MORPHOLOGY The parasite occurs in 3 stage: trophozoite, precyst and cyst Trophozoite(滋养体): it is the Invasive orm of the parasite and is present in the lumen and in the wall of the large intestine. The diameter of most trophozoites falls into the range of 20 to 30um, occasional specimens are small as 10um or as large as 60um. In the intestine and in freshly passed uniformed stools, the parasites actively crawl about, their short, blunt pseudopodia rapidly extending and with drawing. The clear ectoplasm is rather thin but is clearly differentiated from the granular endoplasm. The nucleus is difficult to discern in living specimens, but nuclear morphology may be distinguished after fixing and staining with iron- hematoxylin(铁苏木素) The nucleus is spherical and is about one sixth to one fifth the diameter of the cell. A karyosome is located in the center of the nucleus, and delicate achromatic fibrils radiate from it to the inner surface of the nuclear membrane. Chromatin is absent from a wide area surrounding the karyosome but is concentrated in granules or plaques on the inner surface of the nuclear membrane. This gives the appearance of a dark circle with a bulls-eye in the center. The nuclear membrane itself is quite thin(Figl) Food vacuoles(artiE) are common in the cytoplasm of active trophozoites and many contain host erythrocytes(aL IfI ER)in sample from diarrheic stools. Red blood cells may be ingested but do not often appear in chronic infections. The haematophagous trophozoites are the characteristic features of the invasive amoebae Cyst(E E): In a normal asymptomatic infection, the amoebae are carried out in formed stools. As the fecal matter passes posteriad and becomes dehygrated zk), the ameba is stimulated to encyst. Cysts are neither found in the stools of patients with dysentery nor formed by the ameba when they have invaded the tissues of the host. Trophozoite passed in stools are unable to encyst. At the onset of encystment(包囊形成) the trophozoite disgorges any undigested food it may contain and condenses into a sphere, called the precast(包囊前期). A precast is so rich in glycogen(BE RR)that a large glycogen vacuole may occupy most of the cytoplasm in the young yst. The chromatoid bars(拟染色体) that form typically are rounded and ends. The bars may be short and thick, thin and curved, spherical or very irregular in shape, but they do not have inter-like appearance found in E. coil B Fig I-I-I Trophozoites of Entamoeba histolytica(A: Line drawing; B Stain specimen
21 MORPHOLOGY The parasite occurs in 3 stage: trophozoite, precyst and cyst. Trophozoite(滋养体): it is the invasive form of the parasite and is present in the lumen and in the wall of the large intestine. The diameter of most trophozoites falls into the range of 20 to 30μm, occasional specimens are small as 10μm or as large as 60μm. In the intestine and in freshly passed, uniformed stools, the parasites actively crawl about, their short, blunt pseudopodia rapidly extending and with drawing. The clear ectoplasm is rather thin but is clearly differentiated from the granular endoplasm. The nucleus is difficult to discern in living specimens, but nuclear morphology may be distinguished after fixing and staining with iron-hematoxylin (铁苏木素). The nucleus is spherical and is about one sixth to one fifth the diameter of the cell. A karyosome is located in the center of the nucleus, and delicate, achromatic fibrils radiate from it to the inner surface of the nuclear membrane. Chromatin is absent from a wide area surrounding the karyosome but is concentrated in granules or plaques on the inner surface of the nuclear membrane. This gives the appearance of a dark circle with a bull’s-eye in the center. The nuclear membrane itself is quite thin (Fig1) Food vacuoles (食物泡) are common in the cytoplasm of active trophozoites and many contain host erythrocytes (红血球) in sample from diarrheic stools. Red blood cells may be ingested but do not often appear in chronic infections. The haematophagous trophozoites are the characteristic features of the invasive amoebae. Cyst（包囊）: In a normal asymptomatic infection, the amoebae are carried out in formed stools. As the fecal matter passes posteriad and becomes dehygrated (脱水), the ameba is stimulated to encyst. Cysts are neither found in the stools of patients with dysentery nor formed by the ameba when they have invaded the tissues of the host. Trophozoite passed in stools are unable to encyst. At the onset of encystment (包囊形成), the trophozoite disgorges any undigested food it may contain and condenses into a sphere, called the precyst (包囊前期). A precyst is so rich in glycogen (糖原) that a large glycogen vacuole may occupy most of the cytoplasm in the young cyst. The chromatoid bars (拟染色体) that form typically are rounded and ends. The bars may be short and thick, thin and curved, spherical or very irregular in shape, but they do not have splinter-like appearance found in E. coil. A B Fig Ⅱ－Ⅱ－1 Trophozoites of Entamoeba histolytica (A: Line drawing; B Stain specimen)
