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Bacterium SIZE MATTERS: Deer ticks -- vectors of Borrelia Burgdorferi -- are half the size of the common dog tick, which is not associated with Lyme disease. As a new generation of adolescent deer tick enjoys its first blood meal, scientists in the United States and abroad continue to focus their research efforts on understanding and preventing Lyme disease. Ticks infected with the bacterium Borrelia burgdorferi cause more than 10,000 cases of Lyme disease in the U.S. each year, according to the Centers for Disease Control and Prevention. Scientists specializing in immunology, epidemiology, parasitology, and even rheumatology are all pursuing lines of research they hope will lead to a better understanding of how B. burgdorferi causes Lyme disease and how people can best avoid infection. Researchers studying disease prevention cover a lot of ground, whether they're crawling through the underbrush tracking infected ticks in the field or conducting large-scale clinical trials to determine the efficacy of Lyme vaccines in humans. At the same time, immunologists and microbiologists are trying to determine how B. burgdorferi interacts with the human immune system to cause Lyme disease. Armed with funding from the National Institutes of Health, Lyme researchers are trying to unravel the basic biology of B. burgdorferi-its natural ecology, its genome sequence, and its slippery coat of surface proteins, among other areas of investigation. The life cycle of the Lyme disease vector-the deer tick Ixodes scapularis-is fairly well known. In the spring, nymphal deer ticks that survived the winter seek out tasty mammals for their first postlarval feast. Though adult female I. scapularis ticks prefer deer, the poppy-seed-sized nymphs will also feed on white-footed deer mice or humans. And any tick that fed last season on a mouse infected with B. burgdorferi-a corkscrew-shaped bacterium known to microbiologists as a spirochete-may pass Lyme disease on to a human this year. VEXING VECTOR: Ixodes scapularis, the carrier of Lyme disease -- the most common insect-borne disorder in the U.S. In the northeastern United States, particularly in Rhode Island, Connecticut, and Westchester County and Long Island n New York, scientists estimate that half the deer-tick population may harbor agents that cause illness, including Lyme disease and ehrlichiosis-another bacterial infection characterized by flu-like symptoms (story on page 16). Lyme disease is named after a community on the coast of Connecticut. The first cases in the U.S. were identified in 1975 by physicians Allen Steere and Stephen Malawista, researchers at Yale University School of Medicine. Scientists, however, believe that B. burgdorferi has been around for thousands of years. Examining well-preserved museum samples, parasitologist Sam Telford of the Harvard School of Public Health and David Persing at the Mayo Foundation in Rochester, Minn., have identified B. burgdorferi spirochetes in North American white-footed deer mice collected in the late 1800s. UNANSWERED QUESTIONS: Exactly how the Lyme spirochete causes disease in humans is unknown, observes Texas' Alan Barbour. But if the spirochete has lived in North America as long as humans, why didn't Lyme disease surface on the continent before the 1970s? Did B. burgdorferi become more infectious? No one knows. In fact, many fundamental questions about how the spirochete causes disease in humans are unanswered, says Alan Barbour of the University of Texas Health Science Center in San Antonio, author of Lyme Disease: The Cause, the Cure, the Controversy (Johns Hopkins University Press, 1996). At the most fundamental level, scientists have yet to determine exactly how B. burgdorferi causes disease in humans. By studying how the immune system reacts to the spirochete, immunologists and microbiologists may begin to understand why some infected people experience mild symptoms and others wind up with a chronic illness. EHRLICHIOSIS AND LYME DISEASE: PEAS IN A POD? Although most cases of Lyme disease respond to antibiotic treatment, occasionally the symptoms persist, and even worsen. A number of researchers are investigating why some patients wind up with chronic Lyme disease. Co-infection may have something to do with it, notes Sam Telford, a parasitologist at the Harvard School of Public Health in Boston. According to Telford, when they feed on infected white-footed mice, deer ticks can pick up the bacterium that causes human granulocyte ehrlichiosis (HGE), the spirochete that causes Lyme disease, or the protozoan