疟原虫 Malaria Parasites(Plasmodium)
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<big><big><strong>Introduction</strong></big></big> | |||||||||
1 Malaria is one of the five major parasitic diseases in our country. Each year, more than 1,000,000 children die of malaria in Africa. | |||||||||
Caption: Two women recovering from malaria at Laquintinie hospital in Douala, Cameroon. Patients often have to share beds. | |||||||||
Caption:Navrongo hospital,Ghana: A mother waits patiently in the malaria ward by the bedside of her young child who is recovering from malaria. | |||||||||
2 Four species affecting humans: | |||||||||
P. vivax(P.v) causes benign tertian malaria. P. falciparum(P.f) causes malignant tertian malaria. P. malariae(P.m) causes quartan malaria. P. ovale(P.o) causes tertian malaria. | |||||||||
3 taxonomy: Protista>Protozoa>Apicomplexa>Sporozoa>Eucoccidiida>Plasmodidae>Plasmodium | |||||||||
<big><big>Life Cycle</big> </big> | |||||||||
The malaria parasite life cycle involves two hosts. During a blood meal, a malaria-infected female Anopheles
mosquito inoculates sporozoites into the human host. After initial replication in the liver (exoerythrocytic schizogony), the parasites undergo asexual multiplication in the erythrocytes (erythrocytic schizogony). Multiplication of the blood stage parasites is responsible for the clinical manifestations of the disease. In the blood, some parasites differentiate into sexual erythrocytic stages (gametocytes). The gametocytes, after ingestion by an Anopheles
mosquito during a blood meal, undergo a sporogonic cycle yielding sporozoites. Inoculation of the sporozoites into a new human host perpetuates the malaria life cycle.
Of note, in P. vivax and P. ovale, a dormant stage (hypnozoites) can persist in the liver and cause relapses by invading the bloodstream weeks, or even years later. | |||||||||
<big><big>Morphology</big></big> P.v morphology
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P.f morphology
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Ring forms (early trophozoites) P. vivax thin smear, showing early trophozoites. P. vivax thick smear, early trophozoites. | |||||||||
Late trophozoites P.vivax thin smear. | |||||||||
Schizonts P. vivax thin smear. P. vivax thin smear. | |||||||||
Male gametocytes Female gametocytes | |||||||||
Exflagellation | |||||||||
Zygote(No pic available now)
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Ookinete
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Oocyst
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Sporozoite | |||||||||
Exo-erythrocytic forms(EEF) 红外期裂殖体 Pre-erythrocytic schizont in liver. | |||||||||
<big><big><strong>Pathogenesis</strong></big></big> | |||||||||
1 Incubation period: P.v. 14-17 days P.f. 8-12 days | |||||||||
2 An attack occurs because of the sudden liberation of merozoites, malarial pigment and RBC debris into the blood stream. | |||||||||
Three stages of each paroxysm | |||||||||
(1) The cold stage (chill) lasting for 30min-1hr | |||||||||
(2) The hot stage(fever) lasting for 1-4hr (P.v. P.f. and P.o. once every other day; P.m. once every 2 days. ) | |||||||||
(3) Sweating stage 1-2hr | |||||||||
4 Recrudescence and Relapse | |||||||||
P.f and P.m. have only recrudescence, but, P.v. and P.o. both have relaps and recrudescence. | |||||||||
remnant of parasites in RBC result in recrudescence | |||||||||
hypnozoites cause relapse | |||||||||
5 Anemia; splenomegalia and fatal malaria-cerebral malaria caused by P.f. (small vessels are plugged) | |||||||||
<big><big><strong>Diagnosis</strong></big></big> | |||||||||
A. Clinical symptoms and history | |||||||||
B. Microscopic examination of blood. | |||||||||
1 Thin film and Thick(Giemsa's stain) | |||||||||
To master the morphology of parasites and changes of infected red cells | |||||||||
