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Chapter 51 : The Fluorescent In Situ Hybridization Test in Comparison with Culture for Detection of in Water Samples

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The Fluorescent In Situ Hybridization Test in Comparison with Culture for Detection of in Water Samples, Page 1 of 2

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Abstract:

This chapter evaluates the fluorescent in situ hybridization (FISH) test with a specific 16S rRNA-targeted probe for detecting the presence of viable in water. The results of this FISH test are available after 6 h (without inactivation) or 24 h (with activation). A large number of water samples, mostly from hot water systems, were tested with both test varieties and results were compared with those obtained with the culture method. FISH is a detection technique using a specific fluorescence-labeled DNA probe targeting the rRNA of the cells of the selected microorganism(s). Effect of activation on the result of the FISH test for detecting in water samples is discussed in the chapter. Interpretation of the FISH test results, in terms of hygienic significance, is even more complicated than such interpretation of colony counts. On the basis of the FISH results following conclusions can be drawn: (i) FISH allows detection of in samples of water and biofilms within 24 h after sampling, but activation is needed, (ii) in many cases the numbers of cells detected with FISH are much higher than numbers detected with the culture method, and (iii) further improvement of the FISH test, e.g., a level of detection similar to the culture method, is needed.

Citation: Wullings B, Voogt R, Veenendaal H, van der Kooij D. 2002. The Fluorescent In Situ Hybridization Test in Comparison with Culture for Detection of in Water Samples, p 263-266. 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.ch51

Key Concept Ranking

Legionella pneumophila
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Culture Methods
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Figures

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FIGURE 1

Number of detected with FISH (F) compared with numbers detected with the culture method (CM). The detection limits (2,000 cells/liter in the FISH test; 50 CFU/liter for the culture method) are indicated. Also, ratios between numbers of cells detected with FISH and with the culture method are shown. Symbols on the lines, indicating the detection levels, represent samples in which the number was less than or equal to these limits.

Citation: Wullings B, Voogt R, Veenendaal H, van der Kooij D. 2002. The Fluorescent In Situ Hybridization Test in Comparison with Culture for Detection of in Water Samples, p 263-266. 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.ch51
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References

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1. Amann, R. I.,, B.J. Binder,, R. J. Olsen,, S. W. Chrisholm,, R. Devereux,, and D. A. Stahl. 1990. Combination of 16S rRNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations. Appl. Environ. Miaobiol. 56:19191925.
2. Amann, R. I.,, W. Ludwig,, and K.-H. Schleifer. 1995. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Miaobiol. Rev. 59:143169.
3. Edelstein, P. H. 1981. Improved semi-selective medium for isolation of Legionella pneumophila from contaminate clinical and environmental specimens. J. Clin. Miaobiol. 14:298303.
4. Giovannoni, S.J.,, E. F. DeLong,, G.J. Olsen,, and N. R. Pace. 1988. Phylogenetic group-specific oligodeoxynucleotide probes for identification of single microbial cells. J. Bacteriol. 170: 720726.
5. Grimm, D.,, H. Merkert,, W. Ludwig, K-H. Scheiffer, J. Hacker, and B. C. Brand. 1998. Specific detection of Legionella pneumophila: construction of a new 16S rRNA-targeted oligonucleotide probe. Appl. Environ. Miaobiol. 64:26862690.
6. Hay, J.,, D. V. Seal,, B. Billcliffe,, and J. H. Freer. 1995. Non-culturable Legionella pneumophila associated with Acanthamoeba castellanii: detection of the bacterium using DNA amplification and hybridization. J. Appl. Bacterial. 78:6165.
7. Maiwald, M.,, K. Kissel,, S. Srimuang,, M. von Knebel Doeberitz,, and H.-G. Sonntag. 1994. Comparison of polymerase chain reaction and conventional culture for the detection of Legionellas in hospital water samples. J. Appl. Bacteriol. 76:216225.
8. Moter, A.,, and U. B. Gobel. 2000. Fluorescence in situ hybridization (FISH) for direct visualization of microorganisms. J. Microbial Methods 41:85112.
9. Okpara, J.,, M. Maiwald,, M. Borneff,, J. Windeler,, and H.-G. Sonntag. 1996. Evaluation of a new version of the EnvironAmp™ Legionella kit for the detection of Legionellae in water samples by the polymerase chain reaction. Zentralbl. Hyg. 198: 502513.

Tables

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TABLE 1

Detection of in 124 water samples using FISH and the culture technique

Citation: Wullings B, Voogt R, Veenendaal H, van der Kooij D. 2002. The Fluorescent In Situ Hybridization Test in Comparison with Culture for Detection of in Water Samples, p 263-266. 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.ch51
Generic image for table
TABLE 2

Effect of activation on the result of the FISH test for detecting in water samples

Citation: Wullings B, Voogt R, Veenendaal H, van der Kooij D. 2002. The Fluorescent In Situ Hybridization Test in Comparison with Culture for Detection of in Water Samples, p 263-266. 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.ch51

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