Key Anthrax Virulence Factor Discovered
THURSDAY Jan. 24, 2008 -- Researchers have discovered how the anthrax bacterium protects itself from the immune system's biochemical assault.
The results reveal not only a novel aspect of anthrax virulence, they also suggest a new target, known as bacterial nitric oxide synthase (bNOS), for fighting the pathogen, said study leader Evgeny Nudler, a professor of biochemistry at the New York University School of Medicine.
"That is the obvious continuation of our work," said Nudler, "to find small molecule inhibitors of bacterial nitric oxide synthase that would not touch its human counterpart. If we are lucky, we will find this inhibitor that would potentially work as an antibiotic."
One expert thought the finding could have therapeutic potential.
"bNOS can definitely be a therapeutic target," said Philip Tierno Jr., director of clinical microbiology & immunology in the Departments of Microbiology & Pathology at New York University Medical Center. "Clearly they showed that it should be a target, and if that somehow could be met, they may have knocked out the virulence of the organism, as they showed in the mouse model."
"But it may be more complicated than the model suggests, and there may be more factors at play than they suggest," Tierno added. "Things may not be a simple as it seems, but it's certainly a step in the right direction and should be explored, and might offer a solution, especially because anthrax could be a viable candidate for a terrorist."
The results were reported in this week's issue of the Proceedings of the National Academy of Sciences.
According to Nudler, this study stems from earlier research in which his team determined that bNOS (an enzyme that synthesizes nitric oxide) protects some bacteria from the stress of oxidizing chemicals, including the hydrogen peroxide and nitric oxide that immune cells called macrophages use to kill pathogens. That research involved Bacillus subtilis, a close relative of the anthrax pathogen, Bacillus anthracis.
In the present study, the team found that when they mutated the bNOS gene in the anthrax pathogen, the resulting bacteria were far more susceptible to oxidative stress, less able to germinate within macrophages and considerably less deadly in mice.
Those observations, said Nudler, were not unexpected. However, their magnitude was.
"The surprise was it was so dramatic," he said. "We expected some effect." But with a decrease in virulence of almost 1,000-fold, he explained, "This implies bNOS is an essential virulence factor in anthrax."
The results present an apparent paradox, that the bacteria must synthesize their own nitric oxide to survive the nitric oxide attack inflicted by the immune system. But Nudler said it is all a matter of timing.
"Bacteria produce nitric oxide immediately upon germination, in the first two hours of infection," he explained. "Macrophages produce their own much later, 12 hours post-infection. So, the bacteria takes advantage of its own nitric oxide, as a preemptive strike to protect itself against future nitric oxide attack."
Indeed, he noted that during spore formation, anthrax pathogens store some bNOS, so it will be available immediately upon germination, when the bacteria are most vulnerable.
Bacterial "spores" are seed-like structures the bacteria form to protect themselves against unfavorable environmental conditions. Inhalation of anthrax spores (as occurred within the U.S. Postal Service in late 2001) is especially dangerous, as the spores are engulfed by macrophages, inside which they germinate and escape to spread the infection.
Now Nudler and his team are searching for small molecule inhibitors that can selectively target bNOS while leaving its human counterpart enzyme alone.
Given the differences between the two proteins, he said, "There is a great chance we can find a specific molecule that can inhibit it."
Another potential biothreat breakthrough was also published in the same issue of PNAS. Yoshihiro Kawaoka, of the University of Wisconsin, Madison, and colleagues described a new, potentially safer way to grow ebola virus in culture -- an advance that could enable researchers to handle the pathogen in less-restrictive facilities and thus facilitate wider research into both its life cycle and the development of antiviral therapeutics.
For more on anthrax, visit the U.S. Centers for Disease Control and Prevention.
Posted: January 2008