Involvement Of Response Regulator OmpR In Virulence Control Of Yersinia Pseudotuberculosis In Silkworm, Bombyx Mori

The silkworm Bombyx mori has been widely used for scientific studies as a model for lepidopteran insects because of its large body size, easy rearing, and a large number of described mutants. The silkworm is the first lepidopteran insect that have been sequenced the complete genome. Study of the interaction between silkworm and pathogens not only provides scientific basis for prevention of silkworm diseases, but also provides new targets in biological control of pests.In the present study, we investigated the mortality and immune responses of silkworms injected with wild-type or ΔompR Yersinia pseudotuberculosis to find out the functions of response regulator OmpR during invading Bombyx mori.Initially we detected the mortality of silkworm larvae infected with wild-type or ΔompR Y. pseudotuberculosis. Our results showed that the mortality of ΔompR-injected larvae was higher than which of larvae injected with wild-type bacteria. ΔompR grows more slowly than the wild-type bacteria when cultured in Luria-Bertani medium but more quickly inside the silkworm larvae. These results indicated that the response regulator OmpR acts to reduce the virulence of Y. pseudotuberculosis in B. mori.Next we found that the antibacterial activity of hemolymph from silkworm larvae infected with wild-type Y. pseudotuberculosis was higher than which of the ΔompR-injected ones at 12 and 24 hours post injection(hpi). The results of Tricine-SDS-PAGE coupled with LC-MS/MS, and quantitative real-time PCR showed that the expression levels of Attacin-2, Cecropin-B6 and Gloverin-2 trigged by wild-type Y. pseudotuberculosis were significantly higher than that was trigged by ΔompR bacteria.We also found that phenolosidase activity of hemolymph from ΔompR-injected larvae were significantly lower than from wild-type injected larvae at 6, 12 and 24 hpi. Spontaneous melanization activity of hemolymph from ΔompR-injected larvae at 12 hpi was apparently inhibited, indicating that melanization in silkworm was suppressed by ΔompR Y. pseudotuberculosis.Finally, we examined the survival rate of C57BL/6 mice after intravenously injection with wild-type or ΔompR Y. pseudotuberculosis. All the wild-type injected mice died in 4 days, but no death was observed in ΔompR-injected mice in 15 days. This result indicated that response regulator OmpR in Y. pseudotuberculosis positively involves in the virulence in mice.Taken together, we found that response regulator OmpR negatively controls the virulence of Y. pseudotuberculosis in B. mori but positively in C57BL/6 mice. This finding may provide new insight in high efficiency and low toxicity bacterial pesticide development.