Nosema bombycis (Microsporidia) suppresses apoptosis in BmN cells (Bombyx mori).

Nosema bombycis (N. bombycis, Nb) is a fungus-related and obligate intracellular parasite that causes chronic pebrine disease in the silkworm. After infecting the host, spores obtain energy from host cells and survive for several days. This symbiosis between the pathogen and the host cell suggests that N. bombycis prevents apoptosis and reactive oxygen species (ROS) production of host cells to create the optimal environmental conditions for its growth and development. In this study, different methods were used to prove that N. bombycis suppressed apoptosis in BmN cells. Flow cytometry analysis results showed that spores suppressed apoptosis of BmN cells at 2 and 5 days after infection (P < 0.05). Compared with actinomycin D (ActD) treatment, apoptosis of BmN cells was apparently reduced after spore infection (P < 0.01). Forty-eight hours after infection, the ROS production of BmN cells was down-regulated compared with that after ActD treatment for 6 h. Furthermore, N. bombycis prevented the formation of apoptosomes by down-regulating the expression of apaf-1 and cytochrome C. In addition, N. bombycis also up-regulated the expression of buffy. Western blot analysis demonstrated that spores decreased the level of host cytochrome C at 48 and 98 h post infection. Thus, our results suggested that N. bombycis inhibited the mitochondrial apoptotic pathway of the host cells to create an optimal environment for its own survival.

Detection of an internal translation activity in the 5' region of Bombyx mori infectious flacherie virus.

The 5' untranslated region plays an important role in positive-sense single-stranded RNA virus translation initiation, as it contains an internal ribosome entry site (IRES) that mediates cap-independent translation and is applied to simultaneously express several proteins. Infectious flacherie virus (IFV) is a positive-sense single-stranded RNA virus; however, the IRES function is still not proved. To investigate whether the sequences of IFV contain IRES activity, a series of bicistronic reporter (DsRed and enhanced green fluorescent protein) recombinant baculoviruses were constructed to infect the insect cells and silkworm using the Bombyx mori baculovirus expression system. Results showed that the upstream 311, 323, 383, 551, and 599 nt have IRES activity except for the 155-nt region in BmN cells. More importantly, the tetraloop structure containing region between 551 and 599 nt appeared to be responsible for the enhanced IRES activity in different insect cell lines and silkworm. These results indicated that the IRES activity is not species specific and tissue specific. Therefore, our findings may provide the basis for the simultaneous expression of two or various different genes under the same promoter in baculovirus expression system.