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Journal of Clinical Microbiology, January 2002, p. 205-209, Vol. 40, No. 1
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.1.205-209.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Evaluation of a Novel Heminested PCR Assay Based on the Phosphoglucosamine Mutase Gene for Detection of Helicobacter pylori in Saliva and Dental Plaque

C. Goosen,1 J. Theron,2 M. Ntsala,2 F. F. Maree,1 A. Olckers,3 S. J. Botha,4 A. J. Lastovica,5 and S. W. van der Merwe1*

Department of Internal Medicine,1 Department of Microbiology and Plant Pathology,2 Department of Human Genetics,3 Department of Stomatological Research, University of Pretoria, Pretoria 0002,4 Department of Medical Microbiology, University of Cape Town, Cape Town 7925, South Africa5

Received 23 February 2001/ Returned for modification 13 June 2001/ Accepted 3 October 2001

A novel heminested PCR protocol was developed for the specific detection of Helicobacter pylori at low copy numbers. A set of primers specific for the phosphoglucosamine mutase gene (glmM) of H. pylori produced a 765-bp fragment that was used as template for the heminested primer pair delineating a 496-bp fragment. By using agarose gel electrophoresis for detection of the heminested PCR-amplified products, amplification of H. pylori genomic DNA was achieved at concentrations as low as 0.1 pg, equivalent to 5 x 102 bacteria. A study was subsequently undertaken to evaluate the heminested PCR for detection of H. pylori in dental plaque and saliva. Specimens collected from 58 individuals were cultured, and PCR was subsequently performed on the oral cultures. Identification of H. pylori in the same series of saliva and dental plaque specimens was carried out with PCR using a primer pair specific for the H. pylori urease B gene and by the heminested PCR assay. The identity of the amplified products was confirmed by DNA sequencing. Our results demonstrate that the heminested PCR assay was specific for detection of H. pylori, yielding no false-positive results, and that H. pylori had a low prevalence (approximately 3%) in specimens obtained from the oral cavity.

* Corresponding author. Mailing address: Department of Internal Medicine, P.O. Box 1649, Faerie Glen, Pretoria 0043, South Africa. Phone: 27 12 664 0197. Fax: 27 12 664 0202. E-mail: svdm{at}intekom.co.za.

Journal of Clinical Microbiology, January 2002, p. 205-209, Vol. 40, No. 1
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.1.205-209.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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