that causes human babesiosis, an infection of red blood cells. And ticks with multiple infections can pass along all three diseases with a single bite (S. Telford et al., Proceedings of the National Academy of Sciences, 93:6209-14, 1996). "People don't realize that the tick that transmits Lyme disease also carries two other organisms because it feeds on infected white-footed mice," says Telford. "But I don't see why people are so surprised, because we've seen this kind of co-infection in Europe for many years," he adds. Telford and his colleagues collected Ixodes scapularis ticks from the backyard of a Nantucket patient with HGE. They found that an outdoor enthusiast walking through the woods for an hour could encounter perhaps a dozen ticks infected with Borrelia burgdorferi, the agent responsible for Lyme disease, and another four ticks that carried HGE. That same stroller might also run into two ticks that harbor both disease-causing bacteria. Ehrlichiosis and babesiosis share the flu-like symptoms characteristic of Lyme disease-the muscle aches, fever, headache, and fatigue. In addition, patients with babesiosis may also experience chills, night sweats, even depression. But patients infected with B. burgdorferi develop a distintive circular rash around the site of the tick bite. Still, because the risk of co-infection is high, Telford suggests that physicians who suspect that a patient has contracted one of these tick-borne diseases test for all of them. Without a proper diagnosis, physicians will be unable to treat these distinct diseases, notes Telford. Although each can be cured by antibiotics, Lyme disease responds best to penicillins, ehrlichiosis to tetracycline, and babesiosis to antimalarial medications. "These illnesses may linger because they're never diagnosed. So the disease is not treated at all," he says. Left untreated, chronic Lyme disease can damage the heart and nervous system. And ehrlichiosis can be fatal in some cases. Co-infection with a second tick-borne agent may also explain why Lyme disease varies in severity from patient to patient, says Telford. In a five-year study of 1,156 people living on Block Island in New England, Telford and his collaborators found that patients with both Lyme disease and babesiosis show more symptoms and are sick longer than patients with either infection alone (P. Krause et al., JAMA-Journal of the American Medical Association, 275:1657-60, 1996). Co-infection with babesiosis or HGE may mean the difference between feeling under the weather and being unable to get out of bed for a patient with Lyme disease, says Telford. An ounce of prevention beats a trip to the pharmacist for a cure. Avoiding tick bites remains the most reliable way to avoid Lyme disease, ehrlichiosis, and babesiosis, stress Lyme disease researchers. Telford prescribes insect repellents and nightly tick checks. -K.H. As the most common tick- or insect-borne disease in the U.S., Lyme disease research is well supported by the federal government. As a whole, NIH spent about $13.5 million on Lyme disease research in 1995, with the bulk of the funding coming from the National Institute for Allergy and Infectious Diseases. This level is expected to increase to just over $14 million in 1996. Though most cases of B. burgdorferi infection can be eliminated easily by a short course of antibiotics, Lyme disease can be disabling if left untreated. Scientists are searching for answers to how the relatively small numbers of bacteria (1,000 microbes per milliliter in infected humans) can cause excessive inflammation, and why the spirochetes localize to certain tissues, including the joints, the heart, and the brain. They suspect the key to the mystery lies on the spirochetes' surface, in the collection of proteins that coat its outer membrane. Patients with Lyme disease experience flu-like symptoms-aches, fever, nausea, joint pain. When the spirochetes first get under the skin, they mark their point of entry with a characteristic bull's-eye rash. Left unchecked, the B. burgdorferi bacteria will work their way into the bloodstream and head off to other organs. In late-stage Lyme disease, a month or two after infection, the spirochetes take up residence in the heart or the joints, where they cause inflammation. Others settle in the brain, where the body's immune system can't reach them as easily. Working with the related spirochete B. turicata, Barbour has found that bacteria that wind up in the brains of immune-deficient mice differ by only a single protein from the bacteria that remain in the blood. The protein, which shows homology to