Raman Analysis of RNA Nucleotide Strands From SARS-CoV-2 Variants and Subvariants: A Step Forward in the Definition of "Raman Genome".

Raman spectra of RNA nucleotide strands extracted from 20 SARS-CoV-2 strains, including the original Japanese isolate, Alpha, Beta, Gamma, Delta, Lambda, Theta, Mu, and 12 Omicron subvariants, were systematically collected, deconvoluted, and converted into Raman barcodes. The spectra exhibited common features associated with the RNA backbone, ribofuranose rings, and nitrogenous bases, reflecting the overall structural similarity of the viral RNAs. However, distinct differences were observed in spectral signatures related to beta-D-ribofuranose vibrations, phosphate linkages, and RNA bases sensitive to secondary-structure variations. Conventional principal component analysis provided only limited discrimination among variants and subvariants. In contrast, Raman barcodes captured subtle molecular-scale structural differences and successfully distinguished all investigated strains according to their RNA secondary structures. The analysis identified vibrational markers associated with nucleotide chain length, backbone conformation, and ribofuranose dynamics. Several backbone- and base-related signals exhibited hyperchromic behavior correlated with RNA secondary-structure features, including hairpin content and stem length. These relationships may serve as indicators for the rapid detection of emerging variants. By encoding key spectral signatures, Raman barcodes provide a robust framework for accurate classification of SARS-CoV-2 strains based on the molecular architecture of their RNAs.