Aims: To study the ability of Bacillus thuringiensis subsp. israelensis spores to germinate and subsequently transfer a conjugative plasmid in the intestinal tract of gnotobiotic rats. Methods and Results: Germination was studied by feeding germ-free rats with spores of a B. thuringiensis strain harbouring a plasmid encoding green fluorescent protein (GFP), which enabled quantification of germinated bacteria by flow cytometry. To study in vivo conjugation, germ-free rats were first associated with a B. thuringiensis recipient strain and after 1 week an isogenic donor strain harbouring the conjugative plasmid pXO16 was introduced. Both strains were given as spores and transfer of pXO16 was observed from the donor to the recipient strain. Conclusions: Bacillus thuringiensis is able to have a full life cycle in the intestine of gnotobiotic rats including germination of spores, several cycles of growth and sporulation of vegetative cells. For the first time conjugative plasmid transfer in a mammalian intestinal tract was shown between two B. thuringiensis strains. Significance and Impact of the Study: Strains of B. thuringiensis are used worldwide to combat insect pests, and this study brings new insights into the nature of B. thuringiensis showing the potential of the bacteria to germinate and transfer DNA in the mammalian intestinal tract.