Trophozoite Cyst nested RBC chromatoidal body nudeus karyosome nucleus 10 The precyst rapidly secretes a thin, tough hyaline cyst wall (E*HE)around itself to form a yst. The cyst may be somewhat ovoid or elongate, but usually is spheroid. It is commonly 10 to 16 um wide but may be as small as 5um. The young cyst has only a single nucleus, but this rapidly divides twice to form two- and four-nucleus stages. As the nuclear division proceeds and the cyst matures, the glycogen vacuole and chromatoidal bodies(拟染色体) disappear. In semiformed ools one can find precysts and cysts with one to four nuclei, but quadrinucleate cysts (p4*E Rn)are most commnon in formed stools. The mature cysts can survive outside the host and can fected a new one LIFE CYCLE The life cycle of E. histolytica is simple and is completed in a single host, the man. Man is the main and probably the only natural host of E histolytica acquires infection by ingestion of water and food contaminated with mature quadrinucleate cysts. Man also can acquire the infection directly by ano-genital or oro-genital sexual contact. On ingestion, the cyst excysts(s t)in the small intestine. The cyst wall is lysed berating a single trophozoite wih four nuclei. The trophozoite quickly undergoes a series of cytoplasmic and nuclear div vision to form eight small metacystic trophozoites. These trophozoites are carried by peristalsis through the small intestine to the ileo-caecal area of the large intestine. Here they grow and multiply by binary fission. They then colonise on the mucosal surfaces and in the crypts of the large intestine. Various factors such as the intestinal motility, the transit time, the presence or absence of specific intestinal flora and the diet of the host influence the colonisation of the trophozoites In some individuals, the multiplying trophozoites produce no or little lesion if any in tissue They only feed on the starches and mucus secretions on the surface of mucosa. As trophozoites pass down the colon, they encyst under the stimulus of desiccation and the excreted as cysts with the stool In other individuals infected under similar conditions, the trophozoites may invade the tissue of the large intestine. The factors those lead to invasion of the intestinal tissue are poorly understood. Trophozoites produce characteristic lesions in the colon, through the stages of gelatinous necrosis, abscess and finally ulcer. A large number of trophozoites are excreted along with blood and mucus in the stool In a few cases, erosion of the large intestine may be so extensive that trophozoites gain
22 Fig Ⅱ－Ⅱ－2 Trophozoite and cyst of Entamoeba histolytica The precyst rapidly secretes a thin, tough hyaline cyst wall (包囊壁) around itself to form a cyst. The cyst may be somewhat ovoid or elongate, but usually is spheroid. It is commonly 10 to 16 μm wide but may be as small as 5μm. The young cyst has only a single nucleus, but this rapidly divides twice to form two- and four-nucleus stages. As the nuclear division proceeds and the cyst matures, the glycogen vacuole and chromatoidal bodies (拟染色体) disappear. In semiformed stools one can find precysts and cysts with one to four nuclei, but quadrinucleate cysts (四核包 囊) are most commnon in formed stools. The mature cysts can survive outside the host and can infected a new one. LIFE CYCLE The life cycle of E. histolytica is simple and is completed in a single host, the man. Man is the main and probably the only natural host of E. histolytica. Man acquires infection by ingestion of water and food contaminated with mature quadrinucleate cysts. Man also can acquire the infection directly by ano-genital or oro-genital sexual contact. On ingestion, the cyst excysts (脱囊) in the small intestine. The cyst wall is lysed by intestinal trypsin liberating a single trophozoite wih four nuclei. The trophozoite quickly undergoes a series of cytoplasmic and nuclear divisions to form eight small metacystic trophozoites. These trophozoites are carried by peristalsis through the small intestine to the ileo-caecal area of the large intestine. Here they grow and multiply by binary fission. They then colonise on the mucosal surfaces and in the crypts of the large intestine. Various factors such as the intestinal motility, the transit time, the presence or absence of specific intestinal flora and the diet of the host influence the colonisation of the trophozoites. In some individuals, the multiplying trophozoites produce no or little lesion if any in tissue. They only feed on the starches and mucus secretions on the surface of mucosa. As trophozoites pass down the colon, they encyst under the stimulus of desiccation and the excreted as cysts with the stool. In other individuals infected under similar conditions, the trophozoites may invade the tissue of the large intestine. The factors those lead to invasion of the intestinal tissue are poorly understood. Trophozoites produce characteristic lesions in the colon, through the stages of gelatinous necrosis, abscess and finally ulcer. A large number of trophozoites are excreted along with blood and mucus in the stool. In a few cases, erosion of the large intestine may be so extensive that trophozoites gain