the outer-surface protein OspC from a variety of B. burgdorferi strains, may allow the spirochete to establish the kind of long-lasting brain infection that underlies lingering Lyme disease, he theorizes. According to Barbour, B. burgdorferi lacking its outer-surface proteins (including OspA, OspB, OspC, and OspD) is easily killed by the blood's complement system. Several groups of researchers are currently characterizing which proteins B. burgdorferi loses when it is kept in culture conditions for too long. When the spirochete is raised without contact with a mammalian host, it is unable to infect mice. And, according to microbiologists James Carroll and Frank Gherardini of the University of Georgia, it may also lose OspD and a handful of previously unidentified surface proteins (J.A. Carroll, F.C. Gherardini, Infection and Immunity, 64:392-8, 1996). Many researchers believe that the secret to B. burgdorferi's infectivity and inflammatory capacity lies in the interaction of its surface proteins with the host's immunological system. Yale researcher Stephen Barthold, a veterinarian and professor of comparative medicine who developed the first mouse model of Lyme disease, studies the expression of B. burgdorferi surface proteins throughout various stages of the spirochete's life cycle. He finds that during the early stages of infection, B. burgdorferi avoids immune detection by decreasing its expression of surface proteins or cloaking its expressed surface proteins under a layer of slime. "It's using some sort of stealth-bomber-type mechanism," he says. Or, using another diversionary tactic called blebbing, the spirochete can pinch off bits of its membrane in order to release its surface proteins. Explains Barbour: "It's like a bacterial Star Wars defense program," in which released surface proteins might intercept incoming host antibodies, keeping the spirochete safe from immunological attack. TINY CREATURES: In size order, the three tick feeding stages are nymph, larva, and adult, shown next to a U.S. dime. During the early stages of infection, perhaps while the spirochete is still inside the tick, B. burgdorferi switches on a whole new family of surface proteins that may help it adapt to life in its mammalian host, according to Barthold. These so-called in vivo expressed antigens may help the spirochete invade tissues or elicit inflammation in the joints (S.W. Barthold et al., Laboratory Investigation, 74:57-67, 1996). Although they can offer educated guesses, researchers are still unsure what roles the surface proteins of B. burgdorferi play in infection and disease. A group of protein chemists and crystallographers at Brookhaven National Laboratory in Upton, N.Y., and the State University of New York, Stony Brook, are taking a more structural approach. John Dunn and Cathy Lawson purified and crystallized OspA-a surface protein that B. burgdorferi expresses when it lives in the midgut of the tick. The structure might offer some clues to how the protein functions, says crystallographer Lawson. Further, because the Lyme disease vaccines now being tested in humans contain OspA, Lawson says, "it would be nice to know what the protein looks like." The researchers presented their results at the seventh International Conference on Lyme Borreliosis, held in San Francisco late last month. The biennial meeting draws more than 400 Lyme researchers from around the world. LYME DISEASE RESOURCES American Society for Microbiology 1325 Massachusetts Ave., N.W. Washington, D.C. 20005-4171 Tel: (202) 737-3600 Fax: (202) 942-9340 Executive Director: Michael Goldberg President: Kenneth Bernes E-mail: Call for appropriate address http://www.asmusa.org Journals: Infection and Immunity and Journal of Clinical Microbiology American Lyme Disease Foundation Mill Pond Offices 292 Route 100 Somers, N.Y. 10589 Tel: (914) 277-6970 Tel: (800) 876-LYME http://www.w2.com/docs2/d5/lyme.html E-mail: aldf@computer.net Centers for Disease Control and Prevention U.S. Department of Health and Human Services 1600 Clifton Rd., N.E. Atlanta, Ga. 30333 Tel : (404) 639-3311 Fax information service: (404) 332-4555 You can request any of the following documents by fax: Lyme disease-general information #351701 Lyme disease-symptoms #351702 Lyme disease-treatment/prevention #351703 http://www.cdc.gov/diseases/diseases.html Electronic Newsletter: LymeNet is a monthly electronic newsletter that carries reports on scientific meetings on Lyme disease. To subscribe, send a message to listserv@lehigh.edu. In the body of the message, type: Subscribe LymeNet-L Your Name. USENET group: Anyone interested