Gastrointestinal protozoa (Proceedings)

Giardia

• Flagellated protozoan parasite found in the GIT of animals including man.

• Prevalence: General population cats; <1% to 13.6% (most reports - 10%). Shelter cats: Can be as high as 80% in cats housed in high density rooms. [Worth noting that prevalence data may depend on the sensitivity of the test used: IFA – 14%, IDEXX SNAP Giardia test – 11%]

• Although prevalence can be high, clinical disease is rare, and often not dependent on prevalence – more often dependent on age (young cats, higher incidence of signs).

Life Cycle

• Direct with infection via oral route. Entire development occurs in the GIT. Cysts in feces.

• 2 forms: 1) teardrop-shaped trophozoite (active motile form found in GIT), 2) cyst or dormant resistant stage responsible for transmission.

• Cyst capable of surviving for several months outside host in wet cold condition. Can also survive chlorinated water. Very susceptible to desiccation under dry hot conditions.

Clinical Findings

• Most infections are asymptomatic.

• Diarrhea is main sign. May be acute, short-lived, intermittent, or chronic. Feces pale, malodorous, and steatorrheic. May see weight loss (diarrhea) but rarely inappetence.

Diagnosis

• Clinical signs and tests (CBC, serum chemistry, radiology etc) are non-specific.

• Unequivocal diagnosis relies on finding cysts or trophozoites (or products of these) feces or samples taken from the GIT.

Fecal Smear

• Easy, non-invasive, and very specific but low sensitivity. Trophozoites more likely passed in loose feces, especially in cats.

• Drop of fecal material mixed with a drop of normal saline, cover slipped, examine at 40X. Trophozoites easily identified by rapid forward motion, and concave ventral disc. Trichomonads (the only other organism that looks like Giardia has a rolling motion, no concave disc, single nucleus, and an undulating membrane. Stain organisms with Lugols iodine to confirm.

Zinc Sulfate Concentration Technique (ZSCT)

• Use a 1.18 SG ZnSO4 solution with centrifugation. Needs trained personal to conduct test as cyst shedding (particularly in dogs) is variable (from a few to 50,000/gm of feces in 3 days)

• 1.18 SG ZnSO4 solution may not float heavier eggs (e.g. Taenia spp.) so can use a modified 1.27 SG Sheather sugar solution which will also get Giardia. [Dryden's Modified Sheather's Solution (SG 1.27): 454 gm granulated sugar, 355 ml tap water, 6 ml formaldehyde. Dissolve sugar (gentle heat), check SG (hydrometer), filter (course filter paper) if not clear.]

• Performing 3 tests on fresh fecal samples collected over a 3 to 5 day period (because cysts are shed intermittently) is considered the gold standard. One test is about 70% efficacious, 2 tests are about 93% efficacious, and 3 tests are about between 95 and 100% efficacious.

• Problem:- not very practical in a private practice situation. Cysts may be confused with yeast

• May ship samples refrigerated (cysts live for 2 days at 4C, but will not survive formalin). ZSCT also excellent for nematode eggs.

Fecal ELISA Kits

• ELISA microplate tests – usually conducted in diagnostic laboratories as can run multiple tests at a time. Developed for use in man originally (96% specific, 100% sensitive).

• IDEXX – SNAP® Giardia test - for cats and dogs – in-house – 8 minute test. Compared to ELISA microplate test, this SNAP test is 92% sensitive, and 99.8% specific). Detects a trophozoite protein in feces. Can use frozen samples or feces fixed in formulin. [Worth noting that while 6 of 27 veterinary practices can identify a known positive Giardia sample using flotation techniques, 27 or 27 correctly diagnosed the sample using the IDEXX SNAP Giardia test.] May remain positive for up to 3 weeks post-treatment (according to IDEXX).

Direct Immunofluorescent Test

• Uses fluorescent labeled monoclonal antibodies to detect cysts. As effective as the ZSCT in man.

• Used in both cats and dogs: some studies suggest that it is more sensitive than ZSCT as picks up lower numbers of cysts/gm of feces (certainly the case in cattle).

• Needs to be performed in a diagnostic laboratory (needs a fluorescent microscope).

• Use fresh feces or fix in formulin.

Others

• Duodenal aspirates.

• Impractical to perform endoscopy and duodenal aspirates under general anesthesia to rule out Giardia. However, reasonable to collect a sample during endoscopy for examination.

• Method: flush 10ml of saline through polyethylene tube via endoscope channel, then aspirate immediately. Centrifuge sample (150g, 10min), examine sediment (wet mount or stain).

• Peroral string test.

• Method for collecting duodenal contents from people using a commercially available peroral technique (Entero-Test, HDC Corp, San Jose, Ca). Used in third world countries.

• In tests performed in dogs with Giardia infections, none were positive and one dog developed a string foreign body. Extremely insensitive, impractical, risks string foreign body – don't use!

