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Open Access Research article

Proteomic expression profiling of Haemophilus influenzae grown in pooled human sputum from adults with chronic obstructive pulmonary disease reveal antioxidant and stress responses

Jun Qu16, Alan J Lesse2347, Aimee L Brauer2, Jin Cao16, Steven R Gill56 and Timothy F Murphy2367*

Author Affiliations

1 Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, State University of New York, Buffalo, NY 14260 USA

2 Division of Infectious Diseases, Department of Medicine, School of Medicine and Biomedical Sciences, University at Buffalo, State University of New York, Buffalo, NY 14215 USA

3 Department of Microbiology, School of Medicine and Biomedical Sciences, University at Buffalo, State University of New York, Buffalo, NY 14215 USA

4 Department of Pharmacology & Toxicology, School of Medicine and Biomedical Sciences, University at Buffalo, State University of New York, Buffalo, NY 14215 USA

5 Department of Oral Biology, School of Dental Medicine, University at Buffalo, State University of New York, Buffalo, NY 14215 USA

6 New York State Center of Excellence in Bioinformatics and Life Sciences, 701 Ellicott Street, Buffalo NY 14203 USA

7 Veterans Affairs Western New York Healthcare System, 3495 Bailey Avenue, Buffalo, New York 14215 USA

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BMC Microbiology 2010, 10:162  doi:10.1186/1471-2180-10-162

Published: 1 June 2010

Abstract

Background

Nontypeable Haemophilus influenzae colonizes and infects the airways of adults with chronic obstructive pulmonary disease, the fourth most common cause of death worldwide.Thus, H. influenzae, an exclusively human pathogen, has adapted to survive in the hostile environment of the human airways.To characterize proteins expressed by H. influenzae in the airways, a prototype strain was grown in pooled human sputum to simulate conditions in the human respiratory tract.The proteins from whole bacterial cell lysates were solubilized with a strong buffer and then quantitatively cleaned with an optimized precipitation/on-pellet enzymatic digestion procedure.Proteomic profiling was accomplished by Nano-flow liquid chromatography/mass spectroscopy with low void volume and high separation efficiency with a shallow, long gradient.

Results

A total of 1402 proteins were identified with high confidence, including 170 proteins that were encoded by genes that are annotated as conserved hypothetical proteins.Thirty-one proteins were present in greater abundance in sputum-grown conditions at a ratio of > 1.5 compared to chemically defined media.These included 8 anti-oxidant and 5 stress-related proteins, suggesting that expression of antioxidant activity and stress responses is important for survival in the airways.Four proteins involved in uptake of divalent anions and 9 proteins that function in uptake of various molecules were present in greater abundance in sputum-grown conditions.

Conclusions

Proteomic expression profiling of H. influenzae grown in pooled human sputum revealed increased expression of antioxidant, stress-response proteins and cofactor and nutrient uptake systems compared to media grown cells.These observations suggest that H. influenzae adapts to the oxidative and nutritionally limited conditions of the airways in adults with chronic obstructive pulmonary disease by increasing expression of molecules necessary for survival in these conditions.