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G.I. Infectious Diseases Michael R. Lappin, DVM, PhD, DACVIM Colorado State University Update on Infectious Gastrointestinal Diseases
Clinical problem and differentials. Vomiting is the forceful ejection of stomach and proximal duodenal contents through the mouth. Vomiting can be induced by vestibular, vagal, chemoreceptor trigger zone, or direct input to the emetic center. Diarrhea is a characterized by increased frequency of defecation, increased fluid content of the stool, or increased volume of stool. Markedly increased frequency of defecation, small volume stools, tenesmus, urgency, hematochezia, and mucus are consistent with large bowel diarrhea. Slight increase in frequency of defecation, large volume, melena, steatorrhea, and polysystemic clinical signs are more consistent with small bowel diarrhea. Mixed bowel diarrhea is a combination of characteristics or clinical signs. Gastrointestinal (GI) signs can be the result of primary diseases of the GI system or secondary GI diseases. The secondary GI diseases are generally those of the kidneys, liver, pancreas (pancreatitis or exocrine pancreatic insufficiency), endocrine system (hyoadrenocorticism; diabetic ketoacidosis; hyperthyroidism), or central nervous system. Differential diagnoses for primary GI diseases are often grouped into obstruction (masses, foreign body, and intussusception), dietary intolerance, drugs/toxins (garbage gut), inflammatory gastric and bowel diseases, neoplasia, infectious diseases, and parasites. The primary bacteria associated with gastrointestinal tract disease in dogs and cats include Salmonella spp., Campylobacter spp., Clostridium perfringens, Helicobacter spp., bacterial overgrowth syndrome, bacterial peritonitis, and bacterial cholangiohepatitis. The primary viral agents include parvoviruses, coronaviruses, canine distemper virus, feline leukemia virus, and feline immunodeficiency virus. The primary nematodes are Ancylostoma/Uncinaria, Trichuris vulpis (dogs), Strongyloides, Dirofilaria immitis (vomiting in cats), Toxocara, Toxascaris leonina, Ollulanus tricuspis (cats), and Physaloptera spp. (Table 1). Enteric protozoans include Giardia spp., Cystoisospora spp., Cryptosporidium spp., and Tritrichomonas foetus. The cestodes Taenia, Dipylidium, and Echinococcus generally cause subclinical infection. Diagnostic plan. Occasionally, otherwise healthy dogs or cats with acute vomiting and normal physical examination findings can be handled conservatively by withholding food for 24 hours followed by introduction of a bland food for several days. For all animals with diarrhea with no apparent cause on physical examination, I will perform a fecal flotation, fecal wet mount examination, complete blood cell count (CBC), and rectal cytology if diarrhea is present. While the CBC generally does not lead to a specific diagnosis, the presence of eosinophilia makes inflammatory bowel diseases and parasitism more likely. I perform acid-fast staining of a fecal smear on all animals with diarrhea to assess for the presence of Cryptosporidium parvum oocysts. A wet mount will aid in identifying trophozoites of Tritrichomonas and Giardia. If neutrophils or spirochetes are evident on rectal cytology I recommend fecal culture for Salmonella spp. and Campylobacter spp.. If spore-forming rods consistent with Clostridium perfringens are present in large numbers, fecal enterotoxin assay can be performed to help confirm the diagnosis but this test can be positive in healthy animals as well and so has less than 100% predictive value. A biochemical profile, urinalysis, FeLV antigen assay (cats), and FIV antibody assay (cats) are indicated if secondary GI diseases are on the differential list or if dehydration is present. I generally perform a total T4 on all cats with vomiting or small bowel diarrhea that are greater than 5 years of age. While amylase and lipase are poor predictors of pancreatitis in cats, a pancreatic lipase immunoreactivity assay has now been validated. It can be used to diagnose pancreatitis (increased) in dogs and cats. The positive predictive value is better for acute pancreatitis than chronic pancreatitis. If an animal with suspected pancreatitis has abdominal effusion, assay lipase concentrations in the serum and effusion; if pancreatitis is occurring the effusion lipase is usually greater than serum. Fecal fat assessment with Sudan IV stain can help confirm malabsorption/maldigestion but is not specific for a single disease. If the MCV is low, chronic iron deficiency should be suspected; this occurs almost exclusively with gastrointestinal diseases. A serum iron panel can be used to confirm iron deficiency. Panhypoproteinemia is often associated with gastrointestinal tract disease. Imaging techniques include radiographs, contrast radiographs, and ultrasound. I commonly perform abdominal radiographs in dogs and cats to support my palpation findings. I use contrast radiographs occasionally; often I perform endoscopy or exploratory laparotomy based on abdominal radiographic findings. Ultrasound of the intestinal tract can be hard to interpret and is operator dependent. Diagnosis of gastric foreign bodies and diffuse inflammatory diseases can be made by endoscopy. Endoscopically obtained biopsies are small; I generally take at least 8-10 biopsies from stomach, duodenum, colon, and ileum if possible. Even if a lesion is present, endoscopically obtained biopsies can be falsely negative requiring full thickness biopsies. Gastric biopsies should be placed on urea slants to assess for urease which is found in the cell wall of Helicobacter spp.. DIAGNOSTIC PROCEDURES FOR INFECTIOUS DISEASES Infectious diseases are common causes of vomiting and diarrhea. The following is a brief discussion of the commonly used procedures. Direct smear. Liquid feces or feces that contains large quantities of mucus should be microscopically examined immediately for the presence of protozoal trophozoites, including those of Giardia