The morphological characterization and molecular genetic variation were determined in populations of thief ants, Solenopsis molesta (Say). The genetic variations were elucidated using mitochondrial deoxyribonucleic acid (mDNA) sequences of cytochrome oxidase I. DNA from thief ants was extracted with Qiagen’s Gentra PUREGENE® DNA Isolation Kit using their solid tissue protocol. Polymerase chain reactions (PCR) were run on the extracted DNA using primers Lep-F1 (forward) and Lep-R1 (reverse). The DNA products were concentrated and purified by Microcon Centrifugal Filter Unit YM-100. Purified DNA samples were sequenced at the University of Arkansas Medical Sciences (UAMS) DNA Sequencing Core Facility. The sequences were edited and aligned using Codon Code Aligner. The contigs wee uploaded to www.Phylogeny.fr and phylogenetic trees were produced (Neighbor joining, Maximum likelihood and Bayesian). The trees displayed variation in genetic makeup of the thief ants from various geographic regions and genetic variation corresponded to the morphologic identification. The thief ants collected from different states were separated into three groups. Ants collected from New York, Indiana and one location in Nebraska formed one group identified as S. molesta validiuscula, a second group formed with ants from Louisiana identified as S. carolinensis and the third group consisted of ants from South Dakota, Washington, New Jersey Tennessee, Kansas and two other locations in Nebraska identified as S. molesta molesta