Stanford Report, October 17, 2001
By KRISTA CONGER
Mixing formerly isolated strains of a common human pathogen can rapidly create offspring that are much more lethal than either parent, say Stanford researchers. Their research into the biology of the protozoan parasite Toxoplasma gondii is the first example of such a phenomenon in a non-viral pathogen, and has disturbing implications for scientists trying to predict when and where the next lethal human pathogen might arise. It also indicates that the three main strains of toxoplasma found throughout the world today derive from the relatively recent mixing of two genetically distinct lines.Toxoplasma, which can infect almost any warm-blooded animal, is usually contracted by humans through contact with cat feces or by eating undercooked meat. Although usually harmless, infection can have devastating effects on people with weakened immune systems. Women who become infected for the first time while pregnant also run the risk of miscarrying or bearing a child with birth defects.
Researchers in the laboratory of John Boothroyd, PhD, professor and chair of microbiology and immunology, compared the DNA sequences of many toxoplasma genes from each of the three main strains, as well as several rare ones. Their work, published in the Oct. 5 issue of Science, showed that the genetically distinct strains achieved their unique composition by mixing and matching just two main sets of genes.
"There are two very distinct sequences for every gene, but only two," said Boothroyd. "That’s really surprising. Why just two, and why are they so different?"
An explanation may lie in toxoplasma’s unique lifecycle, which has features of both sexually and asexually reproducing organisms. Unlike humans and many other animals, which have two copies of most genes, toxoplasma has only one. The pathogen reproduces easily in this state, churning out identical copies of itself packaged into cysts embedded in the tissue of an unsuspecting host. The infection is spread when another animal eats the diseased tissue, starting the cycle again.
Cats, however, are special. When toxoplasma infects a feline host, it gains the ability to swap genes between individuals in a process called recombination. If the two individuals are significantly different from one another, recombination can result in unique offspring that may be more fit than either parent. Creating novel pairs of genes from two parents is a tried-and-true way for organisms with two copies of every gene to test the evolutionary waters. But other than in mutant strains of the influenza virus it has not been shown before in an organism like toxoplasma.
If ancestral strains of the parasite were geographically separated, no recombination between strains could occur to generate radically new lines, say the researchers. Allow the two ancestors to mix at exactly the right time in one cat, however, and recombination can theoretically generate more than 10 million new infectious recombinants.
"Toxoplasma is the most common protozoan parasite on Earth that infects warm-blooded animals," said Boothroyd. "Our expectation had been that, given its prevalence, there would be thousands of different strains. We were very surprised when we found several years ago that the number of strains is actually very limited. Now, we’re equally astonished to find that virtually all these strains have one or the other of just two DNA sequences for every gene we look at. "
The existence of a limited number of strains, coupled with the genetic evidence, suggests that this current portfolio of toxoplasma arose from the genetic mixing of two distinct, possibly geographically isolated ancestors. In theory, these could even represent the offspring from an infection of just one cat, said Boothroyd. Out of this mixing came three super-successful strains – two relatively non-virulent and one more deadly – that quickly out-competed their sisters and rocketed to the top of the hierarchy.
To test their hypothesis that recombination could yield offspring with vastly different, and presumably more advantageous, properties than the parents, the team artificially crossed the two non-virulent strains and used 16 of the subsequent recombinants to infect susceptible mice. Fourteen of the 16 offspring remained non-virulent, but two of them were more lethal. In fact, one was more than 1,000 times deadlier than either parental strain.
Because both parents and offspring have only one copy of every gene, such a dramatic divergence from the parental phenotype can result only from novel interactions between new combinations of genes, rather than from inheriting a better copy or combination of copies of any one gene.
"It’s clear that recombination can produce these incredibly fit pathogens without anything else going on," said Boothroyd. As global travel becomes more prevalent and environments are destroyed, scientists have to contend with the fact that humans may be unwittingly facilitating potentially catastrophic blind dates between previously isolated pathogens. "When you allow isolated pathogens to mix, you can get very powerful strains emerging," said Boothroyd.
The researchers are now interested in performing more extensive genetic analysis to determine how many separate crosses actually occurred to produce today’s three main strains from two ancestors. Boothroyd’s laboratory members who participated in the research include post-doctoral researchers Michael Grigg, PhD; Serge Bonnefoy, PhD; and Adrian Hehl, PhD. Yasuhiro Suzuki, PhD, formerly a researcher in the division of infectious diseases and geographic medicine, was also a key contributor.