spp. and Tritrichomonas foetus. A direct saline smear can be made to potentiate observation of these motile organisms. The amount of feces required to cover the head of a match is mixed thoroughly with one drop of 0.9% NaCl. Following application of a coverslip, the smear is evaluated for motile organisms by examining it under 100X magnification. The sample should be fresh. The material for evaluation should be collected from the surface of the fecal material, preferably mucous if present. Alternately, a rectal scraping can be used. Stained smear. A thin smear of feces should be made from all animals with large or small bowel diarrhea. Material should be collected by rectal swab if possible to increase chances of finding white blood cells. A cotton swab is gently introduced 3-4 cm through the anus into the terminal rectum, directed to the wall of the rectum, and gently rotated several times. Placing a drop of 0.9% NaCl on the cotton swab will facilitate passage through the anus, but not adversely affect cell morphology. The cotton swab is rolled on a microscope slide gently multiple times to give areas with varying smear thickness. Following air drying, the slide can be stained. White blood cells and bacteria morphologically consistent with Campylobacter jejuni or Clostridium perfringens can be observed after staining with Diff-Quick or Wright's-Giemsa stains. Histoplasma capsulatum or Prototheca may be observed in the cytoplasm of mononuclear cells. Methylene blue in acetate buffer (pH 3.6) stains trophozoites of the enteric protozoans. Iodine stains and acid methyl green are also used for the demonstration of protozoans. Acid-fast or monoclonal antibody staining of a fecal smear should be performed in cats with diarrhea to aid in the diagnosis of cryptosporidiosis. Cryptosporidium parvum is the only enteric organism of approximately 4 to 6 µ in diameter that will stain pink to red with acid-fast stain. Presence of neutrophils on rectal cytology can suggest inflammation induced by Salmonella spp., Campylobacter spp., or Clostridium perfringens; fecal culture is indicated in these cases. Fecal enterotoxin measurement should be considered for animals with spore-forming rods morphologically consistent with C. perfringens. However, approximately 10% of healthy animals have enterotoxin in stools and so the predictive value of a positive test is not 100% in animals with diarrhea. Fecal flotation. Cysts, oocysts, and eggs in feces can be concentrated to increase sensitivity of detection. Most eggs, oocysts, and cysts are easily identified after zinc sulfate centrifugal flotation. This procedure is considered by many to be optimal for the demonstration of protozoan cysts, in particular, Giardia spp. and so is a good choice for a routine flotation technique in practice. Sugar centrifugation can be used for routine parasite evaluation and may be superior to many techniques for the demonstration of oocysts of Toxoplasma gondii and Cryptosporidium spp.. Giardia cysts are distorted by sugar centrifugation but can still be easily identified. Fecal sedimentation will recover most cysts and ova, but will also contain debris. This technique may be superior to flotation procedures for the documentation of Eurytrema procyonis, the pancreatic fluke. Strongyloides larva may be easier to identify after concentration using the Baerman funnel technique. Culture. Culture of feces for Salmonella spp., Campylobacter spp., and Clostridium perfringens is occasionally indicated in small animal practice. Approximately 2-3 grams of fresh feces should be submitted to the laboratory immediately for optimal results, however, Salmonella and Campylobacter are often viable in refrigerated fecal specimens for 3-7 days. Appropriate transport media should be available through your laboratory. The laboratory should be notified of the suspected pathogen so appropriate culture media can be used. More than 1 culture may be needed to prove infection. Tritrichomonas foetus can be cultured from feces of cats in general practice using a commercially available kit (InpouchTM, Biomed Diagnostics). Some Giardia spp. isolated from cats will grow on culture media, but this technique is not generally performed in small animal practice. Immunologic techniques. Parvovirus, Cryptosporidium parvum, and Giardia spp. antigen detection procedures are available for use with feces. Canine parvovirus antigen assays appear to detect feline parvovirus antigen. An IFA for concurrent detection of C. parvum oocysts and Giardia cysts has been validated for use with dog and cat feces; this assay is commonly available at commercial laboratories. While Giardia spp. antigen assays appear to detect dog and cat genotypes, human Cryptosporidium antigen assays do not. The Giardia assays can be used to increase sensitivity in dogs or cats with diarrhea but should be interpreted in conjunction with results from fecal examination techniques. Serum antibodies against D. immitis can be measured in cat serum but positive test results do not prove current infection or disease induced by D. immitis. FeLV can cause lymphoma and induces the panleukopenia-like syndrome. FIV has been associated with lymphoma and can cause enteritis. Detection of FIV antibodies or FeLV antigen in serum documents exposure, but does not prove that clinical disease is due to the virus. The only way to document that gastrointestinal signs are due to FeLV or FIV is to exclude other known causes. Electron microscopy. Electron microscopy can be used to detect viral particles in feces of dogs and cats with gastrointestinal signs of disease. Approximately 1-3 grams of feces without fixative should be transported to the laboratory by overnight mail on cold packs. Endoscopy or exploratory laparotomy. Ollulanus and Physaloptera rarely pass ova in feces and so frequently are diagnosed only by endoscopy. Diagnosis of diffuse inflammatory diseases can be made by evaluation of endoscopy or surgically obtained tissue samples. Endoscopically obtained biopsies are