2 P.f.: Only Ring forms and gametocytes can be found in blood film. | |||||||||
C. Other methods: Immunologic/Biochemical/Molecular diagnosis.
There are four species of malaria which commonly infect man, P.falciparum, P.vivax, P.malariae and P.ovale. The most important of these is P.falciparum because it can be rapidly fatal. P.vivax is the most commonly seen species. P.malariae is present in Africa, South America and in one area of New Guinea. P. ovale is relatively unusual outside Africa although some cases are now being identified in other regions(eg. Southern States of India). A number of new techniques based on the "dipstick" format, have recently become available for the diagnosis of malaria. These include the ICT-Malaria Pf, OptiMALr and the Determine kits. The methods are based on the principle of the detection of plasmodial histidine rich protein-2 (HRP-2) or parasite-specific lactate dehydrogenase (pLDH) which is present in P. falciparum infections, with a number of reports claiming sensitivities and specificities approaching 100%. Some of these "dipstick" methods have been extended to include screening for other forms of malaria but to date results have not been quite so impressive. In this laboratory we have found these kits to be very useful screening or confirmatory tests, especially when there is difficulty in identifying scanty ring forms in blood films. They are particularly useful out of hours when more junior, less experienced staff are likely to be on duty. However we would like to emphasise, that we regard these methods as additional to the long established method of examining thick and thin blood films (outlined below), which is still regarded as the "gold standard", NOT as replacement methods. Examination of a thick blood film should be the first step, if parasites are seen then the species should be confirmed by the examination of a thin film. Blood is best collected when the patient's temperature is rising. Preparation of thick and thin blood films :- Thick films:- place a drop of blood in the middle of a clean microscope slide and with the corner of a second slide spread the drop until it is about the size of a five cent coin. The thickness should be such that it is just possible to see news print through it. Thin films are made in the standard manner. Allow the films to dry, do not leave on the bench in a laboratory which is not fly proofed otherwise the film will be eaten. <center><table><tbody><tr><th><img height="445" src="http://img.dxycdn.com/trademd/upload/asset/meeting/2013/07/30/A1375172587.jpg" width="491" />
</th></tr></tbody></table></center> When the films are dry, fix and stain the thin films in the conventional manner but be careful about the pH of the stain, a slightly alkaline stain is recommended (pH 7.2) as an acid stain may fail to show the parasites. When only a few thick films are to be stained it is best to use dilute Giemsa stain (1/20), using a staining jar so that the film is in an upright position, this will allow any debris to fall to the bottom of the jar. Do not fix the sample prior to staining. Stain for about 30 minutes, wash gently with clean water and allow to dry. If available use a positive control. When a large number of thick films require staininq, Field's stain is preferred because it is very quick. Field's stain comprises two solutions; a polychrome methylene blue (A) and eosin (B). The solutions are kept in covered staining jars.
