Scientists Identify First Factor That Enables Parasite To Infect Humans

"This is the first pathway we have identified which controls the virulence of this deadly pathogen. Remarkably, it normally acts to limit parasite virulence rather than to increase it," says principal investigator Stephen M. Beverley, Ph.D., the Marvin A. Brennecke Professor of Molecular Microbiology at Washington University School of Medicine in St. Louis.

In the April 13 issue of the journal Science, Beverley’s team reports that the virulence of Leishmania is controlled by the levels of a substance called tetrahydrobiopterin (H4B).

The microscopic parasite infects more than 10 million people in tropical parts of the world, causing an ulcerating disease called leishmaniasis. The disease often is fatal, and no satisfactory vaccines or drug therapies exist.

Like a caterpillar that changes into a butterfly, Leishmania has several stages in its life cycle. In the first stage, it lives in the digestive tract of the sandfly, where it obtains biopterin and then converts it to the H4B needed for growth.

At the end of this stage, the parasite stops growing and transforms into the infectious metacyclic stage, which is now ready to be transmitted when the fly bites a human or other mammal. Once the fly bites, the parasites invade phagocytic white blood cells where they differentiate into yet another stage, the amastigote form, which causes disease.

With postdoctoral fellow Mark Cunningham and colleagues from Colorado State University and the University of Kentucky Medical Center, Beverley set out to learn whether parasites lacking the gene for pteredine reductase 1 (PTR1), the enzyme that converts biopterin to H4B, could infect mice. By knocking out the gene, the investigators lowered the amount of H4B available to the parasite.

Leishmania did more than survive. It became more infectious. After about two weeks, mice that were injected with the mutant microbe had more than 50 times as many parasites in their bodies as mice injected with normal Leishmania. The rapidity of infection and extent of the lesions depends on the number of parasites that enter the second, infective metacyclic stage, and the PTR1-knockout parasites showed a much greater capacity to undergo this transition.

The researchers then exposed normal Leishmania to different concentrations of reduced biopterin in a test tube. They discovered a threshold above which entry into the metacyclic stage decreased rapidly. Below this threshold of exposure, the infectious stage of the parasite was much more likely to form.

"Therefore, falling levels of H4B, which occur normally during the parasite’s development, appear critical to its ability to develop into the stage which infects mammals," Beverley says. "Conversely, high levels keep the parasite in its noninfectious form."

Beverley finds it interesting that lower amounts of this molecule also make Leishmania more deadly. He believes that limiting its ability to infect mammals may be advantageous to the parasite.

"There’s a central tenet of evolutionary microbiology which states that ‘a good parasite does not kill its host too fast,’" he says. "It may be that the levels of H4B that Leishmania normally contain ensure that it won’t kill its host as quickly, giving the parasite time to reproduce and be transmitted."

This research was supported by grants from the National Institutes of Health.

Reference: Cunningham ML, Titus RG, Turco SJ, Beverley SM. Regulation of differentiation to the infective stage of the protozoan parasite Leishmania major by tetrahydrobiopterin. Science, vol. 289, April 13, 2001.