Feeding Rate and Mortality Impact of Crystals and Endospores Versus Crystals - Producing Recombinants of Bacillus thuringiensis Against Cotton Leafworm


  • Saad A. Alotaibi Taif University




Bacillus thuringiensis, Bacillus subtilis, Mortality percentage, Recombinant bioinsecticides, Spodoptera littoralis, Survival


Two Bacillus strains belonging to two species; thuringiensis and subtilis were used in this study. These strains were genetically marking and to be used in conjugation process depending upon the opposite genetic markers to induce recombinants in both strains. This study aimed to evaluate two bioinsecticides induced from recombinants; crystals, crystals + endospores, which derived from two strains of Bacillus and four of their transconjugants resulted from conjugation between Bacillus thuringiensis and Bacillus subtilis, with respect to their toxicity against lepidopterous cotton pest. The results obtained in this study achieved that endospores were effective in reducing feed consumption at the late times of larval age, in contrast, crystals was more effective in reducing feed consumption at the early times of larval age. The results appeared that endospores play a major role in increasing insecticidal activity of bioinsecticides (crystals), this leading to use their in all bioinsecticides. Endospores were effective in reducing the average weight of surviving larvae more than crystals. This indicated that bioinsecticides containing endospores + crystals were more effective than that containing crystals alone. However, most bioinsecticides containing endospores were reduced the average weight of surviving larvae at 48, 72 and 96 hours from treatment. All bioinsecticides containing crystals + endospores were more effective in reducing survival percentage than that containing crystals alone. This achieved that bioinsecticides containing crystals + endospores were more toxic on S. littoralis larvae. The mean number of mortality larvae appeared in treatments with crystals + endospores was more than that caused by bioinsecticides containing crystals alone. In most treatments done in this study the insecticidal activity of crystals + endospores caused higher mortality larvae than that caused by bioinsecticides containing crystals alone.

