Microarray-Based Detection and Clinical Evaluation for Helicobacter pylori Resistance to Clarithromycin or Levofloxacin and the Genotype of CYP2C19 in 1083 Patients

Abstract

Background. Helicobacter pylori (H. pylori) is one of the most frequent and persistent bacterial infections that affect nearly half of the world's population. Antibiotic resistance is a constantly evolving process and local surveillance of antibiotic resistance is warranted to guide clinicians in their choice of therapy. The aim of this study was to establish a microarray-based detection to identify H. pylori infection, clarithromycin and levofloxacin susceptibility, and CYP2C19 genetic polymorphism and guide to potential choice of proton pump inhibitor (PPI), antibiotic administration for tailored H. pylori eradication therapy. Methods. By analyzing the sequence of human genomic CYP2C19⁎2 and CYP2C19⁎3 and mutations within the 23S rRNA and gyrA gene regions conferring clarithromycin and levofloxacin resistance, respectively, we developed a microarray for individual therapy detection of H. pylori infection. Plasmids were established as positive or limit of detection (LOD) reference materials. The specificity and sensitivity of the microarray had been performed. And a total of 1083 gastric biopsy samples were tested and the Kappa value had been calculated between the array and Sanger sequencing. We also analyzed the resistance to clarithromycin and levofloxacin in China, as well as the CYP2C19 polymorphisms. Results. The LOD of detecting H. pylori was 103 CFU/mL and human genome DNA was 2 ng/μL. The detection results of 1083 gastric biopsy samples showed that 691 (63.80%) were H. pylori positive, of which 266 (38.49%) were resistant to clarithromycin, 192 (27.79%) were resistant to levofloxacin, and 61 (8.83%) were resistant to both of them. For the type of CYP2C19 polymorphism, 412 (38.04%) were homozygous fast type (HomEM), 574 (53%) were heterozygous EM (HetEM), and 97 (8.96%) were poor metabolizer (PM). Conclusions. The proposed microarray-based detection has high specificity, sensitivity, and reproducibility for detecting the resistance of clarithromycin or levofloxacin as well as CYP2C19 polymorphism, which may help to improve the clinical eradication rate of H. pylori.

Current methods and prospects of coronavirus detection

“…Microarray is a rapid, high-throughput detection method [ [115] , [116] , [117] ], which can reverse transcription of viral RNA and produce a specific probe labeled cDNA. The cDNA was then loaded into each well and hybridized with solid-phase oligonucleotides on the Microarray, which is then washed to remove the free DNAs.…”

“…In addition, there are many kinds of this method, so it is difficult to establish a unified quality standard, which limits its clinical application. [ 116 ] ELISA At present, ELISA is commonly used in clinical detection of viruses, which is based on the specific binding of antigen and antibody. The procedure of this method is simple, because there is no need to use the second antibody, it can avoid the interaction reaction, high sensitivity and high specificity, and the antigen does not need to be purified in advance, so it can be applied to relatively impure samples, and the data reproducibility is very high.…”

Microarrays and NGS for Drug Discovery

“…Frye et al have developed a DNA microarray capable of detecting all antimicrobial resistance genes found at the National Center for Biotechnology [93]. Furthermore, a microarray has been use to identify Helicobacter pylori resistance to clarithromycin and levofloxacin, as well as to detect CYP2C19 polymorphism [94]. It is reported that this microarray can be used for individual therapy detection as it has high specificity, reproducibility, and sensitivity [78].…”

Development of a DNA microarray assay for rapid detection of fifteen bacterial pathogens in pneumonia

“…The microarray layout is shown in Figure 1a. Microarrays were prepared as previously described by our research group [37].…”