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1 An Introduction to Parasitology
Parasitology is the study of parasites and as such does not include bacterial, fungal or viral parasites. Human parasites are separated into intestinal and blood borne parasites. For a parasite to be defined as intestinal it must have an intestinal life cycle stage, though it may have life-cycle stages in the heart, circulation, lung, tissue, other animals or the environment.
Parasites found in the intestines can be categorised into two groups: Protozoa and Helminths.
Protozoa are single celled organisms. There are four classes of Protozoa commonly found in concentrated faecal samples. These are differentiated by the method of motility. Protozoa include Entamoeba, Giardia, Trichomonas, Cryptosporidium, Isospora, Pneumocystis and Balantidium. There are two diagnostic life-cycle stages commonly seen in parasites - the cyst and the adult trophozoite stage. The trophozoite stage is analysed directly on a slide without concentration. Cysts require concentration. The key diagnostic factor is that Protozoan cysts are typically 5-30 microns in diameter, and as such are smaller than most Helminth eggs. Due to the size they are particularly difficult to see under the microscope if the sample clarity is bad.
The medically important Helminths are nematodes (roundworms), cestodes (tapeworms) and trematodes (flukes). Genera include: Fasciola, Schistosoma, Ascaris, Hookworm, Trichuris, Taenia and Enterobius. The normal stage for examination is the egg stage, although larvae may develop in some organisms (Strongyloides). The diameter of the eggs range from 30-150 microns.
The other major grouping of parasites are known as blood-borne parasites where they are transmitted through an arthropod vector. By far the most important arthropod for transmitting parasitic infections are the mosquitoes. Mosquitoes are known to carry malaria and filarial nematodes. Different types of biting flies transmit African trypanosomiasis, leishmaniasis and several kinds of filariasis.
Most protozoan and helminthic infections that are transmitted by arthropods can readily be diagnosed, on clinical grounds alone, but are usually identified by fairly simple techniques designed to present the presence of the causative parasite by microscopy. Sophisticated techniques are also being employed including highly sensitive and specific simple monoclonal antibody tests, DNA probes and PCR primers.
1.2 Infections acquired through the Gastrointestinal Tract
Many of the infections of the Gastrointestinal tract (GI) are caused by parasites that are cosmopolitan in distribution. Protozoa can be directly infectious for man when they are passed in the faeces into the environment, but helminths require a period of maturation whilst in the soil, where they become infectious. Others such as Taenia saginata require the involvement of an intermediate host during their life cycle.
Infections of the GI tract account for a high proportion of deaths in infants where the standards of hygiene and nutrition are low.
Faecal-oral transmission of the pathogens is the most common mode of GI infections, whereby water, food and hands become contaminated with faecal material which then come in contact with the mouth.
A number of GI infections can reach epidemic proportion, protozoal pathogen Cryptosporidium parvum, has been known to cause the severe water-borne epidemics, even in 1st world countries such as the United States and the UK. Other infections such as amoebiasis or enterobiasis can be more localised, infecting households or institutions.
Some of the rarer, protozoal infections such as the microsporidia are only now being understood as they are appearing as concomitant infections in people with depressed immune responsiveness, e.g. AIDS.
The Amoebae
Amoebae are characterised by possessing clear protoplasm which form pseudopodia. These pseudopodia are the means by which these organisms move and engulf bacteria and red blood cells for feeding purposes. The most common amoebae seen in the intestinal tract are Entamoeba histolytica / dispar,
Entamoeba coli, Entamoeba hartmanni, Endolimax nana and Iodamoeba
bütschlii. All but Entamoeba histolytica are thought to be non-pathogenic. The cysts can be identified in an ethyl acetate concentrate by the addition of iodine to reveal the characteristic inclusions and also by measuring the cyst using an eyepiece graticule. The trophozoites can be seen in a fresh saline preparation of the stool although accurate identification is on a permanently stained faecal smear.
Other links to the amoebae:
Introduction
There are a large number of species of amoebae which parasitise the human intestinal tract. Of these Entamoeba histolytica / dispar is the only species found to be associated with intestinal disease. Although many people harbour this organism world wide, only about 10% develop clinically invasive disease thus the parasite has been shown to present as two very differing clinical presentations.
The commensal or non-invasive luminal form where the parasite causes no signs or symptoms of disease.
