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Journal of Clinical Microbiology, July 2003, p. 3112-3118, Vol. 41, No. 7
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.7.3112-3118.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Identification of cagA Tyrosine Phosphorylation DNA Motifs in Helicobacter pylori Isolates from Peptic Ulcer Patients by Novel PCR-Restriction Fragment Length Polymorphism and Real-Time Fluorescence PCR Assays

Robert J. Owen,1* Sally I. Sharp,1 Stephanie A. Chisholm,1 and Sjoerd Rijpkema2

Laboratory of Enteric Pathogens, Specialist and Reference Microbiology Division, Health Protection Agency, London,1 Division of Bacteriology, National Institute for Biological Standards and Control, Potters Bar, United Kingdom2

Received 31 October 2002/ Returned for modification 2 December 2002/ Accepted 7 March 2003

Cag pathogenicity island-containing Helicobacter pylori (type I) induces signal transduction pathways resulting in tyrosine phosphorylation of proteins adjacent to the site of bacterial adhesion on host gastric epithelial cells. Conventional block PCR-restriction fragment length polymorphism (RFLP) and real-time LightCycler (LC) PCR hybridization assays, validated by direct sequencing, were designed to test for the presence of three nucleotide sequences corresponding to tyrosine phosphorylation motifs (TPMs) A, B, and C in 84 isolates of H. pylori type I from patients in England. Overall, the PCR assays demonstrated that one or more TPMs were present in 62 strains (75%). Motif A was common (71% of strains), whereas motifs B and C were rarer (8% of strains). Strains lacking a TPM were typically vacuolating cytotoxin genotype vacA m2. Motif A was widely distributed in relation to disease severity and was more commonly (but not significantly [P = 0.071]) associated with gastric ulcer than with duodenal ulcer (86 versus 56%). The LC hybridization assay provided a rapid means of detecting all three motifs, but RFLP analysis was more specific for TPM-A. TPMs provide novel additional strain markers for defining cagA variation, including identification of RFLP types within TPM-A. The presence of a particular TPM was not of direct diagnostic value, either singly or in combination, but the higher proportion of TPM-A strains in gastric ulcer patients merits further investigation.

* Corresponding author. Mailing address: Helicobacter Reference Unit, Laboratory of Enteric Pathogens, Specialist and Reference Microbiology Division, Health Protection Agency, 61 Colindale Ave., London NW9 5HT, England. Phone: 44-20 8200 4400. Fax: 44-20 8905 9929. E-mail: rowen{at}phls.nhs.uk.

Journal of Clinical Microbiology, July 2003, p. 3112-3118, Vol. 41, No. 7
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.7.3112-3118.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Mattar, R., Marques, S. B., Monteiro, M. d. S., dos Santos, A. F., Iriya, K., Carrilho, F. J. (2007). Helicobacter pylori cag pathogenicity island genes: clinical relevance for peptic ulcer disease development in Brazil. J Med Microbiol 56: 9-14 [Abstract] [Full Text]  
  • Argent, R. H., Zhang, Y., Atherton, J. C. (2005). Simple Method for Determination of the Number of Helicobacter pylori CagA Variable-Region EPIYA Tyrosine Phosphorylation Motifs by PCR. J. Clin. Microbiol. 43: 791-795 [Abstract] [Full Text]  
  • Backert, S., Schwarz, T., Miehlke, S., Kirsch, C., Sommer, C., Kwok, T., Gerhard, M., Goebel, U. B., Lehn, N., Koenig, W., Meyer, T. F. (2004). Functional Analysis of the cag Pathogenicity Island in Helicobacter pylori Isolates from Patients with Gastritis, Peptic Ulcer, and Gastric Cancer. Infect. Immun. 72: 1043-1056 [Abstract] [Full Text]  


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