Simultaneous diagnosis and physiological profiling of diverse respiratory pathogen infections using multimolecular signatures in infrared spectroscopy.

Acute respiratory infections (ARIs), caused by diverse respiratory pathogens, pose a significant global public health challenge. Accurate and timely diagnosis of respiratory pathogen infections is crucial for effective treatment. Here, we present a versatile diagnostic platform that integrates portable attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy with advanced chemometric methods. Using nasopharyngeal swab samples from two clinical cohorts, we demonstrate the platform's capability to distinguish between infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-COV-2), Mycoplasma pneumoniae, influenza B, and healthy controls. This platform enables simultaneous detection and physiological profiling of respiratory pathogen infections within a single measurement. Infrared (IR) absorbance ratio method is applied to identify meaningful biomarkers that can contribute to more comprehensive clinical diagnosis. Additionally, we propose an important wavenumber range overlapping (IWRO) method to quantify the relative contributions of various biomolecules, including lipids, proteins, nucleic acids, and carbohydrates. These complementary analytical methodologies, based on the multiclass molecular signatures, aid in exploring the infection-induced physiological changes and pathological mechanisms. The diagnostic performance is validated using partial least squares discriminant analysis (PLS-DA), achieving classification accuracies of 95.6% and 90.9% in the two cohorts, respectively. This work significantly expands the application of IR spectroscopy in the simultaneous detection and profiling of diverse respiratory pathogens, providing critical insights into pathogen-host interactions with important implications for future pandemic preparedness.