The pathogenic or invasive form where the parasite invades the intestinal mucosa and produces dysentery or amoebomas and may give rise to extra-intestinal lesions via the blood, mainly to the liver.
Sargeaunt and Williams (1978) conclusively proved that invasive and non-invasive strains of E. histolytica could be differentiated by isoenzyme electrophoresis and the application of molecular biology has confirmed the presence of two distinct species with the same morphological features. The pathogenic or invasive species has retained the name E. histolytica and the non-pathogenic, non-invasive species has been named E. dispar.
![](http://www.soton.ac.uk/~ceb/Diagnosis/Vol1.h1.jpg)
Diagram 1. Life Cycle of Entamoeba histolytica: A protozoan in which its life cycle consists of two stages; cysts and trophozoites
Morphology of Trophozoites
The trophozoites of E. histolytica / dispar recovered from dysenteric stools exhibit ingested red blood cells and clear pseudopodia. Those of E. dispar will have no ingested red blood cells. They can be up to 60m in diameter and motility is rapid and unidirectional. On a permanently stained faecal smear e.g. Trichrome or Iron haematoxylin, the morphological features are more visible. When using Trichrome stain nuclei, chromidial bars, chromatin, red cells and bacteria stain red cytoplasm stains blue-green and background and yeasts stain green. The presence of a small centrally placed karyosome is clearly visible. With Iron haematoxylin, nuclear chromatin and the karyosome will be stained immensely black. The remainder will be varying shades of grey/black. (Fig 1)
موجودة في السيتوبلازم لكريات الدم الحمراء
التشخيص: من براز المريض
Morphology of cysts
Cysts of E. histolytica / dispar are 10 - 15m in diameter and contain 1 - 4 nuclei. Chromatoid bodies are usually present in young cysts as elongated bars with bluntly rounded ends. Glycogen is usually diffuse, but in young cysts it is often present as a concentrated mass, staining reddish brown with iodine. (Fig 2)
Entameoba Histolytica Cyst
الكيس الأميبي الـ هستوليتيكا
مكان الوجود:الأمعاء و خارج الأمعاء
الجزء المعدي: الكيس الرباعي النواة
طريقة العدوى: من البراز إلى الفم
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%201%20vol%201.gif)
Fig 1. Entamoeba histolytica trophozoites in a caecum biopsy, each with pseudopods. (<30µm)
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%202%20vol%201.gif)
Fig 2. Entamoeba histolytica cyst. Present with large nucleus and central (>30m ) Karyosome. Direct examination of bloody mucus stool. (> 30m m)
Clinical Disease
Amoebiasis is an infection usually caused by the pathogenic Entamoeba histolytica / dispar, and is commonly an infection of the colon. It has a world wide distribution where environmental sanitation is poor. The parasite may behave as a commensal (causing no harm to the host) or it may act as a parasite (harming the host). It is a disease of human beings, although some monkeys can become infected and the infection is then transmissable to humans.
Intestinal disease
Patients with intestinal disease may exhibit a number of symptoms including profuse diarrhoea with blood and mucus, fever and dehydration. Amoebic ulcers may develop in the large colon and can also be found in the rectal area. The ulcers are usually "flask shaped" with a small opening on the mucosal surface and a larger area below the surface. Fig 3 illustrates E. histolytica trophozoites in the intestine, resulting in amoebiasis.
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%203%20vol%201.gif)
Fig 3. Entamoeba histolytica trophozoite present in the intestine causing Intestinal amoebiasis. (HES stain). Note in the centre of the picture trophozoites stained red. The nucleus is situated on the right in the amoeba. The central karyosome and the nucleus membrane with its chromatin are distinct.
Hepatic Disease
Trophozoites are transported from the intestine to the liver and liver disease is characterised with abdominal pain, fever, hepatomegaly and tenderness. If the abscess ruptures, there is spreading to the brain, pericardium and other sites. If left untouched the abscess will grow normally until it reaches a surface where it can discharge, e.g. the skin, the peritoneum, the pleural cavity or the pericardium. The stretching of the liver is presumably the main source of the pain.
