Dr. Wargo received his Ph.D. in 2005 from Dartmouth College, studying the regulation of eukaryotic flagellar motility with Dr. Elizabeth F. Smith. For postdoctoral research, he studied microbial pathogenesis with Dr. Deborah Hogan at Dartmouth Medical School and mammalian lung biology with Dr. Laurie Whittaker at UVM, before joining the Department of Microbiology and Molecular Genetics in 2009.

Research in my laboratory is aimed at understanding the role of bacterial catabolism in regulation of both virulent and non-virulence interactions of bacteria with eukaryotes. The focus of current research is to understand the intertwined regulation of bacterial virulence and bacterial catabolism of host-derived compounds during lung infection by Pseudomonas aeruginosa.

The organism and clinical importance:
Pseudomonas aeruginosa is a Gram-negative bacterium nearly ubiquitous in the environment. It is found in freshwater, soils, and man-made water fixtures (particularly faucets and showers), and is the most prevalent antibiotic resistant, Gram-negative, opportunistic pathogen of humans. However, as denoted by the term ‘opportunistic pathogen’, while we are continuously exposed to P. aeruginosa at a low level, healthy individuals do not develop disease. Infection by P. aeruginosa requires some breakdown of the innate immune system: immune deficiency, burns, chemotherapy, or inhibition of physical clearance mechanisms. Lung infection caused by P. aeruginosa is a source of significant morbidity and mortality in ventilated patients and is the primary cause of chronic progressive respiratory failure and death in patients with cystic fibrosis. It is the lung infections caused by P. aeruginosa that my lab studies.

Hypotheses guiding our research:

• Chemical communication between bacteria and eukaryotes alter the interaction.
• Bacteria can use host-derived metabolites to guide behaviors towards colonization, pathogenesis, symbiosis, or commensalism.
• The metabolic flexibility of P. aeruginosa enables it to thrive in the lung.
• Specific virulence genes of P. aeruginosa are induced by the lung environment, in particular, by metabolism of host-derived compounds.
• Development of novel agents to inhibit P. aeruginosa infections within the mammalian lung will require a thorough understanding of the regulation of bacterial virulence within the lung environment.

Below are specific projects currently underway in the laboratory:

The interaction between choline catabolism and P. aeruginosa virulence in the lung
Upon inhalation into the lung, P. aeruginosa comes in contact with the airway surface liquid, which is composed primarily of the phospholipid phosphatidylcholine (PC, ~75% of surfactant by weight). P. aeruginosa secretes phospholipases that can cleave PC, releasing the polar head-group (phosphorylcholine) and the lipid tail (diacylglycerol), both of which can be taken up by the bacterium and utilized as a nutrient source. The phosphorylcholine molecule (ChoP) can be further catabolized to glycine betaine (GB). GB, via the transcription factor GbdR, leads directly to transcriptional induction of one of the major PC-specific phospholipases, PlcH.

We have identified the GbdR transcription factor, the enzymes involved in choline catabolism in P. aeruginosa, and demonstrated the direct transcriptional regulation of plcH by GbdR. However, there remain many related questions, including:

• What other virulence factors are directly regulated by GbdR?
• Are there additional transcription factors that bind GB?
• Since GB can be catabolized, how does the bacterium regulate the balance between catabolism and induction of virulence?

These questions are being addressed using standard molecular genetics techniques, as well as cDNA microarrays, genetic screens, chromatin-affinity purification, and biochemical purifications.

Identification and characterization of novel genes induced by pulmonary surfactant.
Like many bacteria, the majority of open-reading frames in P. aeruginosa are predicted to encode proteins with no known or predicted function. Therefore, in any given genetic screen or microarray experiment, you will identify proteins that appear to be important for your system, but must be fully characterized in order to better understand them. Using a series of microarray experiments examining P. aeruginosa exposure to pulmonary surfactant, we have identified a number of candidate genes that are highly induced by surfactant, but have either have vague predictions of function or no predicted function. The goal of these projects is to exploit the tractability of the P. aeruginosa system (easy genetics, multiple sequenced genomes, mutant libraries) in conjunction with biochemical methods and bioinformatics to determine the function of these proteins of interest, many of which have homologues in other bacteria. Once a function is described, we are interested in understanding the transcriptional or post-transcriptional regulation of any genes that appear to be directly involved in interactions with eukaryotes.

Bacterial phospholipase-mediated alteration of host lung physiology.
The biophysical function of pulmonary surfactant is to lower the surface tension of the airway surface liquid to nearly zero, which prevents our alveoli and respiratory bronchioles from closing during every exhalation. When phospholipases, such as PlcH (described above), hydrolyze PC in pulmonary surfactant, this surface-tension lowering property is lost. The loss of surfactant function due to this, or any other cause, is termed surfactant dysfunction. Our research shows that one of the major physiologic alterations to the mammalian lung during P. aeruginosa infection is surfactant dysfunction. The long term goal of this project is to use animal models of lung infection and small animal ventilation techniques to study the contribution of individual virulence factors to surfactant dysfunction. As we identify the roles of individual virulence factors, we are also using the animal models to study the efficacy of novel therapeutic agents in an effort to develop the next generation of anti-Pseudomonas drugs.

Watters C, Deleon K, Trivedi U, Griswold JA, Lyte M, Hampel KJ, Wargo MJ, Rumbaugh KP. Pseudomonas aeruginosa biofilms perturb wound resolution and antibiotic tolerance in diabetic mice. Med Microbiol Immunol. 2012 Sep 25. [Epub ahead of print]

Fitzsimmons LF, Hampel KJ, Wargo MJ. Cellular choline and glycine betaine pools impact osmoprotection and phospholipase C production in Pseudomonas aeruginosa. J Bacteriol. 2012 Sep;194(17):4718-26

Lovewell RR, Collins RM, Acker JL, O'Toole GA, Wargo MJ, Berwin B. Step-wise loss of bacterial flagellar torsion confers progressive phagocytic evasion. PLoS Pathog. 2011 Sep;7(9):e1002253.

Fitzsimmons LF, Flemer S, Wurthmann AS, Deker PB, Sarkar IN and Wargo, MJ Small molecule inhibition of choline catabolism in Pseudomonas aeruginosa and other aerobic choline catabolizing bacteria. Applied and Environmental Microbiology 2011 Jul;77(13):4383-9

Wargo MJ, Gross MJ, Rajamani S, Allard JL, Lundblad LK, Allen GB, Vasil ML, Leclair LW, Hogan DA.. Hemolytic phospholipase C inhibition protects lung function during Pseudomonas aeruginosa infection. Am J Respir Crit Care Med. 2011 Aug 1;184(3):345-54

Wargo MJ, Hogan DA. Identification of genes required for Pseudomonas aeruginosa carnitine catabolism. Microbiology. 2009 Jul;155(Pt 7):2411-9.

Wargo MJ, Ho TC, Gross MJ, Whittaker LA, Hogan DA.GbdR regulates Pseudomonas aeruginosa plcH and pchP transcription in response to choline catabolites. Infect Immun. 2009 Mar;77(3):1103-11.

All Wargo publications

* indicates equal contribution