Author Biography

Saad A. Alotaibi, Taif University

Biotechnology Department

Faculty of Science


[1] Zakharyan RA, Israelyaynu A, Agabalyan AS, Tatevosyapn E, Akopyans M, Afrikyane K. Plasmid DNA from Bacillus thuringiensis. Microbiol 1979; 48: 226-9.
[2] Thomas CC. Pathogens of invertebrates: Application in biological control and transmission mechanisms, Society for Invertebrate Pathology Meeting Berlin, Germany 1984; Vol. 7: p. 159.
[3] Madigan M, Martinko J. Brock Biology of Microorganisms (11th. ed.), Prentice Hall 2005.
[4] Dean DH. Biochemical Genetics of the Bacterial Insect-Control Agent Bacillus thuringiensis: Basic Principles and Prospects Engineering. Biotechnol Gen Eng Rev1984; 2: 341-63.
[5] Clayton CB, Takhashi Y. Invitation paper (c. p. alexander fund): History of Bacillus thuringiensis berliner research and development. Can Entomol1992; 124: 587-16. http://dx.doi.org/10.4039/Ent124587-4
[6] Xu J, Liu Q, Yin XD, Zhu SD. A review of recent development of Bacillus thuringiensis ICP genetically engineered microbes. Entomol J East China 2006, 15: 53-8
[7] Liu SQ, Shan SP, Xia LQ. Advances on High Performance Insecticide of Bacillus thuringiensis
[J]. Microbiol 2008; 35: 1091-5.
[8] Babu MM, Geetha M. DNA shuffling of Cry proteins. http://www.mrc-lmb.cam.ac.uk/genomes/madanm/articles/ dnashuff.htm
[9] Doi RH. Genetic Engineering in Bacillus subtilis. Biotechnol Gen Eng Rev 1984; 2: 121-55.
[10] Jayaraman KS, Jeffrey LF, Hepeng J, Claudia O. Indian Btgene monoculture: Potential time bomb. Nat Biotechnol 2005; 23: 158. http://dx.doi.org/10.1038/nbt0205-158
[11] Kumar PA, Sharma RP, Malik VS. Insecticidal proteins of Bacillus thuringiensis. Adv Appl Microbiol 1996; 42: 1-43. http://dx.doi.org/10.1016/S0065-2164(08)70371-X
[12] Kumar PA. Insect pest-resistant transgenic crops. In: Advances in Microbial Control of Insect Pests, Upadhyay, R. K. Ed. Kluwer Academic, New York 2003; pp. 71-82.
[13] James C. Global status of commercialized Biotech/GM crops. ISAAA Brief 2006, No. 35, International Service For The Acquisition of Agri-Biotech Applications. (ISAAA), Ithaca, NY, USA http://croplife.intraspin.com/Biotech/global-status-of-commercialized-biotech-gm-crops-2006-isaaa-briefs-no-35/
[14] Bosch D, Schipper B, van der Kleij H, de Maagd RA, Stiekema WJ. Recombinant Bacillus thuringiensis crystal proteins with new properties: possibilities for resistance management. Bio/Technology 1994; 12: 915-8. http://dx.doi.org/10.1038/nbt0994-915
[15] Deml R, Meise T, Dettner K. Effects of Bacillus thuringiensis-endotoxins on food utilization, growth, and survival of selected phytophagous insects. J Appl Entomol 1999, 123: 55-64. http://dx.doi.org/10.1046/j.1439-0418.1999.00312.x
[16] Puntamabeker US, Ranjekar PK. Intergeneric protoplast fusion between Agrobacterium tumefaciens and Bacillus thuringiensis subsp. Kurstaki. Biotechnol Lett 1989; 10: 717-22. http://dx.doi.org/10.1007/BF01044104
[17] Collins CH, Lyne PM. Microbiological Methods. 5th ed. Butterworths, London 1985; pp. 167-81. http://helid.digicollection.org/en/d/Jwho01e/6.html
[18] Toda M, Okubo S, Hiyoshi R, Shimamura T. The bactericidal activity of tea and coffee. Lett Appl Microbiol 1989; 8: 123-5. http://dx.doi.org/10.1111/j.1472-765X.1989.tb00255.x
[19] Sonti RV, Roth JR. Role of gene duplications in the adaptation of Salmonella typhimurium to growth on limiting carbon sources. Genet 1989; 123: 19-28. http://www. genetics.org/content/123/1/19.full.pdf
[20] Ames BN, Lee FD, Durston WE. An improved bacterial test system for the detection and classification of mutagens and carcinogens. Proc Nat Acad Sci USA 1973; 70: 782-6. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC433358/ http://dx.doi.org/10.1073/pnas.70.3.782
[21] Lessl M, Balzer D, Weyrauch K, Lanka E. The mating pair formation system of plasmid RP4 defined by RSF 1010 mobilization and donor-specific phage propagation. J Bacteriol 1993; 175: 6415-25. http://www.ncbi.nlm.nih.gov/ pmc/articles/PMC206749/
[22] Karamanlidou G, Lambropoulos AF, Koliais SI, Manousis T, Ellar D, Kastritsis C. Toxicity of Bacillus thuringiensis to laboratory populations of the olive fruit fly (Dacus oleae). Appl Environ Microbiol 1991; 57: 2277-82. http://aem.asm. org/content/57/8/2277
