Open AccessNanoarray Digital Polymerase Chain Reaction with High-Resolution Melt for Enabling Broad Bacteria Identification and Pheno–Molecular Antimicrobial Susceptibility Test
Author(s) -
Pornpat Athamanolap,
Kuangwen Hsieh,
Christine M. O’Keefe,
Ye Zhang,
Samuel Yang,
TzaHuei Wang
Publication year - 2019
Publication title -
analytical chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.117
H-Index - 332
eISSN - 1520-6882
pISSN - 0003-2700
DOI - 10.1021/acs.analchem.9b02344
Subject(s) - digital polymerase chain reaction , bacteria , polymerase chain reaction , computational biology , nucleic acid , antimicrobial , molecular diagnostics , antibiotics , microbiology and biotechnology , high resolution , identification (biology) , chemistry , biology , genetics , gene , botany , remote sensing , geology
Toward combating infectious diseases caused by pathogenic bacteria, there remains an unmet need for diagnostic tools that can broadly identify the causative bacteria and determine their antimicrobial susceptibilities from complex and even polymicrobial samples in a timely manner. To address this need, a microfluidic and machine-learning-based platform that performs broad bacteria identification (ID) and rapid yet reliable antimicrobial susceptibility testing (AST) is developed. Specifically, this platform builds on "pheno-molecular AST", a strategy that transforms nucleic acid amplification tests (NAATs) into phenotypic AST through quantitative detection of bacterial genomic replication, and utilizes digital polymerase chain reaction (PCR) and digital high-resolution melt (HRM) to quantify and identify bacterial DNA molecules. Bacterial species are identified using integrated experiment-machine learning algorithm via HRM profiles. Digital DNA quantification allows for rapid growth measurement that reflects susceptibility profiles of each bacterial species within only 30 min of antibiotic exposure. As a demonstration, multiple bacterial species and their susceptibility profiles in a spiked-in polymicrobial urine specimen were correctly identified with a total turnaround time of ∼4 h. With further development and clinical validation, this platform holds the potential for improving clinical diagnostics and enabling targeted antibiotic treatments.