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Science  21 Mar 1997:
Vol. 275, Issue 5307, pp. 1736

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  1. Astronomy

    To Catch a WIMP

    1. Andrew Watson

    These heavy, secretive particles—if they exist—could make up the bulk of the universe. Around the world, physicists are setting traps to find out

    If Secretary of the Treasury Robert Rubin announced one day that he could not account for 90% of the $6 billion worth of gold held at Fort Knox, he would have some explaining to do. The world's astronomers have a roughly equivalent problem, only theirs is about mass, not money. For the past 20 years, astronomers have had compelling evidence that there is more to the universe than meets the eye: About 90% of the mass of the universe seems to be invisible. This “dark matter” could consist of dim stars, not bright enough to be seen from Earth. It could simply be neutrinos, infinitesimal particles that zip around the universe in great profusion, rarely interacting with matter. But there is a more exotic alternative, harder to prove but increasingly appealing: that the dark matter is made up of massive particles yet to be discovered, collectively known as WIMPs (weakly interacting massive particles).

    Lighting up dark matter.

    Each spike represents a galaxy in a cluster in Pisces; the mound around them is dark matter with a mass 250 times more than all the galaxies combined.

    G. Kochanski et al./Lucent Technologies' Bell Labs

    This is not just the stuff of theoretical debate. Astronomers are scouring the perimeter of our galaxy for unseen stars and scrutinizing the neutrino to see whether it could make up the mass deficit. But as such searches look less and less likely to offer a full answer to the dark-matter puzzle, researchers are also setting traps for WIMPs, which might be drifting continuously through Earth. This year, what was a trickle of WIMP-hunting experiments will swell to a respectable stream as a new generation of high-technology WIMP detectors joins the effort. A U.S.-based experiment that monitors crystals at close to absolute zero temperature for the tiny pulse of heat that might signal a WIMP was expanded last month, and this month a more sensitive competing experiment will debut at Italy's underground Gran Sasso laboratory. Some 20 other WIMP searches are on their way, aiming to snare these phantoms in everything from grains of tin to tiny droplets of Freon.

    Success is a long shot, the researchers concede. WIMPs are, as their name implies, loath to interact with ordinary matter, and their properties—and thus the best way to trap them—are largely unknown, all of which makes WIMP-hunting “a very difficult business,” says Oxford University physicist Susan Cooper, a leader of the Gran Sasso project. “[A WIMP] doesn't have a clear signature,” she laments. “It's not the kind of experiment that you really like to do, because it is so hard, but I feel we are forced to try because the question is so big.”

    Where's the mass?

    Dark matter may be elusive, but the evidence that it exists is strong. About 2 decades ago, astronomers first began measuring the speeds at which hydrogen clouds orbit the centers of spiral galaxies. The velocities turned out to be too high for the gravitational pull of the visible stars in the galaxies to hold the clouds in their orbits. Astronomers concluded that a large but invisible source of gravitational pull in the outer reaches of these galaxies must also be at work.

    Gravitational lensing provides even more persuasive evidence. Light reaching Earth from distant galaxies is sometimes distorted by the gravitational pull of a cluster of galaxies that lies in the path of the light. By analyzing the distorted image, astronomers can “weigh” the cluster, and “that very convincingly shows that there's lots and lots of dark matter in the cluster,” says Oxford astronomer Will Sutherland.

    At least some of that dark matter, says Sutherland, could consist of the so-called massive compact halo objects (MACHOs), a catchall term embracing brown dwarfs, old white dwarfs, neutron stars, and any other massive and invisibly dull star. Sutherland and colleagues in Australia and the United States are part of the MACHO Collaboration, one of several teams looking for changes in the brightness of distant stars caused by gravitational lensing due to MACHOs in the halo of our own galaxy.

    The MACHO Collaboration announced its discovery of a handful of MACHO candidates last year, but most researchers, even those actively hunting MACHOs, expect that these objects will only make a modest contribution to the total of dark matter, given the dictates of cosmology. Current theories of how matter formed after the big bang limit the amount of normal or “baryonic” matter—the protons and neutrons that make up both normal stars and MACHOs—to about 10% of the mass required to gradually slow the universe's expansion—the type of universe cosmologists currently favor.

    If the cosmologists are to be believed, this means that the rest of the matter must be something more exotic. Neutrinos are one possibility for this nonbaryonic matter. Some theorists speculate that they are endowed with a minute mass, tens of thousands of times smaller than the mass of an electron. Yet, despite decades of trying, experimenters have yet to pin a mass on the neutrino. And cosmological models based on neutrino dark matter predict large groupings of galaxies not seen in the real universe. So many are now turning to WIMPs—heavyweight relics of the big bang—which may match particles predicted by some exotic theories in particle physics such as supersymmetry.

    WIMPs have fewer strikes against them so far than the other dark-matter candidates—except that they are entirely hypothetical. They also provide a unique challenge for experimentalists: building a detector for a particle you do not know for certain exists, with unknown properties. To make matters worse, WIMP detectors risk being flooded with known particles, such as cosmic rays and debris from radioactive decays of atoms around the experiment and in the apparatus itself. Says Cooper: “Any signal that is seen is liable to be a bit shaky, and it's very important to have the possibility of confirming it with different techniques.”

    One technique has already been tested over the last several years in an earlier generation of WIMP searches. It relies on scintillation: the tiny pulse of light given off when an incoming particle strikes an atom in certain crystals, such as sodium iodide. The U.K. Dark Matter Collaboration (UKDMC) runs a scintillation detector in Europe's deepest mine, the Boulby salt and potash mine in northern England, where spurious signals from cosmic rays are at a minimum. The experiment, led by Peter Smith of the Rutherford Appleton Laboratory, consists of a 6-kilogram crystal of sodium iodide watched by a pair of light-detecting photomultipliers. Running since 1994, the detector has established upper limits on the frequency with which WIMPs of various masses knock into the detector.