small; I generally take at least 8-10 biopsies from stomach, duodenum, colon, and ileum if possible. Even if a lesion is present, endoscopically obtained biopsies can be falsely negative requiring full thickness biopsies. In these cases, full thickness biopsies can be made by exploratory laparotomy or by laparoscopy and a key hole biopsy. Gastric biopsies should be placed on urea slants to assess for urease which is found in the cell wall of Helicobacter spp.. The combination of inflammation, exclusion of other causes of inflammation, presence of gastric spiral bacteria, and positive urease testing can be used as a presumptive diagnosis of gastric helicobacteriosis. There is no benefit to performing duodenal aspirates for quantitative bacterial cultures or Giardia trophozoite evaluations in cats; the normal bacterial count range is very broad in cats and Giardia is found in the distal small intestine. Regional enteritis due to feline infectious peritonitis can be confirmed by documenting the organism in tissue after immunohistochemical staining. Polymerase chain reaction As discussed, the diagnosis of Giardia spp. infection is generally made with the combination of fecal flotation techniques, wet mount examination with or without fecal antigen tests or IFA. Fecal PCR assays for Giardia are often falsely negative because of PCR inhibitors in stool and so PCR should not be used as a screening procedure for this agent. However, Giardia spp. PCR can be used to determine whether the infective species is a zoonotic assemblage which is the primary indication for this technique. However, it now appears that assemblage determination should be performed on more than one gene for most accurate results. Genotyping is available at Colorado State University (http://dlab.colostate.edu/; Test code = 437). While Cryptosporidium spp. infection is common, it is unusual to find C. felis or C. canis oocysts after fecal flotation. Acid-fast staining of a thin fecal smear is cumbersome and insensitive. Antigen assays titrated for use with human feces are inaccurate when used with cat or dog feces. Thus, PCR may be aid in the diagnosis of cryptosporidiosis in dogs and cats and has been shown to be more sensitive than IFA in cats. I personally recommend IFA as the screening procedure. Cryptosporidium spp. PCR assays are indicated in IFA negative cats or dogs with unexplained small bowel diarrhea and when the genotype of Cryptosporidium is to be determined (http://dlab.colostate.edu/; Test code = 438). However, C. felis and C. canis infections are common and so positive tests results do not always prove that the agent is the cause of the clinical disease. No drug is known to eliminate Cryptosporidium spp. infections and small animal strains are not considered significant zoonotic agents so PCR is never indicated in healthy animals. PCR assays are also available for detection of DNA of Tritrichomonas foetus, Salmonella spp., Campylobacter spp., Clostridium spp., parvoviruses, and T. gondii and a RT-PCR assay is available for coronaviruses. Trophozoites of T. foetus can often be detected on wet mount examination of fresh feces which can be completed as an in clinic test. DNA of T. foetus can be detected in healthy carrier cats and so positive results do not always prove illness from the organism. Cases with suspected salmonellosis or campylobacteriosis should be cultured rather than assessed by PCR to determine the anti-microbial susceptibility patterns. In dogs, the PPV of Clostridium spp. PCR assays on feces is low and if used, should be combined with enterotoxin assays. Information in cats is currently lacking. There is no current evidence that parvovirus PCR on feces is superior to currently available antigen assays. In one recent study in our laboratory, approximately 40% of healthy cats vaccinated with a modified live FVRCP vaccine were PCR positive for panleukopenia virus DNA in feces one week after vaccination. Thus, the currently used assays cannot differentiate vaccine strains from natural infections which should be considered when making case management decisions. Toxoplasma gondii is only shed for about 7-10 days and millions of oocysts are generally shed during this time making the organism very easy to identify. Thus, PCR assays are usually not needed to diagnosis this infection. Because virus isolation is not practical clinically, RT-PCR is used most frequently to detect coronaviruses RNA in feces. However, positive test results do not differentiate FIP inducing strains from enteric coronaviruses and in one study in our laboratory, there was no association between diarrhea and positive test results for coronavirus in cat feces. INFECTIOUS DISEASE TREATMENT OPTIONS There are multiple drugs used in the treatment of gastrointestinal parasitic infections. For all puppies and kittens, the strategic deworming recommendations for the control of hookworm and roundworm infections from the Centers for Disease Control and the American Association of Veterinary Parasitologists should be followed by veterinary practitioners. (http://www.cdc.gov/ncidod/dpd/parasites/ascaris/prevention.htm). Heavily infected kittens should be administered an anthelmintic at 3, 5, 7, and 9 weeks of age and then periodically monitored or treated. Puppies can be started at 2 weeks of age. If the kitten is not presented to the clinic until 6-8 weeks of age, administer the anthelmintic at least 2-3 times, 2-3 weeks apart. Pyrantel pamoate and fenbendazole are usually effective drugs for use in strategic deworming programs and for the treatment of nematodes causing gastrointestinal tract disease. Albendazole is more likely to cause hematologic side-effects than fenbendazole and so should not be used in cats. Even if anthelmintics for hookworms and roundworms are administered, a fecal flotation should be performed to evaluate for other parasites. Some monthly D. immitis preventatives can help control or eliminate some nematode infections as well as prevent heartworm infection. Use of these