Para Lab Quiz 1

• Stool specimen parasites and parasite stages:

  • Ova of intestinal helminths like Ascaris lumbricoides, Trichuris trichiura, Capillaria philippinensis, Necator americanus, Ancylostoma duodenale, Schistosoma mansoni, Schistosoma japonicum, Diphyllobothrium latum, Taenia solium, Taenia saginata, Rhabditiform larva of Strongyloides stercoralis
  • Cyst & Trophozoite stages of Entamoeba histolytica, Giardia lamblia, Balantidium coli
  • Oocyst of Cryptosporidium parvum
• Sputum specimen parasites:

  • Ova of Paragonimus westermani
  • Trophozoite of Entamoeba histolytica
  • Filariform larva of Necator americanus and Ancylostoma duodenale
  • Larva of Ascaris lumbricoides
• Urine specimen parasites:

  • Ova of Schistosoma haematobium
  • Trophozoite of Trichomonas vaginalis
• Blood specimen parasites:

  • Ring form trophozoites & gametocytes of Plasmodium spp and Babesia microti
  • Microfilaria of Filarial worms like Wuchereria bancrofti, Brugia malayi, Loa loa
• Duodenal aspirate specimen parasites:

  • Trophozoite of Giardia lamblia
• CSF specimen parasites:

  • Naegleria fowleri and Acanthamoeba spp
• Collection, transport & specimen processing:

  • Parasites are often shed intermittently in stool specimens, so multiple specimens are recommended for adequate detection
  • Stool collection protocol typically consists of three specimens collected every other day or a total of three collected in 10 days
  • Medications and substances like barium, bismuth, or mineral oil may interfere with parasite detection
  • Specimens from patients on antibiotics or antimalarial medications should be delayed for 2 weeks following therapy
• Stool specimen preservation:

  • Stool specimens should be collected in a clean, watertight container with a tight-fitting lid
  • Urine should not contaminate stool specimens
  • Stool should not be retrieved from toilet bowl water
  • Specimen container should be labeled with patient's information and collection details
  • Trophozoites are sensitive to environmental changes and should be examined within 30 minutes of passage
• Preservation methods:

  • Formalin: used for recovery of protozoa and helminths, with 5% concentration for protozoan cysts and 10% for helminth eggs and larvae
  • Polyvinyl alcohol (PVA): combined with Schaudinn solution for detecting trophozoites, cysts, and helminth eggs
  • Sodium acetate formalin (SAF): used for concentration techniques and permanent stained smears
• Macroscopic and microscopic examination:

  • Stool specimens should be examined macroscopically for consistency, color, and gross abnormalities
  • Smears can be prepared using NSS or iodine
  • Helminth eggs & larvae are reported under LPO, protozoan cysts and trophozoites under HPO
  • Normal microscopic structures are not reported, only abnormal findings
• Permanent stains:

  • Iron hematoxylin: reveals excellent morphology of the intestinal parasites
  • Wheatly trichrome: used for staining to detect coccidian oocysts
• A shorter technique using a stain is now available for revealing excellent morphology of intestinal protozoa
• In some cases, the nuclear detail of these organisms is stained clearer and sharper than when stained with trichrome
• Color variations among specific parasitic structures and background material are not as distinct as with trichrome
• The most widely used permanent staining method uses reagents with a relatively long shelf life and the procedure is easy to perform
• Kato thick smear:

  • Different from the standard direct smear procedure
  • Uses a larger amount of fecal sample
  • Cellophane strips are used as cover slips instead of glass
  • Glycerine acts as a clearing agent while malachite green provides a green background and reduces brightness of the microscopic field
  • Advantages: economical, applicable for thick-shelled eggs like Ascaris & Trichuris, simple procedure
  • Disadvantages: not suited for watery/liquid stools, not for cyst & trophozoites, not for thin-shelled eggs like those of hookworms
• • A quantitative procedure carried out to determine the degree of infection and to assess the effectiveness of anti-helminthics
• Stoll dilution uses 0.1 N NaOH
• Culture methods are not a common means of detecting parasites, but parasites that can be isolated with culture include E. histolytica, T. vaginalis, Leishmania spp., T. cruzi, and T. gondii
• Harada Mori/Test Tube Culture Method and Copro Culture are culture methods for hookworms & Strongyloides stercoralis
• Concentration techniques aim to increase the possibility of finding protozoan cysts, helminth eggs, and larvae by decreasing the amount of background material and by actual concentration of organisms
• Principle behind all concentration techniques:

  • Sedimentation: parasites settle at the bottom of the tube due to higher specific gravity
  • Flotation: parasites float to the top of the tube as they are lighter
  • Acid ether concentration technique:
  • Method of choice for specimens from animal sources
  • Recommended for the recovery of Trichuris, Capillaria & Schistosoma eggs
  • HCl is used as a clearing agent while ether removes fats
• Formalin ethyl acetate:

  • Most widely used sedimentation technique
  • Provides good recovery of most parasites and is easy to perform
  • Preparation contains more fecal debris than a flotation technique
  • Useful in the recovery of Giardia lamblia cyst & cestode eggs
• Zinc sulfate centrifugal flotation:

  • Uses zinc sulfate with a specific gravity of 1.18 to 1.20
  • Yields a cleaner preparation for microscopic examination
  • Some helminth eggs are very dense and will not float, leading to some parasites being missed
  • Recommended for G. lamblia cyst & H. nana ova
• Sheather’s flotation:

  • Uses sugar solution preserved in phenol
  • Recommended for concentrating oocysts of Cryptosporidium & Isospora
• Brine’s flotation:

  • Uses salt solution, stool is directly mixed with brine without centrifugation
  • Simple, economical, and suited for mass stool examination
  • Not suited for Trematode eggs as they do not float in Brines
  • Hookworm eggs & Schistosome eggs become badly shrunken
• Faust Maloney Egg Hatching:

  • Quantitative test, a miracidial hatching test for Schistosoma
• Thick & Thin Blood Smear:

  • Regarded as the gold standard for Malaria detection
  • Malaria is caused by Plasmodium species (P. falciparum, P. malariae, P. vivax & P. ovale)
  • Best time to collect blood for malaria detection is at night or early in the morning
  • Plasmodium stages seen microscopically in the blood
• Knott’s Concentration Technique:

  • Concentration technique for the diagnosis of Filariasis
  • Consider the periodicity of microfilaria in the collection of blood
  • Diagnostic stage, reagent used
• Several parasites may be detected through examination of blood, including the Plasmodium species, Babesia microti, and the filarial worms i.e. W. bancrofti
• Perianal Swab/Scotch Tape Swab