky ends and these linkers serve as specific anchor points for fragment amplification by PCR. Inclusion of selective bases (encoded in the neighboring region of the native restriction sites) in the primer sequences results in amplification of limited numbers of specific restriction fragments. These can be effectively separated by acryl amide gel electrophoresis and visualized by techniques for detection of haptens incorporated in primers or dNTPs used during amplification (panel B).

Citation: van Belkum A. 2002. Molecular Tools for Epidemiological Investigations into Infections, p 227-236. In Marre R, Abu Kwaik Y, Bartlett C, Cianciotto N, Fields B, Frosch M, Hacker J, Lück P (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555817985.ch43
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1. Benkel, D. H.,, E. M. McClure,, D. Woolard,, J. V. Rullan,, G. B. Miller,, S. R. Jenkins,, J. H. Hershey,, R. F. Benson,, J. M. Pruckler,, E. W. Brown,, M. S. Kolczak,, R. L. Hackler,, B. S. Rouse,, and R. F. Breiman. 2000. Outbreak of Legionnaires' disease associated with a display whirlpool spa. Int. J. Epidemiol. 29: 1092 1098.
2. Den Boer, J. W.,, E. Yzerman,, A. van Belkum,, F. Vlaspolder,, and F. J. M. van Breukelen. 1998. Legionnaire's disease and saunas. Lancet 351: 1056.
3. Formica, N.,, G. Tallis,, B. Zwolak,, J. Camie,, M. Beers,, G. Hogg,, N. Ryan,, and M. Yates. 2000. Legionnaires' Disease outbreak: Victoria's largest identified outbreak. Commun. Dis. Intell. 24: 199 202.
4. Fry, N. K.,, J. M. Bangsborg,, S. Bernarder,, J. Etienne,, B. Forsblom,, V. Gaia,, P. Hasenberger,, D. Lindsay,, A. Papoutsi,, C. Pelaz,, M. Struelens,, S. A. Uldum,, P. Visca,, and T. G. Harrison. 2000. Assessment of intercentre reproducibility and epidemiological concordance of Legionella pneumophila serogroup 1 genotyping by amplified fragment length polymorphism analysis. Eur. J. Clin. Microbiol. Infect. Dis. 19: 773 780.
5. Fry, N. K.,, S. Alexiou-Daniel,, J. M. Bangsborg,, S. Bernarder,, M. Castellani Pastoris,, J. Etienne,, B. Forsblom,, V. Gaia,, J. H. Helbig,, D. Lindsay,, P. C. Luck,, C. Pelaz,, S. A. Uldum,, and T. G. Harrison. 1999. A multicenter evaluation of genotypic methods for the epidemiologic typing of Legionella pneumophila serogroup 1: results of a pan-European study. Clin. Microbiol. Infect. 5: 462 477.
6. Gomez-Lus, P.,, B. S. Fields,, R. F. Benson,, W. T. Martin,, S. P. O'Connor,, and C. M. Black. 1993. Comparison of arbitrary primed polymerase chain reaction, ribotyping and monoclonal antibody analysis for subtyping Legionella pneumophila serogroup 1. J. Clin. Microbiol. 31: 1940 1942.
7. Joly, J. R.,, R. M. McKinney,, J. O. Tobin,, J. Bibb,, I. D. Watkins,, and D. Ramsay. 1986. Development of a standardised subgrouping scheme for Legionella pneumophila serogroup 1 using monoclonal antibodies. J. Clin. Microbiol. 23: 768 771.
8. Jonas, D.,, H. G. W. Meyer,, P. Matthes,, D. Hartung,, B. Jahn,, F. D. Daschner,, and B. Jansen. 2000. Comparative evaluation of three different genotyping methods for investigation of nosocomial outbreaks of Legionnaire's Disease in hospitals. J. Clin. Microbiol. 38: 2284 2291.
9. Knirsch, C. A.,, K. Jakob,, D. Schoonmaker,, J. A. Kiehlbauch,, S. J. Wong,, P. Delta-Latta,, S. Whittier,, M. Layton,, and B. Scully. 2000. An outbreak of Legionella micdadei pneumonia in transplant patients: evaluation, molecular epidemiology and control. Am. J. Med. 108: 290 295.
10. Kremer, K.,, D. van Soolingen,, R. Frothingham,, W. H. Haas,, P. W. M. Hermans,, C. Martin,, P. Palittapnogarnpim,, B. B. Plikaytis,, L. W. Riley,, M. A. Yakrus,, J. M. Musser,, and J. D. A. van Embden. 1999. Comparison of methods based on different molecular epidemiological markers for typing of Mycobacterium tuberculosis complex strains: interlaboratory study of discriminatory power and reproducibility. J. Clin. Microbiol. 37: 2607 2618.
11. McEvoy, M.,, N. Batchelor,, G. Hamilton,, A. MacDonald,, M. Faiers,, A. Sills,, J, Lee,, and T. Harrison. 2000. A cluster of Legionnaires' disease associated with exposure to a spa pool on display. Commun. Dis. Public Health 3: 43 45.
12. Nakachi, N.,, K. Matsunaga,, T. W. Klein,, H. Friedman,, and Y. Yamamoto. 2000. Differential effect of virulent versus avirulent Legionella pneumophila on chemokine gene expression in murine alveolar macrophages determined by cDNA expression array technique. Infect. Immun. 68: 6069 6072.
