The Insects' Deadly Bacterium Receive Reinforcement

In 1911 a bacterium called Bacillus thurigiense (Bt) was discovered, and which produced a toxic crystal proved to be lethal for many insect species.

But recent work of a team from the University of Wisconsin-Madison reports that without the assistance of the native bacteria that colonize the insect gut, Bt is unable to perform its lethal work. Bt is the most environmentally friendly insecticide, used for the control of insects in agriculture, forestry and human health, but also in research of microbial disease in humans and animals.

"The take-home message is that we've shown that the mechanism of killing for Bacillus thuringiensis is facilitated by the normal gut community," says Nichole Broderick, a UW-Madison graduate student. "This is a mechanism that was not previously known."

Bacillus thuringiensis was developed commercially in the 1950s. It is by far the most widely used natural insecticide and the genes that produces Bt's toxic proteins have been introduced into numerous crop plants. Transgenic crops using Bt's genes are the most widespread, cultivated on millions of acres in US alone.

Previously, it was presumed that toxins disrupted the line of insect gut cells and the gut holes led to starvation. Or the bacteria provoked blood poisoning. "What was proposed as a hypothesis in one paper became cited as proven in another and no one seemed to go back to the original literature until now." explains Jo Handelsman, a Howard Hughes Medical Institute Professor in the UW-Madison department of plant pathology.

The new research demonstrates that Bt requires the presence of other bacteria to provoke the lethal effect. All animals - including humans - have, beginning with the birth, colonies of bacteria in their stomach and intestines. For example, the caterpillars have 7 to 20 species of gut bacteria. Humans have between five hundred and one thousand species of micro flora in the intestinal tract. "In moths and butterflies, the complexity is much lower than in mammals, and even some other insects," Broderick explains.

The team cleared the guts of gypsy moth caterpillars, killing all the bacteria with antibiotics. The "clean" were immune to the toxic proteins of BT. "Initially, I was testing the hypothesis that the gut bacteria were actually protecting the moth," she says.

"I found that once they did not have a gut community [of bacteria] I could no longer kill them with Bt."

Large numbers of the nascent moths were fine, even when fed as much as 10 international units of the toxin, 10 times more than ordinarily needed, as long as they were also fed antibiotics. "I think it was by the third fairly large, replicated experiment that we looked at each other and said, 'this is real,'" Handelsman recalls. The Bt toxin could not kill on its own.

Reintroducing Enterobacter species NAB3, Bt's deadly impact was restored. Enterobacter thrive in the hemolymph, multiplying as much as when raised in laboratory broth, whereas B. thuringiensis was wiped out from the blood within six hours. "The gut wall will heal and regenerate itself if there are no gut bacteria there," Handelsman says.

The team used a strain of live transgenic E. coli carrying the Bt toxin to infect caterpillars, a lethal treatment whether or not the gut contained microbes. If the new strain of E. coli was killed before administration, it only killed caterpillars with an intact gut flora. "The significance of the microbial community has been overlooked," Broderick asserts.

"Ultimately, this is a toxin-mediated septicemia (blood poisoning) modulated by the gut community." "Is the Enterobacter moving into the blood? At this point, that's the hypothesis we're pursuing," Broderick notes. "I think it's probably more complex than that. The simple model doesn't always turn out to be the case."

The killing effects of Bt could be enhanced using bacteria known to promote blood poisoning. "The work also raises the possibility that the genes encoding the (Bt) toxins could be deployed more effectively in transgenic crops by exploiting the role of insect-borne bacteria that enhance insecticidal activity," the Wisconsin team writes.

The results could have application in human and animal medicine, because they shed light on the roles of the bacteria in gut infections. Human bacterial infections may account for as much as 10 percent of mortality in US.

"It is thought that the gut is the source of bacteria for a large portion of cases of human septicemia, so if this mechanism is shared by Bt and toxins produced by human pathogens, the implications could be far greater in medicine than in agriculture," Handelsman says.