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Many insects possess endosymbiotic bacteria inside their body, wherein intimate interactions occur between the partners. While recent technological advancements have deepened our understanding of metabolic and evolutionary features of the symbiont genomes, molecular mechanisms underpinning the intimate interactions remain difficult to approach because the insect symbionts are generally uncultivable. The bean bug   R iptortus pedestris  is associated with the betaproteobacterial   B urkholderia  symbiont in a posterior region of the midgut, which develops numerous crypts harbouring the symbiont extracellularly. Distinct from other insect symbiotic systems,   R . pedestris  acquires the   B urkholderia  symbiont not by vertical transmission but from the environment every generation. By making use of the cultivability and the genetic tractability of the symbiont, we constructed a transgenic   B urkholderia  strain labelled with green fluorescent protein ( GFP ), which enabled detailed observation of spatiotemporal dynamics and the colonization process of the symbiont in freshly prepared specimens. The symbiont live imaging revealed that, at the second instar, colonization of the symbiotic midgut  M 4 region started around 6 h after inoculation (hai). By 24 hai, the symbiont cells appeared in the main tract and also in several crypts of the  M 4. By 48 hai, most of the crypts were colonized by the symbiont cells. By 72 hai, all the crypts were filled up with the symbiont cells and the symbiont localization pattern continued during the subsequent nymphal development. Quantitative  PCR  of the symbiont confirmed the infection dynamics quantitatively. These results highlight the stinkbug‐  B urkholderia  gut symbiosis as an unprecedented model for comprehensive understanding of molecular mechanisms underpinning insect symbiosis.

Live imaging of symbiosis: spatiotemporal infection dynamics of a GFP ‐labelled B urkholderia symbiont in the bean bug R iptortus pedestris