Evaluation of optimized Real time-PCR HRM assay and SPR-based biosensor for noninvasive isolation of H. pylori and Clarithromycin resistance 23S-SNP subtype

Abstract

High Resolution Melting analysis is a highly sensitive molecular method, and the plasmonic-based sensor is a convenient test, be clinically elicited, in simultaneous H. pylori control and the prevention of Clarithromycin resistant strains related to SNPs. Comparing gold standard tests, we evaluated HRMA and the SPR-based biosensor in regard to noninvasive forms of H. pylori management, and explored resistance through a possible mechanism by 23S-SNPs.We optimized Realtime-PCR HRM analysis, and SPR-based biosensor. Referring to CTs ± 38, isolation sensitivity was evaluated at 74 %. Compared to the reference-test, results agreements were assessed with a kappa of 43 %, p = 0.001. Within melting plots (differentiation plot), the strains were detectable at Tms ranging from 81.5 to 82.3 (°C). Contributing to phenotypically determined resistant strains for 44 %, there was a wide distribution among resistant variants assessed through Tms of 82, 81.5, 82, 82.3, and 81.7 (°C). A2143C was the only mutation isolated at a specific Tm of 82.3°C; kappa = 8 %, p = 0.3. For sensing analysis, including 100 % specificity, this sensor type was associated with lower sensitivity of 49 %, kappa=19 %, p = 0. 01. For SNP, specificity, kappa value, and p-value of isolation were 100 %, 100 %, and 0.001. Lod was 0.003 ng/mL and 0.03 µg/mL for HRM assay and sensing analysis. In our evaluations, applicable priority of HRM analysis was the higher estimation of sensitivity; its differentiation plot was completely covered the results of strain variability regarding specific SNPs detection. Consistent with the biosensor for isolation of SNP, the development of biosensors is the necessity for noninvasive H. pylori detection.