If films are old or too thick the red cells may not lyse completely in the brief staining time. If this is likely dip the film in clean water for a few seconds or until the haemoglobin has dispersed before staining. Instructions for preparing Field's stain can be found in many laboratory text books. Under the microscope examine the thick film first, using an oil immersion or high dry lens to determine if parasites are present. Be aware of the patient's platelet and leucocyte counts. Malaria is usually associated with a normal or reduced leucocyte numbers. A leucocytosis is only found in terminal cases. Platelet numbers are moderately or markedly reduced in some 80% of patients with malaria. Parasites may appear distorted if the patient has been treated or has had inadequate prophylaxis. Mixed infections are not uncommon. The illustrations show the characteristics of the four species. <center><table><tbody><tr><th><img height="510" src="http://img.dxycdn.com/trademd/upload/asset/meeting/2013/07/30/A1375172600.gif" width="488" />
</th></tr><tr><td><b>Diagnostic points:-</b><br /><ol><li>Red Cells are not enlarged.</li><li>Rings appear fine and delicate and there may be several in one cell.</li><li>Some rings may have two chromatin dots.</li><li>Presence of marginal or applique forms.</li><li>It is unusual to see developing forms in peripheral blood films.</li><li>Gametocytes have a characteristic crescent shape appearance. <br />However, they do not usually appear in the blood for the first four weeks of infection.</li><li>Maurer's dots may be present.</li></ol></td></tr></tbody></table></center> <center><table><tbody><tr><th><img height="433" src="http://img.dxycdn.com/trademd/upload/asset/meeting/2013/07/30/A1375172592.gif" width="492" />
</th></tr><tr><td><b>Diagnostic points:- </b><br /><ol><li>Red cells containing parasites are usually enlarged.</li><li>Schuffner's dots are frequently present in the red cells as shown above.</li><li>The mature ring forms tend to be large and coarse.</li><li>Developing forms are frequently present.</li></ol></td></tr></tbody></table></center> <center><table><tbody><tr><th><img height="334" src="http://img.dxycdn.com/trademd/upload/asset/meeting/2013/07/30/A1375172602.gif" width="488" />
</th></tr><tr><td><b>Diagnostic points :- </b><br /><ol><li>Ring forms may have a squarish appearance.</li><li>Band forms are a characteristic of this species.</li><li>Mature schizonts may have a typical daisy head appearance with up to ten merozoites.</li><li>Red cells are <b><u>not</u></b>
enlarged.</li><li>Chromatin dot may be on the inner surface of the ring.</li></ol></td></tr></tbody></table></center> <center><table><tbody><tr><th><img height="430" src="http://img.dxycdn.com/trademd/upload/asset/meeting/2013/07/30/A1375172586.gif" width="488" />
</th></tr><tr><td><b>Diagnostic points :-</b>
<ol><li>Only found in Africa.</li><li>Red cells enlarged.</li><li>Comet forms common (top right)</li><li>Rings large and coarse.</li><li>Schuffner's dots, when present, may be prominent.</li><li>Mature schizonts similar to those of P. <i>malariae</i>
but larger and more coarse.</li></ol></td></tr></tbody></table></center> | |||||||||
<center><table><tbody><tr><td><font><b><i>Plasmodium</i>
species</b></font></td><td><font><b>Stages found in blood</b></font></td><td><font><b>Appearance of Erythrocyte (RBC)</b></font></td><td><font><b>Appearance of Parasite</b></font></td></tr><tr><td><font><i>P. falciparum</i>
</font><p> </p></td><td><font>Ring</font>
<p> </p></td><td><font>normal; multiple infection of RBC more common than in other species</font></td><td><p><font>delicate cytoplasm; 1-2 small chromatin dots; occasional appliqué (accollé forms)</font></p></td></tr><tr><td><font>Trophozoite</font>
<p> </p></td><td><font>normal; rarely, Maurer's clefts (under certain staining conditions)</font></td><td><font>seldom seen in peripheral blood; compact cytoplasm; dark pigment</font></td></tr><tr><td><font>Schizont</font>
<p> </p></td><td><font>normal; rarely, Maurer's clefts (under certain staining conditions)</font></td><td><font>seldom seen in peripheral blood; mature = 8-24 small merozoites; dark pigment, clumped in one mass</font></td></tr><tr><td><font>Gametocyte</font>
<p> </p></td><td><font>distorted by parasite</font></td><td><font>crescent or sausage shape; chromatin in a single mass (macrogametocyte) or diffuse (microgametocyte); dark pigment mass</font></td></tr><tr><td><font><i>P. vivax</i></font>
