Chapter 19 Reptile Protozoa
Many species of protozoa are found in reptiles. Some may be commensal or nonpathogenic organisms, and others are pathogens. The two most clinically important diseases are cryptosporidiosis and amebiasis. Each of these parasites may infect a wide range of reptilian species and are easily transmitted because of direct life cycles.
Taxonomy and Life Cycle
Cryptosporidiosis is caused by parasites of the apicomplexan genus Cryptosporidium, which has a worldwide distribution. Thirteen species of Cryptosporidium are recognized, two of which, C. serpentis and C. saurophilum, are recognized as pathogens of reptiles.31 Nonreptilian Cryptosporidium spp. are not infective for reptiles.11 Additional species likely exist that have not been completely described at this time. Infected snakes, lizards, and chelonians may become subclinical carriers or diseased.3,17,29
The following life cycle is described for Cryptosporidium parvum in a mammalian host. The actual life cycle of C. serpentis has not been described. Infection occurs when the animal ingests a thick-walled, sporulated oocyst that excysts within the lumen of the stomach or intestine, producing four sporozoites, which penetrate the mucosal epithelial cells and develop into trophozoites inside the parasitophorous vacuoles. The trophozoites undergo asexual divisions to form merozoites. The merozoites invade nearby cells and either undergo asexual reproduction of more merozoites or form type II meronts, which in turn form the sexual stages, microgamonts and macrogamonts. After fertilization a zygote is formed and may develop into a thin-walled oocyst that is autoinfective or a thick-walled oocyst that is passed from the host. The cysts are infective immediately on passing from the host.7
Lizards and chelonians often exhibit poor growth or appetite, weight loss, or other, nonspecific findings, as well as postprandial regurgitation and passing of undigested food items.10,17,27 Snakes may be subclinically infected for years but capable of shedding the organism intermittently. Clinical signs of cryptosporidiosis in snakes may include anorexia, lethargy, intermittent or chronic regurgitation of undigested prey several days after feeding, chronic weight loss, and firm midbody swelling caused by gastric hyperplasia.3 Enteritis with severe inflammatory infiltrates without clinical or histologic signs of gastritis has been seen in wild-caught, rough green snakes (Opheodrys aestivus) and a garter snake (Thamnophis sirtalis). Sudden death was the presenting sign in these snakes.2 Three iguanas (Iguana iguana) presented with aural-pharyngeal polyps.28
Diagnosis may be challenging because oocysts are shed intermittently.
Cryptosporidium spp. oocysts 4 to 8 μm long may be detected on light microscopy in unstained specimens of feces, mucus, or gastric lavage samples, although they may be confused with oocysts of other coccidia. Detection may be enhanced by the use of an acid-fast stain. Further increases in sensitivity may be achieved through the use of concentration techniques, such as Sheather’s sugar flotation. The acid-fast stain and sugar flotation techniques have been published elsewhere.6,7 The use of a commercially available immunofluorescent antibody (IFA) stain (Merifluor, Meridian Bioscience, Cincinnati, OH 45244) increases the sensitivity of the test approximately 16-fold over acid-fast staining for C. serpentis.14 The Merifluor IFA reacts weakly with the type of Cryptosporidium sp. typically found in geckos.12 A positive finding on fecal or gastric wash samples does not necessarily indicate infection with C. serpentis or C. saurophilum. A negative finding may merely indicate lack of shedding.
In reptiles with mild subclinical infections, gastric lavage provides a better sample than feces or mucus. The lavage sample is best performed 3 days after feeding the snake. The increased gastric metabolism after a feeding is thought to increase Cryptosporidium metabolism, resulting in more parasites available for detection.16
It is not possible to determine whether cysts obtained from feces or gastric lavage are nonreptilian Cryptosporidium sp. cysts passing through the gastrointestinal (GI) tract, and therefore not infective to the host, or whether the cysts are C. serpentis or C. saurophilum, and thus represent an infection. Histopathology samples collected by gastric biopsy indicating that the gut epithelial cells are infected are necessary to prove the snake has cryptosporidiosis. Gastric biopsy may be performed using endoscopy or surgery.
Necropsy and Histopathology
The pathologic changes seen grossly have been covered extensively in the literature and include increase in the diameter of the stomach, edematous rugae, thickening of the gastric mucosa, decreased lumen size, mucosal petechiae, and focal necrosis in predominantly gastric cases and mild to severe enteritis in other cases.2,3 Histologically, in infected animals, the organisms are seen on the brush border of the epithelial cells. There may be little damage to the architecture in subclinical cases, or substantial changes may occur, including loss of the brush border, hyperplasia and hypertrophy of gastric glands, proliferation of gastric mucous cells, edema and inflammation of the submucosa and lamina propria, reduced luminal diameter, and inflammation of the mucosal layer.3
In reptiles with intestinal infection, few gastric lesions may be seen. A marked infiltrative enteritis is usually present, with abundant parasites in the enterocytes.2,17,27
Cryptosporidiosis is extremely difficult to eradicate. Anticoccidial drugs are not effective at eliminating Cryptosporidium at nontoxic doses in humans or animals. Some pharmacologic agents may reduce the number of organisms or eliminate fecal shedding, but the organism is still present in the GI tract.6,15
The most promising treatment for reptilian cryptosporidiosis to date has been the use of hyperimmune bovine colostrum (HBC); however, this product is not commercially available. HBC eliminated shedding but did not eliminate the organisms from the stomachs of snakes or the intestines of geckos within a 6-week treatment period. However, a longer treatment period may have succeeded in eliminating the organisms. HBC did eliminate infection in savanna monitors.12,13,18 Further work is needed in the area of treatment of cryptosporidiosis for all species.
Prevention and Management
The cysts are extremely hardy and resistant to standard disinfection procedures, including sodium hypochlorite (bleach) and povidone-iodine. Heat above 60°C, thorough desiccation, or freezing may be used for appropriate nonorganic materials. Ammonia and 10% formalin were effective after 18 hours of contact.4 At the Maryland Zoo in Baltimore, during the years of Cryptosporidium research, ammonia foot baths were used at the door of each reptile quarantine room; ammonia was used for disinfection of tools and equipment after removal of organic matter; separate tools were used for each room in quarantine; and the keepers were knowledgeable and well trained regarding prevention of transmission of Cryptosporidium organisms. Despite this, transmission occurred from one group of snakes to another snake housed in a different room.2
The safest way to eliminate cryptosporidiosis from a collection is through strict quarantine protocols with rigorous testing procedures and elimination of infected individuals. Reptiles with subclinical infections may appear healthy but serve as a source of infection for other reptiles for many years. Reptiles should not be euthanized based on the presence of Cryptosporidium in fecal or gastric samples without confirmation of actual disease in the stomach or intestine by biopsy and histopathology. It is difficult to justify the euthanasia of reptiles testing positive for cryptosporidiosis in quarantine unless the entire current reptile collection has previously been tested and found negative.
Endangered reptiles with cryptosporidiosis that are not considered candidates for euthanasia, even if infected, should be kept strictly isolated from noninfected reptiles. Periodic testing and treatment when shedding may help prolong the quality of life and breeding potential of these endangered reptiles and may reduce transmission to the rest of the collection. There is no evidence of vertical transmission; eggs or offspring produced and removed from the adult environment immediately should have a reduced chance of infection.