CLS08 - Parasites

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  1. Two general stages of many Protozoa?  Usually acquired how?  Exceptions?
    • Trophozoite (moving) and cyst/spore (encapsulated stages
    • Usually fecal/oral except Apicomplexa (AKA Sporozoa) eg Plasmodium and Babesia
  2. What are the common/acceptable intestinal tract specimens (5)?  What are common procedures?
    • Stool in a sterile container
    • Duodenal drainage
    • Duodenal capsule technique
    • Sigmoidoscopy
    • Recovery of tapeworm scolex/proglottids
    • common procedures: O&P exam
    • Direct wet mount, concentration, permanent stain
  3. What is the common fecal collection system (w/ alternatives)? Common timeline/method?
    • Two-vial system
    • 5-10% formalin for immunoassays and concentration
    • *or MIF for direct wet mount/conc
    • *or SAF for permanent smears/conc
    • 3:1 PVA:stool for perminant stained smears
    • *or Modified PVA
    • *or SAF
    • Traditional method: collect 3 stools ONE PER DAY over 3-9 day period
    • Increases chance of detecting intermittent shedding
    • Alternative method: combine all three samples and examine at once
    • faster, but dilution may cause a missed positive
  4. When do non-intestinal samples require prevervation? Why?
    • If there is a delay (almost always is) then samples must be PREVENTED FROM DESSICATING.
    • Sputum, urine, and aspirates are all fine because they provide a moist environment, but tissue, Urogen spec, blood, etc should be preserved
  5. What are 2 stat parasite situations (re transport/evaluation)
    • CNS specimens for amoeba (Naegleria)
    • Blood films for malaria
  6. What are 5 common parasite sample processing procedures used to aid in microscopic detection of organisms / what do they do?
    • Formalin/Ethyl acetate sedimentation: concentrates eggs, larvae, oocysts, and spores
    • Trichome stain: distinguishes cysts and trophozoites
    • Modified iron hematoxylin stain: demonstrates cysts and trophozoites
    • Modified acid-fast stain: highlights Coccidia
    • Modified trichome stains: differentiate microsporidia
    • weber-green, ryan-blue, kokoskin (hot method)
  7. Parasitic life stage and correlation to stool consistency
    • More formed = more cysts
    • More watery = more trophozoites
  8. Why must parasitic stool samples be free of water or urine?
    • Water can contain free-living organisms
    • Urine and kill motile organisms
  9. Formalin vs MIF pros/cons
    • Formalin Pro: overall storage, easy to prepare, long shelf life, use in immunoassays
    • Formalin con: doesn't preserve trophozoites, not good for perm stains
    • MIF pro: preserves trophs, cysts, larvae, helminth eggs
    • MIF con: not usually used for perm stains or immunoassays, not as convenient
  10. PVA vs modified PVA vs SAF pros/cons
    • PVA pro: best method for preserving morphology, can ship
    • PVA con: MERCURY (health hazard) - no longer recommended
    • modPVA pro: no mercury, can use conc and stain
    • modPVA con: inconsistent staining (misID)
    • SAF pro: conc and stain, easy to prepare, long shelflife, immunoassays
    • SAF con: poorly adhesive (albumin coated slides recommended)
