Structural Physiology of the Cryptosporidium Oocyst Wall

Cryptosporidium parvum is a significant cause of human diarrheal disease. This parasite is the causative agent of numerous outbreaks of waterborne diarrheal disease. C. parvum oocysts can survive for many months in water and are resistant to several disinfectant treatments. The prolonged survival of oocysts and the resistance to disinfectants is attributed to the presence of a thick oocyst wall. Very little is known, however, about the biochemical composition and structural...
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Cryptosporidium parvum is a significant cause of human diarrheal disease. This parasite is the causative agent of numerous outbreaks of waterborne diarrheal disease. C. parvum oocysts can survive for many months in water and are resistant to several disinfectant treatments. The prolonged survival of oocysts and the resistance to disinfectants is attributed to the presence of a thick oocyst wall. Very little is known, however, about the biochemical composition and structural physiology of the oocyst wall. The overall goal of the project was to develop probes to identify and assess the roles of proteins, carbohydrates, and lipids in the integrity and resistance of the oocyst wall to disinfectants. The specific objectives were to: (1) develop probes to identify and assess the role of proteins, carbohydrates, and lipids in maintaining integrity of the oocyst wall; and (2) determine the effects of disinfectants on biochemical components of the oocyst wall. The approach was to focus on a few well-characterized molecules or reagents that have been shown to be particularly useful and specific for certain components, and use these molecules and reagents in conjunction with biochemical treatments known to affect certain classes of molecular structure. In this manner, the research team hoped to identify structures and/or molecules that have particular relevance to the unique resistance possessed by this organism. During the course of this project, they were able to accomplish the objectives of developing probes such as antibodies for protein components, lectins for carbohydrate components, and fluorogenic probes for lipid components. The researchers were able to use these probes to study the effect of disinfectants such as chlorine and ozone on these components. Proteins Using antibodies as probes for proteins, the researchers studied the Cryptosporidium oocyst wall protein (COWP) and antigens recognized by monoclonal antibodies 2C3 and Cellabs. COWP was resistant to chlorine and ozone treatment. However, oocyst wall proteins recognized by monoclonal antibodies 2C3 and Cellabs are susceptible to chlorine but resistant to ozone treatment. Carbohydrates Using lectins and glycosidases as probes for carbohydrates, the research team found that Galactose and N-acetyl galactosamine are present in oocyst walls. Ozone and chlorine do not affect the integrity of the oocyst carbohydrates. Lectins that reacted with oocyst wall carbohydrates agglutinated oocysts and induced excystation of intact oocysts in suspension. The monosaccharide composition of carbohydrate components was confirmed by high pH anion exchange chromatography. Lipids Using fluorogenic lipid probes, the research team found that phospholipids and sphingolipids are present in oocyst walls whereas cholesterol is not. For the three lipid classes examined, the following hierarchy appears to hold for the lipids of C. parvum oocyst walls: sphingolipids >phospholipids >>cholesterol. Chlorine and ozone permeabilize the oocyst wall, facilitating uptake of phospholipids. The goal of this study was to develop probes to identify and assess the roles of proteins, carbohydrates, and lipids in the integrity and resistance of the oocyst wall to disinfectants. Using these probes, the research team identified specific components of proteins, carbohydrates, and lipids of the oocyst wall and studied the effect of the disinfectants on them. Based on these results alone, however, no immediate recommendations for drinking water treatment can be made. It is hoped that the information gained from this study can be used in the future to design specific strategies directed at detecting and eliminating Cryptosporidium from drinking water supplies. Originally published by AwwaRF for its subscribers in 2004. This publication can also be purchased and downloaded via Pay Per View on Water Intelligence Online - click on the Pay Per View icon below