Cryptosporidium Parvum (a protozoan) in Drinking Water

Cryptosporidium is the genus of a protozoan organism which causes the parasitic infection, Cryptosporidiosis. It exists in either the free-swimming (trophozoite) form or the oocyst (dormant) form. Cryptosporidium parvum is now recognized as a human pathogen which can cause severe diarrheal illness. Contaminated food can also cause infections.

Members of the genus Cryptosporidium are placed taxonomically within the phylum Apicomplexa, order Eucoccidiorida, suborder Eimeriorina, and family Cryptosporidiidae. Various Cryptosporidium species infect epithelial surfaces, especially those along the gut, and can be found in a wide range of vertebrates, including humans. Cryptosporidium parvum infects the small intestine of an unusually wide range of mammals, including humans (Tyzzer, 1912). Ingestion of drinking water contaminated with viable Cryptosporidium oocysts, the environmentally resistant form of the organism, is the major mode of transmission. Cryptosporidium is introduced into the water supply via animal excreta containing oocysts. Important reservoirs of the oocysts include cattle, sheep, and pigs. Cryptosporidium is also found in wild animal populations. The organism is more prevalent in ruminants such as deer, elk, moose, and caribou and is primarily found in neonates of these species. Person-to-person transmission is common, especially in child daycare settings.

Direct contact with infected animals, especially calves and lambs, can cause illness in exposed persons. For example, Holstein calves that were infected orally at 4 days of age each typically shed 25 million oocysts of C. parvum per day but no oocysts were detected until 4 days post-infection (DPI). Peak oocyst production occurred 6-8 days following exposure and as few as 2 billion and as many as 20 billion oocysts were collected during a single 24-hr period from calves during peak oocyst shedding. A single calf can easily produce 50 billion oocysts within a period of one week.

Life Cycle

The life cycle of C. parvum (see image) begins with the ingestion of the sporulated oocyst, an infective package (the oocyst) of spores (sporozoites) and the resistant stage found in the environment. Each oocyst contains multiple sporozoites (mobile sporelike forms), which exit from a suture located along one side of the oocyst. The preferred site of infection is the ileum (part of the small intestine), and sporozoites penetrate individual epithelial cells in this region. Parasites reside on the luminal (inner) surface of the intestine, enclosed by a thin layer of host-cell cytoplasm. A unique, desmosome-like attachment organelle (how the sporozoite attaches to an intestinal cell), plus accessory folding of the parasite membranes, develop at the interface between the parasite proper and the host cell cytoplasm. This attachment organelle is sometimes referred to as the "feeder organelle." Multiple fission (=merogony; =schizogony; asexual reproduction) occurs, resulting in the formation of 8 merozoites (yet another form of the parasite; they are capable of sexual/asexual reproductions) within the meront (merozoite). These meronts are termed Type I meronts and rupture open, releasing free merozoites. Once these merozoites penetrate new cells, they undergo merogony to form additional meronts. Type I merozoites are thought to be capable of recycling indefinitely and, thus, the potential exists for new Type I meronts to arise continuously. It is thought that some Type I merozoites are somehow triggered into forming a second type of meront, the Type II meront, which contains only 4 merozoites.

Once liberated, the Type II merozoites appear to form the sexual stages. Some Type II merozoites enter cells, enlarge, and form macrogametes (=macrogametocyte; the female form). Others undergo multiple fission once inside cells, forming microgametocytes (the male form) containing 16 non-flagellated microgametes. Microgametes rupture from the microgametocyte and penetrate macrogametes, thus forming a zygote. A resistant oocyst wall is then formed around the zygote (the only diploid stage in the life cycle), meiosis occurs, and 4 sporozoites are formed in the process. Formation of sporozoites is termed sporogony. These (thick-walled) oocysts are passed in the feces and into the environment. Approximately 20% of the oocysts produced in the gut fail to form an oocyst wall and only a series of membranes surround the developing sporozoites. These "oocysts," devoid of a wall, are sometimes termed "thin-walled oocysts." It is believed that the resulting sporozoites produced from thin-walled oocysts can excyst (rupture) while still within the gut and infect new cells. Thus, C. parvum appears to have two autoinfective cycles: the first by continuous recycling of Type I meronts and the second through sporozoites rupturing from thin-walled oocysts.

Development

The development of Cryptosporidium occurs more rapidly than many textbooks imply, and each generation can develop and mature in as little as 12-14 hours. Due to the rapidity of the life cycle, plus the autoinfective cycles, huge numbers of organisms can colonize the intestinal tract in several days. The ileum soon becomes crowded and secondary sites are often infected, such as the duodenum and large intestine. In immunosuppressed individuals, parasites can sometimes be found in the stomach, biliary and pancreatic ducts, and respiratory tract. Diarrhea, weight loss, and abdominal cramping are clinical signs of the disease and in immunosuppressed individuals electrolyte imbalance may occur.

The prepatent period, which is the interval between infection and the first appearance of oocysts in the feces, is generally 4 days (3 days in heavy infections). Patency, which is the length of time oocysts are shed in the feces, generally lasts 6-12 days in immunocompetent individuals but may be prolonged in immunosuppressed patients. Infections are most common in young children, immunocompromised individuals, AIDS/HIV patients, chemotherapy patients, and organ transplant patients.Symptoms of Cryptosporidiosis usually appear within two to ten days after ingestion of the parasite. Symptoms include watery diarrhea, headache, abdominal cramps, nausea, vomiting and low grade fever. These symptoms may lead to weight loss and dehydration. In otherwise healthy individuals, symptoms usually last from one to two weeks, at which time the immune system is able to stop the infection. In the immunocompromised, the infection may continue and become life-threatening. At present, there is no effective drug therapy for Cryptosporidiosis.

Treatment

Water treatment for Cryptosporidium relies on properly designed and operated filtration systems. Chlorine disinfection of the organism is ineffective, as it has been shown that even one oocyst can withstand pure bleach (50,000 ppm chlorine) for 24 hours and still cause an infection, but UV / Ozone Disinfection with filtration is very promising. Filter systems usually consist of several filters. A "roughing filter" containing a 5µ - 10µ (micron) cartridge filter is installed to remove any large diameter sediments, such as iron sediments, sand, salt , etc. Downstream from the roughing filter, a "polishing filter" containing a <1 µm (micron) absolute cartridge filter is installed to remove small particles, including Cryptosporidium, from the water. Most reputable water system vendors are currently recommending a filter porosity of <1 µm to submicron or membrane filters to remove Cryptosporidium cysts and trophozoites (merozoites) from drinking water. Testing for Groundwater Sources and Potential for Surface Water Influence.

Testing

Testing procedures are available for detecting Cryptosporidium oocysts in both raw and treated drinking water. The testing procedure involves filtering a large volume of water through a 1 micron, yarn-wound, polypropylene filter. The filter is then treated to remove any oocysts which may be present and the sample is concentrated. A monoclonal antibody to Cryptosporidium is added to the sample to bind to oocyst wall antigens. The reaction can be visualized by the addition of fluorescein isothiocyanate (FITC) - conjugated anti-immunoglobulin, and scanning with an epifluorescence microscope.