Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Jun;178(11):3099–3105. doi: 10.1128/jb.178.11.3099-3105.1996

Cloning and expression of the first anaerobic toxin gene from Clostridium bifermentans subsp. malaysia, encoding a new mosquitocidal protein with homologies to Bacillus thuringiensis delta-endotoxins.

F Barloy 1, A Delécluse 1, L Nicolas 1, M M Lecadet 1
PMCID: PMC178058  PMID: 8655486

Abstract

A gene (cbm71) encoding a 71,128-Da mosquitocidal protein (Cbm71) was obtained by screening a size-fractionated XbaI digest of total genomic DNA from Clostridium bifermentans subsp. malaysia CH18 with two gene-specific oligonucleotide probes. The sequence of the Cbm71 protein, as deduced from the sequence of cbm71, corresponds to that of the 66-kDa protein previously described as one of the mosquitocidal components of C. bifermentans subsp. malaysia. Cbm71 shows limited similarities with Bacillus thuringiensis delta-endotoxins, especially in the four first conserved blocks. However, Cbm71 was not immunologically related to any of the Cry toxins and thus belongs to a novel class of mosquitocidal protein. The cbm71 gene was expressed in a nontoxic strain of B. thuringiensis, and Cbm71 was produced during sporulation and secreted to the supernatant of culture. Trichloroacetic-precipitated supernatant preparations were toxic for mosquito larvae of the species Aedes aegypti, Culex pipiens, and Anopheles stephensi.

Full Text

The Full Text of this article is available as a PDF (495.2 KB).

