The objectives of this study were to develop anther-derived dihaploid lines from two Maryland tobacco (Nicotiana tabacum L.) cultivars, ‘MD 609’ and ‘MD 341’, and to compare the performance of the dihaploids with their parental sources. Haploid production per anther was 2.8 and 1.0 for MD 609 and MD 341, respectively. The average dihaploid production per haploid was approximately 4.0 for both populations. Seventeen and 23 dihaploid lines from MD 609 and MD 341, respectively, were grown in the field and then evaluated with their parental cultivars for 11 agronomic and chemical traits in a 2-year study (1986 and 1987). Six weekly growth increments were measured for the MD 341 dihaploid population in 1986. Parental genotypes appeared to have a significant effect on dihaploid performance. MD 609 dihaploids demonstrated no significant yield reductions, but they were lower in price and quality index, had fewer leaves, and flowered later than the MD 609 cultivar. MD 341 dihaploids were 5.6% lower yielding, had longer internodes, and flowered later than the MD 341 cultivar. Average weekly growth rates for MD 341 and its dihaploids were similar throughout the growing season. Significant variation for several traits was observed among the dihaploids in both populations. Therefore, the small reduction in performance of anther-derived dihaploids in the two Maryland tobacco populations suggests that anther culture is a viable technique for developing homozygous germplasm lines and for genetic studies in Maryland tobacco.