Dominant point mutation in a tetraspanin gene associated with field-evolved resistance of cotton bollworm to transgenic Bt cotton

Abstract

Extensive planting of crops genetically engineered to produce insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) has suppressed some major pests, reduced insecticide sprays, enhanced pest control by natural enemies, and increased grower profits. However, rapid evolution of resistance in pests is reducing these benefits. Better understanding of the genetic basis of resistance to Bt crops is urgently needed to monitor, delay, and counter pest resistance. We discovered that a point mutation in a previously unknown tetraspanin gene in the cotton bollworm (Helicoverpa armigera), a devastating global pest, confers dominant resistance to Cry1Ac, the sole Bt protein produced by transgenic cotton planted in China. We found the mutation using a genome-wide association study, followed by fine-scale genetic mapping and DNA sequence comparisons between resistant and susceptible strains. CRISPR/Cas9 knockout of the tetraspanin gene restored susceptibility to a resistant strain, whereas inserting the mutation conferred 125-fold resistance in a susceptible strain. DNA screening of moths captured from 23 field sites in six provinces of northern China revealed a 100-fold increase in the frequency of this mutation, from 0.001 in 2006 to 0.10 in 2016. The correspondence between the observed trajectory of the mutation and the trajectory predicted from simulation modeling shows that the dominance of the mutation accelerated adaptation. Proactive identification and tracking of the tetraspanin mutation demonstrate the potential for genomic analysis, gene editing, and molecular monitoring to improve management of resistance.

Disruption of nicotinic acetylcholine receptor α6 mediated by CRISPR/Cas9 confers resistance to spinosyns in Plutella xylostella

“…The current study demonstrates the efficient generation of Pxα6 subunit modification via the CRISPR/Cas9 system in P. xylostella , but it is likely that the editing system can be further optimized. In this work, the proportion of surviving neonates developed to adults (60%) in G 0 was similar to that from H. armigera and S. exigua (52–69%) . However, the hatching rate in G 0 was only 4.5% of the injected eggs, which is obviously lower than that from H. armigera and S. exigua based on the reported studies.…”

“…In contrast, a number of publications have examined target gene mutation mediated resistance. To date, at least ten studies (five from D. melanogaster , three from H. armigera and two from S. exigua ) edited nAChRα6 , RyR , CHS1 , CAD , ABCA2 , TSPAN1 and P‐gp genes . Overall, the CRISPR/Cas9 manipulate techniques are already advanced in the model insect D. melanogaster and agricultural pest H. armigera , and this system provides a useful genome editing tool for other interesting insect species.…”

“…The CRISPR/Cas9 system has been successfully applied in a number of insect species, and has recently been used to investigate insecticide resistance mechanisms . To date, by using CRISPR/Cas9 coupled with non‐homologous end joining (NHEJ) and/or homology directed repair (HDR) approaches, the following mutation(s) edited in the nAChRα6 subunit (G275E and P146S of D. melanogaster ), chitin synthase 1 ( CHS1 , I1015M/F/L of D. melanogaster ), ryanodine receptors ( RyR , G4946E/V & M4790I of D. melanogaster and Spodoptera exigua ), cytochrome P450 gene ( CYP9M10 , KO in Culex quinquefasciatus ), cadherin ( CAD , KO in Helicoverpa armigera ), P‐glycoprotein ( P‐gp , KO in S. exigua ), ABCA2 (KO & KI in H. armigera ), and tetraspanin ( TSPAN1 , KO & KI in H. armigera ) were confirmed in vivo to be involved in insect susceptibility to chemical insecticides or Bacillus thuringiensis Cry toxins …”

Molecular characterization of DNA methyltransferase 1 and its role in temperature change of armyworm Mythimna separata Walker

“…Therefore, we attributed this phenomenon to one of the following two reasons: Elder instar larvae were refractory to RNAi due to the activity of dsRNase (Guan et al, 2018); or MsDnmt1 is not sensitive to RNAi, because genes in the central nervous systems are not easily knocked down (Terenius et al, 2011). Obviously, more work needs to be done to illustrate the function of MsDnm1 in M. separata by other up‐to‐date technologies including CRISPR–Cas9 (Jin et al, 2018).…”

“…Dnmt1 contains three isoforms, Dnmt1o, Dnmt1s , and Dnmt1 , and is considered to be responsible for the maintenance of methylation (Auclair & Weber, 2012); Dnmt2 shows weak DNA methylation activity and is involved in transfer RNA methylation (Tatematsu, Yamazaki, & Ishikawa, 2000); in contrast, the Dnmt3 family includes Dnmt3a and Dnmt3b , which participate in de novo methylation, and are therefore considered de novo methyltransferases (Auclair & Weber, 2012). The de novo methyltransferases Dnmt3a and Dnmt3b are all essential for genome regulation and mammalian development (Jin et al, 2018; Steffen et al, 2018; Zhang et al, 2018). In contrast to mammals, some insects, including the silkworm Bombyx mori and the locusts Schistocerca gregaria and Tribolium castaneum , may only have Dnmt1 and Dnmt2 , indicating that Dnmt1 may play a role in maintaining methylation and de novo methylation (Glastad, Hunt, Yi, & Goodisman, 2011; Lyko & Maleszka, 2011).…”