    Meanwhile, in the Gran Sasso laboratory beneath the Apennine mountains of central Italy, the DAMA group, led by the University of Rome's Rita Bernabei, has a similar 115.5-kilogram sodium iodide detector. To improve their chances of capturing a WIMP, the team is planning to scale up to 1 ton of sodium iodide in the near future. And both DAMA and the U.K. group are currently working on new detectors with liquid xenon in place of the sodium iodide crystals, which is expected to improve sensitivity.

    View this table:

    Small is beautiful

    Many of the new detectors, however, are based on a different strategy which should raise the odds of catching a WIMP: detecting an incoming WIMP not by scintillation but by the energy it deposits in the detector material. “The amount of energy, by room temperature thermal standards, is absolutely insignificant,” explains Tom Shutt of the University of California, Berkeley. But if you cool your crystal to a temperature of 0.020 kelvin, a single particle depositing a few kiloelectron volts will cause a millionth-of-a-degree temperature rise, “which, it turns out, is quite measurable,” Shutt says.

    One set of these detectors, called bolometers, is already keeping watch for WIMPS. The Cryogenic Dark Matter Search (CDMS) experiment, headed by Berkeley's Bernard Sadoulet, has had two detectors stationed a few meters underground at an old particle-accelerator complex at Stanford University since last September and added a third detector just weeks ago. Running at a temperature of a few tens of millikelvins, they consist of between 100 and 200 grams of crystals of germanium or silicon. An incoming WIMP knocks into a crystal nucleus, which recoils and creates heat.

    To distinguish between WIMPs and cosmic rays, particles that constantly rain down on Earth from space, the CDMS detectors can also detect the charged particles—electrons or nuclei—dislodged in the crystal by a particle impact. “That's very important, because essentially all the radioactive background interacts with the electrons, and the signal that we are looking for interacts with nuclei,” says Sadoulet. “In the end, we identify the amount of charge produced by each particle that strikes our detector, and thus distinguish WIMPs from background [gamma-ray] photons,” adds Shutt. Soon, says Sadoulet, the experiment will move to the Soudan mine in Minnesota, where it will be better shielded from cosmic rays.

    By the time it does, a host of other cryogenic experiments should be under way. Makoto Minowa and his colleagues at the University of Tokyo are building a WIMP detector based on measuring the temperature rise in ultracold lithium fluoride. “An array of eight pieces of 20-gram lithium fluoride bolometers is now ready for measurement,” says Minowa, and it will soon be installed in the Nokogiri-yama underground laboratory in Tokyo. And the French EDELWEISS (Experience pour Detecter les Wimps en Site Souterrain) collaboration, is building a germanium bolometer in the Fréjus underground laboratory in the Alps, which, like the Berkeley device, will be sensitive to both the charge signal and the temperature rise from an impinging particle.

    Telltale transitions

    Lighter, slower WIMPs would deposit less heat in a bolometer, a possibility that has encouraged some groups to develop new, more sensitive thermometers. One is the closest competitor to the Berkeley experiments, the CRESST (Cryogenic Rare Event Search with Superconducting Thermometers) project run by Oxford's Cooper and her collaborators from Munich. Currently being nursed into action at Gran Sasso, the detector consists of four 262-gram sapphire crystals with a small rectangular patch of tungsten film stuck to each one. The very low temperature to which the detector is cooled lies just below the temperature at which tungsten undergoes a transition from a normal conductor into a superconductor. At this temperature, tungsten's electrical resistance is extremely sensitive to temperature, so the tiny temperature rise produced by an impacting WIMP will result in a detectable change in the film's resistance. The detector should be able to sense just 500 electron volts, the energy of a single soft x-ray photon. “We're specifically most sensitive to low-mass WIMPS,” says Cooper, “complementing many other experiments.”

    A team of physicists from Paris and Saragossa, Spain, is building a smaller version of CRESST, dubbed ROSEBUD (Rare Object Searches with Bolometers Underground), in the Canfranc underground laboratory in the Spanish Pyrenees, with data taking expected to commence next year. But there are other ways of exploiting superconductivity to catch a WIMP. The Swiss ORPHEUS team is building a detector containing billions of tin granules, each just 30 micrometers in diameter. A total of more than a kilogram of the grains are cooled to the superconducting transition temperature for tin and swathed in a magnetic field, according to ORPHEUS leader Klaus Pretzl of the University of Bern. An incoming WIMP will strike a nucleus in a granule and warm it up. “When the granule is heated up, it then makes a transition from the superconducting phase to the normal conducting phase … [creating a magnetic signal which] can be detected with a magnetometer,” says Pretzl. The team expects to install the detector beneath the city of Bern at the end of the year.

    One advantage of the magnetic “thermometer” strategy, says Tom Girard of the University of Lisbon, is its ability to reject spurious signals from natural radioactivity. A WIMP will strike only a single grain of tin, while radioactive background will cause transitions in a streak of grains, giving a larger blip in the magnetometer, he says. Girard and other researchers from Saragossa, Lisbon, and Paris are mounting a magnetic experiment resembling ORPHEUS, called SALOPARD, which will be installed in the Canfranc laboratory in a year or so. “We estimate the ability to reject 97% of the background contribution,” says Girard.

