Biosystematic studies of Psyttalia species (Hymenoptera: Braconidae), parasitoids of fruit-infesting Tephritids in Africa were conducted to resolve their taxonomy and develop species-specific characters to facilitate their reliable use as biological control agents. These involved the use of morphology, morphometrics, cross mating and Amplified restriction fragment length polymorphism. The populations studied included P. concolor, Psyttalia cosyrae, P. phaeostigma, P. lounsburyi, and Psyttalia samples from Rurima, Ruiru, Mrima Hill, Shimba Hills (locations in Kenya) and populations from Nkolbisson, Cameroon (P. perproximus), Tafo (Ghana) and Pietermaritzburg (South Africa). Additional samples included P. concolor reared from C. cosyra larvae and P cosyrae reared from C. capitata larvae. Morphological comparison of measurements of ovipositor, ovipositor sheath and hind tibia, showed ratios of ovipositor/tibia and ovipositor sheath/tibia distinctly differentiating Psyttalia cosyrae and P. phaeostigma from all other populations. Separation into natural groups was also achieved using flagellomere counts. Parasitoids reared on host larvae other than their natural hosts showed significant changes in linear measurements as well as body colour within one generation of rearing. Morphometric studies on 25 wing measurements in all thirteen (13) populations, assigned populations of unknown identity to groups based on variances. Comparison of populations showed veins enclosing the submarginal cell together with their adjoining veins as the main variables accounting for differences in these populations. Cross mating studies were conducted between both morphologically distinct species, and morphologically similar species. Once mating was successful, viable female offsprings were produced in all crosses up to F2, indicating the absence of post-copulatory isolating mechanisms, and suggesting the inadequacy of the biological species concept alone in separating species in this genus. Total genome comparison by AFLP in P. concolor, P. cosyrae, P. perproximus, P. lounsburyi and populations from Rurima, Ruiru, Shimba Hills and Tafo, detected variations in DNA. Psyttalia concolor, Rurima and Ruiru shared the highest number of similar loci, followed by P. perproximus, Shimba Hills and Tafo, while P. cosyrae and P. lounsburyi stood out as separate entities. The separation also matched the groupings defined by morphometric analyses and thus, lent genomic support to the clusters. The study was able to separate the populations into their natural groupings, which supports earlier conclusion from cross mating data that the populations from Rurima and Ruiru are morpho-species of Psyttalia concolor sensu stricto (i.e. Psyttalia cf. concolor), and P. cosyrae is a different species. The population from Shimba Hills was found to be similar to P. perproximus and that from Tafo in the morphological as well as DNA studies and they were clustered together by morphometric analysis. It is therefore suggested as belonging to P. perproximus, and is hitherto a new occurrence record of P. perproximus from Kenya and East Africa. Body colour and size were found to be determined by host-larvae characteristics and it does not make these characters alone reliable for identification of parasitoids. This is especially important in post-release sampling surveys to ascertain establishment of parasitoids in new environments where they may adapt to new host species. The populations are also characterized based on outcome of the different taxonomic tools used.