Bacillus thuringiensis strains isolated from Latin American soil samples that showed toxicity against three Spodoptera frugiperda populations from different geographical areas (Mexico, Colombia, and Brazil) were characterized on the basis of their insecticidal activity, crystal morphology, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of parasporal crystals, plasmid profiles, and cry gene content. We found that the different S. frugiperda populations display different susceptibilities to the selected B. thuringiensis strains and also to pure preparations of Cry1B, Cry1C, and Cry1D toxins. Binding assays performed with pure toxin demonstrated that the differences in the toxin binding capacities of these insect populations correlated with the observed differences in susceptibility to the three Cry toxins analyzed. Finally, the genetic variability of the three insect populations was analyzed by random amplification of polymorphic DNA-PCR, which showed significant genetic diversity among the three S. frugiperda populations analyzed. The data presented here show that the genetic variability of S. frugiperda populations should be carefully considered in the development of insect pest control strategies, including the deployment of genetically modified maize in different geographical regions.Fall armyworm, Spodoptera frugiperda (J. E. Smith), is a worldwide pest of economic importance for different crops. This species has a highly polyphagous feeding behavior, which includes the consumption of different cultivated plants, such as maize (Zea mays L.), cotton (Gossypium hirsutum L.), and rice (Oryza sativa L.). To date, the most common method for controlling this pest relies on the use of synthetic insecticides such as methomyl, carbaryl, and cypermethrin (10), in spite of the damage they cause to the environment and to nontarget organisms. Two distinct strains of S. frugiperda, one associated with maize and the other with rice, have been already identified in the United States (17, 19). The detection and characterization of genetic diversity among insect populations is a critical issue for the improvement of pest management strategies, since the evolution of resistance to insecticides among insect populations is dependent on the frequency of resistant alleles, the inheritance of resistance, the relative fitness cost, and the gene flow.Bioinsecticides are viable alternatives for insect control in agriculture, and among them, Bacillus thuringiensis is the most widely used. B. thuringiensis is compatible with sustainable and environmentally friendly agricultural practices. This bacterium produces insecticidal proteins (Cry protoxins) during sporulation as parasporal crystals, which are highly specific to their target insects; safe for humans, other vertebrates, and plants; and biodegradable (12). Moreover, recombinant DNA technology using cry genes has developed insect-resistant transgenic plants that are used extensively for cotton, corn, and rice production, among others (23).Information regarding the suscept...
The in vitro activity of a new quinolone, T-3262 [A-60969; DL-7-(3-amino-1-pyrrolidinyl)-1-(2,4-difluorophenyl)-6-fluoro-l-,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic acid monohydrate], was compared with those of ciprofloxacin, ofloxacin, ceftazidime, imipenem, and gentamicin. T-3262 inhibited 90% of isolates of the family Enterobacteriaceae at a concentration of s0.25 ,ug/ml. It was two to four times more active than ofloxacin and similarly or slightly less active than ciprofloxacin. Ninety percent of isolates of Pseudomonas aeruginosa were inhibited at 0.5 ,Lg/ml. It was 4-to 8-fold more active than ciprofloxacin and 8-to 16- There has been continued interest in the synthesis of new quinolones. Although many of the new agents of the quinolone class have excellent in vitro activity against members of the family Enterobacteriaceae and Pseudomonas aeruginosa, activity against some gram-positive species and against many anaerobic species is variable (3,6,8). Compound T-3262, also referred to as A-60969, is a new trifluoronaphthyridine quinolone. The chemical structure is DL-7-(3-amino-1-pyrrolidinyl)-1-(2,4-difluorophenyl)-6-fluoro-1-,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic acid monohydrate.We have compared the activity of this new agent with those of other quinolones, ceftazidime, imipenem, and gentamicin and have determined the effect of various assay conditions on its activity. MATERIALS AND METHODSThe compound designated T-3262 was a gift from Toyama Chemical Co., Ltd., Tokyo, Japan, and that designated A-60969 was provided by Abbott Laboratories, Abbott Park, Ill. All other agents were obtained from their respective manufacturers. Quinolones were prepared as published previously (1, 6
Bacillus thuringiensis subsp. israelensis is the most widely used microbial control agent against mosquitoes and blackflies. Its insecticidal success is based on an arsenal of toxins, such as Cry4A, Cry4B, Cry11A, and Cyt1A, harbored in the parasporal crystal of the bacterium. A fifth toxin, Cry10Aa, is synthesized at very low levels; previous attempts to clone and express Cry10Aa were limited, and no parasporal body was formed. By using a new strategy, the whole Cry10A operon was cloned in the pSTAB vector, where both open reading frames ORF1 and ORF2 (and the gap between the two) were located, under the control of the cyt1A operon and the STAB-SD stabilizer sequence characteristic of this vector. Once the acrystalliferous mutant 4Q7 of B. thuringiensis subsp. israelensis was transformed with this construct, parasporal bodies were observed by phase-contrast microscopy and transmission electron microscopy. Discrete, ca. 0.9-m amorphous parasporal bodies were observed in the mature sporangia, which were readily purified by gradient centrifugation once autolysis had occurred. Pure parasporal bodies showed two major bands of ca. 68 and 56 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. These bands were further characterized by Nterminal sequencing of tryptic fragments using matrix-assisted laser desorption ionization-time of flight mass spectrometry analysis, which identified both bands as the products of ORF1 and ORF2, respectively. Bioassays against fourth-instar larvae of Aedes aegypti of spore-crystal complex and pure crystals of Cry10Aa gave estimated 50% lethal concentrations of 2,061 ng/ml and 239 ng/ml, respectively. Additionally, synergism was clearly detected between Cry10A and Cyt1A, as the synergistic levels (potentiation rates) were estimated at 13.3 for the mixture of Cyt1A crystals and Cry10Aa spore-crystal complex and 12.6 for the combination of Cyt1A and Cry10Aa pure crystals.
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