Intestinal protozoa

Microbiology

• Protozoans are eukaryotic unicellular organisms that have organelles
• Many have a trophozoite form (active, motile, feeding form), as well as a cyst form (vegetative, difficult-to-kill)
• Stains:
  • Iron hematoxilin, which stains nuclei on a black background
  • Acid-fast stain, which highlights red Cryptosporidium/Cyclospora/Cystoisospora
  • Can combine both of the above

Classification

From largest to smallest family

• Amoebae
  • Entamoeba histolytica/dispar (microscopically indistinguishable, but E. dispar is non-pathogenic)
  • Entamoeba harmanni and Entamoeba coli (non-pathogenic)
  • Endolimax nana
  • Iodamoeba butchlii
• Flagellates
  • Giardia lamblia
  • Dientamoeba fragilis (no cystic form)
  • Chilomastix mesnili
• Ciliates
  • Balantidium coli
• Coccidia: small, may be seen as acid-fast
  • Cryptosporidium species (~5 μm)
  • Cyclospora cayetanensis (~10 μm)
  • Cystoisospora belli (previously called Isospora) (~30 μm)
  • Sarcocystis hominis
• Microsporidia

Identification

• Some have classic appearance:
  • Giardia trophs look like an owl
  • Balantidium coli is huge
  • Trichomonas and Chilomastix trophs have flagellae and are tear-drop shaped
  • Cystoisospora belli looks like an eye or halved egg
  • Cyclospora is thick-walled and looks like a golf ball
  • Cryptosporidium is the smallest (4-6 µm) and may have crescent-shaped sporozoites inside
  • Sarcocystis is quite large (15 µm), non-acid-fast, and looks like 2 joined ovals

Amoebae

• In order to differentiate the larger Amoebae, need to consider the size, nucleus characteristics, and other features
  • Does is stain acid-fast? If yes, it is one of the "sporas"
  • What kind of nucleus is it?
    • Entamoeba: dark ring of peripheral chromatin, with a defined central nucleus and relatively clear perikarysomal space
    • Non-Entamoeba: more blob-like, less well defined
    • Granulated: big dots, suggests Dientamoeba fragilis trophozoite
  • How many nuclei?
    • Trophozoites are mostly mono-nucleated, cysts may be polynucleated
    • Dientamoeba has 2 nuclei in trophozoite form
  • What is the average size?
    • Need a measurement microscope
• Other stuff
  • Large vacuole suggests Iodamoeba butschlii
  • Uneven chromatin suggests Entamoeba coli
  • Chromatoidal bars suggests Entamoeba polecki, Entamoeba histolytica, or Entamoeba hartmanni

Organism	Nucleus type	No. nuclei	Size	Other features
E. histolytica	Entamoeba type	2-4	> 10 µm	Chromatoidal bars
E. coli	Entamoeba type	5-8+	> 10 µm
E. hartmanni	Entamoeba type		< 10 µm	Chromatoidal bars
E. polecki	Entamoeba type but 1/3 cell width		> 10 µm	Chromatoidal bars, but may have glycogen mass
Iodamoeba	Blob-like	1		Large vacuole
Endolimax	Blob-like	2-4

Coccidia

• No serology available
• Culture difficult or impossible
• May be seen on histopathology of biopsy specimens

Organism	Size of oocyst	Stool O&P	Stool antigen	Acid-fast	Other stain
Cryptosporidium species	4-6 μm	+	+	+	immunofluorescence
Cyclospora cayetanensis	8-10 μm	+	–	+
Cystoisospora belli	23-33 x 10-19 μm	+	–	+
Microsporidia species	1-3.5 μm	±	–	–	chromotrope, calcofluor