A rapid and reliable method for the evaluation of Bacillus thuringiensis native strains against Tecia solanivora (Povolny) (Lepidoptera : Gelechiidae)

Abstract

Tecia solanivora (Povolny) (Lepidoptera: Gelechiidae)- Potato tuber moth (Polilla guatemalteca), is a new pest in Colombia, that causes big losses to the crop. Because commercial insecticides have not provided consistent control, biological control using entomopathogenic microorganisms appear as a promissory alternative. An efficient method for the evaluation of native strains of Bacillus thuringiensis as biological control of T. solanivora was developed. An artificial diet consisting of mashed potatoes. agar, antioxidants, and preservatives was standardized. This diet was time stable, was consumed by the larvae, and did not affect the bacteria. Bioassays were performed using the artificial diet under laboratory conditions of 20°C temperature, 75% relative humidity, and 12 hours photoperiod. Reference and natives strains used were selected according to molecular characterization. These strains presented cry genes IA(b), IB, and IC which codified for d-endotoxines with insecticidal activity against Phtorimaea operculella, a related lepidopteran potato pest. In order to select a reference strain to be used in bioassays, reference strains HD-1 and HD-137 were used at two concentration levels measured as total protein (150 and 200 µg/ml) of a bacterial suspension, obtained from a culture grown in L agar medium, during 15 days and 28°C, and presenting a big quantity of spores and crystals which was determined by microscopic analysis. The diet was inoculated with the mentioned concentrations and was infested with a first instar larva of T. solanivora. Larvae mortality was registered every 24 hours for 120 hours. The highest mortality percentage was obtained with reference strain HD-1, which showed 92% larvae mortality at 96 hours. Native strains Bt.-40, Bt- I 19, Bt-285, Bt-536 and Bt-538, and reference strain HD-1 as positive control were evaluated for insecticidal activity as a unique concentration of 100 lig of total protein/ml. Among all native strains. Bt-40 was selected because it demonstrated the highest insecticide activity with 72% larvae mortality at 120 hours. Other bioassays using bacterial suspensions corresponding to 50, 75, 100, 125, 150 i_tg of total protein/m1 were performed, under the mentioned conditions in order to determine lethal concentrations. Statistical analysis PROBIT, showed a lethal concentration 50 (LC₀ ) for reference strain of 56.9 pg/mland 95.45 µg/ml for Bt-40 native strain. This work allowed the standardization of a rapid, easy, and reliable method for the evaluation of the insecticidal activity of Bacillus thuringiensis against Tecia solanivora.