CP Exam 4 Notes PDF

CLINICAL PATHOLOGY I - Lecture Overview of Protozoan Parasites & The Flagellates Giardia spp. and Tritrichomonas spp. The Protozoans Single-celled organisms Most are free-living Some occur as parasites in mammals...

CLINICAL PATHOLOGY I - Lecture Overview of Protozoan Parasites & The Flagellates Giardia spp. and Tritrichomonas spp. The Protozoans Single-celled organisms Most are free-living Some occur as parasites in mammals Only very small percentage of protozoa associated with disease Some flourish in face of diarrhea, and often mistaken as cause of diarrhea Protozoan Family Tree Flagellates o Giardia spp.** o Tritrichomonas foetus. Ciliates o Balantidium coli ** Amoebae o Entamoeba histolytica ** o Entamoeba invadens o Entamoeba muris Coccidians o Eimeria spp. o Cystoisospora spp. o Sarcocystis spp.** o Cryptosporidium spp.** o Toxoplasma gondii ** The Flagellates Giardia spp.: A Protozoan Flagellate Species ▪ Giardia intestinalis or Giardia duodenalis** (originally known as G. lamblia): Mammals (including humans), birds and reptiles ▪ Giardia muris: Rodents and birds ▪ Giardia agilis (formerly ranae): Frogs **ZOONOTIC Not That Black and White ▪ Probably other species? ▪ Perhaps they are all the same? ▪ All animals are susceptible: Mammals (including humans), reptiles, birds, frogs, etc. 10/19/2019 1  o Generally not a problem in livestock and wild ungulates o Horses can become infected, but rarely exhibit clinical signs Two Distinct Life Stages Trophozoites ▪ Active, motile, feeding form located in small intestine ▪ Teardrop-shaped flagellate: Motile and binucleate (“tennis racket with eyes”) ▪ Not always found in feces: Transform into infective cysts in colon before passing out in feces ▪ Outside body, this form is incapable of causing infection and will soon die Cysts ▪ Walled, dormant, transmission stage ▪ Trophozoites encyst while passing through colon ▪ Usually phase shed in feces; transmission stage from one host to another ▪ Immature cyst contains two nuclei ▪ Mature, infective cyst contains four nuclei which will become four trophozoites in next DH ▪ Can survive for days to months if environment cool and moist ▪ Can be a common problem in some localities (like Colorado) ▪ Cysts are infective immediately upon passage Life Cycle = Direct ▪ Transmission is by ingestion of food or water contaminated with infective cysts (oral transmission) ▪ Following ingestion, trophozoites emerge from cyst and colonize small intestine ▪ Trophozoites attach to mucosa of small intestine and damage cellular structure ▪ Damaged cells are unable to perform absorptive function, thus causing diarrhea ▪ Often subclinical ▪ Some % of cats and dogs harbor Giardia spp. without any clinical signs Clinical Signs ▪ Acute OR chronic ▪ Stool normal OR abnormal ▪ Small bowel diarrhea: Pale, large volume, gassy, semi-formed ▪ Steatorrhea: Excessive amount of fat in feces; malabsorption syndrome ▪ Weight loss and/or failure to gain ▪ Incubation period ~ 5 days ▪ Prepatent period ~ 1 to 2 weeks Diagnostic Dilemma ▪ May have normal stools with Giardia spp. visible on fecal exam 10/19/2019 2  ▪ Cysts may be excreted intermittently ▪ Results may be “NOO”, yet have Giardia: Must have three consecutive NOO fecals before diagnosis can be deemed truly NOO ▪ One of more difficult cysts to identify (especially if you see it rarely) Diagnosis via Cysts ▪ Cysts are most likely to be seen in semi-formed or formed stool ▪ Using ZnSO4 as flotation solution, may perform o Standard flotation technique o Centrifugal flotation technique ▪ Lugol’s iodine will facilitate identification ▪ Cysts are small, non-motile oval organisms ▪ Cysts are smaller than trophozoite form Diagnosis via Trophozoites ▪ More likely to be found in fluid feces ▪ Perform direct smear: Examine fresh fecal smear using normal saline as diluent (if needed) ▪ Lugol’s iodine may assist in visualizing