bThe entomopathogenic bacteria Xenorhabdus nematophila and Photorhabdus temperata subsp. temperata suppress insect immune responses by inhibiting the catalytic activity of phospholipase A 2 (PLA 2 ), which results in preventing biosynthesis of immune-mediating eicosanoids. This study identified PLA 2 inhibitors derived from culture broths of these two bacteria. Both X. nematophila and P. temperata subsp. temperata culture broths possessed significant PLA 2 -inhibitory activities. Fractionation of these bacterial metabolites in the culture broths using organic solvent and subsequent chromatography purified seven potent PLA 2 inhibitors, three of which (benzylideneacetone [BZA], proline-tyrosine [PY], and acetylated phenylalanine-glycine-valine [FGV]) were reported in a previous study. Four other compounds (indole, oxindole, cis-cyclo-PY, and p-hydroxyphenyl propionic acid) were identified and shown to significantly inhibit PLA 2 . X. nematophila culture broth contained these seven compounds, while P. temperata subsp. temperata culture broth contained three compounds (BZA, acetylated FGV, and cis-cyclo-PY). BZA was detected in the largest amount among these PLA 2 compounds in both bacterial culture broths. All seven bacterial metabolites also showed significant inhibitory activities against immune responses, such as phenoloxidase activity and hemocytic nodulation; BZA was the most potent. Finally, this study characterized these seven compounds for their insecticidal activities against the diamondback moth, Plutella xylostella. Even though these compounds showed relatively low toxicities to larvae, they significantly enhanced the pathogenicity of Bacillus thuringiensis. This study reports bacterial-origin PLA 2 inhibitors, which would be applicable for developing novel insecticides. Insect immune responses consist of innate cellular and humoral components (2). Upon pathogen infection, pattern recognition receptors recognize nonself and the recognition signal is transferred by immune mediators to effector tissues, such as hemocytes and fat body (7). Eicosanoids form a chemical group of oxygenated polyunsaturated C 20 fatty acids, which are derived from arachidonic acid, a catalytic product of phospholipase A 2 (PLA 2 ) on phospholipid (PL) substrate (3). Two types of eicosanoids, prostaglandins and leukotrienes, are known to mediate the nonself recognition signals against various insect pathogens, including bacteria, fungi, viruses, and parasitoid eggs in insects (38). Inhibition of PLA 2 prevents eicosanoid biosynthesis and suppresses immune responses in response to microbial pathogens (23, 37).Xenorhabdus nematophila and Photorhabdus temperata subsp. temperata are insect pathogens that are symbiotic to the entomopathogenic nematodes Steinernema carpocapsae and Heterorhabditis megidis, respectively (1, 13, 25). Infective juvenile-stage nematodes enter target insect hemocoel through natural openings (such as mouth, anus, and spiracles) and then release their symbiotic bacteria from the nematode intestine. The releas...

Bacillus thuringiensis (Bt) is a bacterial biopesticide against insect pests, mainly lepidopterans. Spodoptera exigua and Plutella xylostella exhibit significant decreases in Bt susceptibility in late larval instars. To enhance Bt pathogenicity, we used a mixture treatment of Bt and other bacterial metabolites which possessed significant immunosuppressive activities. Mixtures of Bt with culture broths of Xenorhabdus nematophila (Xn) or Photorhabdus temperata ssp. temperata (Ptt) significantly enhanced the Bt pathogenicity against late larval instars. Different ratios of Bt to bacterial culture broth had significant pathogenicities against last instar P. xylostella and S. exigua. Five compounds identified from the bacterial culture broth also enhanced Bt pathogenicity. After determining the optimal ratios, the mixture was applied to cabbage infested by late instar P. xylostella or S. exigua in greenhouse conditions. A mixture of Bt and Xn culture broth killed 100% of both insect pests when it was sprayed twice, while Bt alone killed less than 80% or 60% of P. xylostella and S. exigua, respectively. Other Bt mixtures, including Ptt culture broth or bacterial metabolites, also significantly increased pathogenicity in the semi-field assays. These results demonstrated that the Bt mixtures collectively names "Bt-Plus" can be developed into potent biopesticides to increase the efficacy of Bt. (Plutella xylostella)에서 보듯 종령기에 접어들면 Bt 감수성이 현격하게 낮아져 방제효율이 낮아진다. 본 연구는 Bt의 병원력을 제고시키기는 데 목적을 두고, 이를 위해 곤충의 면역반응을 억제시키는 곤충병원세균의 대사물질을 이용하여 Bt의 혼합제를 개발하려 했다. 곤충병원세균 (Xenorhabdus nematophila (Xn) 또는 Photorhabdus temperata ssp. temperata (Ptt))의 배양액을 다양한 농도로 Bt에 혼합하여 파밤나방과 배추 좀나방 종령 유충에 처리하였을 때 Bt 병원력을 현격하게 증가시켰다. 세균 배양액으로부터 동정한 다섯 가지의 화합물을 Bt와 혼합하여 처리하 면 Bt 병원력을 또한 증가시켰다. 최적의 방제 효과를 나타낼 수 있는 Bt와 세균배양액 또는 대사물질의 혼합비율과 살포량을 결정한 후 배추 포 장에 발생한 배추좀나방과 파밤나방 종령 유충에 대해 처리하였다. Bt와 Xn 배양액을 2 회 이상 살포하면 배추좀나방과 파밤나방에 대해 100% 의 방제효과를 나타냈으나, Bt 단독으로 처리하면 각각 약 80%와 60% 이하의 방제효과를 나타내었다. Ptt 배양액 및 세균 대사물질을 포함한 다 른 Bt 혼합제들 또한 뚜렷한 병원력 증가를 간이포장실험에서 보여주었다. 이러한 Bt 혼합제를 통틀어 "비티플러스"라 명명하였으며, 이는 Bt 약 효의 단점을 극복한 효과적 미생물 살충제로 개발 가능성을 보여주었다.

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