Dr.T.V.Rao MD

 Entamoeba histolytica
was first described by Lambl in 1859 and Losch established it pathogenic nature in 1875 in a dysenteric patient is St.Petersberg  Councilman and lafleur in 1981 described amoebic liver abscess.  Schauudinn ( 1903 ) differentiated pathogenic and nonpathogenic types of Amoebae

 Amebiasis (am-e-BI-asis) is a disease caused by a one-celled parasite called Entamoeba histolytica (ent-a-ME-ba his-to-LI-ti-ka). Although it is more common in people who live in tropical areas with poor sanitary conditions

Amoebiasis a Major Health Problem
 Amoebiasis is estimated to cause 70,000
deaths per year world wide Symptoms can range from mild diarrhea to dysentery with blood and mucus in the stool. E. histolytica is usually a commensals organism. Severe Amoebiasis infections (known as invasive or fulminant amoebiasis) occur in two major forms. Invasion of the intestinal lining causes amoebic dysentery or amoebic colitis.

Trends of Amoebiasis

Transmission of Amebiasis
 Amoebiasis is transmitted
by fecal contamination of drinking water and foods, but also by direct contact with dirty hands or objects as well as by sexual contact. Additionally, geophagy is a common route of infection in certain cultures.
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Nature of the disease
 Symptoms are usually gastrointestinal including
diarrhoea, vomiting, abdominal pain or discomfort and fever. Symptoms take from a few days to a few weeks to develop and manifest themselves, but usually it is about two to four weeks. Most infected people are asymptomatic but this disease has the potential to make the sufferer dangerously ill, especially if there is any suggestion of immunocompromised.

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Events on Amoebiasis

Trophozoites of E.histolytica

Trophozoites and Cystic stages

Cystic stage - E.histolytica

Amoebiasis causes Epithelial damage

Numerous Eosinophilic spherical structure within necrotic area.

Tissue showing Amoebic infection
 The spherical
structure (Trophozoites) has one basophilic nuclei about the size of RBC’s. Note some RBC's are phagocytosed by the Trophozoites (erythrophagocytosis)

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Virulence factors
 Trophozoites of E.histolytica interact with host through a
series of steps 1 Adhesion of target cell, phagocytosis and cytopathic effect 2 E.histolytica induces both Humoral and cell mediated immune responses. 3 Virulence factors – In many circumstances lumen dwelling Amoeba may be asymptomatic 4 Causes disease only when invade the Intestine 5 Virulence is associated with secretion of Cysteine proteniase which assists the organism in digesting the extracellular matrix and invading tissues

Cysteine proteinase Complement factor C3
 It is observed
Cysteine proteinase produced by invasive strains of E.histolytica inactivates the complement factor C3 and are thus resistant to Complement mediated lysis.

Cysteine proteinase virulent factor
 Cysteine proteinase is an
important virulent factor  Its presence makes E.histolytica is resistant to complement mediated lysis  Can cleave the extracellular structural matrix and degrade fibronectin and laminin, as well as type I collagen.  In this process basement membrane is degraded and leads to invasion

 Lectin binding
Zymodeme analysis, genome specific DNA analysis and staining with Monoclonal antibodies have been successfully used as markers to identify invasive strains of E.histolytica

Types of Zymodemes
 Based on
Electrophoretic mobility E.histolytica strains are classified into 22 Zymodemes  However only 9 are invasive

Invasive x Noninvasive strains
 The invasive and non
invasive strains may appear identical may represent two distinct species  1 Invasive strain – E.histolytica  2 Non invasive strains reclassified as E.dispar.

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Host Factor Contributions
 Several factors contribute to influence
infection 1 Stress 2 Malnutrition 3 Alcoholism 4 Corticosteriod therapy 5 Immunodeficiency 6 Alternation of Bacterial flora

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Risk Factors
 People in developing countries that have     

poor sanitary conditions Immigrants from developing countries Travellers to developing countries People who live in institutions that have poor sanitary conditions HIV-positive patients Men who have sex with men
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 No symptoms (in the
majority of cases),  Vague gastrointestinal distress,  Dysentery (with blood and mucus).

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How the Amebiasis Manifests
 Most cases of amebiasis have very mild
symptoms or none.  More severe infection may cause fever, profuse diarrhea, abdominal pain, jaundice, anorexia, and weight loss.  In severe cases, it can lead to development of abscesses (pockets of amoebae and inflammatory cells) in the liver or, more rarely, the brain.

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Clinical symptoms are Vague
 Wide spectrum, from asymptomatic
infection ("luminal amebiasis"), to invasive intestinal amebiasis (dysentery, colitis, appendicitis, toxic mega colon, amebomas), to invasive extra intestinal amebiasis (liver abscess, peritonitis, pleuropulmonary abscess, cutaneous and genital amoebic lesions).

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Diagnosis of Amebiasis

 Fresh stool: wet mounts and
permanently stained preparations (e.g., trichrome).  Concentrates from fresh stool: wet mounts, with or without iodine stain, and permanently stained preparations (e.g., trichrome). Concentration procedures, however, are not useful for demonstrating Trophozoites.

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Diagnosis of Amebiasis
 Diagnosis of amebiasis can be very difficult. One
problem is that other parasites and cells can look very similar to E. histolytica when seen under a microscope. Therefore, sometimes people are told that they are infected with E. histolytica even though they are not. Entamoeba histolytica and another ameba, Entamoeba dispar, which is about 10 times more common, look the same when seen under a microscope

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 This is the traditional
means of diagnosing the disease—one simply looks at a sample of stool under a microscope. Because E. histolytica is not always found in every stool sample, several samples from different days may be needed. Sometimes red blood cells that have been ingested by the parasite are visible.