<p> </p></td><td><font>Ring</font>
<p> </p></td><td><font>normal to 1</font><font>1/4</font><font> X, round; occasionally fine Schüffner's dots; multiple infection of RBC not uncommon</font></td><td><font>large cytoplasm with occasional pseudopods; large chromatin dot</font></td></tr><tr><td><font>Trophozoite </font><p> </p></td><td><font>enlarged 1</font><font>1/2</font><font>-2 X; may be distorted; fine Schüffner's dots</font></td><td><font>large ameboid cytoplasm; large chromatin; fine, yellowish-brown pigment</font></td></tr><tr><td><font>Schizont</font>
<p> </p></td><td><font>enlarged 1</font><font>1/2</font><font>-2 X; may be distorted; fine Schüffner's dots</font></td><td><font>large, may almost fill RBC; mature = 12-24 merozoites; yellowish-brown, coalesced pigment</font></td></tr><tr><td><font>Gametocyte </font><p> </p></td><td><font>enlarged 1</font><font>1/2</font><font>-2 X; may be distorted; fine Schüffner's dots</font></td><td><font>round to oval; compact; may almost fill RBC; chromatin compact, eccentric (macrogametocyte) or diffuse (microgametocyte); scattered brown pigment</font></td></tr><tr><td><i><font>P. ovale</font></i>
<p> </p></td><td><font>Ring</font>
<p> </p></td><td><font>normal to 1</font><font>1/4</font><font> X, round to oval; occasionally Schüffner's dots; occasionally fimbriated; multiple infection of RBC not uncommon</font></td><td><font>sturdy cytoplasm; large chromatin</font></td></tr><tr><td><font>Trophozoite </font><p> </p></td><td><font>normal to 1</font><font>1/4</font><font> X; round to oval; some fimbriated; Schüffner's dots</font></td><td><font>compact with large chromatin; dark-brown pigment</font></td></tr><tr><td><font>Schizont</font>
<p> </p></td><td><font>normal to 1</font><font>1/4</font><font> X, round to oval, some fimbriated, Schüffner's dots</font></td><td><font>mature = 6-14 merozoites with large nuclei, clustered around mass of dark-brown pigment</font></td></tr><tr><td><font>Gametocyte </font><p> </p></td><td><font>normal to 1</font><font>1/4</font><font> X; round to oval, some fimbriated; Schüffner'dots </font></td><td><font>round to oval; compact; may almost fill RBC; chromatin compact, eccentric (macrogametocyte) or more diffuse (microgametocyte); scattered brown pigment </font></td></tr><tr><td><font><i>P. malariae</i></font>
<p> </p></td><td><font>Ring </font><p> </p></td><td><font>normal to 3/4 X </font></td><td><font>sturdy cytoplasm; large chromatin </font></td></tr><tr><td><font>Trophozoite</font>
<p> </p></td><td><font>normal to 3/4 X; rarely, Ziemann's stippling (under certain staining conditions) </font></td><td><font>compact cytoplasm; large chromatin; occasional band forms; coarse, dark-brown pigment </font></td></tr><tr><td><font>Schizont </font><p> </p></td><td><font>normal to 3/4 X; rarely, Ziemann's stippling (under certain staining conditions) </font></td><td><font>mature = 6-12 merozoites with large nuclei, clustered around mass of coarse, dark-brown pigment; occasional rosettes </font></td></tr><tr><td><font>Gametocyte </font><p> </p></td><td><font>normal to 3/4 X; rarely, Ziemann's stippling (under certain staining conditions) </font></td><td><font>round to oval; compact; may almost fill RBC; chromatin compact, eccentric (macrogametocyte) or more diffuse (microgametocyte); scattered brown pigment </font></td></tr></tbody></table></center> | |||||||||
<big><big>Tteatment</big></big> | |||||||||
1 Three principles: | |||||||||
P. falciparum. Uncomplicated malaria (where patients can take oral therapy) can be treated with one of three regimens:
Choice of regimen is based on :-
In uncomplicated cases in which nausea and vomiting preclude oral therapy, quinine dyhidrochloride 10 mg salt/kg base can be given I.V. in 5% w/v dextrose or normal saline as a 4-hour infusion 8-hourly until the patient can take medication by mouth. Severe malaria. (where patients have coma, jaundice, renal failure, hypoglycaemia, acidosis, severe anaemia, high parasite count, hyperpyrexia) is ideally treated in an intensive care or high dependency unit where patients can be monitored closely both clinically and biochemically. Intravenous quinine is the treatment of choice but rapid injection can lead to hypotension, dysrhythmias and death. In patients who have not received quinine in the previous 48 hours, one of two regimens can be used:
Electrocardiographic monitoring can be done if available but is not essential unless additional cardiac risks are present.
