The presence and distribution of various species of canine hookworms in Africa are poorly known. The main objective of this study, therefore, was to identify the hookworm species present in canine faecal samples from Morogoro, Tanzania, using molecular techniques. Faecal samples from 160 local dogs were collected and hookworm positive samples processed to recover larvae for further molecular characterization. DNA was extracted from pools of larvae from individual samples (n = 66), which were analysed subsequently using two different molecular approaches, polymerase chain reaction-linked restriction fragment length polymorphism (PCR-RFLP) and species-specific PCR coupled with Sanger sequencing. The PCR-RFLP technique detected only the presence of the ubiquitous Ancylostoma caninum in the 66 samples. However, by species-specific PCR coupled with Sanger sequencing we identified ten samples with A. braziliense, two with Uncinaria stenocephala and five with A. ceylanicum. Thus, all four known species of canine hookworms were identified in Morogoro, Tanzania. To our knowledge this is the first report of the detection of the presence of U. stenocephala and A. ceylanicum in Africa using molecular techniques. In addition to their veterinary importance, canine hookworms have zoonotic potential and are of public health concern.