Skip to main content
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1997 Jul;63(7):2716–2721. doi: 10.1128/aem.63.7.2716-2721.1997

Identification and characterization of a previously undescribed cyt gene in Bacillus thuringiensis subsp. israelensis.

A Guerchicoff 1, R A Ugalde 1, C P Rubinstein 1
PMCID: PMC168567  PMID: 9212418

Abstract

Mosquitocidal Bacillus thuringiensis strains show as a common feature the presence of toxic proteins with cytolytic and hemolytic activities, Cyt1Aa1 being the characteristic cytolytic toxin of Bacillus thuringiensis subsp. israelensis. We have detected the presence of another cyt gene in this subspecies, highly homologous to cyt2An1, coding for the 29-kDa cytolytic toxin from B. thuringiensis subsp. kyushuensis. This gene, designated cyt2Ba1, maps upstream of cry4B coding for the 130-kDa crystal toxin, on the 72-MDa plasmid of strain 4Q2-72. Sequence analysis revealed, as a remarkable feature, a 5' mRNA stabilizing region similar to those described for some cry genes. PCR amplification and Southern analysis confirmed the presence of this gene in other mosquitocidal subspecies. Interestingly, anticoleopteran B. thuringiensis subsp. tenebrionis belonging to the morrisoni serovar also showed this gene. On the other hand, negative results were obtained with the anti-lepidopteran strains B. thuringiensis subsp. kurstaki HD-1 and subsp. aizawai HD-137. Western analysis failed to reveal Cyt2A-related polypeptides in B. thuringiensis subsp. israelensis 4Q2-72. However, B. thuringiensis subsp. israelensis 1884 and B. thuringiensis subsp. tenebrionis did show cross-reactive products, although in very small amounts.