trophozoite, but it kills organism, eliminating any motility that may assist in identification of Giardia spp. ▪ Dried smears can be stained with Wright’s or Giemsa stain to confirm presence of trophozoites Diagnosis via ELISA test (IDEXX) ▪ Read and follow all directions ▪ For detection of Giardia spp. antigen in feces of dogs and cats ▪ Feces may be used fresh, previously frozen, or refrigerated for up to 7 days Treatment ▪ Metronidazole (Flagyl®) ▪ Fenbendazole (Panacur®) Bathe pet thoroughly following treatment Prevention ▪ Sanitation: Giardia spp. is transmitted only by fecal/oral contamination o Outdoor areas are difficult to treat successfully o Indoor surfaces ▪ Clean, dry kennels will inhibit cyst survival ▪ Lysol®, chlorine bleach, and iodine will kill cysts ▪ Avoid contamination of food and water ▪ Avoid drinking untreated water in mountain streams (filter, boil or treat w/ iodine) ▪ Vaccine available from Ft. Dodge 10/19/2019 3  o Separate vaccines for dogs and cats o “Approved by USDA as an aid in prevention of disease and reduction in severity of shedding caused by Giardia spp.” Public Health Considerations ▪ Giardia spp. is zoonotic ▪ Carrier state in animals should be treated to avoid human exposure ▪ 7% of world’s human population harbor Giardia spp. Tritrichomonas spp. and Trichomonas spp.: Other Protozoan Flagellates Tritrichomonas spp. and Trichomonas spp. ▪ Tritrichomonas foetus o Venereal disease of cattle o Diarrhea in cats ▪ Trichomonas vaginalis: Venereal disease of humans ▪ Trichomonas gallinae: Oral lesions in birds Tritrichomonas foetus in cattle ▪ Life cycle is direct; transmission by sexual exchange of seminal or vaginal fluids ▪ Venereal disease o Cows: Clinical signs include vaginitis, placentitis, abortions, and decreased fertility o Bulls: Infection is subclinical Diagnosis ▪ Culture and identification of organism from o Preputial wash o Uterine or vaginal discharges o Fluid from aborted fetus ▪ Characteristically pear-shaped trophozoites with 3anterior flagella, undulating membrane, and trailing flagellum along free edge Treatment ▪ Cull bull and rest cows from all breeding activity for 4 to 5 months ▪ Metronidazole; not 100% effective ▪ Vaccine available; not highly effective Tritrichomonas foetus in cats ▪ Life cycle is direct; transmission believed to be fecal-oral ▪ Organism located in colon (sometimes ileum and cecum) ▪ Clinical signs o Large-bowel diarrhea: Loose to semi-formed, blood and mucus, malodorous o Flatulence o Tenesmus o Fecal incontinence 10/19/2019 4  o Weight loss o Fever ▪ More commonly diagnosed in cats less than two years of age ▪ More common in densely-populated conditions; e.g., catteries Diagnosis ▪ Direct fecal smear w/ 0.9% NaCl o Fresh feces that has not been refrigerated o Characteristically pear-shaped trophozoites with 3 anterior flagella, undulating membrane, and trailing flagellum along free edge o Can be mistaken for Giardia spp. trophozoites, but motility is different ▪ Tritrichomonas foetus: Erratic, forwardly progressive movement ▪ Giardia spp.: “Falling-leaf” movement ▪ Fecal culture: InPouch TF Feline® (BioMed Diagnostics) ▪ PCR: Polymerase Chain Reaction – “gold standard” but cost can be prohibitive Treatment ▪ No approved treatment available; some success with off-label protocols such as ronidazole ▪ Recurrence of diarrhea is common ▪ May spontaneously resolve within two years of onset 10/19/2019 5  CLINICAL PATHOLOGY I – Lecture Amoebae and Cilia Entamoeba spp. and Balantidium coli Single-celled protozoan parasites ❖ Amoebic: Entamoeba spp.