The Cypripedium japonicum is a species of orchid, distributed mainly in the temperate regions of the Northern Hemisphere. In Korea, C. japonicum is an extremely rare species and class I endangered plant. However, little is known about the plant-microbe interaction and its ecology, especially rhizosphere bacterial communities in terms of structure and diversity in C. japonicum. In this study, the diversity of bacterial communities was investigated in C. japonicum rhizosphere and bulk soil. Proteobacteria, Acidobacteria and Actinobacteria were the most abundant groups in both rhizosphere and bulk soil. Significantly Proteobacteria and Actinobacteria abundantly inhabited in the rhizosphere when compared with the bulk soil. Conversely, significantly more unclassified bacteria were detected in the rhizosphere soil than in the bulk soil. This work increases our understanding of the rhizosphere bacterial diversity on class I endangered plant, C. japonicum.

During the last century, the overuse of synthetic chemical insecticides led to problems associated with natural environment such as harmful effects on nontarget organisms and development of resistance. Therefore, the application of insect pathogenic microorganisms including Bacillus thuringiensis (Bt), baculoviruses, and entomopathogenic fungi has been an environmentally favorable methods for insect control. However, their use has been limited by several factors such as narrow host spectrum and slow speed of insecticidal activities. A number of approaches have been taken to overcome these defects of microbial insecticides. A novel recombinant baculovirus, NeuroBactrus, was constructed to develop an improved baculovirus insecticide with additional beneficial properties, such as a higher insecticidal activity and improved recovery, compared to wild-type baculovirus. For the construction of the NeuroBactrus, the Bt cry1-5 crystal protein gene was introduced into the Autographa californica nucleopolyhedrovirus (AcMNPV) genome by fusion of polyhedrin-cry1-5-polyhedrin under the control of the polyhedrin promoter. In the opposite direction, an insect-specific neurotoxin gene, AaIT, from Androctonus australis was introduced under the control of an early promoter from Cotesia plutellae bracovirus by fusion of a partial fragment of orf603. The Polyhedrin-Cry1-5-Polyhedrin fusion protein expressed by the NeuroBactrus was not only occluded into the polyhedra, but also activated by treatment with trypsin, resulting in an approximately 65-kDa active toxin. The NeuroBactrus showed a high level of insecticidal activity against Plutella xylostella larvae and a significant reduction in the median lethal time (LT50) against Spodoptera exigua larvae compared to those of wild-type AcMNPV. Expression of the foreign proteins (Bt toxin and AaIT) was continuously reduced during the serial passage of the NeuroBactrus. These results suggested that the NeuroBactrus could be recovered to wild-type AcMNPV through serial passaging. The cry gene from Bt, encoding the Cry protein, has been recently introduced into crops to generate transgenic plants that are resistant to pest insects. Through the 3D structure prediction and accompanying mutagenesis study for the Mod-Cry1Ac, 7 and 16 amino acid residues from domain I and II, respectively, responsible for its insecticidal activity against larvae of S. exigua and Ostrinia furnacalis were identified. We used site-directed mutagenesis to improve the insecticidal activity of Mod-Cry1Ac, resulted 34 mutant cry genes. These mutant cry genes were expressed, as a polyhedrin fusion form, using a baculovirus expression system. The expressed proteins were 95 kDa and SDS-PAGE analysis of the recombinant polyhedra revealed that expressed Cry proteins was occluded into polyhedra and activated stably to 65 kDa by trypsin. When the insecticidal activities of these mutant Cry proteins against to larvae of P. xylostella, S. exigua and O. furnacalis were assayed, they showed higher or similar insecticidal activity compared to those of Cry1Ac and Cry1C. Especially, Mutant-N16 is considered to have the potential for the efficacious biological insecticide since it showed the highest insecticidal activity.

A total of 240 weaned pigs (Landrace ×Yorkshire × Duroc, 22±3 d of age, 5.16±0.90 kg initial body weight) were used to study the effect of feeding level of microbial fermented soya protein on their blood hematology, enzymes and immune cell populations. The microbial (Aspergillus oryzae + Bacillus subtilis) fermented soya protein (FSP) was used. Pigs were allotted to four dietary treatments, each comprising of 4 pens with 15 pigs. Basal diets consisted of 15% soya bean meal (Control diet); while for treatment diets SBM was replaced with 3, 6 and 9% FSP. The experimental diets were fed from 0 to 14 day after weaning and then a common commercial diet was fed from 15 to 35 day. Blood was collected on 14 and 35 day of experiment and analyzed for hematology, plasma aspartate transaminase (AST), plasma alanine transaminase (ALT) and immune cell populations. Increasing the level of FSP in the diet of pigs linearly decreased distribution of red blood cells (P<0.01), MPV (P<0.05), MO (P<0.05), EO (P<0.05) and BA (P<0.05) on d 14. Linear and quadratic decrease in the RBC (P<0.05), Hb (P<0.05), HCT (P<0.01), PLT (P<0.001) and EO (P<0.05) and linear increase in the MCHC (P<0.001), MPV (P<0.05), WBC (P<0.05) and NE (P<0.05) on d 35 was noted. Pigs fed with 6% FSP had lower (P<0.05) levels of AST and ALT on d 14, while the levels of ALT and AST on d 35 did not differ among the dietary treatments. Thus the results suggest that microbial fermented soya protein affected the hematological indices, immune cell populations and plasma enzymes in weaned pigs.