Poster (Painel)
49-1Absence 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 São Paulo, Brazil.
Autores:Rodrigo Buzinaro Suzuki (HEMOCENTRO - FAMEMA - Department of Genotyping / UFABC - Center of Natural and Human Sciences) ; Cristiane Maria Almeida (FAMEMA - Department of Molecular Biology) ; Márcia Aparecida Sperança (UFABC - Center of Natural and Human Sciences)


Background. Considering the importance of H. pylori in the etiology of gastric diseases and increasing treatment failure attributable to selection of resistant strains, before antibiotic prescription, susceptibility testing is of great clinical relevance. Tetracycline is generally included in bacterium eradication regimens after unsuccessful classical first line therapy. H. pylori tetracycline high resistance is mainly due to AGA926-928TTC 16S rDNA nucleotide substitutions which are prevalent in Brazilian resistant isolates. Antibiotic susceptibility has regional epidemiological characteristics and tests based on H. pylori culture present limitations to be used in clinical practice. Thus, this work aimed to determine H. pylori primary resistance to tetracycline using a molecular approach directly on gastric biopsies based on amplification of a 16S rDNA 545bp fragment by polymerase chain reaction and HinfI restriction fragment length polymorphism (PCR/RFLP). We investigated 68 peptic ulcer diseases (PUD) and 327 chronic gastritis (CG) patients consecutively attended at Hospital das Clinicas of Marilia, São Paulo, Brazil, with H. pylori histological positive diagnostic and 16S rDNA 545bp PCR amplification. Results. The 545bp 16S 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 a CG and a PUD patient showed absence of the conserved 16S rDNA HinfI restriction site. BLASTN sequence analysis of four amplicons (two conserved and two with unpredicted HinfI restriction patterns) revealed a 99% homology to H. pylori 16S rDNA from African, North and South American bacterial isolates. A nucleotide substitution abolished the conserved HinfI,/i> restriction site in the two PCR fragments with unpredicted HinfI RFLP, resulting in an EcoRI restriction site. Conclusions. The PCR/RFLP assay employed is suitable to be used in routine practice to detect high 16S rRNA dependent tetracycline resistant genotype directly from biopsy specimens. H. pylori AGA926-929TTC 16S rDNA gene substitutions were not found in our population. More researches are required to investigate if H. pylori tetracycline resistance has a different genetic background in our region and if the nucleotide substitutions of the uncultured H. pylori 16S rRNA partial sequence have biological significance.

Palavras-chave:  Helicobacter pylori, Helicobacter pylori 16S rDNA polymorphis, Helicobacter pylori 16S rDNA, Nucleic acid based diagnostic, Tetracycline resistance