** (some species) Amoeba: Singular Amoebae: Plural ❖ Ciliated: Balantidium coli** Cilium: Singular Cilia: Plural Genus & species o Entamoeba histolytica** and Entamoeba dispar**: Humans, other primates, (dogs and cats) o Entamoeba invadens: Reptiles o Entamoeba muris: Mice and rats o Balantidium coli**: Primates, swine, rats, (dogs and cats) **Zoonotic Life Cycle o Direct o Fecal-oral transmission via contaminated food and water o Humans are a common source of infective cysts for pets o We infect our pets; “reverse zoonosis” o Prepatent period = 1 to 2 weeks o Reproduce asexually via binary fission o E. histolyticaor: binary fission o B. coli: transverse binary fission Two life stages o Trophozoites o Cysts Trophozoites: Actively parasitic ▪ Live in large bowel ▪ Most likely to pass with diarrhea (not stimulated to encyst) ▪ Will not survive outside body of host (exception is B. coli – trophozoites can encyst after passage in feces) ▪ Not infective Cysts ▪ Trophozoites “encyst” in large bowel as they dehydrate, then pass in feces ▪ More likely to pass in formed feces ▪ Infective upon passage 10/25/2018 1  ▪ Contaminate food and water supplies ▪ Once ingested, “excyst” in small intestine ▪ Resulting trophozoites colonize large intestine ▪ Sometimes, Entamoeba spp.** trophozoites secrete proteolytic enzymes that destroy intestinal epithelium allowing organism to enter host tissue o E. dispar: Remains in intestinal wall o E. histolytica: Penetrates intestinal wall and... ▪ Can form large abscesses in intestinal tract that allow parasite to enter blood, move to liver (and other organs) and form abscesses ▪ In extra-intestinal sites, trophozoites cause extensive tissue destruction ▪ **B. coli can invade tissues of intestinal tract resulting in painful ulcerations, but does not disseminate to other organs Extra-Intestinal Sites: E. hystolytica o Liver is most common site of amoebic abscess o Can be found in ❖ Lungs ❖ Spleen ❖ Kidneys ❖ Brain Diseases o Amoebiasis = Infection with Entamoeba spp. o Balantidiasis = Infection with B. coli Clinical Signs o Can be subclinical (especially in pets) o Can cause chronic diarrhea o Can cause acute diarrhea -- aka “Montezuma’s revenge” (i.e., traveler’s diarrhea) “Amoebic Dysentery” o Amoebic dysentery: Intestinal inflammation with abdominal pain, frequent diarrhea with blood and mucus o Enteritis: Inflammation of small intestine o Colitis: Inflammation of colon o Colic: Severe abdominal pain o Tenesmus: Inability or difficulty to defecate (or urinate), accompanied by pain and cramping o Weight loss and anorexia o Dehydration Diagnosis Trophozoites o Can be found in liquid samples o Best to evaluate with direct smear and look for motility o May use iodine stain (Lugol’s), but will kill trophozoite and eliminate motility 10/25/2018 2  o Fecal float will not demonstrate trophozoites Cysts o Rarely seen in dogs and cats o May utilize standard concentration techniques (flotation) to identify o Serologic testing for presence of Ab o Most important technique for recovery and i.d. is permanent stained smear ▪ Trichrome or iron hematoxylin stain ▪ 300 fields examined under oil-immersion Treatment = Metronidazole Prevention o Sanitation and disinfection to limit transmission o Avoid contaminated water and food supplies o Isolate animals from human feces Public Health Concerns o Entamoeba histolytica: Concern for caretakers of non-human primates (reservoir hosts) in zoo or research facilities o Balantidium coli: May be contracted by humans working with non-human primates, pigs, and rats (reservoir hosts) o Amoebiasis and balantidiasis are serious diseases of humans, but dogs and cats are NOT reservoirs for human infection ▪ Most often, dogs and cats acquire disease from owners or other humans ▪ If pet is infected, veterinary professionals should inform owner of possible infection in family members 10/25/2018 3  CLINICAL PATHOLOGY I - Lecture Coccidia (“Apicomplexans”) Another Type of Protozoan Coccidia overview o Intestinal coccidia primarily parasitize small intestine of DH o Microscopic, single-celled, spore-forming parasites o Obligate intracellular parasites: Utilize host’s cells for reproduction and multiplication o Host-specific, with exception of