Larvicidal activities against agricultural pests of transgenic Escherichia coli expressing combinations of four genes from Bacillus thuringiensis.
The genes cry1Ac and cry1Ca from Bacillus thuringiensis subsps. kurstaki HD-73 and aizawai 4J4, respectively, encoding delta-endotoxins against lepidopteran larvae were isolated, cloned and expressed in Escherichia coli, with and without cyt1Aa (encoding cytolytic protein) and p20 (accessory protein) from subsp. israelensis. Nine combinations of the genes under control of an early T7, P A1 inducible promoter, produced the encoding proteins. Toxicities were examined against larvae of three major agricultural pests: Pectinophora gossypiella, Helicoverpa armigera and Spodoptera littoralis. The clones expressing cyt1Aa, with or without p20, were not toxic. The clone expressing cry1Ac (pBt-1A) was the most toxic to P. gossypiella (LC50 of 0.27 x 10(8) cells g(-1)). Clone pBt-1CA expressing cry1Ca and cry1Ac displayed the highest toxicity (LC50 of 0.12 x 10(8) cells ml(-1)) against S. littoralis. Clone pBt-1CARCy expressing all four genes (cry1Ca, cry1Ac, p20, cyt1Aa) in tandem exhibited the highest toxicity to H. armigera (LC50 of 0.16 x 10(8) cells ml(-1)). Cyt1Aa failed to raise the toxicity of these Cry toxins against P. gossypiella and S. littoralis but significantly enhanced toxicity against H. armigera. Two additional clones expressing either cry1Ac or cry1Ca under tandem promoters, P A1 and P psbA (constitutive), displayed significantly higher toxicities (7.5- to 140-fold) than their counterparts with P A1 alone, reducing the LC50 values to below 10(7) cells ml(-1).
Department of Life Sciences, Ben-Gurion University of the Negev, PO Box 653, Be'er-Sheva 84105, Israel.
Vadim
V
Khasdan
Maria
M
Sapojnik
Arieh
A
Zaritsky
A Rami
AR
Horowitz
Sammy
S
Boussiba
Mario
M
Rippa
Robert
R
Manasherob
Eitan
E
Ben-Dov
D000383:Agriculture; D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D001427:Bacterial Toxins / Q000096:biosynthesis* / Q000235:genetics; D004731:Endotoxins / Q000096:biosynthesis* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015870:Gene Expression; D006460:Hemolysin Proteins / Q000096:biosynthesis* / Q000235:genetics; D019476:Insect Proteins / Q000096:biosynthesis* / Q000235:genetics; D007814:Larva / Q000382:microbiology; D007915:Lepidoptera / Q000382:microbiology*; D030781:Organisms, Genetically Modified; D010572:Pest Control, Biological / Q000379:methods*; D011956:Receptors, Cell Surface / Q000096:biosynthesis* / Q000235:genetics
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; C095416:Cry toxin receptors; D004731:Endotoxins; D006460:Hemolysin Proteins; D019476:Insect Proteins; D011956:Receptors, Cell Surface; C030956:insecticidal crystal protein, Bacillus Thuringiensis