1. The TLR51174C>T Polymorphism in Melioidosis Infection
Chief of Medicine Conference 10/2/12
Presented by Amy Dickey, MD
R2, Internal Medicine
Principal Investigator: Eoin West, MD MPH

2. Outline Background: What is melioidosis? What are Toll Like Receptors?
How is the TLR51174C>T polymorphism important in mortality from melioidosis?
Hypothesis – Defining the role of the TLR51174C>T polymorphism in cell signaling.
Data
Conclusions and future directions

3. Melioidosis
Melioidosis – the disease caused by the pathogen Burkholderia pseudomallei
An important cause of pneumonia and sepsis in SE Asia and northern Australia.
The mortality rate from melioidosis can reach 40%.
Figure 1. The global distribution of Melioidosis. NEJM Sept 13, 2012.

4. Flagellin is the TLR5 agonist
Toll Like Receptors are signaling receptors of the innate immune system.
They recognize molecules shared by pathogens called pathogen-associated molecular patterns (PAMPs).
Important antigens containing PAMPs include LPS, flagellin, and lipopeptides
Be sure to clarify that LPS is a TLR4 agonist and that there is no LPS-TLR5 interaction or flagellin-TLR4 interaction.
Figure 2: Toll like receptor signaling pathways

5. The TLR51174C>T non-functional polymorphism improves survival from melioidosis.
The TLR51174C>T polymorphism encodes a premature stop codon in the ligand-binding domain of TLR5.
In a cohort of Thai melioidosis patients, those with the TLR51174C>T polymorphism experienced a 3x decrease in in-hospital mortality from melioidosis infection.
TLR5
TLR5 mutant
C/C
C/T
T/T
Survivors
416 (74.3%)
75 (90.4%)
2 (100%)
Non-survivors
144 (25.7%)
8 (9.6%)
0 (0%)
Table 1: TLR51174C>T genotype and in-hospital mortality

6. Surprisingly, the TLR51174C>T polymorphism modulates cytokine response to both flagellin and LPS.
Figure 4. IL-8 expression
Figure 3. GCSF expression
I would eliminate the legend on the right and instead make your graphs larger. You are going to say what is in the legend.

7. Hypothesis
TLR51174C>T polymorphism modulates the TLR4-dependent innate immune response to B. pseudomallei.
LPS
TLR5
TLR4
TLR5 mutant ? ?
I suggest eliminating the legend
NF-kB
Inflammatory
Cytokines

8. The TLR51174C>T polymorphism increases NF-kB expression in response to LPS.
Ditto re the legend and make the graph larger. Can you abbreviate your heading to say just the essential? Be prepared to spend some time explaining exactly what you are transfecting, stimulating with, and measuring in this slide.
Figure 6. NF-kB activation

9. Surprisingly, the TLR5 polymorphism increases NF-kB expression but decreases IL8 expression in response to LPS.
Same suggestions as for the previous slide. As the NF-KB activation induced by the combo of TLR4/5 is not so reduced compared to TLR4 alone when compared to the last slide, one thing you could do to focus the audience on your main point is just show the TLR4/5 and TLR4/5mut data.
Figure 7. NF-kB activation
Figure 8. IL-8 expression

10. The TLR51174C>T polymorphism attenuates IL8 expression in response to B. pseudomallei, an effect independent of flagellin.
What does this mean? IL-8 expression in response to B. Pseudomallei is both TLR-dependent and TLR-independent.
Figure 9: IL-8 expression assay

11. Conclusion
Conclusion: TLR51174C>T polymorphism modulates the TLR4-dependent innate immune response to Burkholderia pseudomallei. Specifically, the TLR51174C>T polymorphism alters TLR4-dependent NF-kB activation and IL-8 expression.
Additional Questions: Does the mutant TLR5 or the wild type TLR5 modulate TLR4-dependent signaling? At what level does it modulate signaling?
LPS
TLR5
TLR4
TLR5 mutant
Could you tweak the conclusion to reflect some of the subtleties you uncovered (NF-KB vs IL-8, e.g.? I like your diagram!
NF-kB
Inflammatory
Cytokines

12. Acknowledgements The West Lab Members of the Wurfel and Skerrett labs.
Eoin West, MD MPH
Sudeshna Seal, PhD
Johanna Robertson
Members of the Wurfel and Skerrett labs.
I think this last slide is fine. I have noticed that some scientists list all the people who helped with their work, so you could thank whoever in the lab or adjacent labs helped you out.