Development of Highly Sensitive Drug Resistant Mutation Gene Detection in Helicobacter pylori by MALDI-TOF Mass Spectrometry
Date Issued
2015
Date
2015
Author(s)
Hung, Hsiao-Yi
Abstract
Gastric cancer is the fourth most common cancer and the second leading cause of cancer-related death in the world. Among all risk factor, the infection of H. pylori is presented in approximately 63% of patients. H. Pylori is a spiral-shaped, microaerophilic gram-negative flagellate bacterium contributes to duodenal/gastric ulcer, gastritis and gastric adenocarcinoma. Eradication therapy of H. pylori including antibiotics, PPI (Proton Pump Inhibitor) has emerged as the treatment of choice. Although antibiotics have been useful in the treatment of H. pylori-related benign and malignant gastroduodenal disease, the rate of resistance to standard therapies has increased a result of the widespread use of antibiotics in recent year. Our specific aim is to establish a multiplexed high sensitive nucleotide MALDI-TOF MS to detect H. pylori infection and predict its world. Among all risk factor, the infection of H. pylori 23S rRNA, 16S rRNA, PBP-1, rdxA, gyrA and frxA genes were selected for multiplex panel design. 23S rRNA and gyrA that drug resistant related mutation sites were well known was pioneer for MALDI-TOF MS establishment. Other four genes were sequenced by Sanger method in clinical isolates for identifying high prevalence mutation sites correlated drug resistance for MALDI-TOF MS platform in the future. Up to date, we had established two reaction multiplex gene testing panel for 15 mutations within 2 genes.The following mutations important for testing will be recruited in the panel in the future. The detection limitation of our platform is around 1 copy according to serial dilution of H. pylori genome DNA. Future direction will focus on: 1. Optimization of detection sensitivity and specificity; 2. Identification of drug resistant mutation hot spot among 16S rRNA, PBP-1, rdxAand frxA genes; 3. Feasibility of H. pylori detection in clinical specimens such as biopsy from ulcer lesion or gastric cancer patients. With this platform set up, we can develop non-invasive detection strategy of H. pylori and predict its resistance in clinical specimens.
Subjects
Helicobacter pylori
MALDI-TOF Mass Spectrometry
drug resistant gene
SDGs
Type
thesis
File(s)![Thumbnail Image]()
Loading...
Name
ntu-104-R02424021-1.pdf
Size
23.32 KB
Format
Adobe PDF
Checksum
(MD5):211a58c9291901909c245d80cfd64001