In the present study, a molecular detection of Bartonella sp. and Wolbachia sp. in Ctenocephalides felis (Siphonaptera: Pulicidae) isolated from Canis lupus familiaris from different geographical areas of Spain, Iran and South Africa, and in Stenoponia tripectinata tripectinata isolated from Mus musculus from the Canary Islands has been carried out by amplification of the 16S ribosomal RNA partial gene of Wolbachia sp. and intergenic spacer region (its region) of Bartonella sp. A total of 70 % of C. felis analysed were infected by W. pipientis. This percentage of prevalence was considerably higher in female fleas than in male fleas. Bartonella DNA was not detected in C. felis from dogs, while Bartonella elizabethae was detected and identified in S. t. tripectinata from M. musculus from the Canary Islands representing 43.75 % prevalence. This report is the first to identify B. elizabethae in S. t. tripectinata collected in M. musculus from the Canary Islands. Thus, our results demonstrate that this flea is a potential vector of B. elizabethae and might play roles in human infection. The zoonotic character of this bartonellosis emphasizes the need to alert public health authorities and the veterinary community of the risk of infection.