Cryptosporidium species — cattle

Note:  The taxonomy of Cryptosporidium species is very much a work in progress, primarily because of the increasing replacement of ooocyst morphology by molecular techniques to define species within the genus.  One of the consequences of this greater accuracy is that in the earlier published literature the identifications of species and genotypes might not be consistent with the current understanding of the taxonomy of the parasite.

Cattle around the world are infected at some stage in their lives with Cryptosporidium, and intracellular parasite of enterocytes.  To date, four species of the parasite have been reported from cattle: 1) the potentially zoonotic C. parvum, which inhabits the small intestine, seems to be the commonest species in young calves, and which s the cause of significant disease - cryptosporidiosis - in this age group; 2) C. andersoni, which inhabits the gastric pits in the abomasum, is more common in older animals and seems to be less pathogenic; 3) C. bovis - a recently described species which develops in the small intestine, is more often found in older animals,and is of uncertain clinical significance; and 4) C. ryanae, also recently described, with occurrence and significance similar to C. bovis.  In addition, a newly described species - C. ubiquitum (previously known as the "cervine genotyppe") - has been transmitted experimentally to cattle, and has been found very rarely in people in England and Wales.

The life cycle is Cryptosporidium in cattle is direct and similar to that of Eimeria (even though there is evidence suggesting that Cryptosporidium is a gregarine protozoan rather than a coccidian).  Parasitic development occurs within the microvilli on the luminal surface of enterocytes.  Two cycles of asexual reproduction (merogony) are followed by a cycle of sexual reproduction (gametogony) which results in the production of oocysts, which are sporulated and infective when passed in the faeces of the host.  Two types of oocyst are produced: thick-walled, which leave the host and can infect other animals following ingestion; and thin-walled - which hatch within the intestine and can establish autoinfections. 

The primary clinical sign of cryptosporidiosis is diarrhea, which is seen most commonly in neonatal and young calves.  Affected animals can also show anorexia, dehydration, slowed growth and weight loss.  Mortality is rare, except in some instances in beef calves.  Affected animals will likely take some time to return to normality, but long-term effects on productivity have not been confirmed for any type of cattle. In immunocompetent animals, Cryptosporidium infections are self limiting.  There is no drug available that will reliably prevent, control or treat infections in cattle, although several have been investigated.  Control depends primarily on maintaining a clean environment so that oocysts, which are infective immediately when passed in the faeces of a host and are very resistant to adverse environmental conditions, are not available for ingestion by susceptible animals, especially the very young.

Of the species of Cryptosporidium that are currently known to infect cattle, only C. parvum is potentially zoonotic, meaning that in some circumstances infected cattle are a possible threat to human health.

Phylum: Apicomplexa
Class: Conoidasida
Subclass: Coccidiasina
Order: Eucoccidiorida
Suborder: Eimeriorina

Although Cryptosporidium shares many characteristics with other coccidia (e.g. Eimeria and Isospora), recent research suggests that it might be a gregarine protozoan. These are apicomplexans within the Class Conoidasida but are in the Subclass Gregarinasina rather than the Subclass Coccidiasina, to which are assigned Eimeria, Isospora, Neospora, Sarcocystis, and Toxoplasma. Gregarines are typically parasites of invertebrates. (See: Barta J and Thompson RCA (2006) Trends in Parasitology 22, 463-468).

On the basis of morphological, biological and molecular data, currently 12 species of Cryptosporidium are recognized in mammals, some of which are host-specific, three species in birds, three species in amphibians and reptiles, and three possible species in fish. In addition, approximately 40 genotypes have been identified from vertebrate hosts and, based on the previous history of Cryptosporidium, it is possible that some of these genotypes might be elevated to new species. It is clear that the taxonomy of this parasite is a work in progress, and it is possible that the future will bring re-arrangements of species, genotypes and host specificities, as have occurred in the past. For veterinarians a key issue is host specificity, particularly the ability of Cryptosporidium of animal origin to establish in people.

Surveys in various parts of the world have shown the potentially zoonotic C. parvum, to be the predominant species in cattle. Cryptosoridium andersoni (in the abomasum) and the recently described C. bovis, C. ryanae and C. ubiquitum (the last seen to date only in experimentally infected animals) seem to be less common although this situation might change as their occurrence is explored further. It is important to note, however, that the species composition of Cryptosporidium in cattle varies with location, and age type of animal (beef vs. dairy).

