Malaria Parasites (Plasmodium)

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1 Malaria Parasites (Plasmodium)
Department of Parasitology

2 Introduction 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. 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 P.v and P.f are more important than P.m and P.o.
A few cases of P.o have been reported in China.

4 3. taxonomy: Protista>Protozoa>Apicomplexa>Sporozoa>
Eucoccidiida>Plasmodidae>Plasmodium

5 Life Cycle In mosquito 10days at 25℃
Male and female gametocytes → Male and female gametes → zygotes → ookinetes → oocysts → sporozoites gametogony: stomach lumen gamogenesis sexual reproduction sporogony:stomach wall agamogenesis asexual reproduction

6 Anopheles gambiae: adult female bloodfeeding on human skin

7 In human 1. pre-erythrocytic stage EE (exo-erythrocytic stage) 8 days
       tachy-sporozoites TS        brady-sporozoites BS 2.       ES (erythrocytic stage) 48h schizogony EE and ES result in more merozoites

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11 生活史 1.感染阶段：子孢子 2.感染方式：蚊虫叮咬;输血。 3.致病阶段：红内期原虫 4.寄生部位：肝、红细胞 5.媒介：蚊(为终宿主） 6.诊断阶段：红内期原虫

12 Characteristic of life cycle
Intermediate host : human Final host : mosquito Infective stage : sporozoite（子孢子） Infective mode : mosquito bite skin of human Parasitic position : liver and red blood cells Transmitted stage : gametocytes（配子体） Schizogonic cycle in red cells （红内期裂体增殖周期）: 48 hrs/P.v；36-48 hrs/P. f Sporozoite : tachysporozite（速发型子孢子） and bradysporozite（缓发型子孢子）

13 Morphology P.v morphology P.f morphology
Ring forms (early trophozoites) Late trophozoites Schizonts Male gametocytes Female gametocytes

14 Morphology In human Pre-erythrocytic stage
Ring forms (early trophozoites) Late trophozoites Schizonts Male gametocytes Female gametocytes

15 Pre-erythrocytic schizont in liver
Pre-erythrocytic schizont in liver. These mature in 6-14 days’ time liberation merozoites into the blood stream. Giemsa-colophonium. ×400. Enlarged by 5.4.

16 Plasmodium vivax: Blood Stage Parasites: Thin Blood Smears Fig
Plasmodium vivax: Blood Stage Parasites: Thin Blood Smears  Fig. 1: Normal red cell; Figs. 2-6: Young trophozoites (ring stage parasites); Figs. 7-18: Trophozoites; Figs : Schizonts; Figs. 28 and 29: Macrogametocytes (female); Fig. 30: Microgametocyte (male)  

17 P. vivax Rings in 2 slightly enlarged RBCs; 17 y.o. man with a relapse due to P. vivax (PCR confirmed), 6 months after returning from a visit to Papua New Guinea (specimen contributed by Virginia SHD)

18 Double infection with rings, RBC enlarged and deformed, Schüffner's dots beginning to become visible; 69 y.o. woman

19 Late ring in a RBC with Schüffner's dots; 60 y.o. man

20 Plasmodium vivax: Trophozoites Figs
Plasmodium vivax: Trophozoites Figs. 8-18: Increasingly mature trophozoites of P. vivax

21 Smears from patients: Increasingly mature trophozoites
Smears from patients: Increasingly mature trophozoites. The RBCs are enlarged and deformed, the parasites are ameboid, and the Schüffner's dots vary in intensity

22 Plasmodium vivax: Trophozoites

23 Plasmodium vivax: Schizonts Figs. 19-27: Increasingly mature schizonts

24 P. vivax thin smear. A mature schizont about to rupture
P. vivax thin smear. A mature schizont about to rupture. A clump of malarial pigment can be seen in the center. Giemsa. ×1000. Enlarged by 5.4.

25 Plasmodium vivax: Schizonts Smears from patients: Note that in these patients, the Schüffner's dots are not conspicuous. (This happens in many of the smears received; it is probably related to variability in staining.) 

26 Plasmodium vivax: Gametocytes Fig
Plasmodium vivax: Gametocytes Fig. 28 and 29: Nearly mature and mature macrogametocyte (female); Fig. 30: Microgametocyte (male) Plasmodium falciparum: Gametocytes Figs. 27, 28: Mature macrogametocytes (female); Fig. 29, 30: Mature microgametocytes (male)

27 间日疟原虫雌配子体 mature macrogametocyte (female)

28 间日疟原虫雄配子体

29 恶性疟原虫 Plasmodium falciparum
环状体 Ring form

30 Morphology Male and female gametes → zygotes → ookinetes → oocysts
In mosquito Male and female gametes → zygotes → ookinetes → oocysts → sporozoites

31 雄配子形成 Male gametes

32 ookinetes

33 Ookinete, from the midgut of an infected mosquito. Giemsa. ×800
Ookinete, from the midgut of an infected mosquito. Giemsa. ×800. Enlarged by 5.4.

