Amoeba Study Notes PDF

Revision 1. Parasites that requires host(s) to live are known as: 2. The structure of kinetoplast is seen in a parasite under the phylum: 3. Circulatory system is absent in the: 4. Genus of Plasmodium, Toxoplasma and Cyclospora are classified under the phylum: 5. The only ciliates of human tha...

Revision 1. Parasites that requires host(s) to live are known as: 2. The structure of kinetoplast is seen in a parasite under the phylum: 3. Circulatory system is absent in the: 4. Genus of Plasmodium, Toxoplasma and Cyclospora are classified under the phylum: 5. The only ciliates of human that found in the intestinal tract is: Revision Statetement True OR False 1. Copulatory organs of nematodes parasites include spicule and gubernaculum. 2. Apical complex is necessary for attachment to the host. 3. Cyclophyllidean tapeworm has open-ended uterus. 4.Aberrant parasite is parasite that migrate and reach the site of the host but it cannot live or develop further. 5. Starch granules are artifact that appears as irregular shaped.  CSI153 Clinical Parasitology List and Description of Life cycle, Mode of infection, Disease, Infective Stage and Important Identifying features of Diagnostic Stage. Amoeba Nurhidayah binti Ab. Rahim Mac 2019 Diploma Teknologi Makmal Perubatan, UiTM (Pulau Pinang), Kampus Bertam  Protozoa Intestinal Amoeba Intestinal and atrial Free living Amoeba Entamoeba histolytica flagellates and ciliates Entamoeba coli Naegleria fowleri Endolimax nana Giardia lamblia Acanthamoeba spp. Iodamoeba butschlii Dientamoeba fragilis Blastocystis hominis Trichomonas vaginalis Balantidium coli Blood and tissue sporozoa Blood and tissue flagellates Plasmodium vivax Plasmodium ovale Leishmania spp. Plasmodium malariae Trypanosoma spp. Plasmodium falciparum Plasmodium knowlesi Babesiosis Toxoplasma gondii Pneumocystis carinii  Learning outcomes Students should be able to: Features/ Life cycle characteristic infection and Mode of infection diagnostic stage Diagnosis and treatment Introduction  What phylum for Amoeba?  What the locomotory organs for amoeba?  How many forms of amoeba? What it is? Introduction  Differences between trophozoite and cyst Trophozoite Cyst Amoeba  Is classified under protozoa  Do not have fixed shape - Due to extension of pseudopodia for motility (locomotion) and engulfment food by phagocytosis The cytoplasm is bounded by a membrane - Outer ectoplasm - Inner endoplasm  There are free living and parasitic amoeba  Parasitic amoeba causes amoebiasis or infection due to amoeba - Usually inhabit digestive system  Commonly found in human is Entamoeba, Endolimax and Iodamoeba  Motility: crawling or gliding Amoeba Reproduction - Fusion and budding Budding Cysts - Infective form for vertebrate host - Eg: E. histolytica Multiple fission Binary fission Medically important amoeba Intestinal Free living amoeba amoeba Entamoeba histolytica *Naegleria (pathogenic) fawleri Entamoeba dispar *Acanthamoeba spp. Entamoeba coli Entamoeba polecki Entamoeba hartmanni *Balamuthia mandrillaris Entamoeba gingivalis Endolimax nana Iodamoeba butschii *opportunistic pathogens  Entamoeba histolytica A. Entamoeba histolytica  Distribution - Worldwide - Common in tropical area of Asia, Africa and latin America  Mode of infection - ingestion of water and food containing cysts. - Oral-fecal route  Entamoeba histolytica Morphology - trophozoite Size range 8-65µm (irregular shape) Motility Actively motile/ fingerlike pseudopodia Number of nuclei 1 Karyosome Small and central Peripheral chromatin Fine and evenly distributed Cytoplasm Finely granular Cytoplasmic inclusions Ingested RBC Site Lumen  Entamoeba histolytica Pseudopodia - Finger-like projections - Sudden jerky movement of ectoplasm in one direction, then streaming in of the whole endoplasm Ectoplasm – clear, transparent, refractile Endoplasm – finely granular, having ground glass appearance, contains nucleus, food vacuoles, erythrocytes, leucocytes, tissue debris - Sudden jerky movement of ectoplasm in one direction, then streaming in of the whole endoplasm  Entamoeba histolytica Trophozoites life span - Up to 5 hours at 37°C - Killed by drying, heat, chemical sterilization - Ingestion – destroyed in stomach, cannot cause infection **infection is not Nucleus transmitted by trophozoites - Spherical - 4 – 6 µm - Contains central karyosome, chromatin granules (evenly distributed) - Not clearly seen in the living trophozoites - Stained with iron-hematoxylin  Entamoeba histolytica Entamoeba histolytica-trophozoite Karyosome nucleus endoplasm ectoplasm Iron-hematoxylin staining  Entamoeba histolytica Trichrome staining  Entamoeba histolytica In iodine - direct wet mount Entamoeba histolytica Morphology - cyst Size range 8-22µm Shape Spherical to round Number of nuclei 1 to 4 (immature: 1; mature: 4) Karyosome Small and central Peripheral chromatin Fine and evenly distributed Cytoplasm Finely granular Cytoplasmic inclusions Chromatid bars/rounded ends, Diffuse glycogen mass (seen in immature cyst, dissappear in mature cyst)  Entamoeba histolytica Morphology - cyst  A cyst starts as a uninucleate body.Then the nuclei divides by binary fission to form 2 and then 4 nuclei. Chromatoid body will be used during cell division  Entamoeba histolytica Immature cyst Cysts of E. histolytica in an unstained wet mount of stool. Notice the chromatoid bodies with blunt, rounded ends (arrows), nucleus is not clearly seen.  Entamoeba histolytica Morphology - cyst Cysts of E. histolytica in an iron hematoxylin stain of stool. Notice the chromatoid bodies appear deep blue or black (arrow), glycogen mass appears unstained.  Entamoeba histolytica Cyst Cysts of E. histolytica in an iodine. Glycogen mass appears golden brown.  Entamoeba histolytica Immature cyst Stained with trichrome. Two to three nuclei are visible in the focal plane (black arrows), and the cysts contain chromatoid bar with typically blunted ends (red arrows).  Entamoeba histolytica Life cycle of E. histolytica  Entamoeba histolytica Life cycle  Whole life cycle in only one host  Mature cyst (infective form) ingested through water or food.  When cyst reach lower part of small intestine, excystation occur Multiply to become trophozoites  Trophozoite inhabit in submucous tissue of large intestine and multiple by binary fission  Entamoeba histolytica Encystation of trophozoite occur in lower part of large intestine to become cysts Trophozoite and cyst are passed out in feces Trophozoite can spread from intestine to liver and from liver to lung and brain through the bloodstream by invading the intestinal mucosa  Entamoeba histolytica Mucosa Pathogenesis i) Intestinal Primary amoebiasis (amoebic ulcer dysentery) - incubation period is 1-4 weeks. - it produces ulcerative lesions and bloody diarrhoea called as amoebic dysentery.  ii) Extraintestinal amoebiasis happens when trophozoites enter into deeper layers and carried away to the liver thru the vein. causes hepatic amoebiasis where amoebic liver abscess is formed. liver abscess contains pus that yield trophozoites. from the liver, the trophozoites might enter other organs such as lungs, brain and skin. Liver abscess abscess-fluid composed of leukocytes, bacteria, and cellular debris.  Entamoeba histolytica Liver abscess  Entamoeba histolytica Lab diagnosis i) Intestinal amoebiasis a) Microscopic examination of stool  saline preparation to demonstrate motile trophozoites, and iodine preparation for the study of cysts or dead trophozoites. b) Blood examination  shows leukocystosis leukocytosis: abnormal increase of wbc due to infection  Entamoeba histolytica ii) Hepatic amoebiasis a) Diagnostic aspiration -trophozoites may be demonstrated by microscopy of pus aspirated from hepatic abscess b) Liver biopsy -trophozoites demonstrated in the liver tissue iii) Serological test -antibody detection using ELISA, Collecting pus which is a biochemical technique used to detect antibody or antigen when an infection happens. Aspirate: withdrawal of fluid  Entamoeba histolytica Microscopic identification  DO NOT wrongly identify other intestinal amoeba as Entamoeba histolytica  Other commonly found intestinal amoeba  Entamoeba coli  Entamoeba hartmanni  Endolimax nana  Iodamoeba bütschlii 02/06/2024; Slide 34  Entamoeba histolytica Treatment Metronidazole, nitroimidazole are some drugs used for treatment of amoebiasis.  Entamoeba histolytica Prevention  Avoid faecal contamination of food and water.  Proper disposal of human faeces.  