    Two other groups hope to eliminate spurious signals by exploiting different phase transitions at higher temperatures. Both teams, one in Canada and one a CERN-Lisbon-Paris collaboration, use liquid Freon droplets, a few tens of micrometers in diameter, entombed in a clear polymer gel. At room temperature, above Freon's boiling point, the droplets are confined in an unstable superheated state. If an impacting WIMP deposits enough energy in a Freon droplet, “there is a sudden phase transition during which the droplet is vaporized and expands into a bubble of Freon gas of about 1 millimeter in diameter, which is contained at its location in the polymer,” says Viktor Zacek of the University of Montreal, spokesperson for the Canadian group. The result, says Juan Collar of CERN, a member of the competing team, “is a characteristic audible sound emission that can be picked up when this happens.”

    Such detectors “are totally insensitive to low-energy photons, the main source of background in dark-matter searches,” Collar adds. Most background radiation does not deposit enough energy in a sufficiently short distance for the superheated bubbles to notice. “So life is much easier for the WIMP hunter,” says Collar. Last month, the Canadian team started running a prototype system based on just a few grams of Freon droplets. Larger systems are in the pipeline, to be installed in the Creighton mine in Ontario. The European team, says Collar, plans to install its prototype in a shallow tunnel near Paris this year.

    In spite of all the impressive technology being deployed, there remains the possibility that all these searches may draw a blank. WIMPs could simply be a fix conjured up by astronomers and cosmologists to get their theories to match what they see in the universe around them. But Sadoulet says the history of physics shows that what appears at the time to be a “fix” can later look like prescience. He points to the difficulties faced by Neils Bohr and his contemporaries in the early 1930s as they struggled to understand radioactive beta decay, in which some energy seemed to simply vanish. Bohr proposed dumping the principle of energy conservation, while Wolfgang Pauli proposed that the energy was fleeing in the form of a ghostly new particle, purely hypothetical at the time—the neutrino. The rest is history.

    Andrew Watson is a writer in Norwich, U.K.

  2. Astronomy

    Gamma-Ray Source in Distant Universe?

    1. Govert Schilling,
    2. Susan Biggin

    Discovering where a crime was committed isn't the same as solving it. But it's a good start, especially when one possible crime scene is just around the corner and the other is at unimaginably large distances. The crime, in this case, is a celestial act of violence called a gamma-ray burst—a bright flash of extremely energetic radiation from a mysterious source, at a random position in the sky. Just hours after astronomers had narrowed down the position of one burst, Jan van Paradijs of the University of Amsterdam and the University of Alabama, together with Paul Groot and Titus Galama of Amsterdam, pointed a telescope at the site and spotted what may be the source: a dim galaxy in the far reaches of the universe.

    Scoping out a burst.

    The William Herschel telescope.

    Isaac Newton Group

    “The key to [finding burst sources] is to observe the suspect region with a large optical telescope within 24 hours after the event,” says van Paradijs. If he and his colleagues really have pinpointed a burst source, they will have taken a major step toward solving a decades-old mystery. Over the last 30 years, space-based detectors have picked up hundreds of these gamma-ray flashes, but astronomers disagree about what might be producing them and even where they come from. According to the “local” hypothesis, violent events on neutron stars—the dense relics of massive stars—in the neighborhood of our own galaxy are responsible. The competing “cosmological” hypothesis holds instead that gamma-ray bursts are even more violent events in distant galaxies.

    Neither view could prevail because the poor directional resolution of gamma-ray detectors kept astronomers from linking the bursts to any identifiable objects. That may have changed, thanks to a series of observations beginning 28 February, when the Italian-Dutch satellite Beppo-SAX detected a burst and then spotted a source of x-rays at the burst position. The x-ray observation enabled astronomers to pinpoint the event to a patch of sky less than an arc minute across—about a thirtieth of the width of the full moon (Science, 14 March, p. 1560).

    Only 21 hours after the burst, van Paradijs and his colleagues made an image of the stars and galaxies in this tiny patch of sky with the William Herschel and Isaac Newton telescopes at the Roque de los Muchachos Observatory on the Canary Islands. Eight days later, they made a second image—and found that one point of light in the field had faded. On 13 March, Griet van der Steene of the European Southern Observatory took a close look at the object, by then very faint, with ESO's 3.5-meter New Technology Telescope at La Silla, Chile, and discerned a small, dim galaxy.

    Van Paradijs and his colleagues, who announced their discovery late last week in a circular of the International Astronomical Union, argue that “the position and rapid decline [of the galaxy] contemporaneous with that of the Beppo-SAX x-ray transient indicate that the two are related.” Whether the x-ray source really corresponded to the original gamma-ray burst is a bit less certain, says van Paradijs, but the link “smells good.” That would spell the death of the local hypothesis, he adds, noting that he himself has favored that view in the past. Cambridge University astronomer Martin Rees agrees that the Dutch observations “strongly tilt the balance in favor of the cosmological hypothesis,” although he adds that it would take several more observations of the same kind to settle the question.

    The burden would then be on theorists to explain what could produce a flash of gamma rays so powerful that it can be seen from the far reaches of the universe. Many believe that only the collision of two neutron stars could do the job. Neutron stars orbiting each other lose energy by emitting gravitational waves and gradually spiral together. This fatal attraction inevitably leads to the death of both, in a terrible crime passionelle double.

    But as to how the energy of the cataclysm could be turned into gamma rays, “there are hardly any serious theories,” says Frank Verbunt, an x-ray and high-energy astrophysicist at the University of Utrecht in the Netherlands. It will take more efforts like this one, combining gamma-ray, x-ray, and optical observations, to produce a full picture of the crime.

    Such efforts may be hampered, however, by an ailing Beppo-SAX. Two of the six gyroscopes that stabilize the satellite have failed, one in December and one in January, and a third is faltering. Officials at the Italian Space Agency (ASI) are playing down the problem, insisting that they can control the satellite by other methods if more gyros fail. “It's no big deal. … The mission could even continue without the gyroscope system,” a senior ASI scientist told Science.

