Diversity of indigenous Bacillus thuringiensis isolates toxic to the diamondback moth, Plutella xylostella (L.) (Plutellidae: Lepidoptera)

Abstract

Background Toxins from the Bacillus thuringiensis (Bt) bacterium are employed as an alternative to synthetic pesticides in pest management. The greatest threat to the long-term viability of Bt toxins is resistance evolution in the target pests. Genetic diversity and toxicity of Bt isolates were studied in this work in order to find Bt isolates with novel cry genes. Results In terms of colony morphology, among a total of 60 isolates, 51 isolates had off-white colour colonies with typical fried egg appearance, irregular shape, flat and undulate margin. Different crystal shapes, viz. spherical (88.13%), bipyramidal (49.15%), cuboidal (42.37%), rectangular, and crystals attached to spores (3.38%) were observed among Bt isolates. SDS-PAGE analysis of spore crystal mixture showed the presence of proteins with various molecular weights ranging from 124 to 26 kDa. PCR screening with cry1, cry2, cry9 and vip3A1 primers showed isolates with varied insecticidal gene combinations. Bt isolates containing cry1 genes were found to be abundant (30), followed by cry2 (9) and vip3A1 (9). Cry9 was absent in all the 60 isolates tested. Insecticidal activity of spore crystal mixtures ranged from 0 to 100% mortality. Furthermore, 12 isolates were found to be highly toxic against the larvae of diamondback moth, Plutella xylostella (L.) (Plutellidae: Lepidoptera) with 100% mortality, at 25 µg/ml in leaf disc bioassay. Conclusions The present work established the diversity of Bt isolates and confirmed the importance of continuous exploration of new Bt isolates for novel genes. Further, research needs to be carried out to unveil the hidden potential of these toxic isolates.

Characterization and Evaluation of Indigenous Bacillus thuringiensis Isolate T352 against Fall Armyworm, Spodoptera frugiperda (J.E. Smith)

“…PCR screening for lepidopteran toxic gene families confirmed the presence of cry1, cry1Ab, cry1Ac, cry2, cry2Aa, cry2Ab and vip3A genes in isolate T352 (Table 2). Various studies on indigenous Bt isolates revealed the cry1, cry1Aa, cry1Ab, cry1Ac, cry2Aa, cry2Ab and cry9 genes contributing the toxicity against major lepidopterous insects [24,[35][36][37][38] and the dominance of cry1 and cry2 gene existing together was usually high [39]. The Bt isolate T352 having cry1 and cry 2 along with vip3A gene and toxic to S. frugiperda was in agreement with previous studies [40].…”

“…Similarly, isolate GS4 tested by Patel and Ingle [33] against larvae of Helicoverpa armigera, showed multiple proteins in the range of 88, 54, 97, 175 and 135 kDa indicating the presence of more than one Cry protein and 12 Bt isolates effective against Spodoptera littoralis revealed the protein bands with different molecular weights in a range of 20-130 kDa [34]. Navya et al [35] studied protein profile in 60 indigenous Bt isolates and observed that variation in the molecular weight of Bt protein ranging from 26 to 124 kDa in size.…”

Molecular characterization and toxicity evaluation of indigenous Bacillus thuringiensis isolates against key lepidopteran insect pests

“…accordance with the earlier report by Navya et al (2021), confirming the presence of both cry and vip genes in combinations. Sahin et al (2018) revealed that vip3A1 positive isolates also had cry1 and cry2 genes, which are comparable to our findings.…”

“…There are protein bands of different molecular weights between 30 and > 200 kDa in size observed which need to be characterized. Diversified electrophoretic patterns in Cry protein with molecular weights ranging from 20 to 160 kDa were reported earlier by Navya et al (2021). In the present study, 16 isolates were found to produce one protein band (32.00%) and 16 isolates produce two protein bands (32.00%).…”

“…The Bt isolates investigated appeared as creamy white in colour and fried egg or mucoid type colonies having flat or raised elevation with entire or undulated margins. Navya et al (2021) observed creamy white to off-white colour colonies with fried egg appearance, irregular shape, flat and undulate margin in the Bt isolates screened.…”