Microscopic examination of Stool
 A sample of freshly
collected fecal specimen containing mucous and blood is transferred on a slightly warm slide and covered with cover slip and examined microscopically
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E. histolytica /E. dispar cyst.

E. histolytica/E. dispar cysts stained with trichrome

Specific Diagnosis of active infection should demonstrate Trophozoites

 Motile Trophozoites

throwing pseudopodia and containing red blood cells found in large number  Endoplasm appear bluish or found glass in appearance and nucleus is not visible but faint outline may be observed

Charcot Leyden crystals in stool examination supports the Diagnosis,
 Cysts have smooth and
thin cell wall and contain round, retractile chromotoid bars  Glycogen mass is not visible  RBC’s and pus cells are found in fair number  Charcot Leyden crystals, diamond shaped clear and retractile structures are present in faeces

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 Routinely not used  Trophozoites stains
yellow to light brown, Nucleus is clearly visible with central karyosome Cysts shows a smooth and hyaline appearance, Nucleus is clearly seen and no more than 4 nuclei are present, Glycogen mass stains brown, while chromotoid bars are not stained.

Mucosal Scrapings
 Mucosal scrapings can
be obtained by sigmoidoscopy useful in atypical presentations and may serve as adjunct to conventional stool examination for Ova and cyst  Direct wet mount, a permanently stained smear and immuno stained smears are examined.

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Extra intestinal Amoebiasis
 The specimens are
obtained from Liver, lung, or Brain biopsy samples and subjected to routine Histopathology ( H&E) sections  Giemsa stained touch preparations which will revel Trophozoites in extra intestinal lesions.

Amoebic Liver Abscess
 The pus in liver
abscess appear as red Anchovy sauce like appearance  The material aspirated is likely to contain Trophozoites and may be detected by direct microscopic examination

Serological Diagnosis
 The serological become reactive in invasive     
Amoebiasis 1 Indirect Heamagglutination assay ( IHA ) 2 ELISA 3 Latex agglutination test 4 gel diffusion 5 Counter current Imunoelectrphoresis

 Serological tests remain positive for several years ever after successful

 Cultures are not done routinely  Boeck and Drbohlav’s medium modified

by Laidlaw extensively used for isolation and maintenance of E.histolytica.  Diamonds axenic medium used in studies on Pathogenicty, antigenic characterization and drug sensitivity tests
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Do we need culturing for Diagnosis ?
 Trying to get the
amoeba to grow outside the body is very difficult and unreliable, and is therefore not generally done

Immunity in Amoebiasis
 Infection with invasive
strains of E.histolytica induce both Humoral and cellular response.  Infection offers some degree of protection.

Immunological Tests are not confirmatory of Acute Infections
 When the body is exposed
to an infection, the immune system creates antibodies to fight it off. These can be detected with a blood test, and provide evidence that the person has been infected with E. histolytica. Unfortunately, this test does not distinguish between past and present infection
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Emerging methods in Diagnosis
 These are considered the
most useful tests for detecting E. histolytica. They test directly for the parasite itself by exposing some stool to a strip of paper coated with antibodies. The parasites will stick to the antibodies on the paper. The test distinguishes E. histolytica from other parasites.

Treating Amebiasis.
 Frequently, either metronidazole (Flagyl) or
tinidazole (Fasigyn) are used to treat Amebiasis. If this does not work, Chloroquine, emetine, and dehydroemetine can be used. Eliminating cysts in carriers who do not have symptoms is accomplished with diloxanide furoate (Furamide), iodoquinol (Yodoxin), and paromomycin. Nitazoxanide is a newer drug that shows promise against not only E. histolytica but many other parasites as well.

Treating extra intestinal Amoebiasis
 Amoebic abscess is
treated similarly to dysentery, with antibiotics. Sometimes surgical drainage may be performed, but this is usually to rule out other (bacterial) causes of abscess. It is also performed if an abscess is about to, or has already ruptured.

Preventing Amoebiasis
 Drink only bottled or boiled (for 1 minute) water, or
carbonated (bubbly) drinks in cans or bottles. Fountain drinks and any drinks with ice cubes are not safe. Water can be made safe by filtering it through an "absolute 1 micron or less" filter and dissolving iodine tablets in the filtered water.  Avoid fresh fruit or vegetables that were peeled by someone else.  Avoid milk, cheese, or dairy products that may not have been pasteurized.  Avoid anything sold by street vendors.

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Food safety
 Thoroughly cook all raw foods.  * Thoroughly wash raw
vegetables and fruits before eating.  * Reheat food until the internal temperature of the food reaches at least 167º Fahrenheit.  Wash your hands before preparing food, before eating, after going to the toilet or changing diapers, after smoking or after using a tissue or handkerchief.

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Personal Hygiene
 Wash hands thoroughly
with soap and hot running water for at least 10 seconds after using the toilet or changing a baby's diaper.  Clean bathrooms and toilets often. Pay particular attention to toilet seats and taps.  Avoid sharing towels or face washers.

 Vaccines are being developed and tested
for the treatment of Amebiasis. The vaccine is a modified version of the proteins expressed on the surface of E. histolytica. A study in rodents found that the vaccine prevented the formation of liver abscesses, but much more research is needed to determine if these vaccines are useful and safe in humans

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Created for Awareness to Medical and Paramedical workers in Developing World
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