Feasibility study demonstrating that enzymatic template generation and amplification can be employed as a novel method for molecular antimicrobial susceptibility testing

Abstract

BackgroundAntimicrobial Susceptibility Testing (AST) is a methodology in which the sensitivity of a microorganism is determined via its inability to proliferate in the presence of an antimicrobial agent. Results are reported as minimum inhibitory concentrations (MICs). The present study demonstrates that measurement of DNA polymerase activity via Enzymatic Template Generation and Amplification (ETGA) can be used as a novel means of determining the MIC of a microbe to an antibiotic agent much sooner than the current standardized method.MethodsTime course analysis of ETGA is presented from bacterial cultures containing antibiotic agents and compared to the end-point results of standard macrobroth method AST.ResultsMIC determinations from ETGA results at 4, 6, and 22 hours are compared to the MICs from the standard method and the results are shown to be in agreement. Additionally, reliable AST analysis using ETGA can be performed on bacteria harvested directly from spiked blood cultures.ConclusionsAST analysis with ETGA is shown to be equivalent to AST analysis using gene-specific qPCR assays against the measured microbe. Future development of this novel method for performing AST in a clinical setting is discussed.

Rapid Phenotypic Antibiotic Susceptibility Testing of Uropathogens Using Optical Signal Analysis on the Nanowell Slide

“…Numerous molecular methods using genomic markers have been exploited primarily for microbial identification, but also for AST. Molecular ASTs are generally faster than traditional methods as they usually do not require a previous culturing step ( Sodowich et al, 2013 ; Kostić et al, 2015 ). However, their dependence on the detection of genetic markers restricts molecular ASTs to the current knowledge of resistance mechanisms.…”