Absence of Helicobacter pylori high tetracycline resistant 16S rDNA AGA926-928TTC genotype in gastric biopsy specimens from dyspeptic patients of a city in the interior of Sao Paulo, Brazil

Author(s)

Suzuki, Rodrigo Buzinaro ^{[1, 2]} ; Almeida, Cristiane Maria ^[3] ; Speranca, Marcia Aparecida ^{[1, 3]}

Affiliation

^[1] Univ Fed Abc, Ctr Nat & Human Sci, BR-09210170 Santo Andre, SP - Brazil

^[2] Marilia Med Sch, Dept Genotyping, Marilia, SP - Brazil

^[3] Marilia Med Sch, Dept Mol Biol, Marilia, SP - Brazil

• Document type: Journal article
• Source: BMC GASTROENTEROLOGY; v. 12, p. 49, MAY 17 2012.
• Web of Science Citations: 3

Abstract

Background: Treatment effectiveness of Helicobacter pylori varies regionally and is decreasing worldwide, principally as a result of antibiotic resistant bacterium. Tetracycline is generally included in second line H. pylori eradication regimens. In Brazil, a high level of tetracycline resistance (TetR) is mainly associated with AGA926-928TTC 16 S rDNA nucleotide substitutions. As H. pylori culture is fastidious, we investigated the primary occurrence of H. pylori 16 S rDNA high level TetR genotype using a molecular approach directly on gastric biopsies of dyspeptic patients attending consecutively at Hospital das Clinicas of Marilia, Sao Paulo, Brazil. Methods: Gastric biopsy specimens of 68 peptic ulcer disease (PUD) and 327 chronic gastritis (CG) patients with a positive histological diagnosis of H. pylori were investigated for TetR 16 S rDNA genotype through a molecular assay based on amplification of a 16 S rDNA 545 bp fragment by polymerase chain reaction and HinfI restriction fragment length polymorphism (PCR/RFLP). Through this assay, AGA926-928TTC 16 S rDNA TetR genotype resulted in a three DNA fragment restriction pattern (281, 227 and 37 bp) and its absence originated two DNA fragments (264 and 281 bp) due to a 16 S rDNA conserved Hinf I restriction site. Results: The 545 bp 16 S rDNA PCR fragment was amplified from 90% of gastric biopsies from histological H. pylori positive patients. HinfI RFLP revealed absence of the AGA926-928TTC H. pylori genotype and PCR products of two patients showed absence of the conserved 16 S rDNA HinfI restriction site. BLASTN sequence analysis of four amplicons (two conserved and two with an unpredicted HinfI restriction pattern) revealed a 99% homology to H. pylori 16 S rDNA from African, North and South American bacterial isolates. A nucleotide substitution abolished the conserved HinfI restriction site in the two PCR fragments with unpredicted HinfI RFLP, resulting in an EcoRI restriction site. Conclusions: H. pylori AGA926-928TTC 16 S rDNA gene substitutions were not found in our population. More research is required to investigate if H. pylori TetR has a different genetic background in our region and if the nucleotide substitutions of the uncultured H. pylori 16 S rRNA partial sequences have biological significance. (AU)

• FAPESP's process: 05/04087-7 - Investigation of tetracycline primary resistance in Helicobacter pylori obtained from gastric biopsies of individuals from Marilia – SP
• Grantee: Cristiane Maria Almeida
• Support Opportunities: Scholarships in Brazil - Scientific Initiation