Label-free Raman Spectrometric Detection and Differentiation Between Bacteria Staphylococcus Aureus and Neisseria Gonorrhoeae

Abstract

This work reports on the application of Raman spectroscopy in distinguishing rapidly between Staphylococcus aureus and Neisseria gonorrhoeae bacterial strains. The method was also explored in segregating between drug-responsive and drug-resistant Staphylococcus aureus strains. Currently, such tasks take several days and involve the use of labels in methods such as Polymerase Chain Reaction (PCR), Verigene Gram-positive blood culture (BC-GP) assay, and Alere’s BinaxNOW. The samples were applied onto conductive silver paste-smeared glass slides then excited with a 785 nm laser and the Raman scattered radiation was collected, recorded, and analyzed. The Raman spectral data analyses were done using ANOVA and PCA which were able to reveal a distinction between Staphylococcus aureus and Neisseria gonorrhoeae. The bands that were uniquely identified to Staphylococcus aureus had their wavenumbers centered at 800cm-1, 846cm-1, and 1028cm-1. These bands displayed high-intensity variations in ANOVA and high-loading signals in PCA. They are attributed to Tyrosine (800cm-1) and proteins (846, 1028cm-1), and their vibrational bands were tentatively assigned to C-N stretch, ring breathing, and C-C breathing respectively. Raman spectra of Neisseria gonorrhoeae exhibited unique bands centered at 635cm-1, 710 cm-1, 941 cm-1, 1232 cm-1 and 1320 cm-1 which were attributed to Tyrosine(635 cm-1), Polysaccharides (710 cm-1), protein (941 cm-1), Amide III, adenine, polyadenine (1232 cm-1) and DNA (1320 cm-1). Their vibrational assignments are the aromatic ring skeletal, CH2 Rock, C-N ring, and NH2 respectively. These two sets of bands can be used as the biomarker bands for Staphylococcus aureus and Neisseria gonorrhoeae. For the drug-resistant and the drug-responsive strains of Staphylococcus aureus bacteria, the peaks that were found to have high variations in terms of intensities in ANOVA and high loading signals in PCA and associated with thedrug resistant strain were centered at 720cm-1, 950cm-1, and 1320cm-1. These bands are attributed to polysaccharides (720cm-1), proteins (950cm-1), and DNA (1320cm-1), while their vibrational assignments are CH2 Rock, C-C ring breathing, and NH2 respectively. Similarly, the bands identified to the drug-responsive strain are 805cm-1 (proteins), 1035cm-1 (proteins), and 1443cm-1 (phospholipids, deformation mode of proteins), and vibrationally assigned to C-N stretch, C-C ring breathing, and CH2 in that order. These results indicate that the Raman spectroscopy method is capable of rapidly identifying the biomarker bands that can be used in the diagnosis of bacteria strains in hospitals, hence the time used in diagnosis is significantly reduced