entrance into the radicles of the portal vein and are carried away to the liver where they multiply Depending upon the complex interaction of various host and parasitic factors, the trophozoites produce suppurative amoebic liver abscess preceded by non-suppurative infection of the liver Mature Cysts Multiplication A-Non Invasise Colonization Infective Stage B Intestinal Disease s Diagnostic Stage Extra-Intestinal Disease Fig lI-l-3 Life cycle of E. histolytica, Adapted from parasite image library of CDC, USA The mature cysts(成熟包囊) excreted in the feces are the infective forms. They unlike the trophozoites which degenerate within minutes, may remain viable for weeks or months in suitable moist environment. Cysts of E. histolytica can remain viable and infective in a moist, cool environment for at least 12 days, and in water they can live up to 30 days. They are rapidly killed by putrefaction, desiccation, and temperatures below -5C and above 40C. They can withstand passage through the intestine of flies and cockroaches. The cysts are resist ant to levels of chlorine normally used for water purification These cysts cause infection in other susceptible through faecal contamination of water and vegetables or direct faecal-oral contact and the PATHOGENESISAND CLINICAL MENIFESATTIONS Pathogenic mechanisn: Amoebiasis(阿米巴病) is a disease caused by potentially pathogenic strains of E. histolytica. These pathogenic amoebae cause invasive amoebiasis through
23 entrance into the radicles of the portal vein and are carried away to the liver where they multiply. Depending upon the complex interaction of various host and parasitic factors, the trophozoites produce suppurative amoebic liver abscess preceded by non-suppurative infection of the liver. Fig Ⅱ－Ⅱ－3 Life cycle of E. histolytica, Adapted from parasite image library of CDC, USA The mature cysts (成熟包囊) excreted in the feces are the infective forms. They unlike the trophozoites which degenerate within minutes, may remain viable for weeks or months in suitable moist environment. Cysts of E. histolytica can remain viable and infective in a moist, cool environment for at least 12 days, and in water they can live up to 30 days. They are rapidly killed by putrefaction, desiccation, and temperatures below -5℃ and above 40℃. They can withstand passage through the intestine of flies and cockroaches. The cysts are resistant to levels of chlorine normally used for water purification. These cysts cause infection in other susceptible persons through faecal contamination of water and vegetables or direct faecal-oral contact and the cycle is repeated. PATHOGENESISAND CLINICAL MENIFESATTIONS Pathogenic mechanism: Amoebiasis ( 阿米巴病) is a disease caused by potentially pathogenic strains of E. histolytica. These pathogenic amoebae cause invasive amoebiasis through
the sequential stages of: a) Adherence of trophozoites on the surface of the large intestine. b) Invasion of the large intestine by the amoebae, and finally c) Resistance of the amoebae to variour effector mechanisms of the host Initially, the slow transmit of intestinal contents in the caecum and sigmoid colon helps the amoebae to invade these sites. the slow transit of intestinal contents allows amoebae to come in contact with the colonic mucosa for a longer time, thereby bringing a change in the intestine flora ( R)that may facilitate invasion. It has recently been demonstrated that the gut-associated bacterial flora affect invasiveness of the amoebae to a great extent Adherence of amoebae to the intestinal mucosa is mediated by a surface lectin (RR )of ne amoebae known as galactose( Gal) or N-acetyl-O-galactosamine(gal NAC) inhibitable surface lectin( the Gal/gal NaC lectin半乳糖/乙酰氨基半乳糖凝集素). After adherence, trophozoites kill target cells in the intestinal mucosa, only by direct contact and also by secreted cytotoxins(=l 胞毒素). The cytolysis occurs within20 minutes of the amoebic adherence E. histolytica also secretes numerous proteolytic enzymes(水解蛋白酶) that appear to be involved in various pathogenic processes. Cathepsin B proteinase(组蛋白酶) is respondible for dissolution of extra cellular matrix containing cells and tissue components. Amoebic glycosidases (糖苷酶) such as B- . glucosaminidase(氨基葡糖苷酶) and a surface membrane-associated neuraminidase(神经氨酸苷酶) cause degradation of mucos membrane of the colon or alteration of membrane glycoproteins on cell surfaces of the target cell Resistance of the parasite to variety of host effector mechanisms, both specific and non-specific, contributes to the persistence of infection in the intestine Host immunity in amoebiasis may be non-immune defence mechanism, and specific immunity. Non-immune defence mechanisms play an important role in resistance against invasion amoeba. Gastric acid barrier(胃酸屏障) kills amoebic trophozoites, and rapid intestinal transit reduces the time for amoebae to colonise on the intestinal mucosa. also colonic mucin inhibits amoebic adherence to epithelial cells The specific immunity involves both humoral and cell mediated immunity. Humoral Immunity(体液免疫) appears to be responsible for elimination of the amoebae from the intestine and subsequent resistance to re-infection. Cell mediated immunity probably has a role in limiting invasive amoeba and resisting a recurrence after therapeutic cure. Host resistance to initial amoebic invasion of the intestinal mucosa does not appear to involve cell-mediated mechanisms as evidenced from the lack of severity of the amoebic disease in AlDs cases Pathogenesis