Protocol for isolating, culturing, and artificially infecting the Arsenophonus endosymbiont of Nilaparvata lugens

Endosymbionts play a crucial role in insect physiology and adaptation. Here, we present a protocol for isolation, cultivation, and artificial infection of the Arsenophonus endosymbiont of Nilaparvata lugens. We describe steps for symbiont isolation, in vitro culture maintenance, genetic modification, and host reinfection. This protocol enables controlled studies of symbiont-host interactions and provides a foundation for symbiont-based pest management strategies. For complete details on the use and execution of this protocol, please refer to Cai et al.¹.