Specimens collected in or near Pensacola Bay, Florida matching the descriptions of Luidia lawrencei and Luidia clathrata (the congener from which L. lawrencei was recently split) were compared to determine whether their recent taxonomic separation is supported by differences in sediment preference, salinity tolerance and COX-1 mtDNA sequences. Luidia clathrata has a preference for smaller grain sizes, while no statistically significant preference was found for Luidia lawrencei, and no significant difference was found between the species. Luidia clathrata is more tolerant of lower salinity based on the righting response than Luidia lawrencei, especially at salinities lower than 25 g kg−1. The COX-1 comparison returned over 99% homology among individuals of the two species. While sediment preference and salinity tolerance results indicate differences in response to habitat dissimilarities, the COX-1 genetic result is strong evidence against the recently proposed separation of the species. In light of the COX-1 result, we interpret the sediment and salinity results as long-term acclimation responses.