The use of Raman spectroscopy in the epidemiology of methicillin-resistant Staphylococcus aureus of human- and animal-related clonal lineages

AbstractIn order to perform a cost-effective search and destroy policy for methicillin-resistant Staphylococcus aureus (MRSA), a quick and reliable typing method is essential. In an area with a high level of animal-related MRSA ST398, pulsed field gel electrophoresis (PFGE) typing and spa-typing are not sufficient to discriminate between co-incidental findings and true transmission of MRSA. This study is the first to retrospectively show the performance of Raman spectroscopy in 16 well-documented outbreaks. We analysed 525 isolates, 286 MRSA ST398 and 239 from other PFGE clusters with Raman spectroscopy. When epidemiologically linked isolates from the outbreaks were analysed with PFGE as the reference standard, Raman spectroscopy correctly identified 97% of cases that were indistinguishable from the index case. With Raman cluster analysis, the most dominant distinction was between MRSA ST398 and other MRSA of human clonal lineages. Within MRSA ST398, 22 different Raman clusters were identified. Raman typing correctly identified an ST398 (spa type t567) outbreak in a hospital setting. No direct correlation was observed between Raman clusters and spa types. We conclude that Raman spectroscopy is a quick and reliable method of MRSA typing, which can be used in outbreak settings and it is comparable to PFGE, with the added advantage that PFGE non-typeable isolates can also be readily typed using the same sample preparation protocol

Detection and differentiation of causative organisms of onychomycosis in an ex vivo nail model by means of Raman spectroscopy

BackgroundOnychomycosis is worldwide the most prevalent infection of the nail. It is mainly caused by the dermatophytes Trichophyton rubrum and Trichophyton mentagrophytes and to a lesser extent Trichophyton tonsurans. The yeast Candida albicans and the mould Scopulariopsis brevicaulis can also cause onychomycosis. Management of these nail conditions may require appropriate treatment methods and therefore the identification of the causative species can be of importance. However, the determination of agents causing onychomycosis is still not optimal. ObjectivesTo detect and differentiate causative organisms of onychomycosis in an ex vivo nail model by means of Raman spectroscopy. The work focusses is on the discriminative power of Raman spectroscopy for detection of differences between T. rubrum, T. mentagrophytus and T. tonsurans on human nail and distinguishing these dermatophytic from the non-dermatophytic species S. brevicaulis and C. albicans. MethodsRaman spectra (200/sample) were taken from 50-m human nail slices infected with T. rubrum, T. mentagrophytus, T. tonsurans, S. brevicaulis or C. albicans using a 2500 High-Performance Raman Module and 785-nm diode laser. Processed spectra were analysed by sorting the correlation matrix and presented as dendrogram and heat map. Raman spectra from suspended dermatophytic microconidia were taken for mutual comparisons. ResultsSpectral differences between the dermatophytes T. rubrum, T. mentagrophytus and T. tonsurans (635-795, 840-894, 1018-1112, 1206-1372, 1566-1700/cm) and the non-dermatophytes S. brevicaulis and C. albicans (442-610, 692-758, 866-914, 1020-1100, 1138-1380,1492-1602/cm) growing on nail were confirmed by clustering correlation showing two main clusters. Dissimilarities between tested dermatophytes were also found with T. rubrum being most different. Raman spectra of the dermatophytic microconidia varied over the whole tested 400-1800/cm range. ConclusionImportant dermatophytic and non-dermatophytic agents of onychomycosis growing on ex vivo human nail can be distinguished specifically and non-invasively by Raman spectroscopy

Rapid Identification of Mycobacteria by Raman Spectroscopy▿

A number of rapid identification methods have been developed to improve the accuracy for diagnosis of tuberculosis and to speed up the presumptive identification of Mycobacterium species. Most of these methods have been validated for a limited group of microorganisms only. Here, Raman spectroscopy was compared to 16S rRNA sequencing for the identification of Mycobacterium tuberculosis complex strains and the most frequently found strains of nontuberculous mycobacteria (NTM). A total of 63 strains, belonging to eight distinct species, were analyzed. The sensitivity of Raman spectroscopy for the identification of Mycobacterium species was 95.2%. All M. tuberculosis strains were correctly identified (7 of 7; 100%), as were 54 of 57 NTM strains (94%). The differentiation between M. tuberculosis and NTM was invariably correct for all strains. Moreover, the reproducibility of Raman spectroscopy was evaluated for killed mycobacteria (by heat and formalin) versus viable mycobacteria. The spectra of the heat-inactivated bacteria showed minimal differences compared to the spectra of viable mycobacteria. Therefore, the identification of mycobacteria appears possible without biosafety level 3 precautions. Raman spectroscopy provides a novel answer to the need for rapid species identification of cultured mycobacteria in a clinical diagnostic setting

Rapid Identification of Mycobacteria by Raman Spectroscopy▿

A number of rapid identification methods have been developed to improve the accuracy for diagnosis of tuberculosis and to speed up the presumptive identification of Mycobacterium species. Most of these methods have been validated for a limited group of microorganisms only. Here, Raman spectroscopy was compared to 16S rRNA sequencing for the identification of Mycobacterium tuberculosis complex strains and the most frequently found strains of nontuberculous mycobacteria (NTM). A total of 63 strains, belonging to eight distinct species, were analyzed. The sensitivity of Raman spectroscopy for the identification of Mycobacterium species was 95.2%. All M. tuberculosis strains were correctly identified (7 of 7; 100%), as were 54 of 57 NTM strains (94%). The differentiation between M. tuberculosis and NTM was invariably correct for all strains. Moreover, the reproducibility of Raman spectroscopy was evaluated for killed mycobacteria (by heat and formalin) versus viable mycobacteria. The spectra of the heat-inactivated bacteria showed minimal differences compared to the spectra of viable mycobacteria. Therefore, the identification of mycobacteria appears possible without biosafety level 3 precautions. Raman spectroscopy provides a novel answer to the need for rapid species identification of cultured mycobacteria in a clinical diagnostic setting

Epidemiology of Staphylococcus aureus harboring the mecA or Panton-Valentine leukocidin genes in hospitals in Java and Bali, Indonesia

Data of Staphylococcus aureus carriage in Indonesian hospitals are scarce. Therefore, the epidemiology of S. aureus among surgery patients in three academic hospitals in Indonesia was studied. In total, 366 of 1,502 (24.4%) patients carried S. aureus. The methicillin-resistant S. aureus (MRSA) carriage rate was 4.3%, whereas 1.5% of the patients carried Panton-Valentine leukocidin (PVL)-positive methicillin-sensitive S. aureus (MSSA). Semarang and Malang city (odds ratio [OR] 9.4 and OR 9.0), being male (OR 2.4), hospitalization for more than 5 days (OR 11.708), and antibiotic therapy during hospitalization (OR 2.6) were independent determinants for MRSA carriage, whereas prior hospitalization (OR 2.5) was the only one risk factor for PVL-positive MSSA carriage. Typing of MRSA strains by Raman spectroscopy showed three large clusters assigned type 21, 24, and 38, all corresponding to ST239-MRSA-SCCmec type III. In conclusion, MRSA and PVL-positive MSSA are present among patients in surgical wards in Indonesian academic hospitals. Copyrigh