Taxonomic keys and original descriptions were used to identify 26 Xiphinema americanum-group populations from Florida comprising X. georgianum (eight populations), X. citricolum (six), X. floridae (six), X. laevistriatum (five) and X. tarjanense (one). Principal component analysis of a subset of 19 morphometric characters accorded with the species designations; discriminant analysis of six characters assigned 93% (111 of 119) of the specimens to the correct putative species. A phylogeny of these populations estimated from analyses of rDNA sequences (ITS and D2D3) was also congruent with species designations from taxonomic keys and PCA. The D2D3 sequences revealed very little intraspecific variation whereas each population sampled produced a unique ITS sequence. Intraspecific variation in the suites of character code values from polytomous keys resulted mainly from minor discrepancies between population character means and reported character ranges for the species. We show that, for these taxa, species delimitation based on the requirement that sister taxa evolve autapomorphies distinguishes intraspecific variation from phylogeny and, as applied to molecular characters, delimits the same taxa as those predicted by morphological keys and PCA.