Molecular diagnostics for infectious disease

Molecular diagnostic methods have been used in clinical and environmental settings for over three decades. Conventional approaches, which often involve long processing times and limited sensitivity, are gradually being replaced by molecular tests that offer improved analytical performance. Nucleic acid testing (NAT), particularly PCR-based methods and their variants such as quantitative PCR (qPCR) and reverse transcription PCR (RT-PCR), have demonstrated higher sensitivity and specificity than conventional assays. Recent developments in next-generation sequencing (NGS), including metagenomic and whole-genome sequencing, have also enhanced the identification of complex pathogens and resistance mechanisms. Point-of-care testing (POCT) is another advancement, supported by newly developed isothermal amplification (IA) techniques such as loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA), which allow rapid detection in decentralized settings. In addition, newer approaches such as droplet digital PCR (ddPCR) and CRISPR-based diagnostics are very promising due to their potential to provide accurate nucleic acid detection in low amounts of target. Aptamer-based sensors are also under extensive research and development for their application in diagnosis of infection. This review outlines both traditional and recent advanced molecular diagnostic tools, with a focus on their application to infectious disease detection.