Recognition of pathogens by the host cell is of paramount importance for the initiation of an immune response and clearance of the pathogen. Intracellular and cytosolic bacteria must secure entry into the host cytoplasm in order to engage activation of cytosolic immune sensors and the inflammasome. However, microbial ligands from extracellular pathogens are also detected by cytosolic innate immune sensors. We discuss our latest findings on the role of inflammasomes and innate immune regulatory proteins in the recognition of bacteria. We also highlight novel anti-microbial host defense system mediating cytosolic release of bacterial ligands for sensing by pattern-recognition receptors and inflammasomes.