Bioengineered Bugs A 'Brisk Trade' In Applied and Basic Entomology

if (n == null) The Scientist 9[18]:13, Sep. 18, 1995 Research Beyond the Fruit Fly By Karen Young Kreeger Of all the animal species described thus far by scientists, insects account for nearly five-sixths. Nearly 1 million are already identified, with taxonomists estimating another 1 million to go. Despite their ubiquity on Earth, the study of insects has only recently been married to the explosion of molecular biological tools that have come on the scene in the last 15 years. Sidebar: Resources for Entomologists David A. O'Brochta, an associate professor at the University of Maryland Biotechnology Institute's Center for Agricultural Biotechnology, observes that the increasing interest in the molecular biology of insects stems mostly from their medical and agricultural significance: "I think the genetic-engineering technology developed in my and other labs is going to be for the purposes of studying insects because they're pests or are beneficial. This technology will be used for making them better at what they do -- a better honeybee, for example -- or making them do less of what they do -- biting people or transmitting diseases." Molecular biological tools, such as DNA fingerprinting, gene insertion, and the polymerase chain reaction (PCR) are being applied to control and monitor such species as the Mediterranean fruit fly and malaria-carrying mosquitoes. These techniques are also being used for more basic studies in ecology and systematics. Enough applications have accumulated over the past several years to prompt Marjorie A. Hoy, Fischer, Davies, and Eckes Professor for Biological Control at the University of Florida, to write an introductory textbook on the subject: Insect Molecular Genetics: An Introduction to Principles and Applications (San Diego, Academic Press, 1994). The fruit fly Drosophila melanogaster has long been the workhorse of molecular entomology. "In 1981, the ability to introduce genes into the genome of Drosophila melanogaster adults was first reported [G.M. Rubin, A.C. Spradling, Science, 218:384-53, 1982; A.C. Spradling, G.M. Rubin, Science, 218:341-47, 1982]," says O'Brochta. "Since that time that technology has led to an absolute explosion in fruit fly genetics." "In a sense, molecular entomology is one of the most developed fields because we know so much about Drosophila," remarks Hugh Robertson, an associate professor of entomology at the University of Illinois. "But, on the other hand, it's one of the most undeveloped fields because we know so little about any other insect. So the conundrum is, how do we extend what we know about Drosophila to other insects?" O'Brochta maintains that once the 'bottleneck' of developing a means for introducing genes into the chromosomes of insects is overcome, "we can expect an explosion in basic and applied studies of insects" other than fruit flies. Researchers like O'Brochta and Robertson are using short fragments of DNA called transposable elements -- more commonly referred to as jumping genes -- as the vehicles for the introductions. The late Barbara McClintock won the 1983 Nobel Prize in physiology or medicine for her discovery of these elements in corn plants. Jumping genes generally move within the genome of an individual, within the same chromosome, or between different chromosomes. Most of these genes have such wanderlust-laden names as mariner and hobo. "The elements themselves are little pieces of DNA that encode for a couple of proteins. These cut open a chromosome and splice it back together. It is the space created by the cut into which the elements insert themselves," explains O'Brochta. "We can take a gene that we want to integrate into a chromosome and hook it up to the transposable element. Think of the ends of the element as wheels. If we stick the wheels on either end of the DNA we want to insert, everything between the wheels will act like the transposable element and integrate into the chromosome." O'Brochta relates that the basic thrust of his lab has been to identify transposable elements in non-Drosophila insects and to take known transposable elements -- for example, from Drosophila -- and determine if those can be used as gene-transfer agents in other insect species. Robertson has found that mariner elements are widespread in the insect world, detecting similar forms of mariner in more than 400 arthropod species, the phylum to which insects belong. "We've now found hundreds and hundreds of them in species of insects and, more recently, in other animals," he says. "We and others have found them in nematodes, flatworms, hydras, and humans. The human one is very old and completely defective, probably getting into our genome while we were still prosimians about 60 [million] to 80 million years ago. Now it's quintessential junk DNA." Investigations are just getting under way to apply the lab research on jumping genes directly to use in agricultural crops. Hoy and colleagues are developing a more efficient way to deliver genes into predatory mites. She points out that these mites -- members of the same phylum as insects -- are used to control their crop-munching cousins, the spider mites. Hoy and postdoc James Presnail developed a technique for injecting plasmid DNA into the ovaries of predatory mites. Although these experiments are at a preliminary stage, the researchers hope to eventually insert pesticide-resistant genes into the mites. "What we need is a more efficient method of getting DNA