During germplasm explorations within Zambia in 1984, seven Cucumis accessions were collected that could not be identified to species. Two of the accessions were studied in-depth. Based on phenotypic characters, they were closest to Cucumis pustulatus. In ITS analyses of all available Cucumis species and the accessions, the two accessions grouped with 100% bootstrap support in a clade comprising C. anguria, C. dipsaceus, C. insignis, and C. pustulatus. The accessions differed from these four Cucumis species by the following characters: plants pilose, male inflorescences paniculate with 6–30 flowers, staminate-flower calyx lobes linear, pistillate-flower pedicels long and cylindrical, and a genetic difference of 4 base pairs in their ITS sequences. Controlled crosses were made within and between the two accessions and between each of them and C. pustulatus. Fruits were produced from crosses made within and between the two accessions, but attempts involving C. pustulatus were unsuccessful. The two accessions are described as a new species, Cucumis zambianus. Six of the seven unidentified Cucumis collections from Zambia are identified as C. zambianus herein. They were all collected in the northwestern corner of Zambia, but the new species is also expected to occur in eastern Angola and southern Zaire.