Bacillus thuringiensis (Bt) is a gram-positive spore-forming soil bacterium that is distributed worldwide. Originally recognized as a pathogen of the silkworm, several strains were found on epizootic events in insect pests. In the 1960s, Bt began to be successfully used to control insect pests in agriculture, particularly because of its specificity, which reflects directly on their lack of cytotoxicity to human health, non-target organisms and the environment. Since the introduction of transgenic plants expressing Bt genes in the mid-1980s, numerous methodologies have been used to search for and improve toxins derived from native Bt strains. These improvements directly influence the increase in productivity and the decreased use of chemical insecticides on Bt-crops. Recently, DNA shuffling and in silico evaluations are emerging as promising tools for the development and exploration of mutant Bt toxins with enhanced activity against target insect pests. In this report, we describe natural and in vitro evolution of Cry toxins, as well as their relevance in the mechanism of action for insect control. Moreover, the use of DNA shuffling to improve two Bt toxins will be discussed together with in silico analyses of the generated mutations to evaluate their potential effect on protein structure and cytotoxicity.
Crop losses caused by nematode infections are estimated to be valued at USD 157 billion per year. Meloidogyne incognita, a root-knot nematode (RKN), is considered to be one of the most important plant pathogens due to its worldwide distribution and the austere damage it can cause to a large variety of agronomically important crops. RNA interference (RNAi), a gene silencing process, has proven to be a valuable biotechnology alternative method for RKN control. In this study, the RNAi approach was applied, using fragments of M. incognita genes that encode for two essential molecules, heat-shock protein 90 (HSP90) and isocitrate lyase (ICL). Plant-mediated RNAi of these genes led to a significant level of resistance against M. incognita in the transgenic Nicotiana tabacum plants. Bioassays of plants expressing HSP90 dsRNA demonstrated a delay in gall formation and up to 46% reduction in eggs compared with wild-type plants. A reduction in the level of HSP90 transcripts was observed in recovered eggs from plants expressing dsRNA, indicating that gene silencing persisted and was passed along to first progeny. The ICL knock-down had no clear effect on gall formation but resulted in up to 77% reduction in egg oviposition compared with wild-type plants. Our data suggest that both genes may be involved in RKN development and reproduction. Thus, in this paper, we describe essential candidate genes that could be applied to generate genetically modified crops, using the RNAi strategy to control RKN parasitism.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.