Microscopy
Where amoebic dysentery is suspected, the laboratory should be informed that a "hot stool" is being supplied so that it can be examined within twenty minutes of being passed. On cooling the amoebae stop moving which then become very difficult to identify. Direct microscopy should be done by mixing a small amount of the specimen in 0.9% sodium chloride solution. This permits detection of motile trophozoites of Entamoeba histolytica / dispar and can also provide information on the content of the stool i.e. the presence of leucocytes and red blood cells. One search e.g. primarily for cysts, not for amoebae, several stool samples are required to be examined, by direct microscopy and a sensitive concentration technique. Three negative stool samples are required before is can be accepted that there is no amoebic infection. Microscopic examination of an amoebic abscess aspirate e.g. in the liver or lungs, may reveal haematophagous trophozoites. It must be examined immediately by mixing a drop of warm saline with some aspirated pus on a microscope slide.
Serology
If visceral or hepatic amoebiasis is suspected serological tests should be done as microscopic methods do not always reveal the characteristic trophozoites. The tests of choice are indirect fluorescent antibody test (IFAT), counter immunoelectrophoresis (CIEP) and enzyme linked immunosorbent assay (ELISA)
The search for E. histolytica / dispar is mainly carried out in Europe and North America, as there is a natural concern to ensure that patients, even in the absence of symptoms are not harbouring parasites that may lead to serious complications later on.
Other links to Entamoeba histolytica
Entamoeba coli
Introduction
Entamoeba coli is a non-pathogenic amoeba with world wide distribution. Its life cycle is similar to that of E. histolytica but it does not have an invasive stage and does not ingest red blood cells.
Morphology of Trophozoite
The trophozoite is larger than that of E. histolytica ranging from 15-50m in diameter. It exhibits blunt pseudopodia with sluggish movement. A permanently stained preparation shows a nucleus with a moderately large eccentric karyosome with the chromatin clumped on the nuclear membrane. The cytoplasm appears granular containing vacuoles with ingested bacteria and other food particles. (Fig 4)
Morphology of Cysts
Cysts of E. coli are 15 - 30m in diameter and contain 1 - 8 nuclei with irrecgular peripheral chromatin: karyosomes not central. Chromatoid bodies are not frequently seen but when present they are usually splinter-like with pointed ends. Glycogen is usually diffuse but in young cysts is occasionally found as a well-defined mass, which stains reddish brown with iodine. (Fig 5)
Laboratory Diagnosis
Laboratory diagnosis is made by finding the characteristic cysts in an iodine stained, formol-ether concentration method or by detecting the characteristic trophozoites in a wet preparation or a permanent stained preparation.
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%204%20vol%201.gif)
Fig 4. Entamoeba coli Trophozoite
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%205%20vol%201.gif)
Fig 5. Entamoeba coli cysts.The cell wall is not visible as it is a trophozoite, Number of cysts with distinct walls and so will be changing shape continuously nuclei. (18 - 30m m) (Junod technique) to form pseudopods. (17m m) (MIF Stain)
Entamoeba hartmanni
Introduction
Entamoeba hartmanni is a non-pathogenic amoeba with world wide distribution. Its life cycle is similar to that of E. histolytica but it does not have an invasive stage and does not ingest red blood cells.
Morphology of trophozoites
Morphology of the trophozoites is similar to those of E. histolytica / dispar but they do not contain ingested red blood cells and the motility is less rapid. (Fig 6)
Morphology of cysts
Cysts of E. hartmanni 7-9m m in diameter and contain 1 - 4 nuclei. Chromatoid bodies are usually present in young cysts as elongated bars with bluntly rounded ends. Glycogen is usually diffuse, but in young cysts it is often present as a concentrated mass, staining reddish brown with iodine. (Fig 7)
Laboratory Diagnosis
Laboratory diagnosis is made by finding the characteristic cysts in an iodine stained, formol-ether concentration method or by detecting the characteristic trophozoites in a wet preparation or a permanent stained preparation.
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%207%20vol%201.gif)
Fig 6. Entamoeba hartmanni trophozoite
in stool smear.(Gomori-Wheatley stain). No distinct wall, cytoplasm is poorly coloured. . Distinct nucleus showing a central karyosome
and a thick chromatin.
![](http://www.soton.ac.uk/~ceb/Intergifs1/fig%206%20vol%201.gif)
Fig7. Entamoeba hartmanni cysts (Junod technique) 2 small cysts present (7µm) with distinct walls and vacuolizedcytoplasm.
المصدر
http://www.soton.ac.uk/~ceb/Diagnosis/Vol1.htm