Mechanically ventilated surgical patients have a variety of bacterial flora that are often undetectable by traditional culture methods. The source of infection in many of these patients remains unclear. To address this clinical problem, the microbiome profile and host inflammatory response in bronchoalveolar lavage samples from the surgical intensive care unit were examined relative to clinical pathology diagnoses. The hypothesis was tested that clinical diagnosis of respiratory tract flora were similar to culture positive lavage samples in both microbiome and inflammatory profile. Bronchoalveolar lavage samples were collected in the surgical intensive care unit as standard of care for intubated individuals with a clinical pulmonary infection score of >6 or who were expected to be intubated for >48 hours. Cytokine analysis was conducted with the Bioplex Pro Human Th17 cytokine panel. The microbiome of the samples was sequenced for the 16S rRNA region using the Ion Torrent. Microbiome diversity analysis showed the culture-positive samples had the lowest levels of diversity and culture negative with the highest based upon the Shannon-Wiener index (culture positive: 0.77 ± 0.36, respiratory tract flora: 2.06 ± 0.73, culture negative: 3.97 ± 0.65). Culture-negative samples were not dominated by a single bacterial genera. Lavages classified as respiratory tract flora were more similar to the culture-positive in the microbiome profile. A comparison of cytokine expression between groups showed increased levels of cytokines (IFN-g, IL-17F, IL-1B, IL-31, TNF-a) in culture-positive and respiratory tract flora groups. Culture-positive samples exhibited a more robust immune response and reduced diversity of bacterial genera. Lower cytokine levels in culture-negative samples, despite a greater number of bacterial species, suggest a resident nonpathogenic bacterial community may be indicative of a normal pulmonary environment. Respiratory tract flora samples were most similar to the culture-positive samples and may warrant classification as culture-positive when considering clinical treatment.