Discovery and characterization of Sip1A: a novel secreted protein from Bacillus thuringiensis with activity against coleopteran larvae

Abstract

Bioassay screening of Bacillus thuringiensis culture supernatants identified strain EG2158 as having larvicidal activity against Colorado potato beetle (Leptinotarsa decemlineata) larvae. Ion-exchange fractionation of the EG2158 culture supernatant resulted in the identification of a protein designated Sip1A (secreted insecticidal protein) of approximately 38 kDa having activity against Colorado potato beetle (CPB). An oligonucleotide probe based on the N-terminal sequence of the purified Sip1A protein was used to isolate the sip1A gene. The sequence of the Sip1A protein, as deduced from the sequence of the cloned sip1A gene, contained 367 residues (41,492 Da). Recombinant B. thuringiensis and Escherichia coli harboring cloned sip1A produced Sip1A protein which had insecticidal activity against larvae of CPB, southern corn rootworm (Diabrotica undecimpunctata howardi), and western corn rootworm (Diabrotica virgifera virgifera).

Bacterial Vegetative Insecticidal Proteins (Vip) from Entomopathogenic Bacteria

“…It should be mentioned that another secreted insecticidal protein from B. thuringiensis, named Sip, has been reported. This protein shares no homology with the Vip proteins and should not be mistaken for one of them (10).…”

Insecticidal activity of Bacillus thuringiensis crystal proteins

“…Also included are families of highly related crystal proteins produced by other bacteria: Cry16 and Cry17 from Clostridium bifermentans (Barloy et al, 1996(Barloy et al, , 1998, Cry18 from Bacillus popilliae (Zhang et al, 1997), Cry43 from Paenibacillus lentimorbis (Yokoyama et al, 2004) and the binary Cry 48/Cry49 produced by Bacillus sphaericus (Jones et al, 2008). Other crystalline or secreted pesticidal proteins, such as S-layer proteins (Peña et al, 2006;Guo et al, 2008a), vegetative insecticidal proteins (Estruch et al, 1996), and secreted insecticidal proteins (Donovan et al, 2006) are not included in this review. Neither are genetically altered crystal proteins, except those that were modified through single amino acid substitutions (e.g., Lambert et al, 1996).…”

Draft Genome Sequence of Bacillus thuringiensis NBIN-866 with High Nematocidal Activity

“…It shows toxicity to certain insect species (5, 6) and has been widely used as a biopesticide for controlling agricultural pests. B. thuringiensis can produce multiinsecticidal metabolites, like insecticide crystal proteins (7), cytotoxin (8), vegetative insecticidal protein (9), secret insecticidal protein (10), thuringiensin (11), zwittermicin A (12), Mtx-like toxin (13), Bin-like toxin (14), and more. The diversity of toxins is paralleled by diverse pesticidal activity.…”