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Non-Invasive Colorimetric Magneto Loop-Mediated Isothermal Amplification (CM-LAMP) Method for Helicobacter pylori
Khotchawan Bangpanwimon1, Pimonsri Mittraparp-Arthorn1,2, Kanchana Srinitiwarawong1
1Department of Microbiology, Division of Biological Science, Faculty of Science, Prince of Songkla University, Hat-Yai, Songkhla 90110, Thailand.
Abstract:
More than half the world's population is thought to be infected with Helicobacter pylori. Although the majority of infected people are asymptomatic, H. pylori infection may cause gastric ulcers and deadly gastric cancer. Owing to the difficulty and invasiveness of current routine culture and diagnostic methods, a highly sensitive and specific noninvasive assay for H. pylori is of interest. This study highlighted the design and performance of a colorimetric magneto loop-mediated isothermal amplification (CM-LAMP) assay to detect H. pylori in spiked saliva samples. LF primers were coated on magnetic nanoparticles by carbodiimide-induced immobilization and functionally used for solidphase amplification. During the LAMP reaction at 66°C, biotin-tagged FIPs were incorporated into LAMP amplicons. The colorimetric signal developed after the addition of NeutrAvidin horseradish peroxidase conjugate (NA-HRP) and ABTS. None of the tested microorganisms, including closely related bacteria, was shown positive by the CM-LAMP assay except H. pylori isolates. This novel platform was highly specific and 100-fold more sensitive (40 CFU/ml or 0.2 CFU per reaction) than the PCR and conventional LAMP assays for the detection of H. pylori in spiked saliva. Our results demonstrated the feasibility of using this noninvasive molecular diagnostic test to detect H. pylori in saliva samples.
Insights
A new colorimetric magneto loop-mediated isothermal amplification (CM-LAMP) assay offers a highly sensitive and specific noninvasive method for detecting Helicobacter pylori (H. pylori) infection in saliva. This advancement surpasses PCR and conventional LAMP in sensitivity.
Area of Science:
- Microbiology
- Molecular Diagnostics
- Biotechnology
Background:
- Helicobacter pylori infection affects over half the global population.
- H. pylori is linked to gastric ulcers and gastric cancer.
- Current diagnostic methods for H. pylori are invasive and difficult.
Purpose of the Study:
- To develop a sensitive and specific noninvasive assay for H. pylori detection.
- To evaluate the performance of a novel colorimetric magneto loop-mediated isothermal amplification (CM-LAMP) assay.
- To assess the feasibility of using saliva samples for H. pylori diagnosis.
Main Methods:
- Design and implementation of a colorimetric magneto loop-mediated isothermal amplification (CM-LAMP) assay.
- Utilizing magnetic nanoparticles for solid-phase amplification of H. pylori DNA.
- Colorimetric signal development using NeutrAvidin horseradish peroxidase conjugate and ABTS.
Main Results:
- The CM-LAMP assay demonstrated high specificity, detecting only H. pylori isolates among tested microorganisms.
- The assay achieved 100-fold greater sensitivity than PCR and conventional LAMP.
- The limit of detection was as low as 40 CFU/ml or 0.2 CFU per reaction in spiked saliva samples.
Conclusions:
- The CM-LAMP assay is a highly sensitive and specific noninvasive method for H. pylori detection.
- Saliva samples can be effectively used for molecular diagnosis of H. pylori infection.
- This novel platform shows promise for improved H. pylori diagnostics.