34 Oocysts

35 Oocysts of P. falciparum in midgut of an infected mosquito
Oocysts of P. falciparum in midgut of an infected mosquito. Oocysts appear as circular bodies. Fresh preparation. ×100. Enlarged by 5.4.

36 Sporozoites

37 Clinical Manifestation and Pathogenesis
1. Incubation period: P.v days P.f days 2. An attack occurs because of the sudden liberation of merozoites, malarial pigment and RBC debris into the blood stream.

38 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

39 致病 Pathogenicity mechanism 发作 Paroxysm (attack of malaria)
----疟疾的一次典型发作表现为寒战、高热和出汗退热3个连续阶段。发作是由红内期的裂体增殖所致。 mechanism ----成熟裂殖体胀破红细胞后，裂殖子及红细胞碎片进入血流，部分被巨噬细胞、中性粒细胞吞噬，刺激这些细胞产生内源性热原质，它和疟原虫的代谢产物共同作用于宿主下丘脑的体温调节中枢，引起发热。随着血内刺激物被吞噬和降解，机体通过大量出汗，体温逐渐恢复正常，机体进入发作间歇阶段。

40 Process ----to shows a succession of 3 stages
----由于红内期裂体增殖是发作的基础，发作具有周期性，与红内期裂体增殖周期一致。 p.v. 48 hrs; P.f./36 to 48 hrs ;P.m./72 hrs Process to shows a succession of 3 stages ⑴.寒战 The cold stage (chill), lasting for 30 min to 1 hr. ⑵.发烧 The hot stage (fever), 1 to 4 hrs. (3).出汗 Sweating stage， 1 to hrs. Characteristic ----(1).周期性 periodic (2).重复性 repeated (3).规律性 regular

41 3. Recrudescence and Relapse
        Recrudescence occurs when the blood schizonticide does not eliminate all parasites from the blood stream, either because the dose was inadequate or because the parasite is resistant to the drug. Relapse occurs in P vivax and P ovale infections after the delayed development of liver- stage parasites that have not been treated adequately with a tissue schizonticide. P.f and P.m. have only recrudescence, but, P.v. and P.o. both have relapse and recrudescence.

42 Remnant of parasites in RBC result in recrudescence
Hypnozoites in liver cause relapse

43 4. Anemia; splenomegaly and fatal malaria-cerebral malaria caused by P
4. Anemia; splenomegaly and fatal malaria-cerebral malaria caused by P.f. (small vessels are plugged)

44 P.f: schizogony takes place in the capillaries of the internal organs, the infected red cells tend to adhere to one other and the small vessels may become plugged. This may produce several fatal results: Cerebral malaria Renal failure Serious anemia Acute respiratory distress syndrome ARDS Shock

45 贫血 anemia ---- 疟疾发作数次后可出现贫血，以恶性疟为甚。贫血的原因有： （1）疟原虫直接破坏红细胞外。 （2）脾功能亢进，吞噬大量正常的红细胞。 （3）免疫病理损害（怎样损害红细胞）。 （4）骨髓造血功能受抑制。 问题：疟疾患者的贫血程度常超过疟原虫直接破坏红细胞的程度，为什么？

46 脾肿大 Splenomegaly ---- 初发患者多在发作3～4天后，脾开始肿大，长期不愈或反复感染者，脾肿大十分明显。 主要原因是脾充血和单核-巨噬细胞的增生。

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51 恶性疟病人（巨脾） Tropical splenomegaly syndrome. This young Malaysian girl had a protuberant abdomen due to a large spleen extending to the pelvis. She showed high antibody levels to P. falciparum and raised Ig M. It is believed that the condition results from repeated infection with malarial parasite along with an abnormal immunological response.

52 恶性疟疾(凶险性疟疾)Malignant malaria
----凶险型疟疾绝大多数由恶性疟原虫所致。常见于无免疫力或因各种原因延误诊治的疟疾患者，因血中原虫数量剧增而出现凶险症状。 常见的有脑型、超高热型等，多表现为持续高热、抽搐、昏迷、重症贫血、肾功能衰竭等，来势凶猛，若不能及时诊治，死亡率很高。

53 恶性疟病人肝、脑标本切片 Section of liver from a case of P.  falciparum infection. The Kupffer cells (macrophages) are loaded with black pigment, which is malarial pigment (haemozoin). H and E. ×400. Enlarged by 23.4.

54 Brain from a case of P. falciparum infection
Brain from a case of P. falciparum infection. The cortex has a brownish colour and minute haemorrhages can be seen in the region of the basal ganglia. In this case death occurred due to haemorrhages and necrosis in the CNS (cerebral malaria)

55 Section of brain from a case of P. falciparum infection
Section of brain from a case of P.  falciparum infection. The infected cells adhere to the endothelial lining of the blood vessels. In small blood vessels this leads to occlusion and necrosis. H and E. ×400. Enlarged by 23.4.

56 Same tissues observed in polarized light
Same tissues observed in polarized light. The malarial pigment inside the red cells shines making it easily visible. H and E. ×400. Enlarged by 23.4

57 恶性疟病人脾脏 Spleen from a case of P. falciparum infection. The deposition of malarial pigment makes it almost black in colour.