in joining a discussion group on Lyme disease can log onto sci.med.diseases.lyme Understanding when each surface protein gets expressed may be important for developing an effective vaccine for Lyme disease. The OspA vaccine can combat infection only when OspA is present on the surface of B. burgdorferi, when the spirochete is still in the gut of the tick. Once the tick takes a blood meal, the spirochete gets rid of its OspA in preparation for the transfer to its mammalian host. Thus, the OspA-based vaccines cannot prevent Lyme disease once an animal, or a human, has been infected with B. burgdorferi. In fact, the OspA vaccine, which protects mice against a challenge with the infective spirochete, destroys B. burgdorferi while it's still inside the feeding tick, says Barthold. Two large-scale, Phase III clinical trials of OspA vaccines in humans are being conducted by researchers at Connaught Laboratories in Swiftwater, Pa., and SmithKline Beecham in Philadelphia. Some 10,000 individuals received injections of lipidated OspA protein; the researchers will soon analyze the data to determine whether the vaccine protected recipients against Lyme disease. The results from the Phase II clinical trials indicate that the vaccine is safe, but may cause local irritation at the site of the injection. This irritation may be one of the problems facing researchers developing a Lyme vaccine. Because the disease can be cured with antibiotics or simply avoided by limiting exposure to ticks, Barbour thinks that the public may demand a vaccine that has absolutely no side effects. To further complicate matters, researchers don't know how often people would need to be re-vaccinated. To remain protected, a person must have a certain level of circulating OspA antibody in the blood, says Steere, now at the New England Medical Center and Tufts University School of Medicine in Boston. BIRTH CONTROL: Yale's Durland Fish advocates tick population management as a disease-prevention strategy. But that opens questions about who should get vaccinated. "Does Uncle Charlie get vaccinated when he visits you in Westchester County [in New York]?," asks Durland Fish, an entomologist and epidemiologist at Yale. "It'd be like preparing for a trip [overseas]." While several groups of researchers continue to pursue a Lyme vaccine, others think that the key to preventing the disease lies in controlling the ticks themselves. "The bottom line? You need to get rid of the ticks," says Fish, who preaches the power of pesticides. "It amazes me that people would prefer to have a foreign antigen injected into their bodies rather than have a chemical sprayed onto their lawns," he says. With tick territories expanding along with deer populations, Fish advocates using insecticides to stop ticks from moving into new areas. Other researchers at the University of Rhode Island are developing biologicals-searching for ways to use the ticks' natural enemies, including certain fungi, to keep their population under control. Nature may present researchers with other clues for tick control. In the Northeast, tick populations fluctuate naturally-rising and falling from year to year. Why? "We don't have a clue," acknowledges Fish. But the number of reported cases of Lyme disease seems to correlate with number of infected ticks, he notes. After spending 11 years tracking B. burgdorferi in mice and ticks on Nantucket, scientists are no closer to being able to make yearly predictions on the potential for incidences of Lyme disease. But Telford says that the number of cases of Lyme disease does correlate with the weather: Rain usually keeps people indoors, thus limiting their exposure. While scientists continue to work toward developing effective ways of preventing B. burgdorferi infection through vaccines or tick-population management, they advise the public to use common sense to avoid getting bitten by a hungry deer tick. For those heading for the hills or the woods, they recommend using insect repellents, wearing long sleeves, and tucking trouser cuffs into socks. "It really works," says Telford. In fact, the trouser-tuck is even de rigueur for parasitologists beating the bushes for ticks: "I figure I've crawled through the bushes for eight- or ten-thousand hours looking for ticks," asserts Telford, "and I've never come down with anything." Karen Hopkin is a freelance science writer based in New York City and senior producer for National Public Radio's "Science Friday." She created the "Studmuffins of Science" 1996 calendar and is online at khopkin@npr.org.

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