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Cholera Genome Sequenced
Sequence of cholera genome paves way to new therapy
By Chad Cohen

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Scientists announced Wednesday that they have sequenced the genome of the bacterium that causes cholera, one of humanity's most ancient and deadly scourges.

Cholera bacterium, Vibrio cholerae

The intestinal disease poses serious health risks in developing countries where it killed about 8,500 people last year according to the World Health Organization and made nearly a quarter of a million others ill. The researchers anticipate the new information will have implications for the production of better vaccines, diagnostics and treatments.

"Having the sequence available will facilitate these efforts immensely," says Anthony S. Fauci, director of the National Institute of Allergy and Infectious Diseases in Bethesda, Maryland.

Scientists are also hopeful the sequence will help them understand how organisms evolve to become significant human pathogens. The results were presented at a press conference in Washington and are being published in the current issue of Nature.

Circular representation of the V. cholerae genome: Chromosome 1.

Since the project began, there have been several surprises for the team at The Institute for Genomic Research (TIGR) in Rockville, Maryland, who completed the sequencing using the shotgun procedure. The first came in November 1998, when James Kaper and colleagues at the University of Maryland School of Medicine discovered that the organism, Vibrio cholerae, unlike other pathogens, had not one, but two circular chromosomes—a large chromosome of about three million base pairs and a small one of about one million. The authors suggest that sometime during the organism's evolution, an ancestral species captured a megaplasmid, a large circular piece of DNA. Over time the two chromosomes exchanged genes and both became essential for the organism to survive.

It appears that the extra chromosome gives the organism a more competitive advantage in diverse environments, says John F. Heidelberg of TIGR, especially since others in the Vibrio genus have the two-chromosome structure as well.

Of the bacterium's 3,885 genes, however, 75 percent of them are on the larger chromosome, says Claire M. Fraser, president of TIGR. It seems the larger chromosome also houses the most important genes—genes that enable DNA to replicate, produce proteins, maintain the cell wall, and even cause disease.

Claire M. Fraser, president of TIGR.

About a third of the cholera genes are similar to E. coli, a common and usually harmless bacterium that lines the gut, says Fraser. The scientists did find a new toxin in the cholera genome, called RTX, that gives the organism some bragging rights as having "the largest toxin gene known in existence," says John Mekalanos of Harvard Medical School. The toxin has 4,545 amino acids, which makes it about 15 times larger than the average protein.

The genome of Vibrio cholerae also turned out to be much larger than expected. When sequencing began in 1997, it was thought that the cholera genome was between two and a half to three million base pairs, says Fraser. It turned out to be more than four million. The team had hoped to sequence a sample of a related species for the sake of comparison, but the unexpected size strapped them for funds, according to Fraser. The total cost of the project came to nearly $800,000—about twenty cents a base.

Humans have been plagued with cholera for more than 2,000 years. Its symptoms were described in ancient Sanskrit writings and by the Greek physician Hippocrates. It was a cholera epidemic in London in 1849 that prompted John Snow's famous pump handle experiment, which proved that polluted water could cause disease.

Death's dispensary by George John Pinwell

But cholera isn't just part of history. "It's still around after all these years and it's more widespread than ever," says Kaper. It remains an efficient killer in parts of Asia, Africa and Latin America where good sanitation and safe drinking water is lacking.

When ingested, the cholera bacterium attacks the surface of the small intestine and secretes a toxin that causes the intestine to rapidly excrete fluids, resulting in severe diarrhea. The best treatments involve re-hydration therapy with electrolyte solutions, which can be hard to come by in many areas. While most people have no symptoms or mild diarrhea, in severe cases people can die within a few hours.

Current vaccines only protect about 50 percent of the people, and only for a few months. "We don't know all there is to know about making a good vaccine against cholera," says Mekalanos. The genome sequence allows us to refine vaccines by allowing researchers to systematically select desirable gene candidates while eliminating potentially toxic genes that could make an individual sick.

The sequence could also provide viable drugs. Scientists believe that disabling genes that allow the bacterium to move through its watery environment, for example, whether it's the Bay of Bengal or the human gut, could cripple the bacterium. But until now they haven't been able to find these genes. The genome sequence reveals fifty, says Mekalanos. He's working on a Vibrio cholerae gene chip that can be probed for interesting genes.

It's uncertain whether the sequence will ever be necessary to protect industrialized nations against a new cholera pandemic. Current practices of sanitation and chlorinated water have made cholera a rarity in the United States and Europe for nearly a century. But scientists have already isolated a strain of cholera resistant to chlorine, which demonstrates that the bacterium can change, says Mekalanos. "I'm optimistic. We've seen hundreds of years of microbiology on our side, but it could happen."

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Heidelberg, J.F. et al. DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae, Nature 406, 477-484 (August 3, 2000).

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