In animals with heteromorphic sex chromosomes, the first interspecific genetic incompatibilities to accumulate cause sterility in the heterogametic (XY) sex and are disproportionately X-linked. Why the X chromosome has unique roles in the rapid evolution of gametogenesis and speciation is unclear. To determine the evolutionary and molecular genetic basis of the “large X-effect” in speciation, we have generated an ultra high-resolution genetic map of X-linked hybrid male sterility (HMS) factors between two sister species, Drosophila mauritiana and D. simulans, by introgressing doubly-marked segments of the D. mauritiana X chromosome into a D. simulans genome. This map comprises over 500 overlapping introgressed segments ranging in size from 2 Mb to 100 kb that were genotyped at thousands of markers by light-coverage resequencing. Our genetic mapping data, coupled with data on introgression between these species in nature, provide insights into the consequences of incompatibilities for gene flow. Furthermore, a recent selective sweep in D. mauritiana plausibly associated with a meiotic drive element appears to have prevented the accumulation of linked HMS factors in this region of the X chromosome. We have further refined three regions to a dozen candidate HMS genes, and in one of these regions, we have identified and transgenically validated a new HMS gene. Identification of multiple X-linked HMS genes will allow broad generalizations on the functions of these genes within species, the etiology of HMS, the evolutionary forces involved in interspecific divergence, and the overall molecular basis of the large X-effect.