    Schilling and Biggin are writers in the Netherlands and Italy, respectively.

  3. Virology

    First Genes Isolated From the Deadly 1918 Flu Virus

    1. Elizabeth Pennisi

    The great flu pandemic of 1918 was nothing to sneeze at. Although rarely more than a footnote in modern history books, it killed 20 million to 40 million people worldwide, 675,000 in the United States alone. That's 10 times more than the country lost in World War I, which raged at the same time, and almost twice the total number of U.S. deaths caused by the 15-year-long AIDS epidemic. Now, one of the pandemic's victims, a 21-year-old Army private who had been stationed in Fort Jackson, South Carolina, has given modern researchers their first direct look at the culprit.

    National Library of Medicine

    Global killer.

    The 1918 flu filled hospitals, decreased life expectancy, and (right) damaged lungs.

    Taubenberger et al.

    On page 1793, pathologist Jeffery Taubenberger, molecular biologist Ann Reid, and their colleagues at the Armed Forces Institute of Pathology in Washington, D.C., report that they have isolated pieces of five flu genes from the private's lungs. While the genes collected so far portray the killer as a run-of-the-mill swine flu, its deadly secrets may emerge as Taubenberger and Reid collect more of the viral genome and other researchers apply the same methods to other preserved tissue from the 1918 epidemic.

    This report “probably won't make as big a splash as cloning that sheep, [but] if we can get a handle on what caused those deaths, that is important,” says historian Alfred Crosby, author of the book, America's Forgotten Pandemic: The Influenza of 1918. Knowing what a killer flu virus looks like, for instance, could enable health officials to spot particularly dangerous flu strains as they emerge and “help us prepare for what we need to be prepared for,” says Nancy Cox, a virologist from the Centers for Disease Control and Prevention (CDC) in Atlanta.

    For the current work, Taubenberger and Reid began with several dozen samples of lung tissues taken from soldiers who died of the 1918 flu and preserved in their institute's archives. These, they realized, could be analyzed by modern polymerase chain reaction (PCR) methods, which can amplify extremely small quantities of DNA and RNA, thus providing sufficient amounts of material for sequence analysis. From the beginning, though, the researchers knew success was a long shot.

    Typically, the flu virus infects the respiratory system, replicates, and then is shed into the air via the lungs, all in just a few days. Thus, unless one of the soldiers had died very soon after infection, all traces of the virus would probably have vanished by the time an autopsy was performed. What's more, the flu virus stores its genetic information on single strands of RNA, which is much more susceptible to degradation by cellular enzymes than DNA. “So you have to look for little pieces,” Taubenberger explains. These problems had already proved insurmountable once before. Several years ago, virologist Robert Webster of St. Jude Children's Research Hospital in Memphis, Tennessee, had failed in an attempt to pull genetic material from archived samples.

    To increase their chance of finding viral RNA, the researchers worked for a year refining their PCR procedure. In addition, they carefully examined each of the lung-tissue specimens under a microscope to find ones with indications of viral infections, such as lung air spaces filled with fluid and blood and damaged cells in the bronchial epithelial linings. Of the 28 tissue samples examined, Taubenberger says, the private's “was the only case with all the features of viral pneumonia.”

    His was also the only specimen in which they could identify flu-virus RNA sequences, including segments of five genes: those encoding hemagglutinin and neuraminidase—proteins that help the virus get into a cell—and three structural proteins. From the hemagglutinin and neuraminidase gene fragments, the researchers were able to confirm earlier conclusions about the virus's source.

    Influenza viruses are thought to pass into pigs from birds, usually ducks, that are kept in close proximity to swine in some parts of the world, particularly China. Subsequent genetic changes enable the virus to move from pigs into people. However, there had been some speculation that the 1918 flu virus might have been so deadly because it was an avian virus, and these data indicate that isn't the case.

    Because the flu-virus genes from the private closely resemble those of viruses isolated from pigs, the researchers concluded that his virus had been in the pig population for a while. That is also in line with previous antibody studies of blood taken from people who had lived through the 1918 pandemic, indicating that this killer plague was a classical swine flu. “What this says is we had better watch what's happening in the pig populations of the world,” Webster says, because it seems that ever-changing flu viruses in those animals are the source of new flu viruses in humans. Currently, both the CDC and the World Health Organization devote most of their flu-surveillance efforts to monitoring for new flu strains in people, not animals.

    The work does not support another supposition about the virus, however. Virologists have suggested that the extreme virulence of the 1918 flu virus may have resulted from the same genetic change that enabled an avian flu virus to kill whole flocks of chickens overnight in Mexico 2 years ago (Science, 17 March 1995, p. 1594). The deadly mutation altered a segment of the hemagglutinin protein, probably increasing the virus's ability to infect cells. But Taubenberger's group found no such change in the specimen they examined.

    Hoping to find more clues to these and other mysteries, such as why young adults, usually the most resistant to flu infections, were the hardest hit in 1918, Taubenberger and Reid are continuing to isolate more of the viral RNA from the private's tissue samples. At the same time, they plan to use PCR to make lots of copies of all the viral RNA bits they recover. From these, they will create a cDNA library so as to have an unlimited supply of that genetic material.

    Meanwhile, Taubenberger hopes that his success will inspire others to comb their archives for other possible samples. Already, before learning of the current work, virologist John Oxford of London Hospital Medical College had begun collecting samples from his own institution and from Australia and Prague for a similar analysis. And a Canadian-led team hopes to dig up seven miners who presumably died of the 1918 flu and have been preserved in their graves in the frozen ground near Spitzbergen, Norway.

    Webster worries about the slim chance that those bodies will contain live virus. But it may be worth the risk, he says: “We want to know what killed these people. The potential is there for this kind of virus to return.”