Differential Diagnosis (Other Protozoa)

Tritrichomoniasis – cats

• Enteric pear-shaped motile flagellated protozoa similar to Giardia – inhabits the large intestine of cats, dogs and man.

• Tritrichomonas foetus (cause of infertility/abortion in cattle) – causes diarrhea in cats.

• T. foetus prevalence in cats - ~30% in show cats, but very low in feral or indoor cats.

• Pathogenic factors leading to infected cats developing diarrhea – endogenous bacterial flora, adherence of parasite to host epithelium, and cytotoxin and enzyme elaboration.

• Parasites colonize the terminal ileum, cecum and colon – leads to large bowel diarrhea.

• High population density (catteries, shelters) – may be a risk factor for infection.

• Co-infection with Giardia – common.

• Young cats – usually under 1 yr (range: 3 mths – 13 yrs).

• Diagnosis: Direct fecal smear [Method – dilute fresh feces 50:50 in saline, coverslip, examine at 40 X objective with condenser lowered to increase contrast - low sensitivity] or Fecal protozoal culture – use in-house culture system (In Pouch® TF, Biomed Diagnostics, San Jose, Ca). Giardia will not grow [Method – inoculate with 0.05 gm fresh feces, incubate at room temperature, examine for motile trophozoites daily for 12 days.]

• Distinguish from Giardia (concave ventral disc, spiral forward motion) – T. foetus has jerky forward motion, spindle-shaped, undulating membranes.

• T. foetus trophozoites – are not seen on fecal flotation and will not survive refrigeration.

• Treatment: None specific.

Cryptosporidiosis – Dogs and Cats

• Coccidian protozoan (genus Cryptosporidium) which can cause gastrointestinal disease in dogs and cats (as well as man, calves, and rodents).

• Ubiquitous in nature and have a worldwide distribution.

• Life cycle: enteric (like coccidians). Infection occurs when sporulated oocysts are ingested; sporozoites are released to penetrate intestinal epithelial cells. After asexual reproduction, merozoites are released to infect other cells. Prepatent period: Cats; 5 to 10 days.

• Like humans, infection rarely causes clinical disease in dogs and cats. Nearly all reports are in immunocompromised animals.

• Dogs: virtually all clinical cases have occurred in animals ≤ 6-mths; older dogs excrete oocysts without showing clinical signs. Cats: nearly all immunocompromised.

• Treatment: Paramomycin, tylosin, azithromycin.

Giardia Treatment

• Determine why treatment is important: 1) Zoonotic considerations although no clinical signs [NIH web page states: "an infected family dog with diarrhea may pass the parasite to human family members" – presumably same for cat!] 2) Clinical signs consistent with giardiasis, 3) Prevent spread to naive population, 4) As part of a colony "clean up" policy.

• Determine if the drug to be used is appropriate and safe for use in the patient.

• Consider other intercurrent diseases the patient may have which may benefit from drugs used to treat Giardia, or may prevent the use of certain drugs.

• Consider re-infection after treatment, and how this affects interpretation of drug efficacy results.

Specific Drugs

1. Fenbendazole [Dose: 50mg/kg, PO, q24h for 5 consecutive days].

• Benzimidazole derivative. Inhibits microtubule assembly by inhibiting tubulin polymerization.

• Dogs: 3 consecutive doses were found to be 95-100% efficacious. Cats: treated with above dose, 4 of 8 (50%) became completely negative, and others showed improvement in cyst shedding but not all become negative.

• May not be ideal drug in cats but probably is in dogs.

• Also used to remove whipworms, roundworms, hookworms, and the tapeworm, Taenia pisiformis. No side effects, not teratogenic, and safe in cats.

2. Febantel, praziquantel, pyrantel pamoate combination Drontal-Plus® (Bayer Corp, Shawnee, Kan). [Dose: 5.4-7mg/kg praziquantel, 26.8-35.2mg/kg pyrantel, 26.8-35.2mg/kg febantel in combination (label dose for Drontal-Plus®), q24h, for 3 consecutive doses].

• Febantel is metabolized in the liver to fenbendazole and oxfendazole; most anti-Giardia activity is ascribed to the fenbendazole although a synergistic activity of the other drugs likely.

• Close to 100% efficacy. No side effects reported at above doses by vomiting and non-formed feces reported in dogs receiving 5 times label dose for 3 consecutive days.

• Cats: ½ a Drontal Plus tablet (for small dogs)/kg, PO, q24h, for 5 days is close to 100% eff.

3. Albendazole [Dose: 25mg/kg, PO, q12h for 4 doses].

• Benzimidazole derivative (same mechanism as fenbendazole). > 90% efficacy.

• Suspect of being teratogenic. Effective in cats when used for 5 days.

