Genetic research reveals key to cotton resistance

In the last battle in the endless war between farmers and insects, insects are repressed by adapting to genetically modified crops to kill them.

A new study published in the Proceedings of the National Academy of Sciences (PNAS) identifies a dominant hereditary mutation that confers resistance to artificial cotton in cottonseed worms, one of the most destructive crop pests in the world.

The advanced use of genomics and gene editing in the study marks the beginning of a new era in global efforts to promote a more sustainable fight against pests.

Cotton, corn, and soybeans have been genetically engineered to produce antiparasitic proteins from the widespread soil bacterium Bacillus thuringiensis, or Bt.

Non-toxic to humans and wildlife, including bees, these environmentally friendly Bt proteins are used in spraying by organic growers for over 50 years and in Bt engineering crops planted by Millions of farmers worldwide over a cumulative total of over two billion acres since. 1996.

Entomologists from the University of Arizona, the University of Tennessee, and the Nanjing Agricultural University in China collaborated on this three-part study. Their objectives were to determine the mutation conferring resistance to Bt in the worms of the capsule, to modify a gene of the worm of the capsule to prove that this mutation was the cause of the resistance and to discover how this resistance spread in the cotton fields in China.

"It's a remarkable detective story," said Bruce Tabashnik, professor at the Entomological Department of the AU and co-author of the study. "Without the latest advances in genetic technology, it would not have been possible to find the unique DNA base pair change that causes resistance among the hundreds of millions of base pairs of the worm genome." of the capsule. "

Scientists have known for years that insects can develop resistance to Bt proteins just like conventional insecticides. However, resistance to Bt is inherited recessively in almost all the cases previously studied. This means that insects must have two copies of the resistance gene – one from each parent – in order to feed and survive with Bt culture.

To combat resistance, farmers plant shelters for non-Bt crops, where susceptible insects can thrive. The idea is that rare resistant insects will mate with the most abundant sensitive insects of the refuges, producing offspring that harbor only one copy of the resistance gene. With inherited inheritance recessively, such offspring do not survive on Bt culture.

Although shelters do not completely stop the evolution of resistance, they can delay it considerably, especially when resistance is recessive.

But in China, the newspaper reports that the dominant resistance to the Bolls worm in Bt is on the rise. A single copy of a dominant mutation makes the worm of the capsule resistant.

Since the genetic basis of dominant Bt resistance was unknown, researchers had to examine the entire genome of the capsule worm to find the culprit. By comparing the DNA of resistant and susceptible capsule worms, they reduced the search for 17,000 genes to a region of only 21 genes associated with resistance.

"But only 17 of these genes code for proteins produced by caterpillars," Tabashnik said, explaining that only wormworm caterpillars were feeding on cotton and could be killed by Bt proteins. these 17 genes between the strains, there was only one coherent difference. There was a position where all resistant capsule worms had a pair of DNA bases and all likely capsule worms had a different base pair of DNA. "

This pivotal base pair is in a newly identified gene called HaTSPAN1, which encodes a tetraspanine, a protein containing four segments that cover cell membranes. Although the normal function of HaTSPAN1 is unknown, many other tetraspanins play an important role in cell-to-cell communication. Despite nearly 30 000 previous studies on Bt or tetraspanins, this new study is the first to establish a close link between them.

With the identified mutant base pair, the second challenge was to determine if this unique mutation was causing resistance. To find out, the research team used the CRISPR gene editing tool to modify precisely the HaTSPAN1 gene. When the gene was disrupted in resistant capsule worms, they became completely susceptible to Bt. Conversely, when the mutation was inserted into the DNA of susceptible capsule worms, they became resistant – the evidence that this single base pair change can cause resistance.

The final step was to test the hypothesis that this mutation would contribute to Bt cotton resistance in the field. In researching the mutation in DNA of thousands of conserved mites captured between 2006 and 2016, researchers found that the frequency of mutation was multiplied by 100, from 1 in 1,000 to 1 in 10.

Resistant capsule worms are not yet numerous enough to significantly reduce cotton production in China, but the dominant gene is spreading faster than other resistance genes.

Tabashnik's analysis predicts that if the current trend continues, the resistance of half of China's North China capsule worms will be conferred by this mutation within five years.

"If things continue on the same path, it's the mutation that will cause problems to farmers on the ground," Tabashnik said.

However, it is early enough for Chinese farmers to change tactics and postpone resistance. The paper mentions that they could switch from cotton that produces a single Bt protein to types of cotton grown in the United States and Australia that produce two or three distinct Bt proteins. Tabashnik hopes the new research will boost farmers' sustainability.

"It gives them the information they need to make constructive and proactive decisions before it's too late," Tabashnik said.

By sampling pest populations each year, farmers and researchers may be able to determine the most effective methods of counteracting resistance. Understanding the resistance of the capsule worm has global implications, as it occurs in more than 150 countries and is now threatening to invade the United States.

"It will be interesting to look for this mutation in the worm capsule of cotton from Australia, India and Brazil," said Yidong Wu, professor of entomology at the University of Hawaii. Nanjing Agricultural University, which led research in China.