Species	Primary Host	Other Hosts *
C. canis **	Dog	N: foxes, coyotes, people (very rarely); E: cattle
C. felis	Cat	N: people (very rarely); E: cattle
C. andersoni	Cattle	N: people (very rarely)
C. bovis	Cattle	N: a lamb
C. ryanae	Cattle	None
C. ubiquitum	Cattle ?	N/E: Other ruminants, rodents N: Primates (including people), carnivores
C. xiaoi	Sheep	None
C. suis **	Pig	N: cattle, people (very rarely); E: cattle
C. hominis **	People	N: cattle, goats, a lamb; E: piglets
C. parvum **	Mouse	N: people, cattle, sheep, goats, horses, dogs, pigs, white-tail deer, zoo ruminants
C. muris	Mouse	N: a dog, a pig, many wildlife species, (very rarely); E: people, dogs and cats
C. fayeri	Kangaroo	N: other marsupials, sheep
C. macropodum	Kangaroo	N: other marsupials
C. wrairi	Guinea Pig	None
** species with oocysts that cannot be distinguished morphologically	Table based on Fayer R (2010) Experimental Parasitology 124, 90-97	N - natural infection E - experimental infection

The oocysts of all species and genotypes of Cryptosporidium described to date are round or oval, very small (approximately 4 µm by 6 µm), and it is usually difficult or impossible, even under high power, to see the four sporozoites within each oocyst.  In fresh samples, the oocysts of Cryptosporidium have a distinct pinkish tinge, which facilitates detection and identification to genus.  Slight differences in oocyst size allows differentiation of some species, but this is reliable only with considerable experience, and molecular techniques are required for reliable identification, even to species.

In mammalian hosts, intestinal Cryptosporidium develops within the microvilli on the luminal surface of the enterocytes.  The presence of the parasites causes the microvilli to become distended, and once the oocyst is formed the microvillus is disrupted and the oocyst enters the intestinal lumen.  With a light microscope it is very difficult or impossible to distinguish the various developmental stages of the parasite, but the distended microvilli signal its presence.  With C. andersoni, which develops in the abomasum in the enterocytes lining the gastric pits, the  life cycle stages of the parasite are found in parasitiferous vacuoles at the luminal surface of the cells, and/or in the few microvilli of these cells. 

Among the Cryptosoridium species infecting cattle, the potentially zoonotic C. parvum has a wide host range, whereas currently the host ranges for C. andersoni, C. bovis and C. ryanae are believed to be more restricted (see table in Taxonomy section).  The expanding application of molecular techniques to parasite identification might shift this current understanding.  In addition, a newly described species - C. ubiquitum (previously known as the "cervine genotyppe") - has been transmitted experimentally to cattle, and has been found very rarely in people in England and Wales. 

Cryptosporidium infection occurs in cattle around the world.  C. parvum is very common in (neonatal) young calves, while to date the other species have been found primarily in older animals.

The life cycle of Cryptosporidium is similar to that of Eimeria, but there are important differences.  The life cycle stage of Cryptosporidium infective for mammalian hosts is the sporulated oocyst, each of which contains four sporozoites.  The oocyts are sporulated, and hence infective, when passed in the faeces of the host.  There is, therefore, no requirement for development in the external environment for the oocysts to become infective.

Following ingestion by a suitable host, the oocysts excyst and the sporozoites penetrate epithelial cells, in which they locate in the microvilli (where these are present) in parasitiferous vacuoles between the cell wall and the cytoplasm.  They are, therefore, intracellular but extracytoplasmic.   It is in this location that parasite development continues. For C. bovis in cattle, development occurs in the enterocytes of the gastric pits of the abomasum.   For all species. first the sporozoites (at this stage sometimes called trophozoites) undergo a cycle of asexual reproduction to form merozoites which are contained within (Type I) meronts.  The merozoites are then released, invade additional mucosal cells and undergo a second cycle of asexual reproduction to form either a second generation of Type I meronts, which can continue asexual reproduction, or Type II meronts, the merozoites from which, when released, invade yet more cells and form the sexual stages of the parasite: macrogametes and microgametes.  Fertilization of the macrogametes by the microgametes results in the production of oocysts which sporulate within the host and, as indicated above, are infective when passed from the host.  Typically, most of the oocysts produced are thick-walled and can survive very well in the external environment, but some are thin-walled and relatively fragile and have very poor survival outside the host.  Usually these thin-walled oocysts release their sporozoites in the gut lumen of the host and these can result in (internal) autoinfection, potentially a major clinical problem especially in hosts that are immunsuppressed.  Depending on host and parasite species, the pre-patent period of Cryptosporidium varies between two and 14 days.  

Recent reports indicate that some species of Cryptosporidium (C. parvum from cattle and C. hominis) can complete its entire life cycle - oocyst to oocyst - in cell-free culture medium.  The significance of this perhaps surprising finding, especially as it might affect the epidemiology of the parasite, has yet to be fully explored.

