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Research Paper |
Veterinary Laboratories Agency (Weybridge), New Haw, Addlestone, Surrey KT15 3NB, UK1
Molecular Microbiology and Genomics Consultants, Zotzenheim, Germany2
Author for correspondence: Diane G. Newell. Tel: +44 1932 357547. Fax: +44 1932 357595. e-mail: dnewell.cvl.wood{at}gtnet.gov.uk
The consumption of poultry meat contaminated with Campylobacter jejuni is considered to be a risk factor for human campylobacteriosis. The development of targeted strategies to control campylobacters in broilers would benefit from knowledge of those bacterial factors important in colonization of the avian gut. During preliminary studies it was noted that C. jejuni NCTC 11168 was a poorer colonizer of chickens than strain 81116. This poor colonization could not be fully restored by in vivo passage, suggesting that it was a genetically endowed property of strain 11168. As the genome sequence is available for this strain, the technique of subtractive hybridization was used to identify gene fragments of strain 81116 not present in strain 11168. After two screening cycles, 24 out of 42 clones were identified as having DNA inserts specific for strain 81116. Six of these 24 clones contained gene fragment inserts with similarities to restriction–modification enzymes found in other bacteria. Two inserts had similarity to arsenic-resistance genes, whereas four others had similarities to cytochrome c oxidase III, dTDP-glucose 4,6-dehydratase, 
-glutamyl transpeptidase and an abortive phage-resistance protein. At least some of these genes may be involved with colonization. A further six inserts had weak similarities to hypothetical proteins or to proteins with assigned functions from strain 11168. The remaining six clones had gene-fragment inserts with no database matches. Southern-blot analysis confirmed that strain-dependent variation existed for each of these DNA inserts. These results indicate that subtractive hybridization can successfully identify genes that are absent from the only C. jejuni strain for which the genome sequence is currently available.
Keywords: subtractive hybridization
Abbreviations: AP, alkaline phosphatase; R–M, restriction–modification
The GenBank accession numbers for the sequences reported in this paper can be found in Table 2.
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