Dissecting the impact of enteric viral-bacterial coinfection on the host innate immune response and its implications for pathogenicity

Infection biology has moved more into the focus of both the research community as well as the broad public. This is mainly due to the continuous emergence of resistant bacterial pathogens, alongside a steady slow-down of the development of novel antibacterial treatments, as well as the recent and ongoing pandemic caused by a newly emerged viral pathogen, SARS-CoV-2. In order to be able to continuously provide therapeutic options against pathogenic agents, as well as expand our ability to develop novel treatments quickly, we have to solidify our molecular understanding of how pathogens infect their host cells and which interdependencies occur during infection.A major factor in the clinical outcome of infectious diseases is the occurrence of secondary infections with another pathogen. This has been shown most predominantly for patients suffering from infection with Human Immunodeficiency Virus (HIV). They have a massively increased susceptibility to super-infections with a second pathogen, which can be viral, bacterial, fungal or of other nature. In addition to HIV, other infectious diseases display increased severity due to the occurrence of super-infections, such as Influenza A virus (IAV). Here, staphylococcal or streptococcal co-infections increase disease severity and fatality and are commonly isolated from hospitalized patients. Besides that, Rotavirus and Norovirus have been described as clinically relevant pathogens that occur with co-infections of enteropathogenic bacteria, such as E. coli, which triggers exacerbated symptoms in children, or C. difficile, causing an increased bacterial burden.The understanding of host-pathogen interactions on a cellular and molecular level continues to broaden. This is due to the elucidation of molecular targets for pathogenic effectors, the identification of host signaling cascades within the innate immune response, as well as the characterization of limited resources triggering competition between the pathogen and the host during infection. Yet the same level of knowledge has not been acquired for co-infections of two different pathogens, the study of which has so far mainly occurred on the organismal or systemic level. Some studies pioneer the molecular understanding of the interplay between different pathogens, such as IAV and S. pneumoniae or HIV and M. tuberculosis. Yet a comprehensive picture of the interplay between the co-infecting pathogens, as well as the role of host innate immune response is so far still missing. Through the proposed project, I would like to shed light onto the impact of co-infections in the intestinal tract. More specifically, I will dissect synergies and antagonisms by systematically assessing pairwise infections with a panel of both viral and bacterial enteric pathogens in an unbiased screening approach. Rather than focusing solely on the pathogens, I will examine the role of the host cell, especially with respect to cell death and innate immune response. To achieve this, I will infect immunologically profiled, murine macrophages with a defined and established panel of enteric viruses and bacterial pathogens that have been selected through preliminary work. I will use parallelized, automated fluorescence measurements to quantify cell death and pathogen proliferation, and interactions will be scored using a simple Bliss model.Using the dataset generated from this broad approach, I will then seek to identify molecular interaction points and required processes within the host cell through unbiased proteomic and transcriptomic approaches. Mechanistic hypotheses on how pathogens interact with and within the host and influence innate immune responses will subsequently be formulated and tested using orthogonal biochemical and immunological approaches. Relevant methodologies include analysis of signaling cascades by phosphorylation-specific antibodies, host cell mutants, as well as adequate murine infection models. Furthermore, using small molecule inhibitors, the targetability of the interaction points and hence the disruptability of pathogenic synergies can be assessed. I thereby aim to expand our mechanistic understanding of the host-pathogen-pathogen interface, ultimately identifying targets for the development of novel therapeutic interventions.