Identification of <i>Bacillus thuringiensis</i> Isolates Highly Toxic to <i>Plutella xylostella</i>

Abstract

In this research , we employed 31 Bacillus thuringiensis isolates from the soils collected in the Liang Shui Natural Reserve of Heilongjiang province to identifyhighly toxic to Plutella xylostella , an important Lepidopteran pest in agriculture . The results showed that ten of thirty one Bacillus thuringiensis isolates have high toxicity to Plutella xylostella via the bioassay. Microscopic observation for ten seleted isolates was carried out; the results confirmed that these isolates can generate bi-diamond crystal protein which might be the crystalline toxin. The further genotypic analysis by PCR and SDS-PAGE indicated that the genotypes of the selected 10 Bt isolates were consistent with the genotype of cry 1 class toxin genes which can generate crystal protein 130 kD in molecular weight. Whilst, the growth curves of the ten isolates exhibited that the bacteria would be in the logarithmic growth phase after culturing for 2 hours to 14 hours, in the stationary phase after culturing 16 hours or 30 hours and in the decline phase after culturing 32 hours. Among them, strains S2685-1 and S2737-3 were further accredited to generate the bi-diamond crystal protein of about 130 kD in molecular weight after being cultured for 22 hours and 20 hours, respectively; toxin expressed by the former stabilizes after 26 hour in culture, whereas toxin expressing levels of the later gradually increases and stabilizes after 22 hours. The trials of median lethal concentration (LC50) and median lethal time (LT50) of two dominant isolates, S2685-1 and S2737-3, for Diamondback moth ( Plutella xylostella ) were carried out in this research, and the results showed that the LC50 and LT50 for S2685-1 to diamondback moth were 17.113 ng/mL and 37.343 hours, respectively; while for S2737-3 were 25.782 ng/mL and 36.825 hours, respectively. In contrast, LC50 and LT50 of the reference strain HD-73 to the diamondback moth were 15.414 ng/mL and 35.917 hours. It is obvious that S2685-1 and S2737-3 exhibit strong insecticidal activities in the median lethal concentration and median lethal time against Plutella xylostella . In conclusion, the results of this study that the Bt isolates, S2685-1 and S2737-3, have more prominent insecticidal activities, which might be applied as potential alternative strains in Bt bio-pesticide products as initiate strain instead of standard strain HD-73 strain.