Bacterial conjugative type IV secretion systems (T4SS) are nanomachines that transfer DNAs unidirectionally from a donor cell to a recipient cell in a process called conjugation. They are the main means by which antibiotic resistance genes spread among bacterial populations. Before transferring DNA, they elaborate a long extracellular filament, termed conjugative pilus, which serves as conduit for the passage of DNA from donor to recipient cell. Here we will describe recent advances in the field that have provided mechanistic insights on aspects of pilus biogenesis and DNA transfer by these secretion machineries.