Microbial infections are still among the leading causes of death worldwide. Intracellular pathogens, such as Mycobacterium tuberculosis and SARS-CoV-2, utilize the host intracellular niches to replicate, spread, and evade the immune response and antimicrobials. Chemical genetics screening, i.e. functional assessment of small molecule inhibitors with characterized host targets, presents an unbiased and rapid top-down approach to uncovering host components required for sustaining intracellular infections. We discovered markedly distinct intracellular pathogens exploit the same host pathways during infection, with unique mechanisms of action. I will present our application of host-directed therapy screening, and how it allowed exposing and studying pathogen-utilized CHK2, GSK3, and V-ATPase host pathways during infection.