Investigating the impact of the FIC domain Bartonella quintana effector protein, BepA1, on human innate immune responses
Authors: Aman-Deep Ramela, 
Published: February 2022

Bartonella are known as stealth pathogens that subvert the host immune system, using
an arsenal of pathogenicity factors that enable them to survive and persist in their reservoir
host. The focus of our study is B. quintana, a historically famous pathogen that caused trench
fever in soldiers during World War 1, but today causes an increasing number of urban trench
fever cases in vulnerable populations, such as homeless or immunocompromised groups.
After exposure to B. quintana, the bacteria traverse the skin and enter the bloodstream,
infecting erythrocytes. B. quintana encounter host immune cells in the vascular system, but are
not killed, hijacking these cells to disseminate into the lymphatic system where they eventually
make their way into the bloodstream. How B. quintana modulates host immune responses to
its favour during early infection is very poorly understood due to the current lack of research
surrounding this pathogen.
B. quintana, like many Bartonella species, possesses a Type IV Secretion System, a
large protein complex that is used to inject Bartonella Effector Proteins, Beps, directly into
target host cells. B. quintana encodes six BEP proteins, of which BepA1 and BepA2 are the
best studied. BepA1 contains a FIC domain, which in other pathogens have been shown to
catalyse addition of an AMP moiety to target host proteins, interrupting function and
downstream signalling. This process, AMPylation, is used by various bacterial pathogens to
interfere with actin cytoskeletal signalling to inhibit phagocytosis. However, the AMPylation
function in B. quintana BepA1 remains to be shown. BepA2 has been shown to inhibit
apoptosis once translocated into endothelial cells.
Previous studies in our lab indicated that BepA1 may play a role in suppression of the
innate immune system. Wild type BepA1-transfected HeLa-229 cells showed decreased levels
of IL-6 and IL-8 in comparison to an empty vector control. We thus hypothesised that BepA1
is being translocated into host cells to AMPylate a signalling protein/s to suppress immune
signalling and immune cell recruitment, enabling the survival and persistence of B. quintana
in the bloodstream.
Our study thus focused on better understanding the host-pathogen interactions between
B. quintana and the human innate immune response. We began by broadly characterising the
innate immune responses of human cells transfected with the wild type B. quintana BepA1
protein. HeLa-229 cells were transfected with a vector expressing the BepA1 wild type protein,
or a catalytically dead BepA1 mutant protein. Cells were stimulated with TNF-α, and cytokine
ii
and chemokine expression was evaluated. These observations were validated via qPCR
analysis of the transfected, TNF-α treated cells.
Lastly, we aimed to generate a targeted BepA1_BepA2 B. quintana deletion mutant to
further study the consequences on host-pathogen interactions. Unfortunately, we were unable
to generate this mutant.
Our work provides the first evidence that we know of that suggests a bacterial effector
with a FIC domain is involved in manipulation of cytokine and chemokine expression. Our
work provides a foundation for a better understanding of the early stages of Bartonella
infection and pathogenesis. With this research, we hope to shed light on how B. quintana
hampers immune signalling during infection, leading to its intracellular persistence and
survival in the reservoir host.