products is the easiest way to perform strategic deworming after completion of the vaccine and deworming series. Dipylidium and T. taeniaformis infestations usually are eliminated by praziquantel or espiprantel; fenbendazole is effective for Taenia taeniaformis. Since Echinococcus multilocularis can be a significant zoonosis transmitted to cats by carnivorism, hunting cats in endemic areas should be treated up to monthly. Administration of a pyrantel/praziquantel combination may be effective in these cats since praziquantel is approved for the treatment of Echinococcus and roundworms are also transmitted by carnivorism. Giardia infections often respond clinically to the administration of metronidazole but infection is usually not eliminated. Administration of metronidazole benzoate at 25 mg/kg, q12hr, PO, for 7 days was effective in suppressing cyst shedding to below detectable limits in 26 cats. This is the maximum dose of metronidazole that should be used; CNS toxicity can be induced by overdosing or as a cumulative neurotoxin. Until recently, fenbendazole has not been studied for the treatment of giardiasis in cats. In eight cats with Cryptosporidium spp. and Giardia spp. coninfection, only 50% eliminated the Giardia infection when treated with fenbendazole. I personally use fenbendazole at 50 mg/kg, PO, daily for at least 5 days in dogs or cats with giardiasis. Metronidazole and fenbendazole can be combined in resistant cases. Febantel containing products have been used successfully in dogs and cats. The empirical dog dose is the deworming dose, daily for at least 3 days. The empirical cat dose is 2 small dog tablets per cat, daily for 5 days. Vaccination as immunotherapy has been used successfully in one dog study, but failed in an experimental study in cats. Paromomycin, ronidazole, and nitazoxanide are alternate drugs that could be tried in cases with resistant giardiasis. However, in my experience, dogs or cats with Giardia that fails to respond to metronidazole and fenbendazole have another underlying problem. Multiple drugs have been evaluated for the treatment of cats with T. foetus infections; until recently no drug eliminated infection and diarrhea rarely resolves during the treatment period. Recently ronidazole at 20 mg/kg, PO, q24hr, for 14 days eliminated clinical signs of disease and trophozoites from cats infected with one strain of the organism. In another one small study, administration of metronidazole and enrofloxacin lessened diarrhea in kittens but it is unknown if the organisms infecting those cats was T. foetus. It is possible that some cats with T. foetus have other enteric coinfections and so antihelmintics or drugs with activity against Giardia spp., Cryptosporidium spp., and enteric bacteria like Campylobacter spp. are often prescribed. Paromomycin should be avoided cats with bloody stools because of the potential for being absorbed and inducing renal disease or deafness. Some puppies have recently been shown to be infected by T. foetus. Pentatrichomonas hominis DNA is also sometimes amplified from some dogs or cats with diarrhea. Sequential administration of clindamycin followed by tylosin blocked oocyst shedding and resolved diarrhea in one cat with chronic, clinical cryptosporidiosis. Tylosin (10-15 mg/kg, PO, twice daily) has been apparently successful in lessening diarrhea and oocyst shedding in multiple other cats and dogs with diarrhea that were Cryptosporidium positive. However, infection is not eliminated. Unfortunately, tylosin is very bitter and usually has to be given to cats in capsules. Treatment duration may need to be weeks. In cats with naturally occurring cryptosporidiosis, response to azithromycin has been variable (Lappin MR, unpublished data, 2005). If tried, use 10 mg/kg, PO, weekly for at least 10 days. If responding, continue treatment for at least 1 week past clinical resolution. Paromomycin can be effective for lessening diarrhea and oocyst shedding associated with cryptosporidiosis in cats and also is an alternate anti-Giardia drug. However, this orally administered aminoglycoside may cross the diseased intestinal wall to induce renal insufficiency and should never be used in cats with bloody diarrhea. Nitazoxanide is a new drug being studied for the treatment of Cryptosporidium and Giardia. Little information is available concerning dosages, but we have used 25 mg/kg, PO, q12 hours in some of our research studies. The drug Alinia is available and is labeled for both organisms in humans. The primary side-effect to date has been vomiting and so it should be given with food if used. The Toxoplasma gondii oocyst shedding period can be shortened by administration of clindamycin or sulfadimethoxine. Cystoisospora spp. generally respond to the administration of sulfadimethoxine, other sulfa-containing drugs, macrolides, or ponazuril. Ponazuril is superior to other drugs and can be given at 20 mg/kg, PO, q24 hours twice, or 50 mg/kg, PO, once. If there are multiple puppies or kittens with diarrhea, treatment of all in contact animals should be considered. Since many of the gastrointestinal parasites that infect dogs and cats are transmitted by carnivorism, they should not be allowed to hunt or be fed raw meats. Additionally, infection of by many parasites results from ingestion of contaminated water. Clinical disease in some parasitized animals can be lessened by eliminating stress and providing a quality diet and clean environment. Clostridium perfringens and bacterial overgrowth generally respond to treatment with tylosin, metronidazole, ampicillin, amoxicillin, or tetracyclines. The drug of choice for campylobacteriosis is erythromycin; however, oral administration of tylosin or quinolones is often less likely to potentiate vomiting. Salmonellosis should only be treated parenterally due to rapid resistance that occurs following oral administration of antibiotics. Appropriate antibiotics for the empirical treatment of salmonellosis while awaiting susceptibility testing results include ampicillin