P. vivax. Adult treatment.
The primaquine is preferably started after the chloroquine. When the infection is acquired in New Guinea, 3 tablets of primaquine (22.5mg base) should be given daily for 14 days. In the case of a relapse repeat both chloroquine and primaquine treatment. Up to three relapses may occur before the parasite is finally eliminated. Unfortunately there is no other effective treatment. Etaquine is a newly developed long-acting, potent primaquine-like drug which may be available soon for radical cure. Patients should have their G6PD status checked before primaquine is prescribed. Patients with G6PD deficiency may undergo haemolysis if given a daily dose of primaquine and it is recommended that these patients be given 30-45mg once a week for 8 weeks. Malarone TM may also be used to treat P. vivax malaria but a course of primaquine will still be required to eliminate liver forms. P. malariae, P. ovale. Artemisinins. Artemisinin has been used for many years by the Chinese as a traditional treatment for fever and malaria. It is a sesquiterpene lactone derived from Artemisia annua. Because it is being increasingly used in a number of countries and is both cheap and effective it was decided to include treatment schedules here. However, it is not yet licensed for use in Australia, North America or Europe. Its main value at present is in the treatment of multi drug resistant falciparum malaria. If artemisinin is used to treat vivax malaria it should be accompanied by a course of primaquine. Unless used with a second antimalarial as described below there is likely to be a high recrudescent rate. Side effects have been reported but these are comparatively rare and seldom severe. It is recommended only for treatment not for prophylaxis. Artemisinin (500mg tablets): give 10-20 mg/kg on day 1 (500-1,000 mg) orally then 500mg for 4 days. Then give mefloquine 15mg base/kg or split dose 25mg base/kg. Artemisinin (200 mg suppositories): for severe malaria 600-1200mg stat, 400-600mg after 4 hours then 400-800mg twice daily for 3 days. Give mefloquine as above. Artesunate (50 & 60 mg vials for intravenous use): for severe malaria 120mg I.V. stat. 60 mg at 4, 24 and 48 hours, 50-60 mg on days 3-5. Give mefloquine as above. Dihydroartemisinin (20 mg tablets): first dose 120mg then 60mg daily for 4-6 days then give mefloquine as above. Artemeter (vials for intramuscular use ): for severe malaria 3.2 mg/kg intramuscularly stat then 1.6mg/kg twice daily for 3-7 days, Give mefloquine as above. | |||||||||
2 Medicines | |||||||||
<center><table><tbody><tr><td> </td><td>Quinine</td><td>Chloroquine</td><td>Primaquine</td><td>Pyrimethamine</td></tr><tr><td>Anti-ES</td><td>+++</td><td>+++</td><td>-</td><td>-</td></tr><tr><td>Anti-EE</td><td>-</td><td>-</td><td>++</td><td>++</td></tr><tr><td>Anti-Gametocyte</td><td>-</td><td>-</td><td>++</td><td>++</td></tr></tbody></table></center> | |||||||||
<big><big><strong>Prevention</strong></big></big> | |||||||||
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Caption: The community plays an important role in the successful use of insecticide-impregnated bednets as a means to combat malaria. Householders in Xuyi County, Jiangsu Province await their turn to dip their nets in deltamethrin. | |||||||||
<big><big><strong>Epidemiology</strong></big></big> | |||||||||
World Distribution of Malaria | |||||||||
In China | |||||||||
1 South China-high endemic area: P.f. is prevalent and Mosquito vector: A. mimimus. | |||||||||
2 Central China and Yantze Valley: P.v predominant. Mosquito vector: A. listeri | |||||||||
3 Huang Huai Region: Vector: A.sinensis. |