It is winter in America and mosquito season has long since passed. But the threat posed by mosquito-borne illnesses -- malaria, dengue fever, yellow fever, West Nile virus -- is still serious, and growing, across the world. So even as low temperatures keep these insects from breeding in their beloved boggy puddles, inside entomology departments across the country, green-eyed transgenic mosquitoes are swarming -- bioengineered skeeters that represent the front line of the rush to introduce genetically modified creatures into the natural environment.
In 1897, British scientist Ronald Ross discovered that malaria was spread, or "vectored" by mosquitoes. He was also the first to propose reducing or eliminating the world's mosquito population as a way of controlling the disease. But it wasn't until early WWII forays into chemical warfare -- leading to the discovery of insecticides ferocious enough to take on the insects -- that such an idea became truly feasible.
Since that point, the fight against mosquito-vectored ailments has been a chemical battle. Scientists developed drugs that were very useful against the diseases, and insecticides that were very useful against the mosquitoes that transmitted these diseases. And for a time it looked as though we were winning the fight. Unfortunately, in the past 30 years the rules of our chemical battle have changed. Mother Nature interceded and evolution occurred. The insects are now resistant to the pesticides, and the diseases are now resistant to the drugs.
A number of these diseases are considered the deadliest on earth. Today, principally in Africa and Asia, dengue fever annually infects more than 50 million people and kills 500,000. Malaria infects about 400 million and manages to kill more than a million. More disturbingly, the combination of pesticide and drug immunities along with the rise of global transportation and current climate changes has resulted in mosquito-vectored ailments appearing in places where they have not been seen before. Last year in the United States, dengue appeared for the second time in Hawaii and the first time in the Gulf States, and last summer there were 4,000 reported cases of West Nile and 300 deaths. Malaria has so far failed to make serious new inroads into the United States, but in the slightly purple words of the Malaria Foundation International, "a plague is coming back and we have only ourselves to blame."
To address these concerns, during the past 15 years scientists have been trying to move beyond the chemical paradigm and to a genetic one. The dream has been to build a genetically modified insect, a transgenic mosquito, that is unable to transmit such diseases. This new insect would then be introduced in the wild, thus supplanting malaria carriers with a harmless imposter. Seven teams, both in America and in Europe, demonstrating a collaborative spirit not often found in modern science, have been at work on the project. In recent months they have succeeded in achieving major breakthroughs.
It is now possible to walk into any number of molecular biology labs and peer through a microscope at a mosquito unlike any other in history. The magnified insect shows a feature not found in the wild: a pair of bright, fluorescent green eyes -- the telltale sign of successful genetic modification. These eyes are proof that one of the most scientifically advanced cures for disease ever conceived is feasible. They are also proof that, if we are not exceptionally careful as this research progresses, something could go horribly wrong. We could do irreparable damage to the ecosystem or, worse, create new, more devastating ailments currently unknown to science. One thing is certain: The bioengineered mosquito hangs precariously off the cutting edge of genetic research. How we proceed is likely to set standards for how we mold our world at a time when such moldings are both probable and perhaps practical.
The successful creation of a mosquito modified to be unable to transmit disease occurred in May 2002. Working in a laboratory with a version of malaria that infects mice, geneticist Marcelo Jacobs-Lorena (then at Case Western Reserve University and now at Johns Hopkins) found a way to inhibit the disease's transmission. In recent months, Jacobs-Lorena and other scientists building on his work have taken the process further. They feel that a transgenic mosquito that is immune to malaria and able to live in the wild is at hand.
To understand how this will be accomplished, it's first necessary to know a little bit about the relationship between the malaria parasite and mosquitoes. There are about 2,500 kinds of mosquitoes in the world, but no more than a tiny minority have evolved to feed on humans. As mosquitoes learned to live off humans, malaria -- the most prolific of mosquito-borne diseases -- was learning to live off both.
Besides humans, other versions of malaria infect all sorts of mammals and birds. Up to now all of the transgenic research has been done with the disease's avian or rodent version, but it spreads the same way in every species. Transmission begins when a hungry female mosquito (only the female feeds on blood) drinks her dinner from an infected animal, along the way ingesting the malaria parasite. In a few days' time that parasite travels into the mosquito's mid-gut, where it develops sexually reproductive cells. These cells mate and following fertilization release thousands of malaria sporozoites that make their way into the mosquito's circulatory system, eventually taking up residence inside the salivary gland. The next time that mosquito bites something, malaria goes along for the ride.
Much of the life cycle of malaria was understood by the early portion of the 20th century, but most scientists date the attempt to remake insects into allies in the war against insects to the work that began in the 1930s by the late Barbara McClintock. In 1983, McClintock was awarded the Nobel Prize in medicine for her discovery in corn of the existence of short chains of DNA called "transposable elements" or, more commonly, "jumping genes." A jumping gene is so named because the proteins it encodes can splice open a chromosome, jump inside, and then sew the whole deal back together again. This discovery excites scientists because it's now possible to piggyback other, more helpful DNA -- like DNA that would be useful in the fight against malaria -- on a jumping gene and insert the whole package into a mosquito's genome.