or trimethoprim-sulfa; quinolones are also effective. Helicobacter spp. infections are usually treated with the combination of metronidazole and tetracycline or amoxicillin and metronidazole in dogs. Clarithromycin or azithromycin may be logical choices in cats since the species is often difficult to treat with multiple drugs. Whether to concurrently administer an antacid like famotidine is controversial but seems to lessen vomiting in some dogs and cats. Dogs or cats with apparent bacteremia due to enteric bacteria should be treated with parenteral antibiotics with a spectrum against anaerobic and gram negative organisms. The combination of enrofloxacin with a penicillin or first generation cephalosporin is generally effective. Intravenous metronidazole can also be used. Second generation cephalosporins or imipenem are also appropriate choices. Dogs or cats that have hepatic infections and signs of bacteremia should be treated with antibiotics that kill gram positive, gram negative and anaerobic bacteria as discussed before. Non septic hepatic infections generally respond to amoxicillin, first-generation cephalosporins, or chloramphenicol. Decreasing numbers of enteric flora by oral administration of penicillins, metronidazole, or neomycin can lessen the clinical signs of hepatic encephalopathy. Panleukopenia virus, feline leukemia virus, feline immunodeficiency virus, and coronaviruses are the most common viral causes of gastrointestinal tract disease in cats. Viral diseases are managed by supportive treatment. Make sure to maintain hydration, correct hypoglycemia, and maintain normal potassium concentrations. Use of jugular catheters is superior to leg veins since blood samples can be drawn and CVP can be measured. Based on results in dogs with parvovirus infection, administration of plasma or serum (1 ml/kg) from your hyperimmune blood donor cat may lessen morbidity in cats with panleukopenia due to passive transfer of immunity. This is effective because parvoviruses induce a viremic state; virus particles are complexed by the antibodies transferred passively. Antibiotics effective against gram negative and anaerobic bacteria are commonly indicated. Vaccines are available for the prevention of parvovirus, coronaviruses, and feline leukemia virus infection. Histoplasma capsulatum infection is the most common fungal infection of the gastrointestinal tract of dogs and cats in some regions of the United States. Treatment with itraconazole or fluconazole can be effective. Zoonotic considerations. Infection of people by animal associated enteric agents is usually from contact with feces in the environment, by ingestion of contaminated food or water, or by ingestion of undercooked meat (T. gondii). Contact with infected dogs and cats is an unlikely way for humans to acquire infection. The following guidelines may lessen the risk of transfer of enteric zoonotic agents to people.
Giardia SPP. Infection Update
Are most Giardia spp. infections shared between animals and man? The genus Giardia contains multiple species of flagellated protozoans that are indistinguishable morphologically (1). Infection in dogs and cats are common (2-4). Host specificity was thought to be minimal for Giardia spp., but not all small animal isolates cause disease in human beings. There have been varying results concerning cross-infection potential of Giardia spp.. Human Giardia isolates usually grow in cell culture, animal isolates often do not. Recent genetic analysis has revealed 2 major genotypes in people. Assemblage A (G. duodenalis) has been found in infected humans and many other mammals including dogs and cats. Assemblage B (G. enterica) has been found in infected humans and dogs, but not cats. It appears that there are specific genotypes of Giardia that infect dogs (G. canis; Assemblages C and D). cats (G. felis; Assemblage F) but not people. Accordingly, healthy pets are not considered significant human health risks for HIV infected people by the Centers for Disease Control (www.cdc.gov/hiv/pubs/brochure/oi_pets.htm). The majority of dogs and cats with Giardia spp. infections are infected with animal specific genotypes (5). Assemblage A is occasionally found in pets, especially cats (6). What are the optimal diagnostic tests? The primary diagnostic tests that are available to document Giardia spp. in dog and cat feces include direct smear of feces, direct saline preparation, fecal flotation, fecal antigen testing, fecal immunofluorescent antibody assay, and fecal PCR assay (7,8). These tests can be used alone or in combination in the diagnosis of Giardia spp. infection in dogs or cats with diarrhea. How should I evaluate fresh feces from animals with diarrhea? Fresh, liquid feces or feces that contain large quantities of mucus should be microscopically examined immediately in the clinic for the presence of protozoal trophozoites of Giardia spp. (small bowel diarrhea), Tritrichomonas foetus (large bowel diarrhea), and Pentatrichomonas hominus (large bowel diarrhea). A direct saline smear can be made to potentiate observation of these motile organisms. A 2mm X 2mm X 2mm quantity of fresh feces is mixed thoroughly with one drop of 0.9% NaCl or water. The surface of the feces or mucus coating the feces should be used as the trophozoites are most common in these areas. After application of a coverslip, the smear is evaluated for motile organisms by examining it under 100 magni?cation. Culture (T. foetus), antigen testing (Giardia) or PCR (T. foetus or Giardia) can be used to distinguish between specific organisms. What are my options for fecal flotation? Fecal flotation with zinc sulfate centrifugal flotation technique (specific gravity 1.18-1.20) is one of the optimal techniques for the demonstration of cysts (www.capcvet.org) and is more sensitive for detection of Giardia spp. cysts than cup flotation. Sugar and other salt solutions lead to distortion of cysts but are also sensitive tests. Cysts are shed intermittently and their presence does not correlate to