From Dirt Comes Diversity

Courtesy Marisa Pedulla and Graham Hatfull. SAME BUT DIFFERENT: electron micrograph of mycobacteriophages Barnyard (left), and Cjw1 (above). While morphologically similar, the two phages are quite distinct genomically and have fewer than 3% of their genes in common. When high school teacher Debbie Jacobs-Sera showed up with two of her students at Graham Hatfull's door at the University of Pittsburgh, it took only minutes for the Howard Hughes Medical Institute investigator to put them to work. Hatfull enlisted the teens in his pet project, identifying phage viruses that infect Mycobacteria, including the organism that causes tuberculosis. Hatfull's team, including Jacobs-Sera's brother, fellow HHMI investigator William Jacobs at Albert Einstein College of Medicine in New York, hit the jackpot. These students and others more than doubled the number of genetically characterized mycobacteriophages, bringing the total to 14.1 In their morphology and genetics, the novel phages proved very different from those previously characterized, suggesting extensive diversity. Mycobacterium tuberculosis is one tough bug. It remains dormant in its human host for years, evading immune attack, while a thick outer coat of complex lipids protects it from toxic drugs. Hatfull says he thinks the bacterium's natural enemies, phages, may hold the key to understanding its pathogenicity as well as provide advanced methods for drug screening, diagnosis, and disease control. That's why he and Jacobs want to identify as many as possible. Since phages are ubiquitous, the task isn't that difficult. The team collected soil samples from various locations--New York teen Laura Keenan even dipped her spatula into a monkey pit at the Bronx Zoo--and applied them to Petri dish lawns of Mycobacterium smegmatis, a nonpathogenic relative of M. tuberculosis. The researchers cloned and sequenced DNA from the resulting plaques. They also examined virus particles with an electron microscope. When Hatfull and Jacobs compared the new and old isolates, the physical differences suggested significant population diversity. That conclusion extends to the DNA level. Genome length in the 14 phages varies from 49,000 to 156,000 base pairs. On average, the genomes contain 118 potential genes, or open reading frames, and few are repeats. "There's only one gene between any two of [the 14 phages] with greater than 90% homology," notes Jacobs. Janet Westpheling, a University of Georgia geneticist also working on mycobacteriophages, says she isn't surprised by the diversity. "Soil containing the predominant organisms on earth is where you'll find genetic diversity. That's where the really interesting biology is." The genomes also are highly dynamic, trading DNA sequences within the population and even with host cells. Because of this horizontal gene flow, the genomes are mosaics of interchangeable units consisting of single genes. "Each phage genome appears to be a unique assemblage of exchangeable modules," notes Hatfull. The group postulates that viruses cannot be grouped in simple, hierarchical family trees. Instead, phages are related in a 'reticulate' network marked by various gene exchange events. "Species doesn't apply to this, unless you come up with a new definition," says Hatfull. Horizontal gene flow between phages and their hosts, and perhaps even their hosts' hosts, provides links in a chain of events between viruses, bacteria, and mammalian disease. Two of the phages encode homologs of an immunostimulatory antigen found in M. tuberculosis and Mycobacterium leprae, the bacterium that causes leprosy. Another phage has two homologs of enzymes needed by mammals to synthesize a cofactor of nitric oxide synthase, another weapon against invading bacteria. Perhaps even stranger, Bxz1--one of the Bronx Zoo isolates--encodes a protein with 35% homolgy to the human protein Ro, which is strongly implicated in autoimmune diseases. Mice deprived of Ro develop a lupus-like syndrome.2 The result, together with the discovery that Ro also occurs in at least one bacterium, lends credence to the hypothesis that autoimmune responses may be tied to microbiological challenge. As to future discoveries, Hatfull is circumspect. Given the genome diversity, "we have little or no predictive power to say what we'll find in any new phage." But Laura Keenan knows exactly what she got out of her experience in the lab. Says Keenan, a high school student with a Cell cover story under her belt, "I gained so much, it's hard to quantify." Barry A. Palevitz (palevitz@plantbio.uga.edu) is a freelance writer in Athens, Ga. References 1. M.L. Padulla et al., "Origins of highly mosaic mycobacteriophage genomes," Cell, 113:171-82, April 18, 2003. 2. D. Xue et al., "A lupus-like syndrome develops in mice lacking the Ro 60-kDa protein, a major lupus autoantigen," Proc Natl Acad Sci, 100:7503-8, June 3, 2003. function sendData() { document.frm.pathName.value = location.pathname; result = false if (document.frm.score[0].checked) result = true; if (document.frm.score[1].checked) result = true; if (document.frm.score[2].checked) result = true; if (document.frm.score[3].checked) result = true; if (document.frm.score[4].checked) result = true; if (!result) alert("Please select") return result; } .myradio{background-color : #94bee5} Please indicate on a 1 - 5 scale how strongly you would recommend this article to your colleagues? Not recommended 1 2 3 4 5 Highly recommended Please register your vote

From Dirt Comes Diversity

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