In 1981, building on McClintock's work, Gerald Rubin, a scientist then at the Carnegie Institution of Washington, discovered a jumping gene known as the "P element" in the entomological workhorse, the fruit fly Drosophilia melanogaster. A year later, he and another scientist named Allan Spradling took the next step and, by using P as their Trojan horse, built the world's first genetically modified insect. "It was a huge accomplishment," says Peter Atkinson, an entomologist at the University of California at Riverside and one of the scientists leading the new genetic charge. "They took a gene that gives fruit fly eyes a reddish color, and attached it to P, and then inserted the whole thing into a fruit fly. That fly's offspring were born with reddish eyes and their offspring and so on. The trait was stable and heritable."
Scientists then believed that the P element would be found in other insect species and once found would be useful in manipulating genes in the fight against insect-borne diseases. As it turns out, P doesn't exist in mosquitoes. "The '80s were pretty much a wild goose chase down this road," recounts Atkinson. "Entomologists thought that P was going to be this great breakthrough, but all that got published for a decade was negative results."
But in 1996-97, Atkinson and David O'Brochta, a professor at the University of Maryland, found a jumping gene they dubbed Hermes, for the speedy Greek god and messenger, in houseflies. The hope was that this gene would be workable in a way that P wasn't. And the following year, Anthony James, an insect-geneticist at UC-Irvine, proved just that. By using Hermes, he discovered a way to genetically modify a mosquito. On their fruit flies, Spradling and Rubin had only to look for a change in eye color to see proof that their experiments worked. The altered eye color is what scientists call a genetic marker, but no such marker existed for mosquitoes. The only way to find out if that insect had been genetically modified was to kill and dissect it. In 2000, Atkinson got past the problem by finding a way to insert into mosquitoes the gene that makes jellyfish glow green. Mosquitoes modified in this way produced offspring with glowing green eyes.
Then, last year, Jacobs-Lorena figured out how to insert a gene (along with a fluorescent marker) that would block the receptors in a mosquito's gut to which rodent malaria attached itself. Because the malaria parasite can't find any place to attach itself, it dies before it can reproduce. Because it can't reproduce, it can't infect anything else.
Jacobs-Lorena's work was the culmination of 30 years of effort. "His achievement," says James, "was the 'proof of principle' we've been waiting for. We now know it's possible to build transgenic mosquitoes in a laboratory that can kill off rodent malaria. The question now is, how do we do this with human malaria out in the real world?"
One of the main lessons scientists learned in the pesticide wars of the last century was that both mosquitoes and malaria are highly adaptive. Therefore, despite the importance of Jacobs-Lorena's achievement, everyone involved realizes it's not enough. "In order to ensure success," notes James, the UC-Irvine geneticist, "we need to build a transgenic mosquito that kills malarial parasites in a number of different ways. We need to make sure we can stay a few steps ahead of evolution."
Due to malaria's 10-day gestation period, scientists can attack the parasite at different places. Jacobs-Lorena blocked the receptor that the parasite binds to inside the insect's mid-gut. James is working on the inverse. Over the past six years, he has figured out ways to block a molecule produced by the parasite that allows it to bind to the insect's salivary glands. In that time, he has reduced malaria levels in the mosquito by 99 percent and feels it likely that he can get the level down to zero within the next year.
A very different approach is being undertaken by Alexander Raikhel at the University of California at Riverside. Raikhel has figured out how to boost a mosquito's immune system. A mosquito's system naturally produces certain proteins in the presence of foreign bacteria. But since malaria is a parasite, not a bacteria, it doesn't normally have to deal with those proteins. Raikhel has figured out a way to trick the mosquito into producing them during the period of time when the malaria sits inside the insect's gut. The result is a mosquito with a turbo-charged immune system that turns on every time there's a chance the mosquito can get malaria -- thus killing the disease before it has the chance to spread.
This work is the only such enterprise in existence. Nowhere else are scientists fiddling with genetics in an attempt to stop the spread of disease. But exciting as finding a way to kill animal malaria in the laboratory seems, it's only the beginning of the process. The next few steps are about finding a way to make this work in the jungle.
Even the first of these steps, building a transgenic insect that exists on a par with normal mosquitoes, was thought to be a Herculean task, but in work that has just been completed (and has yet to be published) Jacobs-Lorena says that his transgenic mosquitoes have the same life span and produce the same number of offspring as normal mosquitoes. "This means," says Lorena, "that in laboratory conditions there's no fitness cost to building mosquitoes with an immunity to malaria."