Cryptosporidium spp. o Often subclinical, but can be devastating to host Grammar o Coccidium = Noun; singular o Coccidia = Noun; plural o Coccidial / coccidian = Adjective; “pertaining to coccidia” **** DO NOT confuse this with “cocci” which is a circular bacteria and has absolutely nothing to do with coccidia**** Genus and species to know o Eimeria spp.: Cattle, sheep, poultry, rabbits, horses, birds o Cystoisospora spp.: Dogs, cats, swine (potential PH = mouse) – formerly Isospora spp. o Cryptosporidium spp.: People and a wide variety of other hosts – ZOONOTIC o Sarcocystis spp.: Dogs, cats, people (IH = ruminant, swine, rodent) – ZOONOTIC o Toxoplasma gondii: Cats (IH = virtually any warm-blooded animal) – ZOONOTIC The Disease o Coccidiasis: Infection of coccidial organism with no clinical signs o Coccidiosis: Disease caused by any coccidial organism; i.e., clinical signs are present o Organisms colonize small intestine and cause enteritis Life Cycles o Very complex, many stages, may include more than one host o Variable, depending on genus ⋅ Facultative: Not obligatory; has ability to adjust to circumstances ⋅ Obligatory: Necessary, compulsory; can only survive in particular environment ⋅ Heteroxenous: Requiring more than one host to complete life cycle (indirect) ⋅ Homoxenous: Requiring only one host to complete life cycle (direct) General Information o There are no ▪ Adults ▪ Eggs ▪ Larvae o Undergo both asexual and sexual reproduction o Prepatent period = 1 – 2 weeks 1 10/20/2024 Coccidian Family Tree o Gamete: Reproductive cell ▪ Male = spermatozoon ▪ Female = ovum o Zoite: Functional unit of coccidian life ▪ Tachyzoite: Rapidly dividing life stage in blood and lymph ▪ Bradyzoite: Slowly dividing life stage encysted in tissue ▪ Sporozoite: Infective form found in oocysts following sporulation; invades host cells to begin asexual reproduction o Oocyst: Resistant stage of coccidia; transmission stage Basic Life Cycle o Oocysts shed in feces -- undivided spheres in fresh feces o Oocysts sporulate (divide) in approximately 1 week and become infective o Sporulated oocysts each contain 8 infective sporozoites o Sporulated (infective) oocysts must be ingested by DH to complete life cycle o Following ingestion, oocyst wall is digested releasing sporozoites which invade intestinal cells and begin asexual reproduction Asexual & Sexual Reproduction o Asexual reproduction of sporozoites results in exponential increase in organisms o Host cells destroyed with every round of asexual reproduction o Asexual reproduction stops after fixed number of repetitions (depending on genus of coccidium) and sexual reproduction begins o Male and female gametes fuse to form oocyst o Host cell ruptures to release oocyst which passes in feces of DH Life Cycle Variations Between Genus o Eimeria spp. (homoxenous) ▪ DH = Ruminants, horses, rabbits, birds ▪ Basic life cycle with one important exception Sexual reproduction of Eimeria bovis occurs in cells of large intestine Typically results in bloody diarrhea o Cystoisospora spp. (facultative heteroxenous) ▪ DH = Dogs, cats, swine ▪ Life cycle direct, but is facultative heteroxenous coccidium Mice can act as paratenic host for C. felis and C. canis (organism encysts in mouse’s tissue) Cats and dogs acquire Cystoisospora spp. by ingesting Paratenic host OR Infective oocysts o Cryptosporidium spp. (homoxenous with autoinfection) ▪ Zoonotic ▪ DH = Infects wide variety of hosts; i.e., NOT host-specific 2 10/20/2024  ▪ Autoinfection possible Thick-walled oocysts shed in feces Thin-wall oocysts release sporozoites into lumen of host’s intestine; reinitiating infection ▪ Can be fatal to immunocompromised host o Sarcocystis spp. (Obligatory heteroxenous) ▪ Zoonotic ▪ DH = Dog, cat, human IH = Ruminant, swine, rodent ▪ Indirect life cycle: Oocysts are sporulated and infective to IH upon passage in feces of DH All asexual reproduction occurs