Biological Control 40 (2007) 65–68 www.elsevier.com/locate/ybcon 1049-9644/$ - see front matter  2006 Elsevier Inc. All rights reserved. doi:10.1016/j.biocontrol.2006.09.009 Characterization of Bacillus thuringiensis isolates toxic to cotton boll weevil (Anthonomus grandis) Érica Soares Martins a , Lílian Botelho Praça a , Vinícius Fiúza Dumas a , Joseilde O. Silva-Werneck a , Eduardo Hideki Sone a , Isabel C. Waga a , Colin Berry b , Rose Gomes Monnerat a,¤ a Embrapa Recursos Genéticos e Biotecnologia, SAIN-Parque Rural, Caixa postal 02372, 70770-900-Brasilia-DF, Brazil b CardiV School of Biosciences, CardiV University, Museum Avenue, CardiV CF10 3US, Wales, UK Received 6 April 2006; accepted 15 September 2006 Available online 27 September 2006 Abstract The cotton boll weevil (Anthonomus grandis) is the major cotton pest in the Americas. One of the alternatives for its control is the uti- lization of Bacillus thuringiensis (Bt), an entomopathogenic bacterium characterized by its production of insecticidal crystal proteins. Embrapa Genetic Resources and Biotechnology has a collection of Bacilli in which diVerent isolates of Bt are stored. A method for rear- ing and maintenance of Anthonomus grandis on artiWcial diet in the laboratory was developed, and a robust larval bioassay protocol was established for the selection of B. thuringiensis isolates toxic to boll weevil. After preliminary bioassays performed with 215 isolates, 5 were selected that demonstrated a good level of toxicity and these were analyzed in more detail. The most toxic were S601 and S1806 pre- senting LC 50 (lethal concentration to kill 50% if the larvae) of 0.14 mg/ml and 0.30 mg/ml, respectively. S601 showed an LC 50 value that was half that of the standard B. thuringiensis subspecies tenebrionis and S1806 demonstrated similar values of LC 50 to the standard. S601 contained the cry1B gene and S1806 had the cry4A, cry4B, cry10, cry11, cyt1 and cyt2 genes like B. thuringiensis subspecies israelensis.  2006 Elsevier Inc. All rights reserved. Keywords: Bacillus thuringiensis; Anthonomus grandis; Biological control; Toxicity; cry genes 1. Introduction Cotton is one of the most Wnancially proWtable crops worldwide; in the 1990s its price was higher than those of other important products such as soy, corn and wheat (Ponchio, 2001). In the textile industry, the cotton Wber is recognized as one of the most important and valuable in the market. The extensive plantation of monocultures like soy, corn, cotton, canola and other plants, requires a high investment in insecticides owing to their resident pests, especially from the lepidopteran and coleopteran orders. According to Santos et al. (2003), about US$ 3–5 billion are spent on pesticides per year, and of that US$ 645 million are spent in cotton plantations. Cotton boll weevil, Anthonomus grandis Boheman (Cole- optera: Curculionidae), is a signiWcant pest of cotton pro- duction in the Americas (Busoli et al., 1994; Gallo et al., 2002). In order to decrease damage and loss of production, this pest has been controlled primarily by the use of chemi- cal agents. However, while this has produced eYcient cot- ton weevil control (Martin et al., 1987; Wolfenberger et al., 1997), these chemicals are expensive and may cause nega- tive environmental impacts. Other control methods employed are the use of pheromones (Gallo et al., 2002), early budding varieties, gathering and destruction of fallen Xower buds (Silvie et al., 2001), biological control (Pallini et al., 2006) and integrated pest management (Luttrell et al., 1994). In addition, there is a possibility of producing trans- genic cotton containing a gene or genes to make the plant * Corresponding author. Fax: +55 61 3448 4672. E-mail address: rose@cenargen.embrapa.br (R.G. Monnerat).