Functional characterization of trehalases 1, 2, and 4 in Nosema bombycis (microsporidia)

Pebrine caused by Nosema bombycis infection is one of the most threatening diseases that affect sericulture. Trehalases play important roles in the lives of organisms, such as spore germination, insect flight, and stress resistance. The genome of N. bombycis has four trehalase genes, but their functions in the infection and development of N. bombycis remain unclear. In this study, the functions of N. bombycis trehalase (NbTre) 1, 2, and 4 were investigated. The expression profile results revealed that at the early infection stage, the transcripts of the NbTre genes reached the highest levels at 2 h (NbTre4) and 6 h (NbTre1 and NbTre2), respectively, after infection. After infecting BmN cells, before the spores mature, NbTre1 is located primarily in the sporoplasm; NbTre2 and NbTre4 are secreted into the host cytoplasm and nucleus, respectively, where they may play different roles. In dormant spores, NbTre1 and NbTre4 are stored in the sporoplasm membrane or the spore wall. However, NbTre2 was not detected in the dormant spores of N. bombycis. Silencing the expression of two rather than a single NbTre gene with small interfering RNAs could reduce the replication of N. bombycis more effectively. Further when all four NbTre genes was silenced expressing by RNA interference synchronously, the replication of N. bombycis was suppressed to the maximum extent. These results suggested that these four NbTres play important roles in the proliferation of N. bombycis during the intracellular stage and that there may be complementary and cooperative relationships among the four NbTres.