cry Genes profiling and the toxicity of isolates of Bacillus thuringiensis from soil samples against American bollworm, Helicoverpa armigera

Abstract

Aims: The aim of this study was to search for Bacillus thuringiensis (Bt) harbouring cry1A gene which could effectively control cotton pest, American bollworm, Helicoverpa armigera. Methods and Results: cry gene profiling of 50 Bt isolates showed the presence of cry1, cry2, cry3, cry4, cry7, cry8 and cry9 genes. None of the isolates harboured cry1 gene alone. It was always found in combination with cry3. There was no isolate positive for cry10 gene. Considering isolates with single cry genes, the frequency of cry4 was predominant (22%) followed cry2 (6%), cry3 (4%) and cry8 (2%). Isolates having two cry genes in combination had 14% incidence for cry2 + cry4, 12% for cry3 + cry4 and 10% for cry1 + cry3. The most dominant three gene linkage was cry1 + cry3 + cry4. Further profiling

of cry1 gene showed that cry1K gene was abundantly present in all combina- tions such as cry1A, cry1D, cry1F and cry1I. However, cry1C existed indepen- dent of other subtypes. Finally, the Bt isolates with cry1A were analyzed for 16S

rRNA gene, which showed two distinct groups of isolates on the basis of sequence homology. Bioassays of spore–crystal mixtures of SBS-Bt4, 8, 17, 21 and 26 harbouring cry1 against neonate larvae of H. armigera showed LC50 1288, 1202, 467Æ7, 524Æ8 and 108Æ5 lg ml)1

. The SBS-Bt26 showed fourfold higher toxicity than the cry 1Ac harbouring positive control, HD-73. Conclusions: None of the isolates harboured single cry 1 gene. They were always in combination of two or three genes. A Bt isolate (Bt26) had fourfold higher toxicity against H. armigera larvae compared with the positive control HD 73 and hence can be commercially exploited to control insect pest. Significance and Impact of the Study: The inter relationship between the cry genes content and the toxicity may allow better understanding of Bt ecology.