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Journal of Clinical Microbiology, March 2003, p. 976-980, Vol. 41, No. 3
0095-1137/03/$08.00+0 DOI: 10.1128/JCM.41.3.976-980.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
Heterogeneity of cag Genotypes in Helicobacter pylori Isolates from Human Biopsy Specimens
Maria Luisa Tomasini,1 Stefania Zanussi,1 Michele Sozzi,2* Rosamaria Tedeschi,1 Giancarlo Basaglia,1 and Paolo De Paoli1Microbiology, Immunology, and Virology,1 Gastroenterology Units, Centro di Riferimento Oncologico, IRCCS, Aviano, Italy2
Received 14 August 2002/ Returned for modification 16 September 2002/ Accepted 9 December 2002
The Helicobacter pylori chromosomal cluster of genes known as the cytotoxin-associated gene (cag) island may have different compositions in infecting strains. In this study, we analyzed 150 single colonies obtained from gastric biopsy specimens from 10 patients infected with cagA-positive H. pylori strains and sweep isolates (isolates harvested with sweep in different points of the plate) from 6 patients infected with cagA-negative strains. Three loci in the cag island (cagA, cagE, and virB11) and the conserved gene glmM (ureC) were investigated by PCR. The levels of anti-H. pylori and anti-CagA antibodies in patient sera were also measured. For subjects infected with cagA-negative strains, all sweep isolates were also negative for cagE and virB11, suggesting the complete absence of the cag island. For subjects infected with cagA-positive strains, most of the isolates were positive for all three genes studied, whereas 24.7% of the isolates had a partial or total deletion of the cag island. cagA, cagE, and virB11 were, respectively, present in 87.3, 77.3, and 90% of the colonies. The deletion of virB11 was always associated with the deletion of cagA and/or cagE. H. pylori colonies with different cag genotypes were isolated within a single gastric biopsy specimen from 3 of the 10 patients and were further characterized by random amplified polymorphic DNA (RAPD) analysis and by sequencing of an arbitrarily selected gene segment. Although the colonies had different cag genotypes, their RAPD profiles were highly similar within each patient, and the nucleotide sequences of the selected gene segment were identical. All of the patients had detectable antibodies against H. pylori, and 9 of 10 had anti-CagA antibodies. In conclusion, we show that a single infecting H. pylori strain may include variable proportions of colony subtypes with different cag genotypes. The extension of our analysis to patients with well-characterized gastric diseases may provide significant information on the relationship between cag genotypes and clinical outcomes of H. pylori infections.
* Corresponding author. Mailing address: Division of Gastroenterology and Digestive Endoscopy, Centro di Riferimento Oncologico-National Cancer Institute, Via Pedemontana Occidentale 12, 33081 Aviano (Pordenone), Italy. Phone: 39/0434/659362. Fax: 39/0434/659402. E-mail: msozzi{at}cro.it.
Journal of Clinical Microbiology, March 2003, p. 976-980, Vol. 41, No. 3
0095-1137/03/$08.00+0 DOI: 10.1128/JCM.41.3.976-980.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
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