ABC toxins are tripartite, bacterial, pore-forming toxins found in pathogens of insects and mammals. They are characteristically assembled from three different components (TcA, TcB and TcC) into hetero-oligomeric complexes ranging in mass from 1.5-2.5 MDa. The TcA subunit binds to cellular membranes and introduces a protein translocation pore, through which a cytotoxic effector (TcC) is delivered. TcB forms a protective cocoon - part of which is contributed by the N-terminus of TcC - that encapsulates the cytotoxic effector. Most appear to target gut epithelial cells in a host-specific manner and for some members of the ABC toxin family it appears pH plays an important role in activation. Despite this, it remains to be established whether pH-driven activation occurs in microenvironments at the cell surface or within subcellular compartments (e.g. endosomes) and for some toxins there is evidence that pH alone is insufficient for activation. In addition, despite the recent identification of a glycoprotein receptor for a single ABC toxin family member, the structural determinants of host tropism and receptor recognition remain to be fully elucidated, primarily because no toxin-receptor complex has yet been isolated or structurally characterised. In the absence of this information, we used cryo-EM to solve and study the structure of multiple ABC toxins with diverse host specificity profiles, obtaining new insights into the structural determinants of tropism in this family. We also employed all atom molecular dynamics simulations on representative toxins belonging to different subclasses, classified according to their molecular architecture. These simulations, which include >2,000,000 atoms, study the process of toxin activation in unprecedented detail and together with biophysical measurements of pore formation, suggest differences in the way that YenTc, an ABC toxin with an atypical architecture, is activated. Overall, our results provide important insights into how host specificity and activation mechanisms have diverged within this family, information that is important for understanding mechanisms of virulence, as well in evaluating the potential of these toxins to developed for agricultural pest control and drug delivery applications.