Rapid Raman spectroscopy-based test for antimicrobial resistance
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Abstract
Antimicrobial resistance (AMR) is one of the top global health threats. In 2019, AMR was associated with 4.95 million deaths, of which 1.97 million were caused by drug-resistant infections directly. The main subset of AMR is antibiotic resistance, that is, the resistance of bacteria to antibiotic treatment. Traditional and most commonly used antibiotic susceptibility tests are based on the detection of bacterial growth and its inhibition in the presence of an antimicrobial. These tests typically take over 1–2 days to perform, so empirical therapy schemes are often administered before proper testing. Rapid tests for AMR are necessary to optimize the treatment of bacterial infection. Here, we combine the MTT test with Raman spectroscopy to provide a 1.5 h long test for minimal inhibitory concentration determination. Several Escherichia coli and Klebsiella pneumoniae strains were tested with three types of antibiotics, including ampicillin from penicillin family, kanamycin from aminoglycoside family and levofloxacin from fluoroquinolone family. The test provided the same minimal inhibitory concentrations as traditional Etest confirming its robustness.
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The method provided reliable MIC values for several strains of E.coli and K.pneumoniae and three types of antibiotics, including ampicillin from penicillin family, kanamycin from aminoglycoside family and levofloxacin from fluoroquinolone family.
The table above (table 1) shows that levofloxacin MIC measured by Raman and E-test are similar. Levofloxacin exhibits concentration dependent killing which, in theory would be easier to measure in a rapid 1.5 hour test like the one proposed here. You mention ampicillin was also tested but I don't see that data here. This would be interesting to compare beacause ampicillin pharmacodynamics operate on a time > MIC principle, and so it would be even more impressive to detect resistance with a rapid, 1.5 hour test.
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Figure 3.
It appears that the data point for the absorbance measurement at the highest MFU is missing in (C) -- I think because it is covered by the data point from the Raman data? This nicely shows that the two methods have the same qualitative results, but quantifying the correlation would also be good.
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637 nm
Based on the name of the instrument (RL637), I think the earlier mentions of 633nm should instead be 637nm, as stated here and throughout the rest of the manuscript.
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a peak at 967 cm−1 for 532 nm excitation wavelength and the peaks at 722 cm−1 and 967 cm−1 for 633 nm excitation wavelength are valuable for a quantitative analysis40.
Additionally, if there are two measurable peaks when exciting with 633nm vs. only one measurable peak for 532nm, is there any advantage to exciting with 532nm?
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Optimal wavelengths for MTT and formazan spectra measurements are 532 and 633 nm respectively.633 nm laser provides a resonance Raman spectrum of formazan39. Comparable sensitivities for both MTT and formazan could be obtained using excitation wavelength of 532 nm.
Nice study! I am wondering though if you could you provide an explanation of how the sensitivity for these measurements is "comparable" for both 532nm and 633nm excitations when only 633nm has a resonance effect?
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