Towards Raman-based epidemiological typing of Pseudomonas aeruginosa.

Raman spectra of bacteria can be used as highly specific fingerprints, enabling discrimination at strain level. Pseudomonas aeruginosa strains can be strongly pigmented, making it difficult to obtain high quality spectra of such isolates due to high fluorescent spectral backgrounds. Furthermore, the spectra that could be measured with acceptable quality often showed large spectral variations limiting the reproducibility required for strain level discrimination. P. aeruginosa produces a characteristic yellowish green fluorescent pigment, called pyoverdin. Applying a washing procedure to reduce the amount of fluorescent pigment, enabled the highly pigmented isolates to be measured with sufficient spectral quality. Isolation of the pigment/pyoverdin spectral features, together with spectral scaling methods improved reproducibility. It will be important to analyze the range of the spectral variations that can occur and ensure the correction of all of these factors to obtain the highest reproducibility required for strain level typing.

Optical fingerprinting in bacterial epidemiology: Raman spectroscopy as a real-time typing method.

Hospital-acquired infections (HAI) increase morbidity and mortality and constitute a high financial burden on health care systems. An effective weapon against HAI is early detection of potential outbreaks and sources of contamination. Such monitoring requires microbial typing with sufficient reproducibility and discriminatory power. Here, a microbial-typing method is presented, based on Raman spectroscopy. This technique provides strain-specific optical fingerprints in a few minutes instead of several hours to days, as is the case with genotyping methods. Although the method is generally applicable, we used 118 Staphylococcus aureus isolates to illustrate that the discriminatory power matches that of established genotyping techniques (numerical index of diversity [D]=0.989) and that concordance with the gold standard (pulsed-field gel electrophoresis) is high (95%). The Raman clustering of isolates was reproducible to the strain level for five independent cultures, despite the various culture times from 18 h to 24 h. Furthermore, this technique was able to classify stored (-80 degrees C) and recent isolates of a methicillin-resistant Staphylococcus aureus-colonized individual during surveillance studies and did so days earlier than established genotyping techniques did. Its high throughput and ease of use make it suitable for use in routine diagnostic laboratory settings. This will set the stage for continuous, automated, real-time epidemiological monitoring of bacterial infections in a hospital, which can then be followed by timely corrective action by infection prevention teams.