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Chapter 12.4 : Molecular Methods for Epidemiologic Typing of Microorganisms

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Abstract:

The ability to identify specific strains within a species of pathogen is an important aid in the rational development of effective measures to prevent and control nosocomial infections. The efforts of both microbiologists and hospital epidemiologists are facilitated greatly by the availability of the newer molecular epidemiologic typing techniques. The variety of molecular epidemiologic tools available at present is considerable, and based on current experience the methods that appear to be the most practical and useful for both large- and small-scale epidemiologic studies are the DNA-based methods such as pulsed-field gel electrophoresis (Table 12.1-5). Although these methods clearly have limitations, they generally are a significant improvement over the more conventional typing methods, many of which are too cumbersome, insensitive, and time-consuming to be of practical value for epidemiologic evaluations. It is important to understand that no single technique is universally applicable and that the choice for a particular application is related to the species studied, the scope of the question posed, and the convenience of the technique. The techniques of molecular epidemiology are useful in answering real clinical and infection control questions and are not limited to research uses. Examples include distinguishing between relapse and reinfection in an individual patient and in tracking the spread of an individual strain of a bacterium or fungus within the hospital environment ( ).

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
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Figures

Image of Figure 12.4.3-1
Figure 12.4.3-1

plasmid DNA preparations. Uncut, undigested plasmid DNA; RI, DNA digested with RI; dIII, plasmid DNA digested with dIII. Lanes: C, control strain SM818-73; 1, 2, and 3, isolates collected from a single patient. Isolate 3 does not have detectable plasmid DNA. The report for this group of isolates stated, “Based on restriction endonuclease analysis of plasmid DNA, these strains were grouped as follows: isolates 1 and 2 are identical to each other; isolate 3 is not related to isolates 1 and 2.”

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
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Image of Figure 12.4.4-1
Figure 12.4.4-1

Ribotype profiles of isolates. Lanes: S, molecular size standards; 1 and 6, ribotype A; 2 and 5, ribotype B; 3, ribotype C; 4 and 7, ribotype D. Figure from H. D. Isenberg (ed.). 1998. . ASM Press, Washington, DC.

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
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Image of Figure 12.4.5-1
Figure 12.4.5-1

Restriction fragment length polymorphism of chromosomal DNA digested with I. Switch time ramped from 10 to 90 s (running condition A [ Table 12.4.5-2 ]). Lanes 1 through 7 show different patterns. Lanes 8 and 9, 10 and 11, and 12 and 13 represent three pairs of indistinguishable patterns. This photo was kindly provided by Andreas Widmer.

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
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Image of Figure 12.4.5-2
Figure 12.4.5-2

Restriction fragment length polymorphism of chromosomal DNA digested with SpeI (running condition A [ Table 12.4.5-2 ). Lanes 3 and 4 show identical patterns, while the patterns in lanes 5 and 6 are similar but not identical and represent subtypes of a single strain.

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
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References

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Tables

Generic image for table
Table 12.4.5-1

Suggested enzymes and running conditions for some species

Size range includes the majority of fragments created after digestion with the corresponding enzyme. Some species will have several fragments under 48.5 kbp; however, it is difficult to analyze them when the 48.5-kbp lambda ladder molecular size standard is used. In order to analyze those fragments, use d III-cut lambda as a molecular size standard and/or a lower switch time, such as 5 to 15 s.

See Table 12.4.5-2.

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
Generic image for table
Table 12.4.5-2

Examples of running conditions

All electrophoretic conditions are at 13°C.

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
Generic image for table
Table 12.4.7-1

Rinse solution preparation

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4
Generic image for table
Table 12.4.7-2

Conjugate and substrate preparation

Citation: Garcia L. 2010. Molecular Methods for Epidemiologic Typing of Microorganisms, p 380-409. In Clinical Microbiology Procedures Handbook, 3rd Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817435.ch12.4

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