  4. Biodiversity

    Microbiologists Explore Life's Rich, Hidden Kingdoms

    1. Robert F. Service

    To most people, biodiversity means plants, animals, or maybe insects. These are the organisms that taxonomists' tallies put at the top of the numbers game, with more than 248,000 described species of plants, 750,000 species of insects, and 280,000 species of other animals. But these counts are less a reflection of the true biological richness of life on Earth than of our ability to count what we can see, such as differences in the shapes of leaves and fins, and the colors of feathers.

    Dual view.

    Taxonomists' counts (pie chart) suggest that insects dominate the diversity game, but new analyses reveal that microbes are the real winners.

    Now, a new way of counting species is dethroning plants and animals. Using gene-typing techniques that directly sample and compare gene sequences from different organisms, a new breed of biological classifiers is finding that the million-plus known plants and animals represent just a smidgen of life's genetic diversity. The bulk of the diversity, it turns out, lies not with azaleas, ants, and aardvarks but with single-celled microbes. Researchers have turned up dozens of groups of bacteria, archaea, and single-celled eukarya that are at least as genetically distinct from each other as animals are from plants, enough to qualify them as separate kingdoms of life. Such findings suggest that other such kingdoms and untold millions of individual species may still lie hidden in the microbial world. “It's like we're going into the Amazon basin for the first time,” says David Stahl, a microbiologist at Northwestern University in Evanston, Illinois.

    Although much of the work is still preliminary, already the gene surveys are turning up surprises. Scientists have found, for instance, that some bugs once thought to live only in extreme environments, such as hot springs, in fact, can make a living in decidedly nonextreme waters off southern California. The research also is challenging several long-held assumptions about microbial life, including the notion that many common bugs can be found worldwide.

    This effort is bringing together two types of scientists who have rarely before collaborated—molecular biologists who study the inner workings of individual organisms, and microbial ecologists who study how communities of microbes work in their natural settings. Even before this staggering wealth of diversity came to light, researchers recognized that, as Gary Olsen, a microbiologist at the University of Illinois puts it, “microbes are really the movers and shakers in the world,” providing nutrients that sustain all other forms of life. But with the new molecular techniques, these scientists finally are getting glimpses of how human impacts, ranging from oil spills to fertilizer runoff, may be perturbing microscopic communities. Such changes will not “necessarily [precipitate] the collapse of the biosphere today or tomorrow, but the consequences are global in reach and very real,” says Olsen.

    Cultural bias. Researchers always knew that microbes are plentiful; even ordinary microscopes are powerful enough to show that a pinch of soil can contain 1 billion microbes or more. But characterizing the biodiversity of this thimble-sized rain forest, much less describing its ecological structure, was virtually impossible: Unlike macroscopic creatures, which can be categorized by their shapes, colors, and other features, microorganisms defied taxonomic breakdown. “Two microbes could be as different from each other as a grizzly bear from an oak tree, and you'd never know it,” says Stahl.

    To get some sense of microbial diversity, scientists have traditionally tried to isolate organisms from the field and then grow them up in culture. But only about 1% of bacteria can be grown in culture, says Ken Nealson, an environmental microbiologist at the University of Wisconsin, Milwaukee. This approach has given researchers a “horrendously skewed picture of the diversity of bacteria” and other microorganisms because it essentially ignores organisms that can't be cultured.

    Scientists now are bypassing the petri dish and directly characterizing microbial diversity with sophisticated analyses of DNA borrowed from biology's molecular revolution. While these techniques have their origins several years back, “it's only now that this effort is beginning to enter [a period of] exponential growth,” with many groups becoming involved, says Stahl.

    The most widely used technique for gauging diversity relies on the sequence of about 1500 base pairs in a gene that is common to all cells. The gene codes for the so-called 16S subunit of the ribosome, a chunk of cellular machinery that assembles proteins. The 16S sequences of two related organisms—say, humans and chimpanzees—typically are quite similar, while those from two organisms that have been evolving separately for a long period—say, fungi and cyanobacteria—are more distinct. By comparing the 16S sequence of an unknown microbe to those of its unicellular brethren, researchers can gauge how closely it is related.

    Because the 16S sequence evolves very slowly, the technique is good for making broad distinctions. For instance, University of Illinois evolutionist Carl Woese and his colleagues used this technique in the late 1970s to determine that the evolutionary tree of life is divided into three main trunks—bacteria, archaea, and eukaryota—a picture that gained definitive support last year when researchers compared the full genome sequences of organisms in each of the three domains (Science, 23 August 1996, p. 1043).

    In recent years, researchers around the globe have been filling a common database with 16S sequences from thousands of other organisms, including microbes that have been cultured previously, as well as organisms plucked from backyards and arctic lakes, among other settings. While the main goal of this database, maintained by researchers at the University of Illinois, Urbana-Champaign, is to gauge the evolutionary relationships between organisms, it is also proving to be a handy catalog of microbial diversity. The database shows “it's much more diverse out there than we ever suspected,” says University of Maryland microbiologist Rita Colwell.

    Indeed, on land, virtually no two samples turn up with 16S fragment matches. For example, Erko Stackebrandt and his colleagues at the German Collection of Microorganisms and Cell Cultures in Braunschweig recently sequenced 16S fragments from microbes taken from an Australian forest soil and a German peat bog. Their analysis showed that neither of the two sets of more than 150 sequences included any previously recorded sequences, says Stackebrandt. In a related study, Michigan State University microbiologist James Tiege and his colleagues sampled similar soils in California, South Africa, Chile, and Australia for bacteria that metabolize a chlorine-containing compound called 3-chlorobenzoate and found that the organisms on each continent had different genetic makeups.

