The freshwater amphipod, Gammarus fasciatus, and a population that keys to Gammarus pseudolimnaeusare broadly sympatric in southeastern Virginia. By documenting the successful formation of pre-copulatory mate-guarding pairs between individuals collected from Virginia and New York, we confirmed the occurrence of a G. pseudolimnaeus population strongly disjunct from the previously described range in the Mississippi and St. Lawrence River drainage basins. Gammarus pseudolimnaeus appears restricted to high-quality, spring-fed streams that occur at low density across an increasingly fragmented natural landscape in Virginia. Gammarus fasciatus, however, occurs in lakes and streams of developed landscapes that typically are more degraded, and this species does not co-occur with G. pseudolimnaeus in high-quality habitat. Gammarus pseudolimnaeus reproduces year-round, with adults and juveniles of all size classes continuously present. Gammarus fasciatus reproduces primarily from February through June, at which point the adults die, and by late summer the population consists solely of immature individuals. If the two species overlapped in distribution, the smaller Gammarus fasciatus amphipods would have to compete with adult G. pseudolimnaeus for resources. We hypothesize that this competitively disadvantageous life cycle could account, in part, for the absence of G. fasciatus in high-quality streams occupied by G. pseudolimnaeus.