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March 8, 2004

Research on cholera bacteria focuses on biofilm formation

Bacteria in biofilms are much more resistant than free-floating cells to antibiotics and disinfectants

By Tim Stephens

In regions where cholera is an endemic disease causing periodic seasonal outbreaks, the bacterial pathogen (Vibrio cholerae) lives between outbreaks in aquatic ecosystems such as coastal estuaries.

Cholera photo

In cholera bacteria, the "rugose" variant (above) forms biofilms that are more resistant to environmental stresses than single cells (below)
Photos: F. Yildiz

Cholera photo

Little is known about how it survives and adapts to changes in its aquatic habitat, but one factor is the cholera bacteria's ability to switch from a free-floating "planktonic" mode to the formation of slimy bacterial "biofilms" on various surfaces.

"Biofilms are thought to be a refuge for the organism, where it is better able to survive stressful conditions," said Fitnat Yildiz, an assistant professor of environmental toxicology at UCSC.

Yildiz's lab is investigating the molecular mechanisms that enable cholera bacteria to switch between the two growth modes. She and postdoctoral researcher Catharina Casper-Lindley reported their latest findings in a paper featured on the cover of the March issue of the Journal of Bacteriology.

The researchers have identified a new regulatory gene required for the production of a mucus-like substance--an "exopolysaccharide"--that is involved in biofilm formation. The exopolysaccharide attaches the bacteria in a biofilm to each other and to surfaces.

"It is basically goop that the cells excrete and that forms the matrix of the biofilm," Casper-Lindley said.

The newly discovered gene is the second such regulator of biofilm formation found in cholera bacteria. Yildiz discovered the other regulatory gene in previous work done at Stanford University. The findings suggest that biofilm formation is controlled by a complex regulatory network of molecular signals.

"We are now trying to understand the entire pathway--the interplay between the regulators and other signaling molecules that enables the bacteria to sense and respond to environmental conditions," Yildiz said.

In the laboratory, the two growth modes of V. cholerae correspond to distinctive colony morphologies when the bacteria are grown on a solid agar growth medium in petri dishes. The "smooth" variant does not produce as much exopolysaccharide and does not form biofilms as well as the "rugose" variant, in which the colonies appear corrugated or wrinkled.

The environmental factors that trigger the switch from one growth mode to the other are still unclear, but clues may be found in the regulatory mechanisms Yildiz is studying. With a better understanding of the pathogen's life cycle in its natural environment, researchers will be in a better position to predict and control cholera outbreaks, she said.

In nature, biofilms enable the cholera bacteria to attach themselves to a wide range of surfaces, including plankton and particles of sand. Tiny crustaceans called copepods have been found to carry thousands of cholera bacteria.
"These biofilms form on anything floating in the water, plankton included," Casper-Lindley said.

Biofilm formation is also important in many other infectious bacteria. Bacteria in biofilms are much more resistant than free-floating cells to antibiotics and disinfectants such as chlorine. Similiar molecular mechanisms are probably involved in regulating biofilm formation in different organisms, Yildiz said.

"Whatever we find in cholera bacteria may be applicable to other related pathogenic organisms, because biofilm formation is very important in many bacterial infections," she said.

Yildiz's research is supported by the National Institutes of Health (NIH), the Ellison Medical Foundation, and the UC Toxic Substances Research and Teaching Program. She and other UCSC faculty recently received a $258,000 NIH grant to fund the purchase of a special type of microscope--a confocal microscope--needed to study biofilm formation.

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