    What is the evolutionary reason for all this diversity? It may simply be a function of the plethora of microhabitats in soil, says Edward DeLong, a microbiologist at the University of California, Santa Barbara (UCSB). For instance, while one side of a pebble may be exposed to oxygen, the other may not, thereby fostering the growth of aerobes on one side and anaerobes on the other.

    In any case, this rich diversity evident in land-based studies casts doubt on a basic tenet of microbial ecology—that common microbes travel widely, whether as spores or in dormant states via wind or water. If, as the studies suggest, bugs are endemic to specific regions, the total number of microbial species could be staggering, says Jim Staley, a microbiologist at the University of Washington, Seattle. Still, microbes may prove to be more cosmopolitan than these preliminary surveys suggest, says Stackebrandt. Even though he and his colleagues didn't find exact matches in their samples from Australia and Germany, they did find many that were related, he says. He adds that his team may be encountering a sampling problem. Perhaps, the number of species in each environment is so great that the chance of spotting overlap between two samples is slight.

    Microbes in the ocean, by contrast, may be more cosmopolitan—perhaps because seawater mixes relatively freely. Stephen Giovannoni, a marine microbiologist at Oregon State University in Corvallis, and colleagues recently sampled Atlantic and Pacific waters, and found that of 440 different bacterial genes sequenced, 86% come from just eight phylogenetic groups. But other studies point to a different conclusion. Staley and his University of Washington colleagues sampled waters in ocean waters of the Arctic and Antarctic, and although they found some similar organisms, “we couldn't find any that were the same species,” says Staley.

    A few surprising faces are showing up in these diverse crowds of microbes. One remarkable discovery is that a new class of organisms, known as crenarchaeota, exist in Pacific and Atlantic ocean waters. Crenarchaeota were long thought to live exclusively in extreme environments, such as hot springs where most are anaerobes and depend on sulfur for energy production. In 1992, however, two research groups—Jed Fuhrman and his colleagues at the University of Southern California in Los Angeles, and DeLong, then at the Woods Hole Oceanographic Institution in Massachusetts—isolated a 16S ribosomal sequence from crenarchaeota from several sites off southern California, Oregon, and Massachusetts. And 2 years later, DeLong led a team along with UCSB microbiologist Barbara Prézelin that found that these organisms account for as much as 30% of microbes in the Antarctic oceans. “That really opened people's eyes that [the crenarchaeota] are not just for hot springs anymore,” says Susan Barns, an environmental microbiologist at the Los Alamos National Laboratory in New Mexico.

    Indeed, the crenarchaeota have begun turning up everywhere. In a flurry of reports since December, teams have reported finding crenarchaeota in Illinois marsh sediments, Michigan lake sediments, and Wisconsin soil. The abundance of these organisms suggests that they may play a key role in the cycling of nutrients through the biosphere, says DeLong. But because cool-living crenarchaeota have yet to be cultured, researchers are still struggling to figure that out.

    It is on precisely this question—the ecological function of microbial movers and shakers—that a 16S sequence analysis can fall short. By itself, sequence data do not tell researchers much about how an organism makes a living in a natural setting. Still, sequence similarities between newly discovered organisms and microbes with well-characterized biochemistries can sometimes hint at the function of newcomers. For instance, in a 1994 paper in Applied and Environmental Microbiology, Tiege and his colleagues reported isolating a 16S sequence from an organism found at the bottom of a 25-meter-deep oil-drilling site in Michigan. When the researchers compared the sequence to others in the 16S database, they found that their organism was closely related to a nitrogen-fixing bacteria isolated from plant roots in Pakistan. When the researchers then tested their organism, they found that it too could fix nitrogen.

    As critics of this approach point out, not all organisms with genes for some function are necessarily performing it in the real world. Fortunately, 16S analysis is not the only tool in the molecular toolbox. Stahl and others also are working with several other techniques that they hope will give them a window on not just what newly discovered microbes are doing but how they fit into microbial communities. One scheme relies on fluorescent or radioactive probes designed to bind only to the ribosomal DNA of select organisms. By adding these probes to field samples, researchers can track particular microbes so as to learn about their preferred environments and neighbors.

    Stahl, Wisconsin's Nealson, and colleagues, for example, are using a variety of such probes to link specific organisms in lake sediments with their function. Researchers have long known that microorganisms tend to form layers in many lake sediments, based on, among other things, the compounds they rely on to produce chemical energy. In Lake Michigan sediments, for instance, microbes that use molecular oxygen for respiration lie in the top centimeter of sediment. Other microbes that use nitrate, manganese, iron, sulfate, and CO2 can be found in successively deeper layers.

    But while this general layering is well described, little is known about which species occupy each layer. Now, however, by adding radioactive and fluorescent probes to samples of lake sediment, the Stahl team is beginning to identify the organisms living in the layers. Although the work is still in its initial stages, it already has turned up one surprise, that a plethora of crenarchaeota live in the top sediment layer, again suggesting that these organisms are aerobes well adapted to nonextreme environments.

    In another approach, Gary Sayler and his colleagues at the University of Tennessee, Knoxville, are trying to infer the function of microbes by determining which of their genes have been switched on. To do so, the researchers scour their samples for messenger RNAs (mRNAs), the gene transcripts that tell a cell to construct particular proteins.

    The researchers are especially interested in observing how extraordinarily diverse microbial communities change after an environmental assault, such as an oil spill. With their analytic technique, they have been able to track in soil and water samples the activity of organisms that break down oily, hydrocarbon compounds, such as naphthalene. In normal soil, the researchers found that mRNAs for genes for naphthalene degradation are expressed only at low levels. But following an oil spill, the level of these mRNAs rose as much as 10,000-fold, suggesting that oil-chomping organisms were displacing a host of other types of organisms.

