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. 1986 Oct;52(4):650–653. doi: 10.1128/aem.52.4.650-653.1986

Analysis of mosquito larvicidal potential exhibited by vegetative cells of Bacillus thuringiensis subsp. israelensis.

C J Walther, G A Couche, M A Pfannenstiel, S E Egan, L A Bivin, K W Nickerson
PMCID: PMC239091  PMID: 3777922

Abstract

Vegetative Bacillus thuringiensis subsp. israelensis cells (6 X 10(5)/ml) achieved 100% mortality of Aedes aegypti larvae within 24 h. This larvicidal potential was localized within the cells; the cell-free supernatants did not kill mosquito larvae. However, they did contain a heat-labile hemolysin which was immunologically distinct from the general cytolytic (hemolytic) factor released during solubilization of B. thuringiensis subsp. israelensis crystals. The larvicidal potential of the vegetative cells was not due to poly-beta-hydroxybutyrate. Instead, it correlated with the ability of vegetative cells to sporulate during the bioassays. No toxicity was observed when bioassays were conducted in the presence of chloramphenicol or streptomycin. It is unlikely that the vegetative cells sporulate in the alkaline (pH 9.5 to 10.5) larval guts after ingestion. B. thuringiensis subsp. israelensis is not an alkalophile; we have been unable to grow it in culture at pH values of greater than or equal to 9.5. Moreover, we have been unable to demonstrate formation of a protective capsule. However, bacteria may replicate in the gut fluids of dead or dying mosquito larvae because their alkaline gut pH values drop markedly after exposure to the B. thuringiensis subsp. israelensis crystal toxins.

