Pollen tubes introduce Raspberry bushy dwarf virus into embryo sacs during fertilization processes.

We developed a fertilization method in which pollen tubes entered into embryo sacs without any need to contact surrounding female sporophytic cells by using Torenia fournieri (Torenia) plants under the condition of hindering movement of the virus from a stigma, which is the first infection site leading to systemic infection. When RBDV-infected Torenia pollen grains were used for the developed fertilization method, the virus was transmitted to the seeds by pollen tubes germinating from them. On the other hand, no seeds were infected with the virus when Torenia plants were pollinated with healthy Torenia pollen grains in combination with RBDV-infected raspberry pollen grains, which caused the virus infection in the stigma by penetration of their pollen tubes arrested in its style. Our results indicate that vertical transmission of RBDV by pollen occurs in the transport of the virus into embryo sacs by pollen tubes reaching the embryo sacs.

Penetration of pollen tubes with accumulated Raspberry bushy dwarf virus into stigmas is involved in initial infection of maternal tissue and horizontal transmission.

Torenia fournieri (Torenia) plants were infected with Raspberry bushy dwarf virus (RBDV) by pollination with RBDV-infected raspberry pollen grains. The infected raspberry pollen grains germinated on Torenia stigmas, and then the pollen tubes penetrated into the stigma, even though the pollen tubes were arrested in the styles. In whole-mount in situ hybridization of germinating infected raspberry pollen grains, RBDV accumulated in the tips of the pollen tubes. Tissue blot hybridization of Torenia plants pollinated with infected raspberry pollen grains revealed that the first virus infection site leading to systemic infection is the stigma. When infected raspberry pollen grains that had lost germination capacity were pollinated on Torenia stigmas, RBDV could not infect the stigmas, and no horizontal transmission occurred. These results indicate that penetration of pollen tubes with accumulated RBDV into stigmas is essential in causing the first viral infection in the stigma to lead to systemic infection.