    The tools also are providing insight into the structure of these mini-rain forests. University of Illinois microbial ecologist Lutgarde Raskin and her colleagues, for example, are using fluorescent probes to study the associations between organisms that break down nutrients in waste-water treatment plants. In such plants, as in many more-natural environments such as soils and sediments, numerous types of microbes work in close association. In waste-water plants, microbes often clump together to form granules about 1 millimeter across, says Raskin, and with their probes, she and her colleagues have shown that these granules have a consistent structure, with organisms that get their food from the nutrient-rich water located on the outside and species that feed on each others' waste products lying in successive rings toward the center.

    By all accounts, such studies are causing a fundamental shift in the discipline of microbiology. By giving researchers the means to study specific organisms in the environment, “molecular methods are creating a wonderful bridge” between molecular biologists and environmental scientists, says Sayler. Many researchers believe that this type of cooperation will be critical to deepening researchers' understanding of how microbial species control broad biogeochemical processes, such as the cycling of carbon, nitrogen, and sulfur. “The biosphere is dominated by microbial processes,” says Olsen, and global change, ranging from the buildup of greenhouse gases to new land uses, may be altering these processes.

    “We don't yet know a lot [about the effect of such changes on microbial processes] … although the field is progressing rapidly,” says Woese. And for many researchers, that progress spells excitement. “No one could have convinced me 10 years ago that [microbiology] would be this exciting today,” says Staley. “The next decade is likely to be one of the best yet.”

  5. Physics

    Subatomic Spin Still in Crisis

    1. Andrew Watson

    A new detector on the HERA positron-proton collider in Hamburg, Germany, has confirmed that there really is a crisis in particle physics—and physicists are delighted. The crisis is a decade-old conundrum about the toplike spin of protons and neutrons. These particles are composed of quarks, which themselves spin, but the quarks don't contribute nearly enough spin to explain the total. Results announced last week from the new detector, called HERMES, have confirmed the shortfall—and thus shown that HERMES's new spin-probing technology is working properly. That's welcome news, say physicists, because HERMES, as the most advanced experiment in the field, could eventually track down the missing spin.

    Spin mastered.

    Measuring a scattered positron beam and identifying other debris (hadrons) reveal the neutron's spinning interior.

    Source: HERMES

    “People will mainly be excited by the fact that the experiment is working,” says physicist Richard Milner, spokesperson for the HERMES collaboration. HERMES replaces the solid targets of past experiments with a gas of simple atoms, polarized so that their nuclei spin in the same direction. The arrangement allows polarized positrons (anti-matter counterparts of electrons) from HERA's circular beam to pass through the target time after time, yielding plenty of clean data on the spinning innards of nucleons—protons and neutrons. “They made a major accomplishment in proving this new technique works,” says Yale University physicist Vernon Hughes.

    With HERMES, physicists should eventually be able to probe how much spin is carried by elusive components of nucleons such as gluons (force particles exchanged by quarks) and so-called strange quarks, which don't normally inhabit nucleons but can briefly pop into existence and then vanish. A resolution to the spin crisis could also open new insights into the theory that describes how quarks and gluons behave, called quantum chromodynamics (QCD). The surgical precision of QCD's equations is difficult to translate into ironclad predictions about the interiors of nucleons, so “we need all the help we can get from experiments,” says Massachusetts Institute of Technology theorist Robert Jaffe.

    To tease out the sources of spin, physicists make two measurements, one with the spins of the target nucleons aligned with those of the probing particles, and the other with the target spins reversed. Differences in the numbers of particles scattered hold clues to how spin is distributed in the nucleon. Several major experiments, including the Spin Muon Collaboration at CERN, the European center for particle physics, and projects at the Stanford Linear Accelerator Center (SLAC), have fired polarized probe beams at targets such as solid ammonia or butanol. Solid targets yield more collisions—but also a lot of “noise” from unpolarized nucleons, says HERMES team member Klaus Rith.

    A decade of studies at SLAC and CERN has shown that instead of carrying about 60% of a nucleon's spin, as QCD implies, the quarks account for just half that amount. Now, analyses of the early HERMES data confirm that quarks carry about 25% to 30% of a neutron's spin, estimates Milner—“about a factor of 2 lower than you would expect” from theory.

    Physicists now hope that HERMES's innovative technology will allow it to show where the extra spin is coming from. The gas target—a cloud of spin-polarized hydrogen or helium—yields a cleaner signal than the solid targets of past experiments, and a clever polarization scheme exploits the natural alignment of HERMES's high-energy positron beam.

    As Milner explains, the spin axes of the positrons end up pointing across their flight path as they are accelerated in HERA's 6.4-kilometer ring. Spin studies need probe particles that spin along their direction of travel, however, so HERMES twists the positrons' spin with a huge magnet placed just before the detector. After the detector, a second magnet restores the positrons to their original orientation for experiments on HERA's two other detectors.

    Next week, HERMES physicists will start building a new detector element that will identify particles dislodged from nucleons in the gas by the impinging positron. “The [structure of] those residues … reflects important information about the spin of the original proton and neutron and the way it was put together,” says Jaffe. Particles called kaons, for example, betray the presence of strange quarks inside the nucleon. To identify and analyze these fragments, whose low energy makes them hard to detect, the new instrument will record the Cerenkov light (the optical equivalent of a bow wave) generated when charged particles enter the counter moving faster than the speed of light in the material.

    HERMES won't rule the spin field for long. CERN and SLAC both plan new experiments that will also be able to identify emitted particles. But HERMES, true to its name, may bring the first news that could end the spin crisis.

    Andrew Watson is a science writer in Norwich, U.K.