Cryptosporidium is transmitted among animals by oocysts that are sporulated, and thus infective, when passed in the faeces of the host.  For cattle, faeces containing oocysts can be found in many locations, including bedding, feed and water, and on tools, implements and boots.  This potentially widespread environmental contamination is particularly important in calving areas and where young calves are reared.  In addition, there is evidence that flies and birds can serve as transport hosts for the oocysts.  The widespread occurrence of the potentially zoonotic C. parvum, particularly in dairy calves and their surroundings, is a public health concern.

Aspects of the epidemiology of Cryptosporidium were the subject of a recent 24 month longitudinal study in Maryland of dairy calves removed from their dams immediately after birth.  Most calves were infected with the potentially zoonotic C. parvum during the first week of life, and infection prevalence peaked (at close to 100%) during the second week.  It is believed that the infection was acquired from the dams, the environment and, despite the individual housing, from other calves.  The short pre-patent period (3-6 days), the high oocyst output (100 million oocysts per gram of faeces) is not unusual, and the small number of oocysts (30) required to establish infection, all facilitate this rapid establishment of Cryptosporidium in young, susceptible calves.  Prevalence subsequently declined to close to zero percent by eight weeks of age, likely because of an immune reaction which might or might not provide some protection against subsequent infections..  There was a second peak in prevalence between 16 and 20 weeks of age, and associated with C. bovis and C. ryanae.  By 22 months of age none of the calves were infected.  This pattern - much higher prevalences in younger than in older animals - is typical for Cryptosporidium in its mammalian hosts.  The Maryland study presented no data for oocyst abundance in the faecal samples, but data from a similar study of dairy calves in western Canada showed that age changes in oocyst prevalence and abundance were similar.

Survey data indicate that Cryptosporidium is much less common in beef  than in dairy calves, possibly as a result of different management systems as well as other factors which have not yet been identified.  A recent IFA-based survey of more than 500 beef cows and 600 calves in western Canada during the calving season showed prevalences of 1.1% in cows and only 3.1% in calves.  Clinical disease is also less common in beef calves, although there can be notable calf mortality.

The most significant pathology associated with Cryptosporidium parvum results from the destruction of enterocytes, especially the microvilli, where the parasite develops. As a result of these effects there is stunting of the villi, hyperplasia of the crypt epithelium and increased mucosal permeability, and inflammatory infiltration of the lamina propria. On stained histological sections the developmental stages of the parasite can be seen in distended microvilli, although it is difficult to see details of the various life cycle stages.  The abomasal C. andersoni produces similar effects in the enterocytes lining the gastric pits, but to date this species seems to be relatively non-pathogenic.  Little is known of the pathogenesis of the recently described C. bovis, C. ryanae or C. ubiquitum, though they are likely to be similar to those associated with C. parvum.  Currently available data indicate that the potentially zoonotic C. parvum is the major problem in neonatal and young calves, in which peaks in prevalence and oocyst output occur in the first few weeks of life.  Infections with C. andersoni, C. bovis and C. ryanae are more common in older animals, including yearlings and adult cattle. 

Infection with Cryptosporidium spp. in cattle is likely more common than is clinical disease.  The primary clinical sign in calves infected with C. parvum is diarrhea, although there may also be anorexia, dehydration, slowing growth and weight loss.  Symptoms can first appear when the calves are a week or two old and usually persist for a week or two.  Currently, clinical cryptosporidiosis caused by C. parvum seems to be more common in dairy than in beef calves, a difference that may result, at least in part, from differences in management systems.  Deaths among dairy calves with clinical cryptosporidiosis are rare, but high mortality rates (up to 30%) have been reported in beef calves in western Canada.  Affected animals will likely take some time to return to normality, but long-term effects on productivity have not been confirmed for any type of cattle.  Young calves with enteric disease often harbour a mix of bacterial and viral pathogens, together with Cryptosoridium, and it is sometimes difficult to tease apart the roles of individual pathogens.  In  immunocompetent cattle, infections with Cryptosporidium are usually self limiting as a result of an immune response.  This is important because currently there are no specific treatments that are uniformly effective.