[23] Klanfon AR, De Barjac H. Screening of the insecticidal activity of Bacillus thuringiensis strains against the Egyptian cotton leafworm Spodoptera littoralis. Entomophaga 1985; 30: 177-86. http://link.springer.com/article/10.1007% 2FBF02372251 http://dx.doi.org/10.1007/BF02372251
[24] Ignoffo CM, Hostetter DL, Pinnell RE, Garcia C. Relative susceptibility of six soybean caterpillars to a standard preparation of Bacillus thuringiensis var. Kurstaki. J Econ Entomol 1977; 70: 60-3. http://www.ingentaconnect.com/ content/esa/jee/1977/00000070/00000001/art00018
[25] Inagaki S, Miyasono M, Ishiguro T, Takeda R, Hayashi Y. Proteolytic processing of -endotoxin of Bacillus thuringiensisvar. Kurstaki HD-1 in insensitive insect, Spodoptera litura: Unusual proteolysis in the presence of sodium dodecyl sulfate. J Invert Pathol 1992; 60: 64-8. http://dx.doi.org/10.1016/0022-2011(92)90155-W
[26] Schwinghamer EA, Dudman WF. Evaluation of spectinomycin resistance as a marker for ecological studies with Rhizobium spp. J Appl Bacteriol 1973; 36: 263-72. http://www.ncbi.nlm.nih.gov/pubmed/4270618 http://dx.doi.org/10.1111/j.1365-2672.1973.tb04101.x
[27] Stuart JG, Zimmerer EJ, Maddux RL. Conjugation of antibiotic resistance in Streptococcus suis. Vet Microbiol 1992; 30: 213-22. http://www.ncbi.nlm.nih.gov/pubmed/ 1313622 http://dx.doi.org/10.1016/0378-1135(92)90115-A
[28] Campbell NA, Reece JB, Mitchell. Biology 6th Ed. Benjamin Cummings, Publ. San Francisco 2002. http://freedownloadb. com/pdf/bbiology-b-jdenuno-5937588.html
[29] Pr├╝tz G, Dettner K. Effect of Bt corn leaf suspension on food consumption by Chilo partellus and life history parameters of its parasitoid Cotesia flavipes under laboratory conditions, Entomologia Experimentalis et Applicata 2004; 111: 179-87. http://dx.doi.org/10.1111/j.0013-8703.2004.00166.x
[30] Somashekara R, Udikeri SS, Patil SB, Basavanagoud K. Food consumption indices for spotted bollworm Earias vittella (Fab.) on transgenic cottons expressing one or two Bt genes. Karnat J Agr Sci 2011; 24: 140-2. http://www.inflibnet.ac.in/ ojs/index.php/KJAS/article/view/777
[31] Wang SG, GongYYE, Cui HU, Qing YSHU, Yin W XIA, Illimar A. Effects of Bt rice on the food consumption, growth and survival of chilo suppressaus larvae under different temperatures. Insec Sci 2001; 8: 218-26. http://dx.doi.org/10.1111/j.1744-7917.2001.tb00445.x
[32] Bienvenu T, Daniel CC. Feeding rate and survival of Plutella xylostella (L.) larvae (Lepidopteria: Plutellidae) after intoxication by Bacillus thuringiensis berliner var. kurstaki and var. Aizawai. Arch Phytopath Plant Protec 1997; 31: 201-6. http://dx.doi.org/10.1080/03235409709383228
[33] Felke M, Langenbruch GA, Feiertag S, Kassa A. Effect of Bt-176 maize pollen on first instar larvae of the Peacock butterfly (Inachis io) (Lepidoptera; Nymphalidae). Environ Biosaf Res 2010; 9: 5-12. http://dx.doi.org/10.1051/ebr/2010006
[34] Latham J. The effects of pollen from the GM maize Bt176 on the European common swallowtail butterfly 2006. http://www.lobbywatch.org/archive2.asp?arcid = 6826
[35] Li SY, Fitzpatrick SM. Responses of larval Choristoneura rosaceana (Harris) (Lepidoptera: Tortricidae) to a feeding stimulant. Can Entomol 1997;129: 363-9. http://dx.doi.org/10.4039/Ent129363-2
[36] Navon A, Wysoki M, Keren S. Potency and effect of Bacillus thuringiensis preparations against larvae of Spodoptera littoralis and Boarmia (Ascotis) selenaria. Phytoparasitica 1983; 11: 3-11. http://dx.doi.org/10.1007/BF02980706
[37] Hibbard BE, Frank DL, Kurtz R, Boudreau E, Ellersieck MR, Odhiambo JF. Mortality impact of Bt transgenic maize roots expressing eCry3.1Ab, mCry3A, and eCry3.1Ab plus mCry3A on western corn rootworm larvae in the field. J Econ Entomol 2011; 104: 1584-91. http://dx.doi.org/10.1603/EC11186
[38] Guo JY, Dong H, Wan Fang H. Influence of Bt Transgenic Cotton on Larval Survival of Common Cutworm Spodoptera litura. Chin J Biol Cont 2003; 19: 145-8. http://en.cnki.com. cn/Article_en/CJFDTotal-ZSWF200304001.htm
[39] Barwale RB, VR Gadwal, Zehr U, Zehr B. Prospects for Btcotton technology in India. AgrBioForum 2004; 7: 23-6. http://www.agbioforum.org/v7n12/v7n12a04-zehr.htm