clinical signs of disease. Evaluation of a single fecal flotation has a sensitivity of approximately 70%. Sensitivity increases to > 90% if at least 3 stool specimens are examined. Do currently available antigen ELISA detect dog and cat strains of Giardia? Multiple ELISAs for detection of Giardia antigens in stool are available. In experiments performed in our laboratory, all human and veterinary assays assessed to date have detected G. canis and G. felis. In one of our dog studies (5), all 17 Giardia assemblage C and D isolates were detected by the commercially available ELISA labeled for use with dog and cat feces (SNAP®Giardia, IDEXX Laboratories, Portland, ME). In other studies where we have compared different Giardia antigen assays results to IFA, there can be up to a 5% false positive and 5% false negative rate. Thus, some animals that are Giardia spp. cyst negative on fecal flotation but Giardia antigen positive may have a false positive antigen test. Does the IFA produced for human feces detect Cryptosporidium spp. and Giardia spp. in dog and cat feces. The IFA for simultaneous detection of Giardia spp. cysts and Cryptosporidium oocysts is currently available in most commercial veterinary laboratories and has been shown in our laboratory to identify C. felis, C. canis, G. felis, and G. canis. I use this assay as a first "wave" diagnostic test in dogs and cats with diarrhea (combined with a fecal float, a wet mount examination, and a rectal or fecal cytology) instead of Giardia antigen tests because it detects two important agents with zoonotic potential and because Cryptosporidium spp. are usually not detected on fecal flotation in small animals. When should I use PCR for amplification of Giardia spp. DNA from feces? The sensitivity of the currently available PCR assays are low and should not be used in lieu of fecal flotation or other tests. These assays should only be used if genotyping of the previously detected Giardia spp. is desired. Genotyping is available at Colorado State University; call 970 297 0367 concerning sample submission. Which dogs and cats should be screened for Giardia spp. infections and what tests should I use? In healthy dogs and cats, a fecal flotation should be performed at least once or twice yearly. In dogs and cats with diarrhea, the combination of the direct saline preparation with fecal flotation should be used as the minimum initial screening tests for Giardia infection. Antigen tests or IFA test can be used to increase sensitivity but should not be used in lieu of the other assays. What are the best drugs for Giardia spp. infection? Giardia spp. have specific antimicrobial sensitivity patterns like bacteria and so it is currently impossible to predict which anti-Giardia drug will be effective. Because G. canis and G. felis can be difficult to cultivate and there is little in vitro susceptibility test results available. While there have been multiple drugs used for the treatment of giardiasis in dogs and cats, there are few studies that utilized dose titrations and evaluation of drugs in experimentally infected animals (9-18). In most studies, fecal samples were only assessed for short periods of time after treatment and immune suppression was not induced to evaluate whether infection was eliminated or merely suppressed. Infection with Giardia does not appear to cause permanent immunity and so reinfection can occur, a finding that also hampers assessment of treatment studies. Treatment options currently available or used historically include metronidazole, tinidazole, ipronidazole, ronidazole, fenbendazole, albendazole, pyrantel/praziquantel/febantel, quinacrine, and furozolidone (Table 1). Newer drugs being studied include paromomycin and nitazoxanide. How do I choose which of these drugs to use? If spore-forming rods, morphologically consistent with Clostridium perfringens are concurrently detected with Giardia, use of metronidazole is indicated as this drug is an antibiotic. If there is clinical evidence to suggest concurrent infection with a nematode, like eosinophilia or eosinophils on fecal cytology, fenbendazole or febantel are indicated. Some clinician's currently recommend the combination of metronidazole and fenbendazole. What are the goals of Giardia spp. treatment? The primary goal of treatment is to stop diarrhea. Because healthy pets are not considered human health risks, elimination of infection (which is difficult) is a secondary goal. What do I do if diarrhea continues and Giardia infection is still detected? Giardia spp. can have resistant patterns and so if the first drug fails to clear the infection (cysts or antigen) or resolve the diarrhea, a second drug from an alternate class is indicated. The addition of fiber to the diet may help control clinical signs of giardiasis in some animals by helping with bacterial overgrowth or by inhibiting organism attachment to the microvillus. Immunotherapy with the Giardia vaccine has aided in the elimination of cyst shedding and diarrhea in some infected dogs. However, in a controlled study in 16 experimentally infected cats, vaccination as immunotherapy was ineffective with one strain of Giardia. Probiotic administration may also be beneficial in some animals. In one study, bathing the dog was a beneficial adjunct therapy. In dogs and cats with persistent diarrhea and Giardia spp. infection, a more extensive workup to attempt to diagnoses other underlying diseases is indicated if several therapeutic trials fail. Common underlying disorders include cryptosporidiosis, inflammatory bowel disease, bacterial overgrowth, exocrine pancreatic insufficiency, and immunodeficiencies. Should healthy dogs and cats with Giardia infection be treated? Healthy pets are not considered significant human health risks by the Centers for Disease Control (www.cdc.gov/hiv/pubs/brochure/oi_pets.htm). However, because clinical signs induced by Giardia spp. can be intermittent and since some Giardia spp. may be zoonotic, treatment of healthy infected animals should be