Macrobrachium (Bate, 1868) is a large and cosmopolitan crustacean genus of high economic importance worldwide. We investigated the morphological and molecular identification of freshwater prawns of the genus Macrobrachium in South, South West, and Littoral regions of Cameroon. A total of 1,566 specimens were examined morphologically using a key described by Konan (Diversité morphologique et génétique des crevettes des genres Atya Leach, 1816 et Macrobrachium Bate, 1868 de Côte d'Ivoire, 2009, Université d'Abobo Adjamé, Côte d'Ivoire), leading to the identification of seven species of Macrobrachium: M. vollenhovenii (Herklots, 1857); M. macrobrachion (Herklots, 1851); M. sollaudii (De Man, 1912); M. dux (Lenz, 1910); M. chevalieri (Roux, 1935); M. felicinum (Holthuis, 1949); and an undescribed Macrobrachium species M. sp. To validate the genetic basis of the identified species, 94 individuals representing the species were selected and subjected to genetic characterization using 1,814 DArT markers. The admixture analysis revealed four groups: M. vollenhovenii and M. macrobrachion; M. chevalieri; M. felicinum and M. sp; and M. dux and M. sollaudii. But, the principal component analysis (PCA) separated M. sp and M. felicinum to create additional group (i.e., five groups). Based on these findings, M. vollenhovenii and M. macrobrachion may be conspecific, as well as M. dux and M. sollaudii, while M. felicinum and M. sp seems to be different species, suggesting a potential conflict between the morphological identification key and the genetic basis underlying speciation and species allocation for Macrobrachium. These results are valuable in informing breeding design and genetic resource conservation programs for Macrobrachium in Africa.