  11. What are the 3 O&P examination protocols w/ details?
    • Direct wet mount: fresh stool stained w/ iodine and sealed edges
    • viewed at 100x and 400x (high dry)
    • allows motility visualization, UV exam (cyclospora fluoresces blue)
    • Concentration wet mount: facilitates recovery of cysts, oocysts, spores, eggs, larvae using flotation or sedimentation protocols
    • stain with iodine, view at 100x and 400x
    • Permanent stained smear: MOST IMPORTANT for intestinal protozoa
    • trichrome and iron-hematoxylin - most protozoa
    • modified acid-fast - coccidia
    • trichrome - microsporidia spores
    • view >300 fields on oil immersion
    • *definitive ID reqs conc and perm stain
  12. Describe the flotation method of concentration
    • Requires a flotation fluid (Zinc sulfate or Shether's sugar)
    • mix spec w/ fluid, filter out solids, organisms rise and debris sink
    • *NOTE - walls of eggs/cysts can collapse, and some eggs DO NOT FLOAT
  13. Describe the sedimentation method of concentration
    • requires formalin and ethyl acetate
    • preserved stool passed through filter into tube
    • 10% formalin added until 13mL total
    • centrifuged then decanted
    • sediment used for slides
    • if too much debris on slides add formalin to 10mL and 3mL ethyl acetate and respin/decant
  14. Malaria PCR Lanes
    • S: 50bp ladder
    • 1: P. vivax (120bp)
    • 2: P. malariae (144bp)
    • 3: P. falciparum (205bp)
    • 4: P. ovale (800bp)
  15. define karysome, peripheral chromatin, chromatoidal bars
    • karyosome: spherical chromatin mass in the nucleus (center of bullseye)
    • peripheral chromatin: DNA on nuclear membrane (outer ring of target)
    • chromatoidal bars: refractile chromatin structure (rod)
  16. Flow chart of protozoans (both)
    • Image Upload
    • Image Upload
  17. Entamoeba histolytica - type, infection route, infective/repr stages, disease, treatment, prevention, morphology (troph and cyst)
    • type: amoeba (only true pathogen)
    • route: fecal/oral
    • stages: trophozoite (reproductive) and cysts (infective)
    • disease: abemic dysentary (bloody diarrhea), hepatic abcesses, invasive infections
    • can be fatal if organisms leave intestines and reach lungs, liver, bladder
    • *note - asymptomatic carriers exist
    • treat: metronidazole
    • prevent: heating or chlorination of water
    • troph: oblong, bulls-eye nucleus, ingested RBCs (diff from E. dispar)
    • cyst: 1-4 nuclei (bulls-eyes), chromatoidal bars w/ round ends
  18. How to distinguish E. hartmanni, Endolimax nana, E. coli, and Iodamoeba butschlii from Entamoeba histolytica
    • E. hartmanni: smaller in size
    • Endolimax nana: smaller in size, nuclei have larger karyosome
    • E. coli: larger in size, "dirty" cytoplasm
    • Iodamoeba butschlii: no peripheral chromatin
    • Image Upload
  19. Blastocystis hominis - controversy and morph
    • Role as pathogen is controversial, many labs report and let physician decide
    • Have uniquely stained vacuole that is bright red
  20. Giardia lamblia - type, infection route, infective/repr stages, disease, treatment, prevention, morphology
    • type: flagellate
    • infection route: fecal/oral
    • stages: cyst (infective) and trophozoite (repr)
    • disease: acute infection (self-limiting EXPLOSIVE, foul-smelling diarrhea, nausea)
    • chronic infection (weight loss, fatigue)
    • asymptomatic infection
    • *most common intestinal protozoa in US
    • treatment: metronidazole
    • prevention: cysts resistant to chlorination
    • trophozoite: chill-ass ghost (central binucleated, karyosome, midline axonems)
    • cysts: up to 4 nuclei, flagella is retracted within
  21. Dientamoeba fragilis - type, infective/repr stages, diagnosis, morphology
    • type: flagellates
    • stages: no cyst stage
    • diagnosis: stained smears only
    • morphology: binucleated, disintigrates RAPIDLY after excretion
  22. Trichomonas vaginalis - type, infection route, infective/repr stages, disease, treatment, prevention, morphology
    • type: flagellate
    • infection route: STD
