Courtesy of CDC/Jim Gathany
A decade ago, scientists around the world recognized that despite malaria's tremendous disease burden, research on the topic had stagnated. With funding at low levels, robust molecular biology tools numbered few. Today, genome sequences for
LOOKING AT THE CULPRIT
Before committing money, a projected $30 million (US) at the time, funders supported proof-of-principle pilot studies on two of
Once chromosomes 2 and 3 were underway,34 and the technological and biological hurdles cleared, the project went forward. Investigators used a chromosome-by-chromosome strategy; each center was assigned different chromosomes. "It would've been a very large whole-genome shotgun project for the time," says Gardner, adding that back then it was five to eight times bigger than any shotgun project yet completed. "It seems ridiculous to talk about it like that these days when they can do mammalian genomes by a shotgun process."
Data derived from the Science Watch/Hot Papers database and the Web of Science (ISI, Philadelphia) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age.
Alas, political challenges loomed. As with the Human Genome Project, concerns about access to sequence data surfaced early on. Researchers doing the sequencing feared that chunks of data, if released, would be poached and published by others in incomplete form. Meanwhile, researchers outside of the project wanted immediate access to the genomic data to expedite ongoing research projects.
"There was a fair amount of rancor on the part of community scientists, who felt, 'We've all supported the investment of a lot of money in this project, and now they're generating data and they won't let us see it,"' says David Roos, director of the Genomics Institute at the University of Pennsylvania. Time and money might be wasted on sequencing individual genes that had already been sequenced but not yet released by the genome project. Roos and others addressed these issues by starting the
Access revealed early clues about the parasite's metabolism and pathogenicity. Prior to the genome's completion, researchers identified novel classes of genes involved in antigenic variation, the process by which the parasite evades the host immune system. Researchers also discovered pathways associated with a chloroplast-like organelle called the apicoplast, including an unusual pathway for isoprenoid biosynthesis. Clinical trials targeting this pathway, which is crucial for parasite survival, are now underway.
STARTING ON THE MOSQUITO
Courtesy of CDC/Edwin P. Ewing, Jr., M.D.
Placental tissue revealing the presence of
Started several years after the
Prior to starting the whole-genome shotgun of the 278-megabase
Though more than 10 times larger than
One feature of the mosquito genome proved to be both a benefit and a drawback: its variation. To get sufficient DNA, investigators ground up about 100 mosquitoes, but the strain they used wasn't fully inbred, thus causing significant genetic heterogeneity. As a result, assembly was tricky; for about 7% of the genome, researchers had two versions of the same chromosomal region. This variation had an upside, however. Researchers were able to identify potentially important genomic variants, or single nucleotide polymorphisms (SNPs), about 450,000 of them. These SNPs may serve as markers for genes underlying specific traits such as pesticide resistance, or for classifying wild mosquitoes according to their subspecies.
Holt and colleagues have begun a large-scale study of wild mosquito populations. They'll collect isolates throughout Africa and sequence small chromosome fragments at regular intervals across the genome to estimate overall variation in the population.
Tracking species and subspecies differences could provide insight into how malaria is transmitted. It might help explain, for example, the genetic basis of
The genome also helped investigators isolate three metabolic enzyme families associated with insecticide resistance: carboxyl esterases, cytochrome p-450s, and glutathione S-transferases. One practical implication: If scientists can track mosquitoes and identify those that are resistant, they can avoid spraying, say, DDT on DDT-resistant mosquitoes.
Researchers also recently identified mosquito genes required for the development of the parasite within. The proteins TEP1 and LRIM1 helped the mosquito's immune system fight the parasite6; the proteins CTL4 and CLMA2, discovered directly from the genome data, helped to protect the parasite.7 "We're now trying to understand ... whether they bind to the parasite, for example, mediating its killing or its protection, or whether it's an indirect kind of reaction," says European Molecular Biology Laboratory staff scientist George Christophides, a coauthor of the latter paper. Future work may uncover parasite- and vector-protein interactions.
OTHER SEQUENCING PROJECTS
Meanwhile, more sequencing projects are underway. The sequencing of the mosquito that causes yellow fever,
Despite many scientific strides, the public health problem remains massive. "We realize that today the genome sequence [of