Real-Time PCR detection and quantitation ofHelicobacter pyloriclarithromycin-resistant strains in archival material and correlation with Sydney classification.
BACKGROUND
Helicobacter pylori (H. pylori), infects gastric mucosa causing gastritis. Treatment failure is mainly due to certain genetic changes in the peptidyltransferase loop of 23S rRNA of the microorganism. The aim of the study was to evaluate genetic changes in gastric biopsies ofH. pylori(+) patients that lead to clarithromycin resistance and to correlate them with histology data.


METHODS
A total of 150H. pylori(+) gastric biopsies were studied, taken before and after eradication therapy from 75 dyspeptic patients divided in 2 groups: group A consisted of 25H. pylori(+) triple-therapy resistant patients and group B consisted of 50H. pylori(+) successfully treated patients. Histological classification of theH. pylori(+) gastritis was done according to the Sydney criteria. Genetic material was analyzed with the ClariRes™ RT-PCR bi-probe based assay for the determination of point mutations in the 23S rRNA gene and with a Quantitative-RT-PCR (Q-RT-PCR) method for the quantitation ofH. pylori.


RESULTS
We showed that in 18/ 25 group A patients certain point mutations of 23S rRNA at sites A2142C, A2142G and A2143G had occurred. Nine of these 18 mutated cases (50%) were characterized as mixed infections. Mixed infections in 2/50 patients of group B were also observed. Using Q-RT-PCR, we found that gastric mucosal density ofH. pyloricorrelates well with bacterial colonization. There was a statistically significant association (P<0.005) between the presence of the detectedH. pylorigenetic alterations and inflammation, activity andH. pyloridensity as histologically determined.


CONCLUSIONS
Certain point mutations inH. pylorigenome that affect susceptibility to clarithromycin correlate with histological features of gastritis.
National Organization for Medicines, Micobiology Lab, Athens, Greece (Sofia Gazi).
Sofia
S
Gazi
Department of Pathology, NIMTS Hospital, Athens, Greece (Andreas Karameris).
Andreas
A
Karameris
Private Endoscopic Clinic, Nicosia, Cyprus (Marios Christoforou).
Marios
M
Christoforou
Department of Pathology, University of Ioannina, Greece (Dimitrios Stefanou, Niki Agnantis).
Niki
N
Agnantis
Gastroenterology Unit, Henry Dunant Hospital, Athens, Greece (Theodore Rokkas).
Theodore
T
Rokkas
Department of Pathology, University of Ioannina, Greece (Dimitrios Stefanou, Niki Agnantis).
Dimitrios
D
Stefanou
ClariRes assay; Clarithromycin resistance; Helicobacter pylori infection; Sydney classification; histology; quantitative real-time PCR; real-time PCR