13. Saunders, N. A.,, T. G. Harrison,, T. Haththotuwa,, and A. G. Taylor. 1991. A comparison of probes for restriction fragment length polymorphism (RFLP) typing of Legionella pneumophila serotype 1 strains. J. Med. Microbiol. 35: 152 158.
14. Sessa, R.,, M. Di Pietro,, M. Zamparelli,, and M. Del Piano. 2000. Biofilm formation on the surface of ceramic tiles. New Microbiol. 23: 407 413.
15. Schoonmaker, D.,, T. Heimberger,, and G. Birkhead. 1992. Comparison of ribotyping and restriction enzyme analysis using pulsed field gel electrophoresis for distinguishing Legionella pneumophila isolates obtained during a nosocomial outbreak. J. Clin. Microbiol. 30: 1491 1498.
16. Struelens, M. J.,, N. Maes,, F. Rost,, A. Deplano,, F. Jacobs,, C. Liesnard,, N. Bornstein,, F. Grimont,, S. Lauwers,, M. P. Mclntyre,, and E. Serruys. 1992. Genotypic and phenotypic methods for the investigation of a nosocomial Legionella pneumophila outbreak and efficacy of control measures. J. Infect. Dis. 166: 22 30.
17. Tenover, F. C.,, R. Arbeit,, G. Archer,, J. Biddle,, S. Byrne,, R. Goering,, G. Hancock,, G. A. Hebert,, B. Hill,, R. Hollis,, W. J. Jarvis,, B. Kreiswirth,, B. Eisner,, J. Maslow,, L. K. MacDougal,, M. Miller,, M. Mulligan,, and M. A. Pfaller. 1994. Comparison of traditional and molecular methods for typing isolates of Staphylococcus aureus. J. Clin. Microbiol. 32: 407 415.
18. Tompkins, L. S.,, N. J. Troup,, T. Woods,, W. F. Bibb,, and R. M. McKinney. 1987. Molecular epidemiology of Legionella species by restriction endonuclease and alloenzyme analysis J. Clin. Microbiol. 25: 1875 1880.
19. Van Belkum, A.,, H. Maas,, H. Verbrugh,, and N. van Leeuwen. 1996. Serotyping, ribotyping, PCR-mediated ribosomal 16S-23S spacer analysis and arbitrarily primed PCR for epidemiological studies on Legionella pneumophila. Res. Microbiol. 147: 405 413.
20. Van Belkum, A.,, J. Kluytmans,, W. van Leeuwen,, R. Bax,, W. Quint,, E. Peters,, A. Fluit,, C. Vandenbroucke-Grauls,, A. van den Brule,, H. Koeleman,, W. Melchers,, J. Meis,, A. Elaichouni,, M. Vaneechoutte,, F. Moonens,, N. Maes,, M. Struelens,, F. Tenover,, and H. A. Verbrugh. 1995. Multicenter evaluation of arbitrarily primed PCR for typing of Staphylococcus aureus strains. J. Clin. Microbiol. 33: 1537 1547.
21. Van Belkum, A.,, M. Struelens,, and W. G. V. Quint. 1993. Typing of Legionella pneumophila strains by polymerase chain reaction mediated DNA fingerprinting. J. Clin. Microbiol. 31: 2198 2200.
22. Van Belkum, A.,, W. van Leeuwen,, M. E. Kaufman,, B. Cookson,, F. Forey,, J. Etienne,, R. Goering,, F. Tenover,, C. Steward,, F. O'Brien,, W. Grubb,, P. Tassios,, N. Legakis,, N. Morvan,, N. El Solh,, R. de Ryck,, M. Struelens,, S. Salmenlinna,, J. Vuopio-Varkila,, M. Kooistra,, A. Talens,, W. Witte,, and H. A. Verbrugh. 1998. Assessment of resolution and intercenter reproducibility of results of genotyping of Staphylococcus aureus by pulsed field gel electrophoresis of SmaI macrorestriction fragments: a multicenter study. J. Clin. Microbiol. 36: 1653 1659.
23. Van Leeuwen, W.,, H. Verbrugh,, J. van der Velde,, N. van Leeuwen,, M. Heck,, and A. van Belkum. 1999. Validation of binary typing for Staphylococcus aureus strains. J. Clin. Microbiol. 37: 664 674.
24. Van Leeuwen, W.,, M. Sijmons,, J. Sluijs,, H. A. Verbrugh,, and A. van Belkum. 1996. On the nature and use of randomly amplified DNA from Staphylococcus aureus. J. Clin. Microbiol. 34: 2770 2777.
25. Vogel, J. P.,, H. L. Andrews,, S. K. Wong,, and R. R. Isberg. 1998. Conjugative transfer by the virulence system of Legionella pneumophila. Science 279: 873 876.
26. Wever, P. C.,, E. P. F. Yzerman,, E. J. Kuiper,, P. Speelman,, and J. Dankert. 2000. Rapid diagnosis of Legionnaire's Disease using an immunochromatographic assay for Legionella pneumophila serogroup 1 antigen in urine during an outbreak in The Netherlands. J. Clin. Microbiol. 38: 2738 2739.


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Frequency of usage of various typing procedures for epidemiological analysis of isolates: a recent historical perspective

Citation: van Belkum A. 2002. Molecular Tools for Epidemiological Investigations into Infections, p 227-236. In Marre R, Abu Kwaik Y, Bartlett C, Cianciotto N, Fields B, Frosch M, Hacker J, Lück P (ed), . ASM Press, Washington, DC. doi: 10.1128/9781555817985.ch43

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