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Neorickettsia spp. are bacterial endosymbionts of parasitic flukes (Digenea ) that also have the potential to infect and cause disease (e.g., Sennetsu fever) in the vertebrate hosts of the fluke. One of the largest gaps in our knowledge ofNeorickettsia biology is the very limited information available regarding the localization of the bacterial endosymbiont within its digenean host. In this study, we used indirect immunofluorescence microscopy to visualizeNeorickettsia sp. within several life cycle stages of the digeneanPlagiorchis elegans . Individual sporocysts, cercariae, metacercariae, and adults ofP. elegans naturally infected withNeorickettsia sp. were obtained from our laboratory-maintained life cycle, embedded, sectioned, and prepared for indirect immunofluorescence microscopy using anti-Neorickettsia risticii horse serum as the primary antibody.Neorickettsia sp. was found within the tegument of sporocysts, throughout cercarial embryos (germ balls) and fully formed cercariae (within the sporocysts), throughout metacercariae, and within the tegument, parenchyma, vitellaria, uteri, testes, cirrus sacs, and eggs of adults. Interestingly,Neorickettsia sp. was not found within the ovarian tissue. This suggests that vertical transmission ofNeorickettsia within adult digeneans occurs via the incorporation of infected vitelline cells into the egg rather than direct infection of the ooplasm of the oocyte, as has been described for other bacterial endosymbionts of invertebrates (e.g.,Rickettsia andWolbachia ).

Germs within Worms: Localization of Neorickettsia sp. within Life Cycle Stages of the Digenean Plagiorchis elegans