Differentiation of Gram-Negative Bacterial Aerosol Exposure Using Detected Markers in Bronchial-Alveolar Lavage Fluid

The identification of biosignatures of aerosol exposure to pathogens has the potential to provide useful diagnostic information. In particular, markers of exposure to different types of respiratory pathogens may yield diverse sets of markers that can be used to differentiate exposure. We examine a mouse model of aerosol exposure to known Gram negative bacterial pathogens, Francisella tularensis novicida and Pseudomonas aeruginosa. Mice were subjected to either a pathogen or control exposure and bronchial alveolar lavage fluid (BALF) was collected at four and twenty four hours post exposure. Small protein and peptide markers within the BALF were detected by matrix assisted laser desorption/ionization (MALDI) mass spectrometry (MS) and analyzed using both exploratory and predictive data analysis methods; principle component analysis and degree of association. The markers detected were successfully used to accurately identify the four hour exposed samples from the control samples. This report demonstrates the potential for small protein and peptide marker profiles to identify aerosol exposure in a short post-exposure time frame

Example mass spectra comparing of a portion of the mass range for a Fn-4 hrs sample (A) and a Pa-4 hrs sample (B).

<p>Example mass spectra comparing of a portion of the mass range for a Fn-4 hrs sample (A) and a Pa-4 hrs sample (B).</p

Total number of mononuclear (MN) and poly morphonuclear (PMN) cells counted in the BAL fluid at 4 h, 24 h, and 48 h for Fn and Fn-ATT and at 4 h and 24 h for Pa (A).

<p>Error bars indicate the standard error of the mean (SEM) for each. Percent PMN cells in BAL fluid (B).</p

Measurement of bacterial deposition in colony forming units (CFU) per lung for virulent (Fn) and attenuated (Fn-ATT) <i>F. novicida</i> and <i>P. pseudomonas</i> (Pa) with standard error of the mean (SEM) indicated for each measurement.

<p>Measurement of bacterial deposition in colony forming units (CFU) per lung for virulent (Fn) and attenuated (Fn-ATT) <i>F. novicida</i> and <i>P. pseudomonas</i> (Pa) with standard error of the mean (SEM) indicated for each measurement.</p

Distribution of replicate samples for seven classes of treatment: Control, PA-4 hrs, PA-24 hrs, Fn-4 hrs, Fn-24 hrs, Fn ATT-4 hrs, Fn ATT-24 hrs using two principle components.

<p>Distribution of replicate samples for seven classes of treatment: Control, PA-4 hrs, PA-24 hrs, Fn-4 hrs, Fn-24 hrs, Fn ATT-4 hrs, Fn ATT-24 hrs using two principle components.</p

The class predictions of the full 7 class Degree of Association model (Global Accuracy ∼75.0%).

<p>The class predictions of the full 7 class Degree of Association model (Global Accuracy ∼75.0%).</p