    • stages: no cyst stage, human only host
    • disease: trichomoniasis (many asyptomatic carriers)
    • major complications during preggo, increased HIV risk
    • reinfection common (no long-term immunity)
    • treatment: metronidazole
    • morph: Image Upload
  23. Microsporidia - type, infection route, infective/repr stages, disease, treatment
    • type: obligate, intracellular, spore-forming parasite
    • infect: inhalation or ingestion of spores, AIDS patients greater risk
    • stages: spore (infective stage)
    • disease: diarrhea, fever, malaise
    • treatment: albendazole
  24. Leishmania - stage found in humans, spectrum of disease, treatment
    • stage: amastigote
    • disease: cutaneous lesions, mucocutaneous infections, viscerotropic dissemination, fatal visceral infection
    • treatment: pentavalent antimoney compounds
  25. Trypanosomes - African vector + spp, American vector + spp, stage in humans, diseases
    • African: Tsetse fly (T. brucei gambiense, T. brucei rhodesiense)
    • American: Reduviid (kissing) bugs (T. cruzi) chagas disease
    • stage: trypomastigote
    • disease: african sleeping sickness (brain, CSF), loss of coordination, somnolence
    • cardiac involvement in chagas
  26. Plasmodium general - type, infection route, disease, spp, lab ID
    • type: apicomplexa
    • infection route: mosquitos
    • disease: malaria (2.7m deaths/yr - mostly children)
    • spp: falciparum, ovale, malariae, vivax
    • lab: travel history, fever/chills cycle, cytoplasmic structures help to determine spp
  27. Plasmodium falciparum - RBCs infected, incubation, disease manifestations/complications, treatment, lab diagnosis
    • *most deadly
    • RBC: all ages of RBCs
    • incubation: short, 8-12 days 
    • disease: severe complications related to blood vessel obstruction, CNS complications
    • blackwater fever (usually repeated infection and therapy)
    • treatment: chloroquine (beginning resistance)
    • lab: characteristic ring form in smear
  28. Plasmodium vivax- RBCs infected, incubation, disease manifestations/complications, lab diagnosis
    • RBC: only reticulocytes
    • incubation: medium, 10-17 days
    • disease: splenomegaly
    • severe complications are rare
    • lab diagnosis: schuffner's dots
  29. Plasmodium ovale- RBCs infected, incubation, disease manifestations/complications, lab diagnosis
    • RBC: only reticulocytes
    • incubation: medium, 10-17 days
    • disease: lower frequency and severity
    • spontaneous recovery
    • lab: Schuffners dots
  30. Plasmodium malariae- RBCs infected, incubation, disease manifestations/complications
    • RBC: mostly older
    • incubation: long, 27-40 days
    • disease: regular 72hr cycles 
    • nephrotic syndrome (caused by Ab/Ag complex)
    • spontaneous recovery
    • recrudescence >50 years
  31. Babesia: type, transmission, repr, disease, US spp, morph
    • type: apicomplexa
    • transmission: tick
    • repr: no exoerythrocytic stage (must be inside RBC)
    • disease: similar to malaria
    • spp: B. microti
    • morph: maltese cross in RBC
  32. Cryptosporidium parvum - transmission, disease, outbreaks at...?, lab diagnosis, similar organisms
    • transmission: fecal-oral via oocyst
    • disease: self-limiting GI infection in Immunocomp
    • outbreaks: day care
    • Lab ID: immunofluorescen antibody stain (DFA), or modified acid-fast
    • *note - big outbreak, resistant to chlorine, tiny
    • similar: other apicomplexa coccidia include Cyclospora (infective stage sporozoites) and Isospora (infective stage oocyst)
  33. acanthamoeba - type, disease, lab ID
    • type: free-living amoeba
    • disease: amebic karatitis (contact lenses)
    • likely to lose vision in infected eye
    • can travel to brain, but trophozoites rarely seen in CSF
    • lab ID: corneal scraping wet-preps may show cyst
    • tapwater agar + E. coli lawn = does acanth appear?
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CLS08 - Parasites
2016-04-25 05:29:04
CLS08 Parasites
CLS08 - Parasites
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