A phase variation has been reported in an entomopathogenic bacterium, Xenorhabdus nematophila. Compared to a wild type primary form, a secondary form usually lose several physiological and biochemical characters. This study showed that the phase variation of X. nematophila caused a significant alteration in its immunosuppressive activity and subsequent entomopathogenicity. A secondary form of X. nematophila was detected in laboratory colonies and exhibited significant differences in dye absorption and entomopathogenicity. In addition, the secondary form was different in production of eicosanoid-biosynthesis inhibitors (EBIs) compared to the primary form of X. nematophila. Production of oxindole and p-hydroxypropionic acid was significantly reduced in the culture broth of the secondary form of X. nematophila. The reduced EBI production resulted in significant suppression in the inhibitory effects on a cellular nodule formation and phenoloxidase activity. Culture broth of the primary form of X. nematophila significantly more enhanced the pathogenicity of Bacillus thuringiensis (Bt) than the culture broth of the secondary form. Furthermore, this study developed a high efficient ‘Dual Bt-Plus’ to control both lepidopteran insect pests of Plutella xylostella and Spodoptera exigua by mixing two effective Bt strains along with the addition of potent bacterial metabolites or 100-fold concentrated X. nematophila culture broth.

An entomopathogenic bacterium, Xenorhabdus nematophila (Xn), is symbiotic to a nematode, Steinernema carpocapsae, and exhibits high pathogenicity to lepidoptera insects. Its metabolites released into the bacterial culture broth and also virulent in oral especially when they are treated with Bacillus thuringiensis (Bt). This study devised a high efficacy microbial insecticide by combining Xn culture broth and Bt. Bt kurstaki (Btk) exhibited relatively higher pathogenicity to Plutella xylostella than Spodoptera exigua larvae. By contrast, Bt aizawai (Bta) showed a reverse pathogenicity pattern. Phase Ⅰ type of Xn (XnK1) was isolated from S. carpocapsae Pochun and exhibited high pathogenicity than phase Ⅱ bacteria. Three bacterial mixtures of Bta+XnK1, Btk+XnK1, and Bta+Btk+XnK1 were prepared and analyzed in their target insects. Bta+XnK1 showed higher pathogenicity than those of Bta alone or Btk+XnK1 in P. xylostella. Btk+XnK1 showed higher pathogenicity than those of Btk alone or Bta+XnK1 in S. exigua. Bta+BtK+XnK1 showed high pathogenicity against both P. xylostella and S. exigua.

An entomopathogenic bacterium, Xenorhabdus nematophila, secretes at least eight bacterial metabolites, which have suppressive effects on insect immunity. This study quantified their sequential production during bacterial growth and analyzed their individual immunosuppressive activities against an insect host, Spodoptera exigua. X. nematophila exhibited a typical bacterial growth in both insect host and culture medium, in which eight metabolites were secreted in different time points. At early growth phase (6 to 12 h), Ac-FGV, Cis-cPY, PHPP and indole metabolites were detected in the culture broth. During early growth phase, PHPP was highly potent to inhibit phenoloxidase activity as well as nodule formation. At late growth phase (24 to 48 h), BZA, HPA, PY were detected at 10 – 140 ppm in the culture broth, their metabolites were highly potent to inhibit phospholipase A2 and to induce cytotoxicity to hemocytes. These results suggest that X. nematophila sequentially produces the immune suppressive metabolites, which cooperatively inhibit different steps of insect immune responses.