Dışkı ve Mide Biyopsi Örneklerinden Helıcobacter Pylorı Enfeksiyonunun ve Klaritromisin Direncinin Moleküler Yöntemler ile Saptanması

Abstract

Inal Neşe, Detection of Helicobacter pylori Infection and Clarithromycin Resistance in Stool and Gastric Biopsy Samples by Molecular Methods, Hacettepe University Faculty of Medicine, Department of Medical Microbiology, Thesis In Medical Microbiology, Ankara, 2022. Helicobacter pylori is a bacterium that infects half of the world population and continues to exist in the stomach, unless treated. Many invasive and non-invasive methods have been developed for diagnosis. Since the diagnosis with non-invasive methods is more suitable and easy for the patient, there are many studies to increase the specificity and sensitivity of non-invasive methods. Detection of H. pylori and clarithromycin point mutations from stool samples by polymerase chain reaction (PCR) is a non-invasive method. In cases where endoscopy is difficult to perform or not available as in pediatric patients and pregnant women, and also in the follow-up of H. pylori treatment, a real-time PCR test using stool samples may be effective. The aims of this study were (i) to detect H. pylori infections and the point mutations that lead to clarithromycin resistance by using real-time PCR from gastric biopsy and stool samples (ii) to perform antibiotic susceptibility test, (iii) to compare the results of antibiotic susceptibility tests and the point mutations that lead to clarithromycin resistance. The study included 63 patients whom have undergone gastroduodenal endoscopy. Culture, rapid urease test, histopathological examination, HpSA and gastric biopsy methods were used for the diagnosis of H. pylori. In our study, the combination of two tests or gastic biopsy culture positivity were accepted as the “reference result”. The “Ezplex HP-CLA real-time PCR” (SML Genetree, Korea) diagnostic kit was used for detecting H. pylori and the most common point mutations (A2142G, A2143G) that lead to clarithromycin resistance in the 23S rRNA gene. H. pylori was detected in 30.2% of the gastric biopsy samples by rapid urease test, 38.1% by histopathological examination, and 58.7% by gastric biopsy real-time PCR test. H. pylori was positive in 28.5% by HpSA test. Growth was achieved in 17.5% of 63 specimens by the culture. Antibiotic susceptibility tests were applied by gradient strip test for nine isolates according to EUCAST standards. Resistance to levofloxacin, tetracycline, and amoxicillin was not detected. Clarithromycin resistance was found in three of the isolates and metronidazole resistance was found in three of the isolates. H. pylori was detected positive in 28.5% by stool real-time PCR test. When the reference methods results were compared to “Ezplex” stool real-time PCR; the sensitivity in terms of H. pylori detection was 66.6%, the specificity was 100%, PPV was 100%, NPV was 80%, and the diagnostic accuracy was 85.7%. In the detection of clarithromycin point mutations from stool samples with “Ezplex” real-time PCR, the overall agreement was 65% when compared to the “Ezplex” gastric biopsy real-time PCR results. When the phenotypic antimicrobial susceptibility results were compared to clarithromycin point mutations detected by stool real-time PCR, it was found to be correlated in 66.6%. In this study, it is concluded that real-time PCR from stool samples can be used in diagnosis of H. pylori, especially where invasive tests are hard to perform, like in the pediatric patients, pregnant women and follow-up of treatment. Although the clarithromycin point mutations detection rate from stool samples were not as expected, it is still promising, but more studies should be performed. Keywords: H. pylori, real-time PCR, clarithromycin resistance Supporting Organisation: Hacettepe University Scientific Research Project Unit Project No: THD-2021-1943