But the transgenic mosquito must be stronger than regular mosquitoes. "For us to really control the disease, we still have to find a way to make our transgenic insects have more offspring than wild mosquitoes," says Jacobs-Lorena. Much of this work is being done by Atkinson and James; their computer modeling will be followed by lab studies followed by, ultimately, field studies. "We still need to understand how transposable elements move through a mosquito population," says Atkinson, "and we need to know how to make this more efficient."
What these scientists are looking for is a non-Mendelian mechanism for driving these genes into a wild population: They want something quicker than typical insect birth rates. The ideas being explored include using a jumping gene or attaching the malaria-blocking gene to a virus or bacterium that has the ability to rapidly travel through a wild population. Until this research occurs, no one can say how long it will take to actually eradicate the disease.
Simultaneous to this work, some of the teams are undertaking the switch from mosquitoes that carry animal malaria to mosquitoes that carry human malaria -- a feat not as easy as it sounds. Not only are the mosquitoes that carry human malaria much harder to breed in captivity, but there are also differences between the animal models and the human models. The same gene that blocks malaria in Jacobs-Lorena's mice does not work in humans, although Jacobs-Lorena has reported in another study soon to be published that he believes he's found another gene to accomplish the task.
In experimenting with human malaria, the risks associated with the enterprise become greater. There's no way to build transgenics with a human form of malaria immunity without first breeding insects with the human form of malaria. So now there's a level 3 bio-containment laboratory on the UC-Riverside campus complete with multiple airlocks, electronic passkeys, and drainage systems that dump waste water, not directly into the sewer system, but into a heating chamber that cranks up high enough to boil off anything untoward.
There are sound reasons for the building's Fort Knox approach. One concern is that some of the mosquitoes might escape and people would get malaria. Even more worrisome is a problem addressed in the fall of 2002, in a report published by the National Academy of Sciences titled "Animal Biotechnology: Science-Based Concerns." It is primarily a study of the dangers of genetic modification and its possible impact on the environment. Starting with what we already know, that jumping genes got their name because of their ability to hop around a genome and hop from species to species, the report examines existing laboratory conditions and the amount of work currently done in these labs and then, using a low-to-high scale, ranks the chances that existing transgenic animals could escape, become feral, interbreed with wild populations and potentially produce something that we've never seen before. Across the board, insects rated in the high category.
And even if the Riverside containment lab does its job, and neither of those things occur, the progression of this work to the next stage brings a whole host of other ecological concerns. We know very little about how mosquitoes live in the wild. We don't completely understand how they breed -- meaning everything from how they select certain mates to why they choose to lay their eggs in one puddle of water rather than another. We lack understanding of how seasons affect population size or how wide a territory certain populations inhabit or, critically, how and why genes travel through given populations. Thus we don't yet know all the dangers involved in tinkering with this balance. "This is a high-risk venture with a high-yield outcome," says Collins.
To combat the risk, Collins and James and others have been advocating for just this type of basic research. Back in the mid-'90s, inspired by their anti-malaria transgenic work, insect ecologists at UC-Davis and elsewhere made long strides into understanding everything from mosquito breeding patterns to gene flow. Already, in Kenya, there exists a giant screened-in greenhouse, complete with mud huts and breeding puddles, dubbed the "malariasphere." There scientists are studying population dynamics and transmission rates. Even so, Collins quickly points out that whatever we learn from the malariasphere is a drop in the ocean of what we need to know.
The reasons for our need to know are more than a little frightening. Mosquito-borne ailments are among the most devastating and successful diseases on earth. The chemical paradigm of the last century produced a disease immune to our drugs and an insect immune to our pesticides. What if genetically modified mosquitoes result in yet another boost for disease-carrying insects?
"The chances that we're going to end up making a Frankenstein mosquito are pretty remote," says Collins -- but even he agrees that the possibility exists.
Even if our transgenics don't end up vectoring a super-malaria, James points out there are other things to worry about: "If by making a mosquito unable to transmit malaria, will it suddenly become possible for that insect to vector another disease that, as of yet, is not transmitted by mosquitoes? A disease like AIDS?"
Will we assuage all these fears before releasing transgenic mosquitoes? "There's no way to know exactly what will happen in 10 thousand generations of mosquitoes," says Atkinson. But we do know the threat of mosquito-vectored disease is growing more serious every year, and as that threat rises so does the possibility of a new page in history.
In 77 A.D., Pliny the Elder published "Natural History," his rather imaginative 37-volume attempt to catalog the entire contents of the world. From him, we learn that the artichoke is one of the earth's great monstrosities; that dog-headed people who communicate by barking exist; that on islands off the coast of Germany live a tribe of people whose ears are so large they cover their entire body; and that arsenic, sulfur, caustic soda and olive oil are used to protect crops against pestilence. That last bit of information might seem wan compared to other elements in Pliny's mythic compendium, but it is our first written record of insecticides. And now, two thousand years later, the next time someone sets out to catalog the entire contents of the world, there will be a new entry in that great list: a mosquito that lives a dual life as the direct descendent of two of Pliny's described lineages: both a fantastical creature and a pesticide.
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