inside IH: IH ingests sporulated oocysts, sporozoites develop to tachyzoites, which develop to bradyzoites that encyst in IH tissues All sexual reproduction occurs inside DH: When DH consumes bradyzoites in raw flesh of IH, sporozoites released and sexual reproduction begins Typically not pathogenic to DH, but can cause serious disease in IH: FAD, weight loss, anemia, ataxia, abortion, death o Toxoplasma gondii: (Facultatively heteroxenous) To be discussed in separate lecture -- zoonotic Clinical Signs of Coccidiosis o Often subclinical (coccidiasis), especially in adult animals o Severe outbreaks may occur in ▪ Puppies and kittens in crowded, poorly cleaned catteries, kennels, pet shops ▪ Calves in feedlots o Secondary factors ▪ Plane of nutrition ▪ Overall immune status ▪ Concurrent disease o Each reproductive cycle (asexual and sexual) results in more zoites with more ruptured and dead intestinal cells → enteritis o Diarrhea ▪ Often green, mucoid (bloody in calves) ▪ Can be copious and last for weeks if not treated (results from destruction of intestinal epithelium by hordes of multiplying organisms) o Dehydration o Weakness o Weight loss and/or failure to gain o FAD: Fever, anorexia, depression o Host cell death directly related to degree (severity) of infection o In acute cases, clinical signs can precede shedding of oocysts o Once exposed to specific coccidium, immunity is developed, however..... o Immunity can break with stress Diagnosis o Oocysts found in feces 3 10/20/2024  ▪ Standard fecal flotation ▪ Direct smear o Serologic testing available for T. gondii and Cryptosporidium spp. Treatment o *Not justified without signs of disease* o Sulfadimethoxine only drug approved for treatment of coccidial infections o Coccidiocidal: Severe cases ▪ Kills coccidia ▪ Does not permit immune response o Coccidiostatic: Less severe cases ▪ Stops replication but does not kill coccidia ▪ Permits immune response o Coccidiocidal drugs ▪ Semduramicin ▪ Toltrazuril o Coccidiostatic drugs ▪ Amprolium ▪ Sulfa drugs o Broad spectrum antibiotics for tissue destruction and secondary bacterial infections o Supportive care: Fluids, nutrition, blood transfusion (calves) Prevention o Mature oocysts can survive for years and are resistant to most disinfectants o Proper and consistent sanitation of kennels and pens o Strong ammonia-containing compounds or steam cleaning required to destroy oocysts o Do not feed raw meat to prevent spread of Sarcocystis spp. (and T. gondii) 4 10/20/2024  CLINICAL PATHOLOGY I – Lecture Toxoplasma gondii Another coccidian organism Toxoplasma gondii Enteric coccidian parasite causes toxoplasmosis Zoonotic Cats are only DH Domestic cats (Felis catus) Wild cats Cats are only species that shed T. gondii oocysts Most any warm-blooded animal and many reptiles can serve as IH Life Cycle Obligatory heteroxenous: Requires IH Life cycle categorized as indirect T. gondii can be transmitted from cat to cat via infective oocysts, but this is very rare Cats (and IHs, including people) can be infected Transplacentally: Via tachyzoites Orally: By ingestion of any of three life stages Sporozoites (in oocysts): PP = ~ 21 days** Tachyzoites (in blood / lymph of IH): PP = ~ 21 days (usually only cats infected this way)** Bradyzoites (in tissues of IH): Prepatent period = 3 to 10 days** ** Prepatent periods for DH (cats) only Sporozoites = Develop in oocysts 1 to 5 days after passing in feces of cat Sporulated oocysts resistant and live for years Oocysts are important source of human exposure Tachyzoites = Rapidly dividing stage Found in blood and lymph of affected DH and IH; may pass in milk of lactating females Responsible for clinical signs Bradyzoites = Slowly dividing stage Found in tissues (inside zoitocysts) of affected DH and IH Most common source of infection for cat Two distinct cycles Enteroepithelial Occurs only in cat (DH) Occurs in intestinal