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  1. Aly C. Germination of Bacillus thuringiensis var. israelensis spores in the gut of Aedes larvae (Diptera: Culicidae). J Invertebr Pathol. 1985 Jan;45(1):1–8. doi: 10.1016/0022-2011(85)90041-2. [DOI] [PubMed] [Google Scholar]
  2. Armstrong J. L., Rohrmann G. F., Beaudreau G. S. Delta endotoxin of Bacillus thuringiensis subsp. israelensis. J Bacteriol. 1985 Jan;161(1):39–46. doi: 10.1128/jb.161.1.39-46.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Battisti L., Green B. D., Thorne C. B. Mating system for transfer of plasmids among Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis. J Bacteriol. 1985 May;162(2):543–550. doi: 10.1128/jb.162.2.543-550.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bechtel D. B., Bulla L. A., Jr Electron microscope study of sporulation and parasporal crystal formation in Bacillus thuringiensis. J Bacteriol. 1976 Sep;127(3):1472–1481. doi: 10.1128/jb.127.3.1472-1481.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bernheimer A. W., Grushoff P. Cereolysin: production, purification and partial characterization. J Gen Microbiol. 1967 Jan;46(1):143–150. doi: 10.1099/00221287-46-1-143. [DOI] [PubMed] [Google Scholar]
  6. Dadd R. H. Alkalinity within the midgut of mosquito larvae with alkaline-active digestive enzymes. J Insect Physiol. 1975 Nov;21(11):1847–1853. doi: 10.1016/0022-1910(75)90252-8. [DOI] [PubMed] [Google Scholar]
  7. Green B. D., Battisti L., Koehler T. M., Thorne C. B., Ivins B. E. Demonstration of a capsule plasmid in Bacillus anthracis. Infect Immun. 1985 Aug;49(2):291–297. doi: 10.1128/iai.49.2.291-297.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. HARDWICK W. A., FOSTER J. W. On the nature of sporogenesis in some aerobic bacteria. J Gen Physiol. 1952 Jul;35(6):907–927. doi: 10.1085/jgp.35.6.907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hurley J. M., Lee S. G., Andrews R. E., Jr, Klowden M. J., Bulla L. A., Jr Separation of the cytolytic and mosquitocidal proteins of Bacillus thuringiensis subsp. israelensis. Biochem Biophys Res Commun. 1985 Jan 31;126(2):961–965. doi: 10.1016/0006-291x(85)90279-7. [DOI] [PubMed] [Google Scholar]
  10. Ikezawa H., Yamanegi M., Taguchi R., Miyashita T., Ohyabu T. Studies on phosphatidylinositol phosphodiesterase (phospholipase C type) of Bacillus cereus. I. purification, properties and phosphatase-releasing activity. Biochim Biophys Acta. 1976 Nov 19;450(2):154–164. [PubMed] [Google Scholar]
  11. 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]
  12. Lee S. G., Eckblad W., Bulla L. A., Jr Diversity of protein inclusion bodies and identification of mosquitocidal protein in Bacillus thuringiensis subsp. israelensis. Biochem Biophys Res Commun. 1985 Jan 31;126(2):953–960. doi: 10.1016/0006-291x(85)90278-5. [DOI] [PubMed] [Google Scholar]
  13. Myers P., Yousten A. A., Davidson E. W. Comparative studies of the mosquito-larval toxin of Bacillus sphaericus SSII-1 and 1593. Can J Microbiol. 1979 Nov;25(11):1227–1231. doi: 10.1139/m79-193. [DOI] [PubMed] [Google Scholar]
  14. Nickerson K. W. Purification of Poly-beta-Hydroxybutyrate by Density Gradient Centrifugation in Sodium Bromide. Appl Environ Microbiol. 1982 May;43(5):1208–1209. doi: 10.1128/aem.43.5.1208-1209.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Nickerson K. W., Schnell D. J. Toxicity of cyclic peptide antibiotics to larvae of Aedes aegypti. J Invertebr Pathol. 1983 Nov;42(3):407–409. doi: 10.1016/0022-2011(83)90185-4. [DOI] [PubMed] [Google Scholar]
  16. Pendleton I. R., Bernheimer A. W., Grushoff P. Purification and partial characterization of hemolysins from Bacillus thuringiensis. J Invertebr Pathol. 1973 Mar;21(2):131–135. doi: 10.1016/0022-2011(73)90192-4. [DOI] [PubMed] [Google Scholar]
  17. Pfannenstiel M. A., Couche G. A., Muthukumar G., Nickerson K. W. Stability of the larvicidal activity of Bacillus thuringiensis subsp. israelensis: amino acid modification and denaturants. Appl Environ Microbiol. 1985 Nov;50(5):1196–1199. doi: 10.1128/aem.50.5.1196-1199.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. THORNE C. B., GOMEZ C. G., HOUSEWRIGHT R. D. Synthesis of glutamic acid and glutamyl polypeptide by Bacillus anthracis. II. The effect of carbon dioxide on peptide production on solid media. J Bacteriol. 1952 Mar;63(3):363–368. doi: 10.1128/jb.63.3.363-368.1952. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Taguchi R., Asahi Y., Ikezawa H. Purification and properties of phosphatidylinositol-specific phospholipase C of Bacillus thuringiensis. Biochim Biophys Acta. 1980 Jul 14;619(1):48–57. [PubMed] [Google Scholar]
  20. Temeyer K. B. Larvicidal activity of Bacillus thuringiensis subsp. israelensis in the dipteran Haematobia irritans. Appl Environ Microbiol. 1984 May;47(5):952–955. doi: 10.1128/aem.47.5.952-955.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Thomas W. E., Ellar D. J. Mechanism of action of Bacillus thuringiensis var israelensis insecticidal delta-endotoxin. FEBS Lett. 1983 Apr 18;154(2):362–368. doi: 10.1016/0014-5793(83)80183-5. [DOI] [PubMed] [Google Scholar]
  23. Tyrell D. J., Davidson L. I., Bulla L. A., Jr, Ramoska W. A. Toxicity of parasporal crystals of Bacillus thuringiensis subsp. israelensis to mosquitoes. Appl Environ Microbiol. 1979 Oct;38(4):656–658. doi: 10.1128/aem.38.4.656-658.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Waalwijk C., Dullemans A. M., van Workum M. E., Visser B. Molecular cloning and the nucleotide sequence of the Mr 28 000 crystal protein gene of Bacillus thuringiensis subsp. israelensis. Nucleic Acids Res. 1985 Nov 25;13(22):8207–8217. doi: 10.1093/nar/13.22.8207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. de Barjac H. Une nouvelle variété de Bacillus thuringiensis très toxique pour les moustiques: B. thuringiensis var. israelensis sérotype 14. C R Acad Sci Hebd Seances Acad Sci D. 1978 Mar 13;286(10):797–800. [PubMed] [Google Scholar]

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