• No side effects reported in beagles treated with 30mg/kg q24h for 13 weeks. Reported to cause bone marrow toxicosis with pancytopenia in a dog and cat.

4. Metronidazole: Dogs: Dose of 25mg/kg, PO, q12h for 5 days reported to be 67% effective.

• Cats: Dose of 25 mg/kg, PO, q12h for 7 days reported to be 100% effective.

• Associated with anorexia, vomiting progressing to neurologic signs of ataxia and nystagmus, but usually only at higher doses than those above.

5. Oxfendazole: Dogs: Dose: 11.3 mg/kg, PO, q24h for 3 days. Not used in cats.

• In European, very effective in dogs when used in association with cleaning procedures.

• In the US, only available as a cattle de-wormer (Synanthic Bovine Dewormer Suspension® 9.06%, Fort Dodge, Fort Dodge, Ia).

6. Quinacrine hydrochloride: [Dose: 6.6 mg/kg, PO, q12h for 5 days].

• 100% effective at this dose but complicated with a 50% side effect rate (lethargy, fever which stop 2 to 3 days after treatment stops).

7. Furazolidine: [Dose – Cats: 4mg/kg, PO, q12h for 5 to 10 days].

• High efficacy but side effects include diarrhea and vomiting. Teratogenic.

Vaccination

• Released by Fort Dodge early in 2000.

• Made of inactivated trophozoites

• Pups and kittens immune to giardia challenge 6 and 12mths after 1 injection and booster.

• Reduces cyst excretion on older dogs (non-immune).

• Has been used to treat dogs – given to dogs with chronic infection to reduce load.

• Some studies show that vaccination fails to prevent infection

• Not efficacious in cats already infected.

• Probably little utility for its use except for odd case that does not respond to drugs.

Control

Drugs may not remove organisms for GIT but only inhibit cyst production.

• In wet cold environments, cysts survive for long periods. These, as well as cysts on the hair coat of pet are a source of infection.

• 4 approaches to control:

1. Decontaminate environment by creating "clean areas".

2. Treat animals with highly efficacious drugs, and rotate drugs used.

3. Clean cysts from coats of animals.

4. Prevent re-introduction of infection.

• If only a few animals are involved, move them all out of the facility while it is cleaned. In large facility, create a "clean area" a few cages at a time on a rotation basis once animals are moved out to a holding facility.

• Treat animals with efficacious agents (3 - 5 day course of fenbendazole or Drontal Plus for dogs; a 7 day course of metronidazole for cats) prior to moving: move on the last day of Tx.

• Remove all solid fecal material

• Steam clean facility or clean with quaternary ammonium-containing disinfectants (use at manufacturers recommended concentration – cysts are very susceptible).

• Allow facility to dry thoroughly – facility is now considered "clean".

• Prior to moving animals back in to "clean area", bath with all-purpose pet shampoo, rinse shampoo off coat, then wash coat (especially perianal area) with quaternary ammonium compound, then rinse thoroughly from coat (can irritate mucus membranes if left on for long periods of time – leave on only for 1 minute). Dry hair coat thoroughly, preferably blow dry.

• When return animals to "clean area," treat with a second course of drugs.

• Prevent re-infection of "clean area" by treating and bathing animals prior to introduction.

• Prevent fomite transmission by physically cleaning boots of people entering facility.

Zoonosis

Assemblage A and F have been identified in cats from the USA and Europe, but only D in Australia. Little specific information on dogs but only D to date!!

Reference

Vasilopulos RJ, Mackin AJ, Rickard LG, et al. Prevalence and factors associated with fecal shedding of giardia spp. in domestic cats. J Am An Hosp Assoc 42:424-429, 2006.

Calin EP, Bowman DD, Scarlett JM, et al. Prevalence of Giardia in symptomatic dogs and cats throughout the United States as determined by the IDEXX SNAP Giardia test. Vet Therapeut. 7:199-206, 2006.

Scorza AV, Radecki SV, Lappin MR. Efficacy of a combination of febantel, pyrantel, praziquantel for the treatment of kittens experimentally infected with Giardia spp. J Fel Med Surg 8:7-13, 2006.

Dryden MW, Payne PA, Smith V. Accurate diagnosis of Giardia spp. and proper fecal examination procedures. Vet Therapeut 7:4-14, 2006.

Barr SC. Giardiasis. In: Infectious diseases of the Dog and Cat. Greene CE (ed) 3rd ed. Saunders, Philidelphia. Pp 736-742, 2006.

Barr SC. Giardiasis. In: Canine and Feline Infectious Diseases and Parasitology. Barr SC and Bowman DD (ed). Blackwell, Ames. Pp 259-264, 2006.

Vasilopulos RJ, Rickard LG, Mackin AJ et al. Genotypic analysis of Giardia duodenalis in domestic cats. J Vet Intern Med 21:352-355, 2007.