Field surveys and laboratory experiments were conducted during the season 2009/10 and 2010/11 in witchweed (Striga hermonthica [Del.] Benth.) endemic areas in Sudan to investigate the host specificity of witchweed populations collected from different locations with respect seed germination and haustorium initiation in response to sorghum root exudates and extracts. Field surveys were conducted to collect seeds from witchweed plants growing under their respective hosts; sorghum and millet. A total of fifteen witchweed populations were collected. Tow in vitro experiments were conducted at the Phytopathology Center and Biology Laboratory, Faculty of Agricultural Sciences, University of Gezira, Sudan to study the effects of root exudates and root extracts of sorghum cv. Abu-70, cv. Wad Ahmed and cv. Hakika on percentage of seed germination and haustorium initiation. Treatments (fifteen witchweed populations and three sorghum cultivars) were arranged in a factorial completely randomized design with three replicates. Data were collected and subjected to analysis of variance. Means were separated for significant using Duncan's Multiple Range Test (at p £ 0.5). The results showed that, there were significant differences in seed germination and haustorium initiation of witchweed in response to root exudates and root extracts among sorghum cultivars and among the witchweed populations. However, the highest seed germination and haustorium initiation percentages attained by each of the witchweed population were on their respective hosts. This study suggests two levels of physiological specialization in witchweed in Sudan: intercrop specialization and intra-crop specialization. Moreover, two strains of witchweed are suggested, one specific to sorghum and the other, to millet. The existence of variability and host specificity within witchweed populations are suggested to be based almost entirely on differential response of Striga isolates to root exudates and root extracts from host.