Asymptomatic carriers should be treated properly. 02/06/2024; Slide 36 Exercise A previously healthy 28 years old man who had recently returned from a trip to Mexico, was seen by his family physician for crampy abdominal pain, malaise, slight fever and bloody, muccoid diarrhea. Liquid stool specimens were collected and submitted for culture for enteric bacterial pathogen, as well as parasites. Stool cultures were negative for bacterial pathogens, examination for ova and parasites were positive for motile trophozoites in the saline wet mount, and ameboid trophozoites with finely granular cytoplasm and ingested RBC in the permanent trichrome stain. 1. What intestinal parasite would you consider in making a diagnosis? 2. Is it parasite capable of causing extra-intestinal infection? What organs is most commonly involved? 3. How is this parasite transmitted? 4. Should this patient be treated? How? 5. What the characteristic of ameboid trophozoites?  Entamoeba coli Entamoeba coli  Distribution: Worldwide  Nonpathogenic commencal  Larger that E. histolytica (20-50 um)  Sluggish motility  Life cycle is the same as E. histolytica but not invade tissue (liver, lung, brain)  Inhabit in lumen of large intestine  Always confuse with E. histolytica  Entamoeba coli E. histolytica vs E. coli Entamoeba histolytica Entamoeba coli trophozoites cysts  Entamoeba coli E. histolytica vs E. coli Entamoeba histolytica Entamoeba coli Trophozoite 20-30 mm 20-40 mm Motility Unidirection Multiple direction Cytoplasm - Well separated clear -Not separated ectoplasm and granular - Contain no RBC endoplasm - Contains RBC Nucleus - Smaller central karyosome - Larger eccentric - Membrane: even thickness karyosome - Membrane: uneven thickness Cyst 6-15 mm 15-20 mm Nucleus -1 to 4 unit -1 to 8 unit - central karyosome - eccentric ( not central) 02/06/2024; Slide 40 Eccentric: not in the center  Entamoeba coli Entamoeba coli mature cyst Iodine staining NUCLEUS: Only 5 nuclei are visible, eccentric karyosome (arrow) Glycogen mass : prominent in early stage (immature) Cnromatid bodies : solinter-like and irregular  Entamoeba coli Entamoeba coli-Mature cyst: Trichrome staining. These two images represent the same cyst shown in two different focal planes.  Entamoeba coli Entamoeba coli trophozoite Unstained (nucleus is clearly visible) has large eccentric karyosome, thick nuclear membrane (uneven thickness), coarse granules of chromatin  Entamoeba coli Entamoeba coli trophozoite has large eccentric karyosome, thick nuclear membrane (uneven thickness), coarse granules of chromatin Trichrome staining Endolimax nana  Trophozoite  Small (6-15 mm); clear ectoplasm and granular endoplasm  Contains no erythrocyte  Nucleus: large central karyosome Trophozoite  Sluggish motility  Cyst  Small (8-10 mm); oval; 1-4 nuclei (quadrinucleate)  Nucleus: eccentric, large, irregular karyosome Cyst  Iodamoeba bütschlii Iodamoeba bütschlii Widely distributed, less common than E. coli and E. nana Trophozoite – Small (6-25 mm); ectoplasm not differentiated from endoplasm – Nucleus: large central karyosome Cyst – 8-15 mm; oval; 1 nucleus (uninucleated) – Nucleus: large central karyosome – Contains a large glycogen vacuole (seen under iodine staining)  Iodamoeba bütschlii Cyst Iodine staining: large glycogen vacuole (arrow) Trichrome staining: large central karyosome (black arrow), large glycogen vacuole (red arrow)  Iodamoeba bütschlii trophozoite ectoplasm not differentiated from endoplasm Central karyosome in nucleus Blastocystis hominis Worldwide distribution Size: 2 – 100 µm Yeast-like form Can accumulate starch and then filled in granules Cyst - Large central body - surrounded with thin inner membrane of cyst wall - Several dense inclusions Life Cycle (nonpathogenic intestinal amebae) Exercise - Draw the life cycle of E. histolytica - What the diagnostic stage and infective stage of intestinal amoeba? Exercises Tabulate the differences between E. histolytica, E. coli, Endolimax nana and Iodamoeba butschlii Trophozoites cyst Size size Motility nuclei Psedopodia glycogen mass Cytoplasm chromidial Inclusion Nucleus Karyosome  CSI153 Clinical Parasitology List and Description of Life cycle, Mode of infection, Disease, Infective Stage and Important Identifying features of Diagnostic