Full Text

The Full Text of this article is available as a PDF (998.6 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Agaisse H., Lereclus D. How does Bacillus thuringiensis produce so much insecticidal crystal protein? J Bacteriol. 1995 Nov;177(21):6027–6032. doi: 10.1128/jb.177.21.6027-6032.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Agaisse H., Lereclus D. Structural and functional analysis of the promoter region involved in full expression of the cryIIIA toxin gene of Bacillus thuringiensis. Mol Microbiol. 1994 Jul;13(1):97–107. doi: 10.1111/j.1365-2958.1994.tb00405.x. [DOI] [PubMed] [Google Scholar]
  3. Baum J. A., Malvar T. Regulation of insecticidal crystal protein production in Bacillus thuringiensis. Mol Microbiol. 1995 Oct;18(1):1–12. doi: 10.1111/j.1365-2958.1995.mmi_18010001.x. [DOI] [PubMed] [Google Scholar]
  4. Ben-Dov E., Boussiba S., Zaritsky A. Mosquito larvicidal activity of Escherichia coli with combinations of genes from Bacillus thuringiensis subsp. israelensis. J Bacteriol. 1995 May;177(10):2851–2857. doi: 10.1128/jb.177.10.2851-2857.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ben-Dov E., Einav M., Peleg N., Boussiba S., Zaritsky A. Restriction map of the 125-kilobase plasmid of Bacillus thuringiensis subsp. israelensis carrying the genes that encode delta-endotoxins active against mosquito larvae. Appl Environ Microbiol. 1996 Sep;62(9):3140–3145. doi: 10.1128/aem.62.9.3140-3145.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bourgouin C., Klier A., Rapoport G. Characterization of the genes encoding the haemolytic toxin and the mosquitocidal delta-endotoxin of Bacillus thuringiensis israelensis. Mol Gen Genet. 1986 Dec;205(3):390–397. doi: 10.1007/BF00338072. [DOI] [PubMed] [Google Scholar]
  8. Brown K. L. Transcriptional regulation of the Bacillus thuringiensis subsp. thompsoni crystal protein gene operon. J Bacteriol. 1993 Dec;175(24):7951–7957. doi: 10.1128/jb.175.24.7951-7957.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Delécluse A., Poncet S., Klier A., Rapoport G. Expression of cryIVA and cryIVB Genes, Independently or in Combination, in a Crystal-Negative Strain of Bacillus thuringiensis subsp. israelensis. Appl Environ Microbiol. 1993 Nov;59(11):3922–3927. doi: 10.1128/aem.59.11.3922-3927.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Donovan W. P., Gonzalez J. M., Jr, Gilbert M. P., Dankocsik C. Isolation and characterization of EG2158, a new strain of Bacillus thuringiensis toxic to coleopteran larvae, and nucleotide sequence of the toxin gene. Mol Gen Genet. 1988 Nov;214(3):365–372. doi: 10.1007/BF00330468. [DOI] [PubMed] [Google Scholar]
  11. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  12. Hue K. K., Cohen S. D., Bechhofer D. H. A polypurine sequence that acts as a 5' mRNA stabilizer in Bacillus subtilis. J Bacteriol. 1995 Jun;177(12):3465–3471. doi: 10.1128/jb.177.12.3465-3471.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Höfte H., Whiteley H. R. Insecticidal crystal proteins of Bacillus thuringiensis. Microbiol Rev. 1989 Jun;53(2):242–255. doi: 10.1128/mr.53.2.242-255.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kawalek M. D., Benjamin S., Lee H. L., Gill S. S. Isolation and Identification of novel toxins from a new mosquitocidal isolate from Malaysia, Bacillus thuringiensis subsp. jegathesan. Appl Environ Microbiol. 1995 Aug;61(8):2965–2969. doi: 10.1128/aem.61.8.2965-2969.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Knowles B. H., Blatt M. R., Tester M., Horsnell J. M., Carroll J., Menestrina G., Ellar D. J. A cytolytic delta-endotoxin from Bacillus thuringiensis var. israelensis forms cation-selective channels in planar lipid bilayers. FEBS Lett. 1989 Feb 27;244(2):259–262. doi: 10.1016/0014-5793(89)80540-x. [DOI] [PubMed] [Google Scholar]
  16. Knowles B. H., White P. J., Nicholls C. N., Ellar D. J. A broad-spectrum cytolytic toxin from Bacillus thuringiensis var. kyushuensis. Proc Biol Sci. 1992 Apr 22;248(1321):1–7. doi: 10.1098/rspb.1992.0035. [DOI] [PubMed] [Google Scholar]
  17. Koni P. A., Ellar D. J. Cloning and characterization of a novel Bacillus thuringiensis cytolytic delta-endotoxin. J Mol Biol. 1993 Jan 20;229(2):319–327. doi: 10.1006/jmbi.1993.1037. [DOI] [PubMed] [Google Scholar]
  18. Park H. W., Kim H. S., Lee D. W., Yu Y. M., Jin B. R., Kang S. K. Expression and synergistic effect of three types of crystal protein genes in Bacillus thuringiensis. Biochem Biophys Res Commun. 1995 Sep 14;214(2):602–607. doi: 10.1006/bbrc.1995.2328. [DOI] [PubMed] [Google Scholar]
  19. Ragni A., Thiéry I., Delécluse A. Characterization of six highly mosquitocidal Bacillus thuringiensis strains that do not belong to H-14 serotype. Curr Microbiol. 1996 Jan;32(1):48–54. doi: 10.1007/s002849900009. [DOI] [PubMed] [Google Scholar]
  20. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Schaeffer P., Millet J., Aubert J. P. Catabolic repression of bacterial sporulation. Proc Natl Acad Sci U S A. 1965 Sep;54(3):704–711. doi: 10.1073/pnas.54.3.704. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sekar V., Thompson D. V., Maroney M. J., Bookland R. G., Adang M. J. Molecular cloning and characterization of the insecticidal crystal protein gene of Bacillus thuringiensis var. tenebrionis. Proc Natl Acad Sci U S A. 1987 Oct;84(20):7036–7040. doi: 10.1073/pnas.84.20.7036. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Thiery I., Delécluse A., Tamayo M. C., Orduz S. Identification of a gene for Cyt1A-like hemolysin from Bacillus thuringiensis subsp. medellin and expression in a crystal-negative B. thuringiensis strain. Appl Environ Microbiol. 1997 Feb;63(2):468–473. doi: 10.1128/aem.63.2.468-473.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Thomas W. E., Ellar D. J. Bacillus thuringiensis var israelensis crystal delta-endotoxin: effects on insect and mammalian cells in vitro and in vivo. J Cell Sci. 1983 Mar;60:181–197. doi: 10.1242/jcs.60.1.181. [DOI] [PubMed] [Google Scholar]
  25. Wong H. C., Schnepf H. E., Whiteley H. R. Transcriptional and translational start sites for the Bacillus thuringiensis crystal protein gene. J Biol Chem. 1983 Feb 10;258(3):1960–1967. [PubMed] [Google Scholar]
  26. Yu Y. M., Ohba M., Gill S. S. Characterization of mosquitocidal activity of Bacillus thuringiensis subsp. fukuokaensis crystal proteins. Appl Environ Microbiol. 1991 Apr;57(4):1075–1081. doi: 10.1128/aem.57.4.1075-1081.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES