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Laboratorium voor Microbiologie, Universiteit Gent, B-9000 Gent, Belgium
KeyGene nv, Wageningen, The Netherlands
3Author for correspondence: Paul Janssen. Tel: +32 9 264 5121. Fax: +32 9 264 5346. e-mail: paul.janssen@rug.ac.be
ABSTRACT
We investigated the usefulness of a novel DNA fingerprinting technique, AFLP, which is based on the selective amplification of genomic restriction fragments by PCR, to differentiate bacterial strains at the subgeneric level. In total, 147 bacterial strains were subjected to AFLP fingerprinting: 36 Xanthomonas strains, including 23 pathovars of Xanthomonas axonopodis and six pathovars of Xanthomonas vasicola, one strain of Stenotrophomonas, 90 genotypically characterized strains comprising all 14 hybridization groups currently described in the genus Aeromonas, and four strains of each of the genera Clostridium, Bacillus, Acinetobacter, Pseudomonas and Vibrio. Depending on the genus, total genomic DNA of each bacterium was digested with a particular combination of two restriction endonucleases and the resulting fragments were ligated to restriction halfsite-specific adaptors. These adaptors served as primer-binding sites allowing the fragments to be amplified by selective PCR primers that extend beyond the adaptor and restriction site sequences. Following electrophoretic separation on 5% (w/v) polyacrylamide/8.3 M urea, amplified products could be visualized by autoradiography because one of the selective primers was radioactively labelled. The resulting banding patterns, containing approximately 30-50 visualized PCR products in the size range 80-550 bp, were captured by a high-resolution densitoscanner and further processed for computer-assisted analysis to determine band-based similarity coefficients. This study reveals extensive evidence for the applicability of AFLP in bacterial taxonomy through comparison of the newly obtained data with results previously obtained by well-established genotypic and chemotaxonomic methods such as DNA-DNA hybridization and cellular fatty acid analysis. In addition, this study clearly demonstrates the superior discriminative power of AFLP towards the differentiation of highly related bacterial strains that belong to the same species or even biovar (i.e. to characterize strains at the infrasubspecific level), highlighting the potential of this novel fingerprinting method in epidemiological and evolutionary studies.
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S. C. Jiang, M. Matte, G. Matte, A. Huq, and R. R. Colwell Genetic Diversity of Clinical and Environmental Isolates of Vibrio cholerae Determined by Amplified Fragment Length Polymorphism Fingerprinting Appl. Envir. Microbiol., January 1, 2000; 66(1): 148 - 153. [Abstract] [Full Text]
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P. H. M. Savelkoul, H. J. M. Aarts, J. de Haas, L. Dijkshoorn, B. Duim, M. Otsen, J. L. W. Rademaker, L. Schouls, and J. A. Lenstra Amplified-Fragment Length Polymorphism Analysis: the State of an Art J. Clin. Microbiol., October 1, 1999; 37(10): 3083 - 3091. [Full Text]
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B. Kokotovic, N. F. Friis, J. S. Jensen, and P. Ahrens Amplified-Fragment Length Polymorphism Fingerprinting of Mycoplasma Species J. Clin. Microbiol., October 1, 1999; 37(10): 3300 - 3307. [Abstract] [Full Text]
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I. Geornaras, N. F. Kunene, A. von Holy, and J. W. Hastings Amplified Fragment Length Polymorphism Fingerprinting of Pseudomonas Strains from a Poultry Processing Plant Appl. Envir. Microbiol., September 1, 1999; 65(9): 3828 - 3833. [Abstract] [Full Text]
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 A. Meijer, S. A. Morré, A. J. C. Van Den Brule, P. H. M. Savelkoul, and J. M. Ossewaarde Genomic Relatedness of Chlamydia Isolates Determined by Amplified Fragment Length Polymorphism Analysis J. Bacteriol., August 1, 1999; 181(15): 4469 - 4475. [Abstract] [Full Text]