Selected References

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

  1. Aquino de Muro M., Mitchell W. J., Priest F. G. Differentiation of mosquito-pathogenic strains of Bacillus sphaericus from non-toxic varieties by ribosomal RNA gene restriction patterns. J Gen Microbiol. 1992 Jun;138(6):1159–1166. doi: 10.1099/00221287-138-6-1159. [DOI] [PubMed] [Google Scholar]
  2. Arantes O., Lereclus D. Construction of cloning vectors for Bacillus thuringiensis. Gene. 1991 Dec 1;108(1):115–119. doi: 10.1016/0378-1119(91)90495-w. [DOI] [PubMed] [Google Scholar]
  3. Aronson A. I., Beckman W., Dunn P. Bacillus thuringiensis and related insect pathogens. Microbiol Rev. 1986 Mar;50(1):1–24. doi: 10.1128/mr.50.1.1-24.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Arthur M., Molinas C., Depardieu F., Courvalin P. Characterization of Tn1546, a Tn3-related transposon conferring glycopeptide resistance by synthesis of depsipeptide peptidoglycan precursors in Enterococcus faecium BM4147. J Bacteriol. 1993 Jan;175(1):117–127. doi: 10.1128/jb.175.1.117-127.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Charles J. F., Nicolas L., Sebald M., de Barjac H. Clostridium bifermentans serovar malaysia: sporulation, biogenesis of inclusion bodies and larvicidal effect on mosquito. Res Microbiol. 1990 Jul-Aug;141(6):721–733. doi: 10.1016/0923-2508(90)90066-y. [DOI] [PubMed] [Google Scholar]
  6. Delécluse A., Charles J. F., Klier A., Rapoport G. Deletion by in vivo recombination shows that the 28-kilodalton cytolytic polypeptide from Bacillus thuringiensis subsp. israelensis is not essential for mosquitocidal activity. J Bacteriol. 1991 Jun;173(11):3374–3381. doi: 10.1128/jb.173.11.3374-3381.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. 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]
  8. Delécluse A., Rosso M. L., Ragni A. Cloning and expression of a novel toxin gene from Bacillus thuringiensis subsp. jegathesan encoding a highly mosquitocidal protein. Appl Environ Microbiol. 1995 Dec;61(12):4230–4235. doi: 10.1128/aem.61.12.4230-4235.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Felmlee T., Pellett S., Lee E. Y., Welch R. A. Escherichia coli hemolysin is released extracellularly without cleavage of a signal peptide. J Bacteriol. 1985 Jul;163(1):88–93. doi: 10.1128/jb.163.1.88-93.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gray L., Mackman N., Nicaud J. M., Holland I. B. The carboxy-terminal region of haemolysin 2001 is required for secretion of the toxin from Escherichia coli. Mol Gen Genet. 1986 Oct;205(1):127–133. doi: 10.1007/BF02428042. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  13. Lederberg E. M., Cohen S. N. Transformation of Salmonella typhimurium by plasmid deoxyribonucleic acid. J Bacteriol. 1974 Sep;119(3):1072–1074. doi: 10.1128/jb.119.3.1072-1074.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lereclus D., Arantès O., Chaufaux J., Lecadet M. Transformation and expression of a cloned delta-endotoxin gene in Bacillus thuringiensis. FEMS Microbiol Lett. 1989 Jul 15;51(1):211–217. doi: 10.1016/0378-1097(89)90511-9. [DOI] [PubMed] [Google Scholar]
  15. Nicolas L., Charles J. F., de Barjac H. Clostridium bifermentans serovar malaysia: characterization of putative mosquito larvicidal proteins. FEMS Microbiol Lett. 1993 Oct 1;113(1):23–28. doi: 10.1111/j.1574-6968.1993.tb06482.x. [DOI] [PubMed] [Google Scholar]
  16. Porter A. G., Davidson E. W., Liu J. W. Mosquitocidal toxins of bacilli and their genetic manipulation for effective biological control of mosquitoes. Microbiol Rev. 1993 Dec;57(4):838–861. doi: 10.1128/mr.57.4.838-861.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rao D. R., Mani T. R., Rajendran R., Joseph A. S., Gajanana A., Reuben R. Development of a high level of resistance to Bacillus sphaericus in a field population of Culex quinquefasciatus from Kochi, India. J Am Mosq Control Assoc. 1995 Mar;11(1):1–5. [PubMed] [Google Scholar]
  18. Simonen M., Palva I. Protein secretion in Bacillus species. Microbiol Rev. 1993 Mar;57(1):109–137. doi: 10.1128/mr.57.1.109-137.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Thiery I., Hamon S., Dumanoir V. C., de Barjac H. Vertebrate safety of Clostridium bifermentans serovar malaysia, a new larvicidal agent for vector control. J Econ Entomol. 1992 Oct;85(5):1618–1623. doi: 10.1093/jee/85.5.1618. [DOI] [PubMed] [Google Scholar]
  20. Thiery I., Hamon S., Gaven B., De Barjac H. Host range of Clostridium bifermentans serovar. malaysia, a mosquitocidal anaerobic bacterium. J Am Mosq Control Assoc. 1992 Sep;8(3):272–277. [PubMed] [Google Scholar]
  21. Tinoco I., Jr, Borer P. N., Dengler B., Levin M. D., Uhlenbeck O. C., Crothers D. M., Bralla J. Improved estimation of secondary structure in ribonucleic acids. Nat New Biol. 1973 Nov 14;246(150):40–41. doi: 10.1038/newbio246040a0. [DOI] [PubMed] [Google Scholar]
  22. Yiallouros M., Storch V., Thiery I., Becker N. Efficacy of Clostridium bifermentans serovar Malaysia on target and nontarget organisms. J Am Mosq Control Assoc. 1994 Mar;10(1):51–55. [PubMed] [Google Scholar]
  23. Young M., Minton N. P., Staudenbauer W. L. Recent advances in the genetics of the clostridia. FEMS Microbiol Rev. 1989 Dec;5(4):301–325. doi: 10.1111/j.1574-6968.1989.tb03402.x. [DOI] [PubMed] [Google Scholar]
  24. de Barjac H., Sebald M., Charles J. F., Cheong W. H., Lee H. L. Clostridium bifermentans serovar malaysia, une nouvelle bactérie anaérobie pathogène des larves de moustiques et de simulies. C R Acad Sci III. 1990;310(9):383–387. [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES