Raman microscopic spectroscopy as a diagnostic tool to detect Staphylococcus epidermidis in bone grafts

Abstract

IntroductionRaman microscopic spectroscopyis a new approach for further characterization and detection of molecular features in many pathological processes. This technique has been successfully applied to scrutinize the spatial distribution of small molecules and proteins within biological systems by in situ analysis. This study uses Raman microscopic spectroscopyto identify any in-depth benefits and drawbacks in diagnosing Staphylococcus epidermidis in human bone grafts. Material and Methods40 non-infected human bone samples and 10 human bone samples infected with Staphylococcus epidermidis were analyzed using Raman microscopic spectroscopy. Reflectance data were collected between 200 cm−1 and 3600 cm−1 with a spectral resolution of 4 cm−1 using a Senterra II microscope (Bruker, Ettlingen, Germany). The acquired spectral information was used for spectral and unsupervised classification, such as principal component analysis. ResultsRaman measurements produced distinct diagnostic spectra that were used to distinguish between non-infected human bone samples and Staphylococcus epidermidis infected human bone samples by spectral and principal component analyses. A substantial loss in bone quality and protein conformation was detected by human bone samples co-cultured with Staphylococcus epidermidis. The mineral-to-matrix ratio using the phosphate/Amide I ratio (p = 0.030) and carbonate/phosphate ratio (p = 0.001) indicates that the loss of relative mineral content in bones upon bacterial infection is higher than in non-infected human bones. Also, an increase of alterations in the collagen network (p = 0.048) and a decrease in the structural organization and relative collagen in infected human bone could be detected. Subsequent principal component analyses identified Staphylococcus epidermidis in different spectral regions, respectively, originating mainly from CH2 deformation (wagging) of protein (at 1450 cm−1) and bending and stretching modes of C-H groups (∼2800-3000 cm−1). ConclusionRaman microscopic spectroscopyis presented as a promising diagnostic tool to detect Staphylococcus epidermidis in human bone grafts. Further studies in human tissues are warranted.