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Publisher Summary There is general recognition among evolutionists that no matter how one defines a species, the boundaries between distinct, sexually reproducing species are maintained by barriers to gene flow at individual loci. Isolating barriers have typically been categorized into those operating before fertilization (prezygotic) and after fertilization (postzygotic). Prezygotic reproductive barriers can arise due to the evolution of habitat utilization differences, mating behavior differences, and/or phenological differences between incipient species. Postzygotic isolating barriers encompass hybrid inviability, hybrid sterility and hybrid breakdown. This chapter is concerned with the set of prezygotic barriers that occur in closely related animal species after ejaculation or spawning and before fertilization, resulting in postmating, prezygotic (PMPZ) isolation. The transfer or spawning of sperm sets the stage for the operation of PMPZ barriers to gene exchange. The environment of the female reproductive tract eliminates or kills many male gametes, even conspecific ones, in a variety of ways. These barriers to fertilization by conspecific sperm may also serve as barriers to fertilization by heterospecific sperm. Moreover, in heterospecific pairings, sperm may fail to move appropriately in the female reproductive tract. The importance of chemotaxis in mediating PMPZ isolation varies among major taxonomic groups and appears to be related to whether fertilization occurs internally or externally. In insects, the proteins in the seminal fluid can influence the fertilization process in a number of ways including: the storage and capacitation of sperm; the induction of peristalsis in the female reproductive tract; and the induction of ovulation.