Aggregatibacter actinomycetemcomitans exploits and modulates the immune response by human neutrophils for survival in the anaerobic environment.

Abstract

Aggregatibacter actinomycetemcomitans (Aa) is a Gram-negative facultative anaerobe and an opportunistic oral pathogen, strongly associated with localized periodontitis and other inflammatory diseases. Periodontitis is a chronic inflammation of the periodontium resulting from the inflammatory response of the host towards the dysbiotic microbial community present at the gingival crevice. The host immune response creates a hostile environment for microorganisms; therefore, it is important for Aa to be able to regulate the necessary genes to survive and thrive in such an environment. Aaexpresses several virulence factors such as a cytolethal distending toxin (Cdt), lipopolysaccharide (LPS) and leukotoxin A (LtxA), that allows it to evade the immune response. For example, LtxA, considered a major virulence factor, targets leukocytes by creating pores on their membranes leading to cell lysis. LtxA has also been shown to provide Aa resistance against internalization and killing by neutrophils, however the effect of the toxin on neutrophils at a non-lethal level is often overlooked. At the subgingival pocket Aa is in a tug-of-war scenario where immune cells will secrete molecules such as catecholamines (i.e., epinephrine) to sequester residual metals, so these are not available as nutrients. To compete for such mineral bacteria have evolved different mechanisms, such as two-component systems (TCS) that allow them to assess their environment and adjust accordingly. Aa expresses the QseBC TCS composed of the sensor protein QseC and the response regulator QseB. Previously, catecholamines and iron were identified as the signals that activate the QseBC TCS in Aa, necessary for the organism to acquire iron as a nutrient to survive in the anaerobic environment. However, the source of catecholamines had not been identified at the time. The main objectives presented in this dissertation are a) the characterization of epinephrine interaction with different components of QseBC TCS, b) to characterize the synthesis, storage, and release of catecholamines by neutrophils stimulated with Aa, and c) determination of the effect sublytic levels of LtxA has on neutrophils. Previously, our group showed that QseC, primarily the periplasmic domain of QseC, is required for biofilm formation and virulence. A third gene was found to be co-expressed in the qseBC operon, YgiW, a protein of unknown function, but essential for biofilm growth and virulence. Additionally, it was previously shown that in the presence of Cat-Fe the expression of the enterobactin operon remained unchanged. Among the genes in this operon there is the fepA gene encoding for the enterobactin receptor FepA.