Adhesion of Vibrio cholerae to granular starches

Abstract

Cholera is a severe diarrheal disease caused by specific serogroups of Vibrio cholerae that are pathogenic to humans. Cholera can become epidemic and deadly without adequate medical care. Appropriate rehydration therapy can reduce the mortality rate from as much as 50% of the affected individuals to <1%. Thus, oral rehydration therapy (ORT) is an important measure in the treatment of this disease. To further reduce the symptoms associated with cholera, improvements in oral rehydration solution (ORS) by starch incorporation were suggested. Here, we report that V. cholerae adheres to starch granules incorporated in ORS. Adhesion of 98% of the cells was observed within 2 min when cornstarch granules were used. Other starches showed varied adhesion rates, indicating that starch source and composition play an important role in the interaction of V. cholerae and starch granules. Sugars metabolized by V. cholerae showed a repressive effect on the adhesion process. The possible mechanisms involved are discussed. Comparing V. cholerae adhesion with the adhesion of other pathogens suggests the involvement of starch degradation capabilities. This adhesion to granular starch can be used to improve ORT.

FIG. 1.

Immediate adhesion of V. cholerae to different starches. Vibrio cholerae O1 was grown in 50 ml of Luria-Broth liquid medium in a 250-ml flask with agitation for 16 to 18 h at 37°C. The bacteria were harvested by centrifugation (16,000 × g for 5 min) and washed three times with modified ORS without starch. The bacteria were then diluted to a final concentration of ∼10⁶ CFU · ml⁻¹. The starch-containing ORS (preequilibrated) was mixed at a 9:1 ratio to give a final concentration of 10% starch and approximately 10⁵ CFU · ml⁻¹ (10⁶ CFU to 1 g of starch). The mixture was rotated on an axial spinning agitator, and samples were drawn after 2 min. The samples were spin centrifuged for 30 s at 320 × g. A 100-μl sample of the supernatant was serially diluted and plated on Luria agar plates; after 24 h incubation at 37°C, the numbers of CFU were recorded. Controls containing modified ORS without starch were used to normalize the results (P < 0.05). a, b, and c indicate significant differences (P < 0.05).

FIG. 2.

Adhesion of GFP-tagged V. cholerae to cornstarch. Vibrio cholerae O1, tagged with a plasmid encoding GFP, was grown and mixed with ORS containing cornstarch as described in the legend to Fig. 1. The starch granules were washed three times with the modified ORS and then viewed under light and fluorescence microscopes (magnification, ×1,000). Images were captured by digital camera and processed using Adobe Photoshop 7.0. The left panel shows starch granules with the GFP-tagged bacteria under UV light. The right panel shows the same field under normal light.

FIG. 3.

Effect of low-molecular-weight solutes on V. cholerae adhesion to cornstarch. Vibrio cholerae O1 was grown and mixed with ORS containing cornstarch as described in the legend to Fig. 1, except that the ORS used contained 111 mmol · liter⁻¹ of either glucose, sucrose, trehalose, sodium chloride, xylose, fructose, maltose, or lactose or 50 g · liter⁻¹ dextrins (DE19 and DE6). The mixture was rotated on an axial spinning agitator, and samples were drawn after 2 min. The samples were spin centrifuged for 30 s at 320 × g. A 100-μl sample of the supernatant was serially diluted and plated on Luria agar plates; after 24 h incubation at 37°C, the numbers of CFU were recorded. Controls containing modified ORS without starch were used to normalize the results (P < 0.05). a, b, and c indicate significant differences (P < 0.05).