considered with each owner. Treatment of healthy animals is controversial because all of the drugs have side-effects, animals with normal stools are not considered human health risks, treatment is unlikely to eliminate infection, and re-infection can occur within days. For example, in a recent study of naturally infected healthy dogs, we induced clinical side-effects in 50% of the dogs treated with nitazoxanide or fenbendazole and of the dogs that the treatment protocol was completed successfully, 62.5% were still infected on recheck evaluation (9). Should healthy housemates of Giardia infected dogs and cats be tested? Whether to test all housemates of infected dogs and cats is controversial because all Giardia tests can give false negative results. It may be more financially viable to treat all animals in lieu of testing. Should healthy housemates of Giardia infected dogs and cats be treated? Whether to treat all healthy housemates of infected dogs or cats is controversial because all of the drugs have side-effects, animals with normal stools are not considered human health risks, treatment is unlikely to eliminate infection, and re-infection can occur within days. However, the safety margin of fenbendazole is very wide and so if treatment chosen, this drug seems to be a reasonable choice. Should I follow Giardia test results after treatment? It is currently unknown how long Giardia antigens will persist in feces after successful treatment (resolution of diarrhea). In one of our small studies on treated healthy dogs, four of eight dogs treated with nitazoxanide or fenbendazole were still SNAP®Giardia positive on day 34 after treatment. The AAFP Advisory Panel on Zoonoses recommends attempting to remove the source of infection during the treatment period and performing a fecal flotation (not an antigen test) after Giardia treatment one time, within 2-4 weeks after the end of the treatment period (www.aafponline.org). If the animal is healthy and negative for cysts, retesting is not indicated again until the next scheduled fecal flotation. What should I do with dogs or cats that have normal stool and are Giardia antigen positive, Giardia cyst negative? These animals have either a low grade infection or have false positive antigen test results. To further evaluate the case, the veterinarian can perform an IFA test, 2 additional fecal flotations, or an antigen test from a different manufacturer; if these other test results are negative, the antigen test was likely falsely positive. What can I do to prevent re-infection with Giardia spp. Prevention involves boiling or filtering of water collected from the environment prior to drinking and disinfection of premises contaminated with infected feces with quaternary ammonium compounds (1 minute contact time). Transport hosts should be controlled and treatment of all animals in the environment could be considered. To date, no study has shown the Giardia spp. vaccines licensed for dogs and cats to have lessened Giardia spp. infections in the field and so both vaccines have been classified by AAHA and AAFP as generally not recommended as preventatives. The feline Giardia spp. vaccine was recently discontinued by the manufacturer. Table 1. Drugs used for the treatment of Giardia spp. infections.
C = canine; F = feline; B = canine and feline References
Update on the Diagnosis and Management of Feline Stomatitis Stomatitis is a common and often debilitating disease of cats. Lesions can range from mild inflammation to severe deep ulcerations and proliferative tissue affecting the gingiva, fauces, buccal mucosa and tongue. Treatments are primarily symptomatic with variable success rates. Most cats must be treated life-long and many undergo multiple tooth extractions in an attempt to maintain the cat's appetite and control the oral pain. Despite aggressive therapy, some cats with severe caudal stomatitis do not respond and are euthanized to due weight loss, pain and poor quality of life. Histological evaluation of affected tissues generally reveals infiltrations of lymphocytes and plasma cells. The cause of the syndrome is unknown in most individual cats, but most likely is a combination of a precipitating infectious agent or agents and an abnormal, hyperactive, immune response. Autoimmune reactions and dietary hypersensitivities are also proposed causes. It is likely different forms of the syndrome have different causes. Feline calicivirus is the viral agent that has been implicated most frequently; feline herpesvirus 1 may also be associated with the syndrome in some cats. In one of our recent studies, we could amplify the RNA of calicivirus from approximately 40% of biopsies obtained from cats with chronic lymphocytic plasmacytic-stomatitis (Dowers et al, 2009). However, FCV reverse transcriptase PCR results on oral swabs obtained from cats with and without stomatitis did not correlate to the presence of disease in one small study (Quimby et al, 2007). Stomatitis is more common in cats infected with feline leukemia virus or feline immunodeficiency virus and may relate to activation of other infections as immune suppression develops. Bartonella henselae is a gram-negative organism that causes cat scratch disease in people. Up to 80% of the cats in serologic surveys have serum antibodies suggesting previous or current infection. Approximately 40% of cats tested are blood culture positive and the infection can be persistent. When first discovered in the blood of cats, the organism was felt to cause subclinical infection. However, recently B. henselae has been associated with fever, uveitis, neurologic disease, gingivitis, and lymphadenopathy in some experimentally infected or naturally infected cats. The organism is harbored inside erythrocytes and endothelial cells of cats where it is likely to partially evade the immune response, possibly explaining persistence of infection. It is possible that this persistent infection induces aberrant immune responses resulting in the clinical manifestations. Cat scratch