delivered into the mite genome," contends Hoy. "There are a lot of people working on vectors that would increase the frequency of insertion, like the transposable elements that O'Brochta works with." In May, the United States Department of Agriculture (USDA) and France's Centre National de la Recherche Scientifique (CNRS) signed a far-reaching research agreement to study how to transfer new genes into invertebrates. Alfred M. Handler, a research geneticist with USDA's Agricultural Research Service in Gainesville, Fla., is the U.S. scientific coordinator for the project. Alfred Handler of USDA studies the medfly in an international project. "What's so nice about this project is that you have a group of scientists that normally don't talk to one another," says Handler. The project so far involves about 100 researchers worldwide. Handler left for France in June to work with the French scientific coordinator of the project, insect virologist Max Bergoin, who is the head of the comparative pathology lab at the University of Montpellier, II. Handler and Bergoin's portion of the project is to develop reliable technologies for gene transfer in the Mediterranean fruit fly. "The medfly is not established in the continental U.S., but it has devastated fruit and vegetable crops around the world. Florida and mostly California spend on the level of about $50 million per year just to monitor infestations [and to] eradicate the small infestations that are detected, and on general research and control programs," Handler relates. He is working abroad because the medfly isn't allowed in the continental U.S. "The advantage of me being here is that I can work on the medfly itself," instead of working solely with its DNA, he says. Most of the vectors Handler and colleagues are currently studying are transposable elements, such as the hobo element from Drosophila. "One prospective control strategy using genetically engineered medflies is the release of transgenic individuals that are sensitive to cold stress," notes Handler. "A dominant cold-sensitive gene introduced into the population during spring and summer could cause extensive lethality at the first cold snap." In the overall discipline of virology, insect viruses are currently a little-studied area, despite their potential usefulness as agents for controlling insect pests. "They're very much the Cinderella of the virus world, but interest in them is gaining," maintains Donald Hendry, an associate professor of microbiology at Rhodes University in Grahamstown, South Africa. Hendry studies a virus found in the pine emperor moth, whose larval stage denudes pine trees. His lab is hoping to study the potential for using the virus as a control mechanism, but other methods of insect control such as pesticides are currently more effective. In work initiated as a postdoc in the University of California, Davis, lab of Bruce Hammock, Bryony Bonning -- now an assistant professor of entomology at Iowa State University -- studies a type of baculovirus in hopes of controlling plant-puncturing caterpillars, specifically the European corn borer. "The reasoning behind this research is basically to find alternatives to chemical insecticides," explains Bonning. Left, a genetically engineered baculovirus to control a type of Europpean corn borer. The virus infects a broad range of insect pest species. One successful technique has been to put a gene coding for an insect-selective toxin into the virus. A particularly good toxin has been isolated from scorpion venom. "We put the scorpion-toxin gene into the virus genome, so that when the caterpillar eats the virus, which is sprayed on the leaves of crops, the virus starts to replicate inside the larvae, thus producing the toxin, which quickly kills the larvae," says Bonning. Techniques such as DNA fingerprinting are being used by biomedical researchers and insect ecologists to study a host of questions. For example, USDA researchers are using these techniques to study the migration patterns of Africanized honeybees and corn earworms. Other government agencies are also using fingerprinting techniques to accurately identify malaria-carrying mosquito species. The Department of Defense (DOD) is heavily involved in entomological research because of health concerns for troops stationed abroad. Richard Wilkerson, a research entomologist at DOD's Walter Reed Army Institute of Research, studies the mosquito genus Anopheles, some species of which carry malaria. "I started out just looking at morphological characteristics, but we found different species that look the same," recalls Wilkerson. "So we had to use different tools to tell them apart. That's what prompted me to start using molecular techniques. This is the first step. You can't study the epidemiology of malaria without studying the ecology of the species that spreads it. And you can't translate this information into malarial control until you can separate the species because each one is potentially very different in its habits and life history." Wilkerson has been using a technique called random applied polymorphic DNA, or RAPD, along with PCR to sort out a complex of four species of South American Anopheles mosquitoes. "Because of DNA technology of several sorts, particularly PCR, there are big opportunities to carry out these phylogenetic studies," says Hoy. "It's a brisk trade, so to speak."

Bioengineered Bugs A 'Brisk Trade' In Applied and Basic Entomology

Beyond the Fruit Fly

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