Acinetobacter baumannii is an important nosocomial pathogen that causes a range of infections, including respiratory and urinary tract infections, meningitis, endocarditis, wound infections, and bacteremia. In fact, A. baumannii is now responsible for up to 20% of all intensive care unit infections in some regions of the world with pneumonia being the most common presentation. Additionally, A. baumannii is a considerable cause of infection in combat soldiers and is the most common gram-negative bacillus recovered from traumatic injuries to extremities obtained on the battlefield. The clinical significance of A. baumannii has been propelled by this organism's rapid acquisition of resistance to virtually all antibiotics. Taken together, these facts have established A. baumannii as a significant threat to the health of Veterans. Based on this, we have recently initiated a research program focused on identifying novel targets for therapeutic intervention against A. baumannii. In this application, we describe our discovery of an immune enhancing biologic that has tremendous therapeutic efficacy against A. baumannii pneumonia and can cure this infection in mice. We have made the exciting discovery that transposon mutagenesis of A. baumannii severely attenuates its virulence, regardless of the site of transposon insertion. Moreover, strains that have experienced transposon mutagenesis are capable of curing infections caused by wildtype A. baumannii during co-infection experiments, and this attenuating phenotype does not require that the transposon mutagenized strain is viable. This attenuation requires exposure of a DNA-protein complex on the surface of A. baumannii, and relies on key innate immune signaling pathways in the host. Finally, we have found that transposon mutagenesis of A. baumannii leads to up-regulation of genes encoding for the Type IV secretion system (T4SS) and increased abundance of pilus-like surface appendages visible by electron microscopy. Based on these findings, we propose a model whereby A. baumannii up-regulates its T4SS upon exposure to exogenous DNA, and this secretion system is recognized by the innate immune system of the host to coordinate clearance of the invading pathogen. To test this model we propose a series of three integrated Specific Aims.
In Aim 1 we will define the immune response elicited by A. baumannii T4SS hyperexpressors. Results obtained from these experiments may lead to the rational design of immunomodulatory therapies for the treatment of bacterial pneumonia.
In Aim 2 we will elucidate the mechanism by which transposition alters T4SS expression. In these experiments we will determine the mechanism by which the T4SS is up-regulated upon exposure to foreign DNA and interrogate the impact of the A. baumannii T4SS on DNA transfer and exchange. These results will lay the foundation for studies focused on reducing DNA transfer and preventing the acquisition of antimicrobial resistant determinants by this organism. Finally, in Aim 3 we will determine the broad spectrum efficacy of T4SS-based immune enhancing biologics. A. baumannii transposon mutants exhibit therapeutic efficacy against pneumonia caused by Pseudomonas aeruginosa suggesting that this strategy has broad applicability against a variety of infectious agents. In addition to the potential clinical benefits of this discovery, these proposed experiments will provide us with a tool to study the host-pathogen interaction during infection with the goal of defining a successful immune response to pneumonia caused by Gram negative pathogens.

Public Health Relevance

Acinetobacter baumannii is responsible for up to 20% of intensive care unit infections worldwide, and this organism is a leading cause of infection in combat soldiers wounded during battle. The public health threat of this organism is compounded by its propensity to develop antimicrobial resistance; in fact isolates of A. baumannii have emerged that are resistant to all clinically relevant classes of antibiotics. Taken together, these facts underscore the threat of A. baumannii to the health of United States Veterans. In this proposal, we describe our discovery of an innate immune enhancing biologic for the treatment of A. baumannii pneumonia. We propose to use this reagent as a tool to define the characteristics of a successful immune response to bacterial pathogens of the lung. Taken together, these studies may lay the foundation for the development of a broad-spectrum strategy for the treatment of pneumonia caused by a variety of bacterial pathogens.

Agency
National Institute of Health (NIH)
Institute
Veterans Affairs (VA)
Type
Non-HHS Research Projects (I01)
Project #
5I01BX002482-03
Application #
9275422
Study Section
Infectious Diseases B (INFB)
Project Start
2014-10-01
Project End
2018-09-30
Budget Start
2016-10-01
Budget End
2017-09-30
Support Year
3
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Veterans Health Administration
Department
Type
Independent Hospitals
DUNS #
156385783
City
Nashville
State
TN
Country
United States
Zip Code
37212
Zackular, Joseph P; Moore, Jessica L; Jordan, Ashley T et al. (2016) Dietary zinc alters the microbiota and decreases resistance to Clostridium difficile infection. Nat Med 22:1330-1334
Spraggins, Jeffrey M; Rizzo, David G; Moore, Jessica L et al. (2016) Next-generation technologies for spatial proteomics: Integrating ultra-high speed MALDI-TOF and high mass resolution MALDI FTICR imaging mass spectrometry for protein analysis. Proteomics 16:1678-89
Zackular, Joseph P; Moore, Jessica L; Jordan, Ashley T et al. (2016) Erratum: Dietary zinc alters the microbiota and decreases resistance to Clostridium difficile infection. Nat Med 22:1502
Diaz-Ochoa, Vladimir E; Lam, Diana; Lee, Carlin S et al. (2016) Salmonella Mitigates Oxidative Stress and Thrives in the Inflamed Gut by Evading Calprotectin-Mediated Manganese Sequestration. Cell Host Microbe 19:814-25
Palmer, Lauren D; Skaar, Eric P (2016) Transition Metals and Virulence in Bacteria. Annu Rev Genet 50:67-91
Juttukonda, Lillian J; Skaar, Eric P (2015) Manganese homeostasis and utilization in pathogenic bacteria. Mol Microbiol 97:216-28
Noto, Michael J; Boyd, Kelli L; Burns, William J et al. (2015) Toll-Like Receptor 9 Contributes to Defense against Acinetobacter baumannii Infection. Infect Immun 83:4134-41
Zackular, Joseph P; Chazin, Walter J; Skaar, Eric P (2015) Nutritional Immunity: S100 Proteins at the Host-Pathogen Interface. J Biol Chem 290:18991-8
D'Orazio, Melania; Mastropasqua, Maria Chiara; Cerasi, Mauro et al. (2015) The capability of Pseudomonas aeruginosa to recruit zinc under conditions of limited metal availability is affected by inactivation of the ZnuABC transporter. Metallomics 7:1023-35