Destruxin A

Spodoptera litura is one of the most destructive phytophagous pest infesting cotton, vegetable, oilseed and ber crops around the world. Dextruxin A (DA), is a one of a kind microbial insecticide, which has potent toxins with bioactivity against S. litura larvae. An abnormal wing disc (AWD) protein was identified as a DA toxic effect protein in S. litura SL-1 cells. To better understand the role of the AWD gene of S. litura (SLAWD) it was purified and characterized. The entire coding region of the SLAWD gene was cloned into a pET-32a(+) expression vector and transformed into competent Escherichia coli BL21 (DE3) cells. SDS-PAGE and western blotting analysis and western blotting showed that the best induction conditions were 1 mmol mL(-1) isopropyl-β-D-thiogalactopyranoside (IPTG) for 6 h at 37°C; the molecular weight of the fusion protein was 35.0 kDa. The production of polyclonal antibodies and an enzyme-linked immunosorbent assay (ELISA) showed that the titer of antiserum was 1:25,600; western blotting analysis showed that the recombinant SLAWD was recognized by the anti-SLAWD polyclonal antibody.

Destruxin A is a mycotoxin that is secreted by entomopathogenic fungi which has a broad-spectrum insecticidal effect. Previous transcript and protein profiling analysis showed that destruxin A has significant effects on the expression of serine protease inhibitor genes (serpin-2, 4, 5) in the larvae of Plutella xylostella. In the current study, we aimed to understand the role of serpins under application of destruxin A. We obtained two full-length cDNA sequences of P. xylostella serpins, named serpin-4 and serpin-5, and cloned the serpin-2 gene whose full-length has already been published. Phylogenetic analysis indicated that these two serpin genes were highly clustered with other serpins associated with the immune response in other insects. The temporal and spatial expression of serpin-2, serpin-4 and serpin-5 were determined to be the highest in the fat body and hemolymph of 4th larval stage using qRT-PCR and western blot detection techniques.

Destruxin A (DA), a cyclodepsipeptidic secondary metabolite of the entomopathogenic fungus, Metarhizium anisopliae, is an important anti-immunity agent against insect hemocytes. To understand the mechanism of the molecular responses to DA, fifth-instar larvae of the silkworm, Bombyx mori, were injected with 2 μg of DA. The proteomics of hemocytes were then investigated using two-dimensional electrophoresis and mass spectrometry, and validated qPCR. As a result, a total of 47 differently expressed protein spots were detected and 22 proteins in 26 spots were identified. There are eight immunity-related proteins, including three downregulated proteins (antitrypsin isoform 3, p50 protein, and calreticulin precursor) and five upregulated proteins (C-type lectin 10 precursor, serine proteinase-like protein, paralytic peptide, PPO-1, and PPO-2). Four resistance- and/or stress-related proteins (arginine kinase, carboxylesterase clade H, member 1, aminoacylase, and thiol peroxiredoxin) were upregulated.

Destruxin A (DA) is a cyclo-peptidic mycotoxin from the entomopathogenic fungus Metarhizium anisopliae. To uncover potential genes associated with its molecular mechanisms, a digital gene expression (DGE) profiling analysis was used to compare differentially expressed genes in the hemocytes of silkworm larvae treated with DA. Ten DGE libraries were constructed, sequenced, and assembled, and the unigenes with least 2.0-fold difference were further analyzed. The numbers of up-regulated genes were 10, 20, 18, 74 and 8, as well as the numbers of down-regulated genes were 0, 1, 8, 13 and 3 at 1, 4, 8, 12 and 24 h post treatment, respectively. Totally, the expression of 132 genes were significantly changed, among them, 1, 3 and 12 genes were continually up-regulated at 4, 3 and 2 different time points, respectively, while 1 gene was either up or down-regulated continually at 2 different time points. Furthermore, 68 genes were assigned to one or multiple gene ontology (GO) terms and 89 genes were assigned to specific Kyoto Encyclopedia of Genes and Genomes (KEGG) Orthology.