While history and clinical signs are helpful, in living animals the specific diagnosis of Cryptosporidium is based on the detection of the sporulated oocysts in faecal samples.  In veterinary medicine two main methods are used for detection: 1) direct visualization of the oocysts in a faecal flotation using sugar solution, remembering that the oocysts are very small (approx. 5 µm in diameter) and light, and so float to just below the coverslip, above eggs and oocysts of other parasites; 2) an immunofluorescent antibody (IFA) test kit for detection of oocyst antigens  (Cyst-a-Glo (Waterborne Inc.), that requires a fluorescent microscope); and 3) an enzyme immunoassay SNAP test kit (EIA) (ProsPecT (Oxoid)) for parasite antigen in faeces - in Canada this kit seems to be available only for Giardia.  Methods 1 and 2 can be used after concentration of the oocysts using a sucrose gradient.  Because of the sometimes sporadic output of oocysts, a repeat sample after a few days might increase diagnostic test sensitivity.  Other techniques, including microscopic examination, or acid-fast staining, of a faecal smear are not widely used in veterinary medicine.  Field studies indicate that PCR applied to faeces for the detection of Cryptosporidium oocysts has very high sensitivity and specificity, and molecular techniques are required for reliable identification of species and genotypes.

No drugs are approved for Cryptosporidium in cattle in Canada, although various drugs have been used elsewhere in the world, particularly halofuginone.  A 2009 meta-analysis of treatment trials using this drug in cattle indicates that design faults in the 20 published studies evaluated did not allow determination of the value of halofuginone on the prevalence of infection, diarrhea or mortality. 

The control of Cryptosporidium and cryptosporidiosis in cattle depends primarily on maintaining a clean environment, particularly for young animals, and on remembering that the oocysts are immediately infective when passed by the hosts and are very resistant to adverse environmental conditions.  Particular concerns with cattle are the cleanliness of the calving areas, where the parasite can easily be transmitted from adult animals to calves and from calf to calf, and the possibility of faecal (oocyst) contamination of feed and water.

Among the species of Cryptosporidium infecting cattle, C. parvum is potentially of the greatest public health concern, primarily because it is commonly found in cattle, especially calves, and in people.  It has also been recovered from a range of other domestic animal hosts, as well as wildlife. Cryptosporidium andersoni has been reported very rarely from people, C. hominis occasionally infects cattle, and C.bovis has not (yet) been reported from people (see Taxonomy section).  Examination of 55 Cryptosporidium isolates from infected dairy calves and from people with sporadic cases of cryptosporidiosis in Ontario revealed an array of genotypes, three of which were found in both calves and people.  Recent studies on Prince Edward Island, Canada, found some subtypes of C. parvum in both people and cattle, suggesting the possibility of zoonotic transmission

A similar recent study in England and Wales identified genetically identical (gp60 locus) C. parvum in cattle and in people with diarrhea on seven of eight farms examined. A study of 8,000 Cryptosporidium isolates from people in England and Wales during the period 2000-2003 revealed 45.9% of infections were by C. parvum alone, 49.2% by C. hominis alone, 0.4% by C. parvum and C. hominis, 0.9% by other genotypes, and 3.5% were untypable.  These data have recently been updated for 2004-2006 (n=4509) , and show an increase in the proportion of C. hominis, a decrease in C. parvum, and the first detection of C. ubiquitum (two cases), a newly described species that infects primarily ruminants and was previously referred to as the cervine genotype.

When reviewing reports of possible zoonotic transmission of Cryptosporidium, it is important to consider the date of publication and the then current understanding of the taxonomy of the parasite.  It is also important to remember that species of Cryptosporidium from other hosts can infect people, for example C. meleagridis from birds, C. canis from dogs, and C. felis from cats.  Cryptosporidium is a particular problem in hosts who are immunocompromised, for example people with HIV-AIDS, and this should be born in mind when there is the possibility of contact between these individuals and animals that are likely to be infected.

Probably most human infections with Cryptosporidium result from person to person transmission.  There are, however, examples where epidemiological evidence, sometimes supplemented by molecular identification of the parasite, suggest transmission from animals.  Examples include veterinary students infected from young ruminants used for teaching and school children visiting petting zoos.  People have also acquired the parasite from water, including municipal drinking water (for example 400,000 cases in Milwaukee, Wisconsin in 1993, and 7,000 cases in North Battleford, Saskatchewan in 2001; in both instances the causes were identified as human sewage contamination of the water supply, coupled with sub-optimal water purification), wells, streams, rivers, ponds and lakes, recreational fountains, and especially swimming pools.  People can also become infected from foods (for example, fruits and vegetables) and drinks (for example and raw milk, fresh-pressed apple cider) contaminated with faeces from an infected person or animal.  In only a few of these outbreaks has it been possible to identify animals as the cource of human infection on the basis of epidemiological and/or molecular evidence.  In North America, the Milwaukee outbreak, together with smaller outbreaks associated with recreational water, served to increase awareness of Cryptosporidium and to lead to the introduction of preventive measures (for example, ozonization of drinking water supplies, and hyperchlorination of swimming pools), as well as recognition of the risks associated with contact with infected animals or their environment.

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