# High-Wavenumber Raman Spectroscopy for the Identification and Biochemical Characterization of Microbial Species

## TL;DR

High-wavenumber Raman spectroscopy can accurately identify and characterize microbial species without damaging them.

## Contribution

The study introduces a multitiered classification model and biochemical characterization method using high-wavenumber Raman data.

## Key findings

- High-wavenumber Raman achieved 97.4% accuracy in identifying microbial species.
- Spectral unmixing revealed biochemical components like proteins and lipids for each species.
- Raman dry mass characterization of E. coli matched previously reported values.

## Abstract

Raman spectroscopy
is a powerful tool for microbiological
and infectious
disease research, enabling rapid discrimination of microbial species.
While spectral discrimination has typically been performed using the
fingerprint region (400–1800 cm–1), high
autofluorescence backgrounds can degrade signal quality and decrease
the overall effectiveness. This work investigates the effectiveness
of utilizing the low-background high-wavenumber region (2800–3800
cm–1) to both identify and biochemically characterize
microbial species. High-wavenumber spectra of 14 microbial species
were collected and used to train and validate a multitiered classification
model capable of identifying cell wall type (100%), genus (98.9%),
and species (97.4%) with high accuracy. Additionally, utilizing a
spectral unmixing approach, the relative Raman contributions from
proteins, carbohydrates, nucleic acids, lipids, and cell wall components
were determined for each species, with general trends matching reported
physiological differences. Utilizing a method of converting high-wavenumber
Raman spectral fractions to relative dry mass, a biochemical characterization
of each species was obtained, with the Raman dry mass characterization
of Escherichia coli (E. coli) closely
matching previously reported values. Taken together, these results
demonstrate that high-wavenumber Raman spectroscopy is a feature-rich
technique capable of performing both accurate discrimination and nondestructive
biochemical characterization of microbial species.