It has previously been shown that paternal mitochondrial DNA (mtDNA) can be detected in later generations in Drosophila. To further analyze the paternal transmission of mtDNA, the progeny of two intraspecific and three interspecific crosses were examined in the frequency of the paternal transmission of mtDNA, using closely related species of the melanogaster species subgroup. Types of mtDNA in the progeny of the individual backcrosses of F₁ females were analyzed by selective amplification of paternal mtDNA. More than 100 F₁ females were examined for each backcross. The same type of mtDNA as that of the paternal mtDNA was detected in approximately 20 – 60% of the backcrosses. The present results indicate that paternal leakage occurs in the intraspecific crosses as well as in the interspecific crosses in Drosophila.