[Objectives]  Bacteria of the genus Asaia can symbiotically colonize insects of phylogenetically distant genera and orders, e.g. Diptera, Hymenoptera, Hemiptera and Lepidoptera. Our aim in this study was to investigate the prevalence of Asaia infection in four Lepidopteran species; Prodenia litura (Fabricius), Mythimna separata Walker, Agrotis ypsilon Rottemberg and Mamestra brassicae Linnaeus, and to analyze its mode of transmission within and between species. [Methods]  Asaia infection rates were investigated using the polymerase chain reaction (PCR) with primers targeting the Asaia 16S rDNA sequence. An Asaia-infected P. litura strain was established to study the vertical transmission of Asaia. [Results]  We found that P. litura, M. separata, A. ypsilon and M. brassicae, were all infected with Asaia bacteria with infection rates of 45.8%, 66.7%, 47.2% and 31.0%, respectively. Phylogenetic analysis based on the Asaia 16S rDNA sequences revealed three sequence types: ST1, ST2 and ST3. All four species were infected with the ST1 type indicating a high possibility of horizontal transmission of Asaia among species. Our results also demonstrate the vertical transmission of Asaia from infected P. litura to their offspring. [Conclusion]  Asaia has a high prevalence in the four Lepidopteran species examined and can be both horizontally, and vertically, transmitted.