Harvard scientists use PACE to fast-track discovery of Bt toxins to target super-bugs

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Corn crop, Indiana, US.
Corn crop, Indiana, US.
David Cornwell, Flickr CC

One of the big challenges in modern agriculture is finding new ways to outwit insect pests.

Advances in plant science have led to the development of genetically engineered crop varieties with ‘built-in’ bioresistance to specific insect pests known as 'Bt crops'. 

Until recently, ‘Bt crops’ have proven highly effective at controlling pests including European corn borer, rootworm, corn earworm, tobacco budworm and bollworm. As a consequence of planting Bt varieties, the use of chemical pesticides on many agricultural crops, notably corn, cotton and soybeans, has dropped dramatically.

Over time, however, several species of crop pest have become resistant to the Bt toxins in these crop varieties. Bt toxin resistance costs the US agriculture sector many millions a year in crop loss and damage from resistant insects.

The problem is that finding new Bt strains capable of killing these super-bugs takes time - and meanwhile, farmers are losing crops, money and patience.

Picking up the PACE in the war against super-bugs

Fortunately for farmers, a group of Harvard University scientists led by David Liu, a Professor of Chemistry and Chemical Biology, has developed a technology known as ‘phage-assisted continuous evolution’, or PACE, which can speed up the process of finding new, effective Bt strains dramatically – from several years to just a few weeks.

Using PACE, Professor Lui and a team of researchers have been able to evolve new forms of Bacillus thuringiensis toxin that they believe can be used to combat Bt toxin resistance in an array of crop pests.

Their findings are detailed in the 5 May 2016 issue of Nature, in a paper co-authored by Prof. Lui and graduate student Ahmed H Badran from Harvard's Department of Chemistry and Chemical Biology with Cornell University entomologist Ping Wang and scientists at Monsanto (which produces Bt-toxin crops).

“Our goal in this collaboration was ambitious,” Professor Liu told Phys.org shortly after the paper’s publication. “The key questions were: Can we retarget a Bt toxin to a different insect gut protein by evolving the Bt toxin, and will doing so enable us to kill insects that have become resistant to wild-type Bt toxin? Our hope was to use PACE to help stay ahead of insect resistance.”

The team took just over three weeks to evolve the new, effective Bt toxin proteins, a process that would likely have taken them 150 times longer without the help of PACE.

The new technology enables researchers “to tackle problems that would be difficult to solve by traditional protein-evolution methods”, Prof. Liu told Phys.org.

“In the case of Bt toxin, we evolved new Bt toxins containing dozens of amino-acid changes over 500 generations in 22 days of PACE. To do that many generations of protein evolution with traditional stepwise methods, at the rate of about one generation per week, might take a decade.”

With the help of PACE, the Harvard team recently developed a system that selects for proteins capable of binding to a target protein, Prof. Liu said.

As binding to a protein in the insect’s gut is an essential step in the action of Bt toxin, the system made it possible for Professor Lui and his team to evolve new forms of Bt toxin exceptionally fast.

What is Bt-toxin?

Critical in the development of Bt-toxin (or Bt) crops is a bacterium with insecticidal properties known as Bacillus thuringiensis (Bt).

Initially, Bt bacteria were used to combat European corn borer (Ostrinia nubilalis), a common and destructive corn-crop pest. Bt's effectiveness against this insect pest led to the development of the first commercial Bt-based biopesticide, Sporine, introduced to France in 1938. Since that time, Bt toxins have been used widely for crop-pest control, including in organic agriculture.

By the 1990s, tens of thousands of strains of the bacterium that were toxic to a wide array of insect species had been isolated.

The first corn varieties engineered to incorporate Bt genes became commercially available in 1995.

In the two decades since, GM Bt-toxin-containing varieties have come to dominate US corn and cotton crops: by 2015, 81 percent of corn and 84% of cotton planted across the US were Bt-toxin crops.

Bacillus thuringiensis (Bt) strain 4A4, as viewed at 1000x magnification after gram staining.
Bacillus thuringiensis (Bt) strain 4A4, as viewed at 1000x magnification after gram staining.
Sam L, Flickr CC

Will the Harvard team's new Bt toxins work?

To test their effectiveness, Professor Lui and his team fed the newly evolved toxins to colonies of Bt-resistant insects.

“The resistant insects tolerated about 1,000-fold higher levels of wild-type Bt toxin than normal, sensitive insects,” Prof. Liu explained to Phys.org. “But the evolved Bt toxins kill these resistant insects up to 335-fold more potently than wild-type Bt toxin, thereby restoring almost all of the lost Bt toxin potency.”

This doesn’t mean the new proteins will work forever – eventually, the offending insects will are likely to evolve resistance to the new versions of Bt toxin. However, in PACE, scientists have a tool that enables them to pinpoint crucial steps in toxins’ actions, find new molecules capable of performing them and thereby evolve more new Bt toxins targeting different insect proteins, Prof. Lui said.

Potentially, PACE also makes it possible to develop Bt toxins that target several insect gut proteins simultaneously, making it difficult for targeted pests to put up an effective resistance.

“The hope is that by applying this strategy, we can overcome what's considered to be one of the biggest threats to sustaining the yield gains of modern agriculture,” Prof. Lui said.

Harvard’s Office of Technology Development has granted Monsanto a limited-term exclusive license to use its PACE technology for agricultural applications.

Source: phys.org

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