Callie Effler, a student at Lee University, worked in the lab of Dr. Eric Harvill.

Abstract: Intracellular survival is a common trait among human pathogenic bacteria that has advantages for the bacteria’s protection from the host immune response, persistence, and dissemination within the host. Bordetella pertussis, the gram-negative bacteria that causes whooping cough in humans, is commonly regarded as an extracellular pathogen. However, it has been recovered from macrophages in in vitro experiments, and reported anecdotally in clinical samples. It is unknown what contribution to pathogenicity the intracellular population has, if any, on the host. In this work, our broad objective was to evaluate the impact of B. pertussis’ intracellular survival and its role in pathogenicity. To do so, we planned to identify mutants similar to the wild type in general measured aspects of virulence, but that failed to survive inside of macrophages. A transposon library of B. pertussis UT25 was screened, resulting in the identification of several putative mutants that were deficient in intracellular survival. These strains were further screened for intracellular deficiency as a confirmatory measure, and went through further in vitro assays screening for cytotoxicity, hemolytic activity, resistance to serum complementation, and general fitness (growth). Based upon these assays, mutant strain G4 was the best candidate among those tested for an intracellularly-deficient mutant with similar virulence-related characteristics to the wild type. Preliminary C57 mouse infection studies suggest that the mutant strain behaves similarly to the wild type in vivo, indicating that intracellular survival may not be contributing to virulence. It is hypothesized that intracellular survival may be a phenotypic remnant of an ancestral strain of B. pertussis that transitioned from the environment to a become a human pathogen using this trait.

Margaret Dedloff, a junior at Clarkson University, worked with Dr. Monical Gestal in the lab of Dr. Eric Harvill to study the role of a sigma factor by bacteria in manipulating host immune response.

Abstract:  During the course of infection, some pathogens are able to sense and respond to the host immune system. This can cause prolonged pathogenesis, high transmission rates, and vaccine failure. One group of bacteria known to manipulate the immune response in varied ways is the Bordetella spp. B. pertussis, B. bronchiseptica and B. parapertussis are able to inhibit complement and suppress B and T-cell functions [1].  The BvgAS two-component system is understood to be a regulator of many virulence factors and is known as the master virulence regulon, however this is most likely an oversimplification. We hypothesized that pathogens like bordetellae should be under strong selective pressure to sense and respond to signals in blood and serum, in order to modulate immune defenses. When studying blood and serum responsive genes, we identified a putative sigma factor up-regulated in both conditions, and we hypothesized that this might be a regulator that dictates adaptation to pressure from the immune system. Here we identified the role of this sigma factor, the Bordetella Sigma Factor, or bsr, in manipulating the immune response. Through the use of a B. bronchiseptica bsr knockout and in vitro assays, we have found that bsr interferes with the innate response. bsr inhibits survival in macrophages by changing the dynamics of phagocytosis, causing macrophage death, and causing bacterial death and replication within the macrophage by interfering with cytokine and chemokine expression as well as differentially interacting with TLR receptors. This suggests that subsequent cell recruitment will be different within the host. Our results demonstrate that the bsr gene plays a critical role in Bordetella interaction with the innate immune system. A better understanding of this gene and its function will be valuable in efforts to create successful vaccines and treatments not only for Bordetella spp. but also for other bacterial species.

[1] Gestal M.C., Whitesides L.T., and Harvill E.T. “Integrated Signaling Pathways Mediate Bordetella Immunomodulation, Persistence & Transmission”. In revision, Current Trends in Microbiology.

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