Zoysiasp.) is used as a warm-season turfgrass for lawns, parks, and golf courses in the warm, humid and transitional climatic regions of the United States. Zoysiagrass is an allotetraploid species ( 2n = 4x = 40) and some cultivars are known to easily self- and cross-pollinate. Previous studies showed that genetic variability in the clonal cultivars Emerald and Diamond was likely the result of contamination (seed production or mechanical transfer) or mislabeling. To determine the extent of genetic variability of vegetatively propagated zoysiagrass cultivars, samples were collected from six commercially available zoysiagrass cultivars (Diamond, Emerald, Empire, JaMur, Meyer, Zeon) from five states (Arkansas, Florida, Georgia, North Carolina, Texas). Two of the newest cultivar releases (Geo and Atlantic) were to serve as outgroups. Where available, one sample from university research plots and two samples from sod farms were collected for each cultivar per state. Forty zoysiagrass simple sequence repeat (SSR) markers and flow cytometry were used to compare genetic and ploidy variation of each collected sample to a reference sample. Seventy-five samples were genotyped and an unweighted pair group method with arithmetic mean clustering revealed four groups. Group I ( Z. japonica) included samples of Meyer and Empire11 (Empire sample at location #11), Group II ( Z. japonica Z. pacifica) included samples of Emerald and Geo, Group III ( Z. matrella) included samples of Diamond and Zeon, and Group IV ( Z. japonica) consisted of samples from Empire, JaMur, Atlantic, and Meyer3 (Meyer at sample location #3). Samples of Empire, Atlantic, and JaMur were indistinguishable with the markers used. Four samples were found to have alleles different from the respective reference cultivar, including two samples of Meyer, one sample of Empire, and one sample of Emerald. Three of these samples were from Texas and one of these samples was from Florida. Three of the four samples that were different from the reference cultivar were university samples. In addition, one sample, Empire11, was found to be an octoploid ( 2n = 8x = 80). For those samples that had a fingerprint different from the reference cultivar, contamination, selfing, and/or hybridization with other zoysiagrasses may have occurred.