Stage. Protozoa: Free living amoeba Nurhidayah binti Ab. Rahim April 2018 Diploma Teknologi Makmal Perubatan, UiTM (Pulau Pinang), Kampus Bertam  Learning outcomes Students should be able to: Features/ Life cycle characteristic infection and Mode of infection diagnostic stage Diagnosis and treatment  Protozoa Intestinal Amoeba Free living Amoeba Intestinal and atrial Entamoeba histolytica flagellates and ciliates Entamoeba coli Naegleria fowleri Endolimax nana Giardia lamblia Acanthamoeba spp. Iodamoeba butschlii Dientamoeba fragilis Blastocystis hominis Trichomonas vaginalis Balantidium coli Blood and tissue sporozoa Blood and tissue flagellates Plasmodium vivax Plasmodium ovale Leishmania spp. Plasmodium malariae Trypanosoma spp. Plasmodium falciparum Plasmodium knowlesi Babesiosis Toxoplasma gondii Pneumocystis carinii Free living protozoa Naegleria fowleri  Distribution: World-wide (live  Only species of genus in soil and warm freshwater, Naegleria that infects eg: lakes, rivers and spring) man Naegleria fowleri  Cause primary amoebic meningi encephalitis (infection in a brain and lead to destruction of brain tissue) Free living protozoa Naegleria fowleri Morphology -stage Trophozoite Cyst Amoeboid form Flagellate form 10 – 20 µm 7 - 10 µm Rounded pseudopodia 2 flagella Smooth double wall Spherical nucleus, big Spherical nucleus Spherical nucleus endosome Vacuoles (densely granular) vacuoles vacuoles amoebostomes = engulf RBCs, WBCs Feeding, growing and Can be revert to Dormant, resist replicating form of parasite amoeboid form; unfavorable conditions amoeboflagellate (drying, high concentration of chlorine) Invasive stage, infective form to Shape: pear Shape: spherical; Can human be found in CSF Free living protozoa Naegleria fowleri Cyst Amoeboid Flagellate trophozoite trophozoite *under electron microscope Free living protozoa Naegleria fowleri Life cycle Free living protozoa Naegleria fowleri Pathogenesis Nasal mucosa Olfactory nerves Cribriform plate Meninges, brain Extensive hemorrhage and necrosis is Primary amoebic meningo present in the brain, mainly in the frontal cortex (white arrow) encephalitis (PAM) Incubation period : 2 days to 2 weeks Fever, headache, vomitting, stiff neck, seizure, coma Free living protozoa Naegleria fowleri Laboratory diagnosis a) CSF examination - Gross: cloudy to purulent - Microscopic: prominent neutrophilic leucocytosis, elevated protein, low glucose - Wet film: trophozoites - Immunoflurecent staining: trophozoites b) Culture - Liquid axenic media or non nutrient agar plates coated with Escherichia coli c) Molecular diagnosis - Polymerase chain reaction treatment Amphotericin-B intravenously  Free living protozoa Acanthomoeba spp. Acanthomoeba spp.  A. culbertsoni, A. polyphagia, A. castalleni, A. astromyx  Distribution: world wide (soil and water) stage Trophozoite cyst Active, large, 20 – 50 µm Forming in tissues Spine-like pseudopodia Polygonal double-walled No flagellate stage Highly resistant  Free living protozoa Acanthomoeba spp. *under electron microscope (Left) Acanthamoeba remains in the trophozoite form and divides mitotically under favorable conditions (Right) and while under harsh conditions, the amoeba transforms into a dormant cyst, highly resistant to harsh conditions. This photo was published in 2012. By Ruqaiyyah Siddiqui and Naveed Ahmed Khan  Free living protozoa Acanthomoeba spp. Life cycle  Free living protozoa Acanthomoeba spp. Pathogenesis Immunodeficiency, diabetes, malignancies, malnutrition, alcoholism, SLE Keratitis Hematogenously into CNS, connective tissue Keratitis, Granulomatous amoebic encephalitis (GAE) Granulomatous amoebic encephalitis (GAE)  Free living protozoa Acanthomoeba spp. Laboratory diagnosis a) Diagnosis of amoebic keratitis - Wet mount, histology, culture (nutrient agar): corneal scrapings (cyst) b) Diagnosis of GAE - Brain biopsy, culture and immunofluorescence microscopy - CSF: slightly elevated protein levels or slightly decreased glucose levels. - CT scann of brain: inconclusive finding treatment Biguanide or chlorhexadine with or without diamine agent N. fawleri cyst Acanthomoeba spp. wall shape condition resistant N. fawleri Trophozoite Acanthomoeba spp. Pseudopodia Flagella Function Shape Invasive stage