  6. Evolutionary Biology

    Thanks to a Parasite, Asexual Reproduction Catches On

    1. Martin Enserink

    Asexual reproduction is usually considered a way of life—an evolutionary choice a species makes when the drawbacks of sex outweigh its long-term benefits. Recent research, however, has shown that in some insects, parthenogenesis (in which females give rise only to daughters and no males are born) is more of a sickness than a strategy. And now, in the 22 March issue of the Royal Society Proceedings: Biological Sciences, two Dutch entomologists claim that parthenogenesis may literally be contagious.

    David Ferro/Fran Heyl

    Wolbachia at work.

    H. R. Braig and S. L. O'Neill/Yale University

    All these Trichogramma wasps are female (top) thanks to Wolbachia, seen as a cloudy area in a wasp egg (above).

    The culprit is Wolbachia, a bacterium that infects perhaps 16% of all insect species and has come under intense scrutiny by biologists in recent years because it provides some bizarre examples of how a parasite can alter its host's sex life and reproduction for the parasite's benefit. “It's an exploding field,” says Richard Stouthamer of Wageningen Agricultural University, who did the new study with colleague Menno Schilthuizen. Their work may offer practical benefits in pest control, as well as a whole new perspective on the evolution of reproductive strategies—a topic evolutionary geneticist Laurence Hurst of the University of Bath calls “one of the big problems in biology.” The string of recent studies suggests, he says, that some instances of asexual reproduction may have evolved not for the organism's best interest but because of a parasite: “What [this] work shows is that it may not be adaptive to be asexual at all.”

    Wolbachia spends its life within the ovaries and testes of many arthropod species and is chiefly transmitted from mother insects to their offspring through the egg's cytoplasm. Because sperm are almost empty of cytoplasm, male insects—although they can be infected—are unable to pass on the organism. So Wolbachia prefers females, and it has developed ways of playing havoc with its hosts' sex lives and gender ratios. In wood lice, for example, the bacterium manages to transform infected males into functional females. And in a well-known phenomenon called cytoplasmic incompatibility, Wolbachia prevents some mating pairs from having viable offspring. In flies and mosquitoes, infected males are unable to fertilize uninfected eggs—a combination that wouldn't yield infected offspring and so wouldn't benefit Wolbachia.

    In some wasp species, Wolbachia has eliminated males altogether, by somehow disrupting the first cell division in the wasp's egg. That makes the egg diploid, which in most wasps causes the egg to develop as a female. Researchers have found that in Trichogramma, a genus of minuscule wasps that parasitize moth and butterfly eggs, this condemns the insects to perpetual asexual multiplication. The wasps' asexual state can be “cured” only by treating them with antibiotics or bacteria-killing heat in the laboratory, explains Stouthamer.

    But how does Wolbachia colonize Trichogramma species? The bacterium can jump from one species to another in mosquitoes and flies, a phenomenon called horizontal transmission. But in wasps the infection seemed to be inborn; it was impossible to transfer the bacterium from one wasp to another in the lab. This led researchers to speculate that in Trichogramma, the insect and its parasite might have cospeciated: Whenever a wasp lineage diverged into two species, a new Wolbachia strain would then develop in each isolated wasp species.

    To test this hypothesis, Stouthamer and Schilthuizen collected 20 Trichogramma species that carried Wolbachia and sequenced specific DNA regions in each—about 800 base pairs in the microbe and some 420 base pairs of nuclear DNA from a diagnostic region in the wasp. Then, they drew up two evolutionary family trees, one for the insects and one for the microbe. If wasps and their parasites coevolved, the trees would be very much alike.

    But they weren't. “It's a phylogenetic mess,” says Stouthamer. “You find almost identical Wolbachia in wasp species that are far removed and vice versa.” So cospeciation is highly unlikely. And although researchers couldn't re-create it in the lab, in nature Wolbachia must have leaped from one Trichogramma species to another many times, carrying the parthenogenetic lifestyle with it. “We knew parthenogenesis was curable; now we know it's contagious too,” says Stouthamer. He suggests that the infection spreads in butterfly eggs, Trichogramma's favorite place to breed. When infected and uninfected wasp species share an egg, the infection may jump between species.

    All this may have practical use. “This is very important,” says Wolbachia researcher Henk Braig of the Yale University School of Medicine, “because if there is horizontal transfer, there's hope we might be able in the future to make insects parthenogenetic at our will.” Certain Trichogramma species are already used as effective parasitic weapons against pest insects, and all-female strains would be even better, because only the female wasps parasitize the pest by laying eggs. So Stouthamer says he will keep trying to carry the infection between wasp species.

    Meanwhile, other researchers are trying to unravel the molecular mechanisms by which Wolbachia manipulates its host, with much of the work focusing on the details of cytoplasmic incompatibility. “If we can figure out the mechanism, it will tell us something brand-new and very fundamental about what happens when a sperm enters an egg. That would be a great step forward,” says Timothy Karr of the University of Chicago.

    Nor do researchers know whether Wolbachia really harms its hosts by making them asexual. The highly intimate relationship between the bacterium and its hosts suggests that Wolbachia is on the evolutionary road to becoming a cell organelle, just as mitochondria and chloroplasts have done, says Karr. But Hurst disagrees. “This relationship goes back an awfully long way,” he says, perhaps for tens of millions of years. “If these things are going to evolve to be nice to the host, why haven't they done it already? On the whole, I think they are being quite nasty.”

    Another mystery is how widespread Wolbachia may be. Recently, it has shown up in crustaceans, mites, and even nematode worms, as well as insects. “We're years away from knowing its true distribution across animal and plant taxa,” says Karr. “We could find out that humans have this, for all we know. Anything is possible.”

    Even catching a new kind of sex life.

    Martin Enserink is a science writer in Amsterdam, the Netherlands.

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