58 恶性疟病人胎盘切片 Section of placenta form a case of P. falciparum infection. The maternal blood shows red cells containing parasites and pigment. The foetal blood shows red cells without any parasites. It is generally believed that congenital transmission of malaria does not occur and that malarial parasites are unable to pass the placental barrier H and E. ×400. Enlarged by 5.4.

59 Immunology Premunition

60 Diagnosis 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.

61 诊断 1 病原检查 血涂片姬氏或瑞氏染色法： 取外周血制成厚、薄血膜，经吉氏或瑞氏液染色后光镜检查malaria parasites.
采血时间： 恶性疟：发作时、查环状体，发作数小时后因晚期滋养体寄生虫的红细胞滞留下皮下脂肪及内脏微血管中，不易查见。 间日疟和三日疟：发作后10小时内，太久后原虫数量要下降。

62 实验诊断 Laboratory diagnosis
1．病原检查 （1）血膜染色镜检 最好一张玻片上同时制作厚、薄两种血膜，瑞氏染色、镜检。 Thin film Thick film Question: Which stages are there in the blood film of P.v. or P.f. ? （2）血沉棕黄层定量分析法（了解）

63 主要诊断--外周血涂片检查 厚血膜涂片：原虫变形，且红细胞已溶，鉴别有困难，但原虫集中，易发现
薄血膜涂片：原虫形态结构完整，清晰，可辩认原虫的种类和各发育阶段的形态特征，适用于临床诊断，但虫数较少易漏检。

64 2、免疫学诊断（了解） 循环抗体检测----间接荧光抗体实验、 酶 联免疫吸附实验 循环抗体检测----放射免疫实验、 夹心法酶联免疫吸附实验 3、分子生物学技术 PCR、PROBE

65 Treatment 1. Three principles: * Control of clinical symptoms
* Eradication of gametocytes * Prevent relapse

66 治疗 Treatment Chlorquine 、quinine、artemisinin and artemether----anti-erythrocytic stage drugs. (question: Which stage of plasmodium can these drugs kill?) Primaquine and pyrimethamine ----anti-exoerythrocytic stage drugs. primaquine(anti-exoerythrocytic & gametecyte)

67 2. Medicines - Quinine Chloroquine Primaquine +++ ++ 对抗氯喹株则宜用青蒿素类药物
Pyrimethamine Anti-ES +++ - Anti-EE ++ Anti-Gametocyte 对抗氯喹株则宜用青蒿素类药物

68 菊科植物黄花蒿 Artemisia annua

69 Prevention Chemoprophylaxis Malaria vaccines
-----Chloroquine / pyrimethamine used for prophylaxis of malaria -----Chemotherapy: 1 week before entry into the endemic area ; for 4 weeks after returning from the endemic area. Malaria vaccines

70 Mosquito control (1). Reconstruction of environment: eradicate the breeding places of moquitoes. (2). Spry insecticides: DDVP and so on. (3). Use mosquito nets (dipping in insecticide), screen, or mosquito repellents to protect the person from mosquito bites.

71 Epidemiology A global problem: Each year, million people become ill with malaria and several million died.

72 World Distribution of Malaria

73 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.

74 Ronald Ross Great Britain University College Liverpool, Great Britain – 1932 The Nobel Prize in Physiology or Medicine 1902 "for his work on malaria, by which he has shown how it enters the organism and thereby has laid the foundation for successful research on this disease and methods of combating it"                                                                    

75 Charles Louis Alphonse Laveran
                                                       France Institut Pasteur Paris, France The Nobel Prize in Physiology or Medicine 1907 "in recognition of his work on the role played by protozoa in causing diseases"

76 Julius Wagner-Jauregg
The Nobel Prize in Physiology or Medicine 1927 "for his discovery of the therapeutic value of malaria inoculation in the treatment of dementia paralytica” Austria Vienna University Vienna, Austria

77 小结 1.疟疾是由疟原虫寄生于人体肝细胞和红细胞而引起的一种严重寄生虫病。 2.能寄生在人体的疟原虫有4种，P.v 多见多发；P.f 严重，来势凶，脑型疟。 3.疟原虫需要人-蚊转换宿主后，才能完成生活史；蚊为按蚊属。 4.P.v可有复发，其机理是子孢子有速发型和迟发型，复发是迟发型子孢子所致；P.f无复发。 5.疟疾的典型发作症状是：冷、热、汗。 6.疟疾的临床症状主要是：贫血、肝脾肿大。 7.疟原虫的感染阶段为：红内期无性体（输血）或子孢子（蚊咬）。 8.疟原虫的致病阶段是：红内期无性体。 9.疟疾的诊断：主要是在外周血红细胞内查见疟原虫各期形态，亦可用血清 学和PCR检测协助。 10.P.v全国流行,P.f南方山区; P.f与P.f可混合感染;P.m与P.o则较少见。 11.输血可传播疟疾。