disease in people is associated with a multitude of possible immune-mediated manifestations like fever, uveitis, and lymphadenopathy. There is conflicted evidence linking Bartonella spp. exposure to gingivostomatitis in cats. Serum antibodies can persist for months to years after infection is eliminated and so do not denote current Bartonella spp. infection. In a study of cats with caudal stomatitis in our laboratory, the positive and negative predictive value of Bartonella spp. antibodies in cats with and without stomatitis was poor (Quimby et al, 2007). Use of culture to grow the organism from blood or polymerase chain reaction (PCR) to amplify Bartonella spp. DNA from blood can be used to document infection. Documentation of infection by culture or PCR versus documentation of exposure by detection of serum antibodies is likely to be superior for use in epidemiologic studies as it confirms infection. However, healthy cats can be positive as well limiting the positive predictive value in cats with stomatitis or other clinical manifestations of potential bartonellosis. A complete blood cell count, serum biochemical panel, urinalysis, FeLV antigen test, and FIV antibody test should be completed to evaluate for systemic diseases associated with stomatitis. A dental examination should be performed and abnormal teeth repaired or removed. Biopsy for histopathological examination should be performed, particularly if a mass is present. Squamous cell carcinoma can sometimes appear similar to severe proliferative stomatitis. As mentioned previously, diagnostic tests for feline calicivirus, feline herpesvirus 1, and Bartonella spp. have low predictive value and so whether to perform these tests are controversial. There is no one therapeutic protocol that is effective for every cat (Table 1). After abnormal teeth are repaired or removed during the initial diagnostic workup, antibiotics are generally used to control secondary infections, and potentially disease from B. henselae. Clindamycin is an excellent antibiotic for stomatitis owing to effects against anaerobes and penetration into bone. If administered cold, the liquid formulation available in the United States is often well-tolerated. I use doxycycline frequently in mild cases of feline gingivostomatitis because of efficacy against normal flora, effect against B. henselae, and an anti-inflammatory effect. Doxycycline can be liquefied in tuna flavoring and administered once daily. For those owners that cannot afford the formulation fee, doxycycline tablets or capsules can be administered followed by water or other liquids, administered coated in butter, or administered within pill delivery treats to avoid esophageal retention, esophagitis, and potential for strictures. Azithromycin is an alternate drug with effect against bartonellosis that is also anti-inflammatory. This antibiotic is expensive but can be administered q72hr which is beneficial for cats with extremely painful mouths. Long-term or pulse antibiotic therapy are required in some cats. Analgesic therapy is indicated for most cats. Those with mild disease are usually administered buprenorphine for the first three days after the initial diagnostic workup. Those with severe disease may benefit from the administration of non-steroidal anti-inflammatory agents (NSAIDS). Anti-inflammatory therapy is often used non-specifically. If used, oral administration of prednisolone is preferred, but injectable projects are often needed because of the difficulty associated with administering oral drugs. For some cats, NSAIDS are more effective in controlling pain and also the inflammation. I generally use meloxicam in cats with this syndrome. If used chronically, I follow both renal tests as well as the PCV as gastrointestinal bleeding can occur without vomiting. Resistant cases may respond to administration of cyclosporine at up to 7.5 mg/kg, PO, daily or every other day but controlled data is lacking. Trough blood levels should be checked 2 weeks after starting cyclosporine to make sure that excessive blood levels are not achieved which may activate infectious diseases. The cytotoxic agent, chlorambucil has been tried with variable responses in some cats but can be difficult to administer. Some cats respond to gold salts; 8-10 weeks of an induction period are required, followed by monthly maintenance therapy. In the United States, human interferon alpha products are commonly available. In one recent study, low dose oral interferon therapy (30U/kg, PO, daily) improved quality of life in cats with FIV infections. The effect of oral interferon is thought to be from mediation of inflammatory cytokines and the cytokine may have beneficial effects in cats with other causes of stomatitis as well. Feline omega interferon injected into the affected tissues has been effective in some cats and is available in some countries. Coating the affected tissues with bovine lactoferrin by mixing with milk has about a 15% positive response rate. Bovine lactoferrin can be purchased from Emerson Ecologics in the United States by calling 800-654-4432. Use of a hypoallergenic diet and omega 3/omega 6 fatty acid supplements is beneficial for some cats since the syndrome may result from a dietary hypersensitivity. CO2 laser ablation has been effective for the treatment of some cats but this form of therapy does not have universal acceptance by veterinary dentists in the United States. Up to 80% of affected cats have a positive response to extraction of all teeth in the area of the inflammation. Care should be taken to make sure all dental tissues are removed. It is the opinion of many veterinary dentists in the United States that this form of therapy should be considered early in the course of the syndrome in severely affected cats. The concern is that ineffective medical management prior to tooth extractions can lessen the efficacy of this therapy. Medical treatments frequently attempted for feline idiopathic stomatitis.
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