epithelium Results in sexual reproduction and formation of oocysts Extraintestinal Occurs in DH and IH Occurs throughout body Results in asexual reproduction and formation of encysted bradyzoites in tissues 1 10/27/2024 Enteroepithelial Cycle Occurs only in cats Cat ingests Sporulated oocysts containing sporozoites (rarely) OR IH containing tachyzoites (uncommonly) and/or bradyzoites (commonly) Typically a mouse.... But, most warm-blooded animals can serve as IH Some sporozoites or bradyzoites enter intestinal cells, and.... Reproduce asexually for 5 cycles Then sexual reproduction commences Results in formation of unsporulated oocysts Oocysts are passed in feces Oocysts sporulate in 1 to 5 days and are infective Extraintestinal Cycle Occurs in both cats and IH In cats, cycle occurs simultaneously with enteroepithelial cycle Cat or IH ingests Sporulated oocysts containing sporozoites (most common route for IH) OR IH containing bradyzoites or tachyzoites (most common route for DH) Sporozoites or bradyzoites penetrate intestinal wall and begin asexual reproduction, producing tachyzoites Tachyzoites spread throughout body via bloodstream and lymphatic system Tachyzoites cause clinical signs Invade tissues throughout body Replicate asexually, causing cell death Animal’s immune response cause tachyzoites to encyst in host’s tissues and develop to bradyzoites Bradyzoites persist for life of host (DH and IH) Clinical Signs in Cats Often subclinical Does NOT typically cause GI signs. Clinical signs d/t extraintestinal cycle; i.e., asexual reproduction If clinical signs occur, most healthy cats develop an effective immune response and recover uneventfully FAD Uveitis Weight loss Lungs (“pulmonic toxo”) Liver (“hepatic toxo”) CNS (brain and spinal cord) 2 10/27/2024 Diagnosis by Fecal Flotation Very difficult because... Oocysts are tiny -- 1/8 size of Toxocara spp. ova Focusing “through” sample will assist in visualization Oocysts only shed by DH for 10 to 14 days following exposure to T. gondii T. gondii oocyst cannot be distinguished from many other coccidia Diagnosis Via Serology Blood test to determine if antibodies (Ab) to T. gondii are present Serum titers are most reliable indicator of previous exposure Post-exposure, takes at least one week for Ab levels to become detectable IgM titer: Indicates recent exposure IgG titer: Indicates exposure at some time in past ELISA: Enzyme-Linked Immunosorbant Assay Titers only mean patient has been exposed to T. gondii, NOT that patient is actively infected Cats with positive titer Most likely immune to reinfection with T. gondii Much less likely to pass T. gondii to humans Once acquired, titers generally last lifetime Of course, in cats.... nothing is that simple Positive Titer in Cat Recent exposure (i.e., detectable IgM level) Cat was, but is not now, threat to humans (unless re-exposed to T. gondii) Oocysts previously shed by cat can persist in environment for years If re-exposed, cat will shed fewer oocysts than never-exposed cat OR Past exposure (i.e., detectable IgG level): Presently immune Oocysts previously shed by this cat can persist in environment for years If immunosuppressed or starved, cat could shed oocysts again (whether reinfected or not) Negative Titer in Cat No exposure Cat has no immunity Will shed large numbers of oocysts if exposed OR Recent exposure (i.e., tested prior to production of detectable levels of IgM) Cat is direct threat to humans Will begin to shed oocysts in near future or may already be shedding them OR Past exposure: But Ab did not develop Treatment of Clinically Ill Cats Clinical signs rarely observed and typically only appear during extraintestinal cycle 3 10/27/2024 Medications include Clindamycin Trimethoprim-sulfa +/- corticosteroids? Prevention in Cats Do not feed raw meat to cats: Feed commercial diets only Confine cats indoors to prevent hunting and eating prey 4 10/27/2024