Table 1.
Partial inventory of contemporary, near-future, and long-term alternative methodologies for antimicrobial susceptibility testinga
| Antimicrobial testing technology | Test principle | Needs more than 105 cells | POP or CA | Cost | Automatic or manual | Heteroresistance detection | Real MIC | Test time (h) |
|---|---|---|---|---|---|---|---|---|
| Currently in use | ||||||||
| Agar dilution testing | Growth inhibition on solid medium with antibiotics | Y | CA | L | M | + | Y | >10 |
| Automated testing (Vitek, Phoenix, MicroScan) | Monitoring of growth or substrate conversion in a dedicated machine using optics | Y | CA | L | A | ± | Y/N | <10 |
| Broth dilution testing | Growth inhibition in liquid medium with antibiotics | Y | CA | L | M/A | ± | Y | >10 |
| Chromogenic agars | Metabolic conversion of chromogenic compounds in agar medium | Y | CA | I | M | + | N | >10 |
| Disk diffusion | Measurement of growth inhibition around an antibiotic-containing disk | Y | CA | L | M/A | + | Y/N | >10 |
| Etest | Measurement of growth inhibition around a strip containing an antibiotic gradient. | Y | CA | L | M/A | + | Y | >10 |
| Fluorescent live/dead staining | Microscopy of (non)permeable cells in the presence of fluorescent stains | N | CA | L | M | − | N | <1 |
| PCR gene detection | DNA amplification | N | CA | I | A/M | − | N | <1 |
| Real-time microscopy | Filming bacterial division at the single-cell level | N | CA | L | M | − | N | <1 |
| Near-future alternatives | ||||||||
| Calorimetrics | Detection of heat produced by stressed bacteria | N | POP | NK | A | − | N | <5 |
| Cantilever technology | Weighing bacterial cells by changes in cantilever vibrations | N | POP | NK | A | ± | Y | <5 |
| FACS | Sizing and measuring differential fluorescence between living and dead cells | N | CA/POP | I | A | − | Y/N | <5 |
| Magnetic bead spin | Changes in spin of beads in a magnetic field as a function of the no. of attached bacteria | Y | POP | NK | A | − | N | <5 |
| MALDI-TOF MS | Detection of antibiotic degradation products | Y | POP | L | A | − | N | <5 |
| Microdroplets | Monitoring of growth or substrate conversion in nanoliter droplets | N | POP | NK | A | ± | Y/N | <5 |
| Next-generation sequencing | Sequencing of all cellular DNA and RNA | N | CA/POP | H | A | − | N | >10 |
| Long-term alternatives | ||||||||
| Apoptosis markers | Detection of compounds produced upon programmed cell death | Y | POP | I | M/A | − | N | <1 |
| Bacteriophage amplification | Detection of phage reproduction in living cells only | N | CA | I | M | − | N | <10 |
| Colorimetric detection of cell respiration | Optical detection of substrate or indicator color change at active cell respiration | Y | POP | L | A | − | N | <1 |
| Electronic noses | Direct detection of volatile organic compounds | Y | POP | L | M/A | − | Y | <1 |
| Impedance measurements | Changes in electrical characteristics of suspension with living or dead cells | Y | POP | NK | A | − | N | <5 |
| Infrared spectroscopy | Absorption characteristics of bacteria exposed to IR | N | CA/POP | I | A | ± | N | <5 |
| LC-ESI MS | Proteomics of living/dead cells and resistance proteins | Y | POP | H | A | − | N | <5 |
| Metabolomics (including ROS) | Detection of changes in intracellular composition focused on small molecules | Y | POP | NK | A | − | N | <1 |
| Microsound measurements | Measuring vibrational differences between living and dead cells | N | POP | NK | A | − | N | <5 |
| NMR | Assessment of molecular composition of complex mixtures | Y | POP | NK | A | − | N | >10 |
| Raman spectroscopy | Absorption characteristics of bacteria exposed to laser light | N | CA/POP | L | A | ± | N | <5 |
| RNA sequencing | Definition of gene expression differences by sequencing | N | POP | H | A | − | N | >10 |
a
FACS, fluorescence-activated cell sorting; MALDI-TOF MS, matrix-assisted laser desorption ionization–time of flight mass spectrometry; LC-ESI MS, liquid chromatography-electron spray ionization mass spectrometry; ROS, reactive oxygen species; NMR, nuclear magnetic resonance; Y, yes; N, no; POP, proof of principle; CA, commercially available; H, high; L, low; I, intermediate; NK, not known; +, detects heteroresistance; ±, may detect heteroresistance; −, fails to detect heteroresistance.