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B. Duim, T. M. Wassenaar, A. Rigter, and J. Wagenaar High-Resolution Genotyping of Campylobacter Strains Isolated from Poultry and Humans with Amplified Fragment Length Polymorphism Fingerprinting Appl. Envir. Microbiol., June 1, 1999; 65(6): 2369 - 2375. [Abstract] [Full Text]
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J. Vandenberghe, L. Verdonck, R. Robles-Arozarena, G. Rivera, A. Bolland, M. Balladares, B. Gomez-Gil, J. Calderon, P. Sorgeloos, and J. Swings Vibrios Associated with Litopenaeus vannamei Larvae, Postlarvae, Broodstock, and Hatchery Probionts Appl. Envir. Microbiol., June 1, 1999; 65(6): 2592 - 2597. [Abstract] [Full Text]
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D. M. Olive and P. Bean Principles and Applications of Methods for DNA-Based Typing of Microbial Organisms J. Clin. Microbiol., June 1, 1999; 37(6): 1661 - 1669. [Full Text]
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M. Desai, A. Efstratiou, R. George, and J. Stanley High-Resolution Genotyping of Streptococcus pyogenes Serotype M1 Isolates by Fluorescent Amplified-Fragment Length Polymorphism Analysis J. Clin. Microbiol., June 1, 1999; 37(6): 1948 - 1952. [Abstract] [Full Text]
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J. van Eldere, P. Janssen, A. Hoefnagels-Schuermans, S. van Lierde, and W. E. Peetermans Amplified-Fragment Length Polymorphism Analysis versus Macro-Restriction Fragment Analysis for Molecular Typing of Streptococcus pneumoniae Isolates J. Clin. Microbiol., June 1, 1999; 37(6): 2053 - 2057. [Abstract] [Full Text]
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C. Arnold, L. Metherell, G. Willshaw, A. Maggs, and J. Stanley Predictive Fluorescent Amplified-Fragment Length Polymorphism Analysis of Escherichia coli: High-Resolution Typing Method with Phylogenetic Significance J. Clin. Microbiol., May 1, 1999; 37(5): 1274 - 1279. [Abstract] [Full Text]
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M. Desai, A. Tanna, R. Wall, A. Efstratiou, R. George, and J. Stanley Fluorescent Amplified-Fragment Length Polymorphism Analysis of an Outbreak of Group A Streptococcal Invasive Disease J. Clin. Microbiol., November 1, 1998; 36(11): 3133 - 3137. [Abstract] [Full Text]
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L. Hauben, M. Steenackers, and J. Swings PCR-Based Detection of the Causal Agent of Watermark Disease in Willows (Salix spp.) Appl. Envir. Microbiol., October 1, 1998; 64(10): 3966 - 3971. [Abstract] [Full Text]
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J. G. M. Koeleman, J. Stoof, D. J. Biesmans, P. H. M. Savelkoul, and C. M. J. E. Vandenbroucke-Grauls Comparison of Amplified Ribosomal DNA Restriction Analysis, Random Amplified Polymorphic DNA Analysis, and Amplified Fragment Length Polymorphism Fingerprinting for Identification of Acinetobacter Genomic Species and Typing of Acinetobacter baumannii J. Clin. Microbiol., September 1, 1998; 36(9): 2522 - 2529. [Abstract] [Full Text]
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J. R. Gibson, E. Slater, J. Xerry, D. S. Tompkins, and R. J. Owen Use of an Amplified-Fragment Length Polymorphism Technique To Fingerprint and Differentiate Isolates of Helicobacter pylori J. Clin. Microbiol., September 1, 1998; 36(9): 2580 - 2585. [Abstract] [Full Text]
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P. Boerlin, S. Chen, J. K. Colbourne, R. Johnson, S. De Grandis, and C. Gyles Evolution of Enterohemorrhagic Escherichia coli Hemolysin Plasmids and the Locus for Enterocyte Effacement in Shiga Toxin-Producing E. coli Infect. Immun., June 1, 1998; 66(6): 2553 - 2561. [Abstract] [Full Text] [PDF]
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A. van Belkum, S. Scherer, L. van Alphen, and H. Verbrugh Short-Sequence DNA Repeats in Prokaryotic Genomes Microbiol. Mol. Biol. Rev., June 1, 1998; 62(2): 275 - 293. [Abstract] [Full Text] [PDF]
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