changes E. histolytica is almost unique among the amoebae of humans in its ability to hydrolyse host tissue. Once in contact with the mucosa, the amoebae secrete proteolytic enzymes(蛋白水解酶) which enable them to pentrate the epithelium and begin moving deeper. The intestinal lesion usually develops initially in the cecum, appendix, or upper colon and then spreads the length of the colon. The number of parasites builds up in the ulcer, increasing the speed of mucosal destruction. The muscularis mucosae(粘膜肌层) is somewhat of a barrier to further progress,and the pockets of amoebae form, communicating with the lumen of the intestine through a slender duct like ulcer. The lesion may stop at the basement membrane or at the musculars mucosae and then begin eroding laterally, causing broad, shallow areas of necrosis(3E). The tissues may heal nearly as fast as they destroyed, or the entire mucosa may become pocked. These early lesions usually are not complicated by bacteria invasion, and there is little cellular response by the host. In
24 the sequential stages of: a) Adherence of trophozoites on the surface of the large intestine. b) Invasion of the large intestine by the amoebae, and finally c) Resistance of the amoebae to variour effector mechanisms of the host. Initially, the slow transmit of intestinal contents in the caecum and sigmoid colon helps the amoebae to invade these sites. The slow transit of intestinal contents allows amoebae to come in contact with the colonic mucosa for a longer time, thereby bringing a change in the intestine flora (区系) that may facilitate invasion. It has recently been demonstrated that the gut-associated bacterial flora affect invasiveness of the amoebae to a great extent. Adherence of amoebae to the intestinal mucosa is mediated by a surface lectin (凝集素)of the amoebae known as galactose(Gal) or N-acetyl-O-galactosamine(gal NAC) inhibitable surface lectin (the Gal/gal NAC lectin 半乳糖/乙酰氨基半乳糖凝集素). After adherence, trophozoites kill target cells in the intestinal mucosa, only by direct contact and also by secreted cytotoxins (细 胞毒素). The cytolysis occurs within 20 minutes of the amoebic adherence. E. histolytica also secretes numerous proteolytic enzymes (水解蛋白酶) that appear to be involved in various pathogenic processes. Cathepsin B proteinase (组蛋白酶) is respondible for dissolution of extra cellular matrix containing cells and tissue components. Amoebic glycosidases (糖苷酶) such as β-glucosaminidase (氨基葡糖苷酶) and a surface membrane-associated neuraminidase (神经氨酸苷酶) cause degradation of mucos membrane of the colon or alteration of membrane glycoproteins on cell surfaces of the target cell. Resistance of the parasite to variety of host effector mechanisms, both specific and non-specific, contributes to the persistence of infection in the intestine. Host immunity in amoebiasis may be non-immune defence mechanism, and specific immunity. Non-immune defence mechanisms play an important role in resistance against invasion amoeba. Gastric acid barrier (胃酸屏障) kills amoebic trophozoites, and rapid intestinal transit reduces the time for amoebae to colonise on the intestinal mucosa. Also colonic mucin inhibits amoebic adherence to epithelial cells. The specific immunity involves both humoral and cell mediated immunity. Humoral immunity (体液免疫) appears to be responsible for elimination of the amoebae from the intestine and subsequent resistance to re-infection. Cell mediated immunity probably has a role in limiting invasive amoeba and resisting a recurrence after therapeutic cure. Host resistance to initial amoebic invasion of the intestinal mucosa does not appear to involve cell-mediated mechanisms as evidenced from the lack of severity of the amoebic disease in AIDs cases. Pathogenesis changes: E. histolytica is almost unique among the amoebae of humans in its ability to hydrolyse host tissue. Once in contact with the mucosa, the amoebae secrete proteolytic enzymes (蛋白水解酶), which enable them to pentrate the epithelium and begin moving deeper. The intestinal lesion usually develops initially in the cecum, appendix, or upper colon and then spreads the length of the colon. The number of parasites builds up in the ulcer, increasing the speed of mucosal destruction. The muscularis mucosae (粘膜肌层) is somewhat of a barrier to further progress, and the pockets of amoebae form, communicating with the lumen of the intestine through a slender, duct like ulcer. The lesion may stop at the basement membrane or at the musculars mucosae and then begin eroding laterally, causing broad, shallow areas of necrosis (坏死). The tissues may heal nearly as fast as they destroyed, or the entire mucosa may become pocked. These early lesions usually are not complicated by bacteria invasion, and there is little cellular response by the host. In
older lesions usually the amoebae, assisted by bacteria, may break through the muscularis mucosae, infiltrate the submucosa(粘膜下层), and even penetrate the muscle layers and serosa. This enables trophozoites to be carried by blood and lymph to ectopic sites throughout the body where secondary lesions then form. A high percentage of deaths result from perforated colons with concomitant peritonitis (HSL3). Surgical repair of perforation is difficult because a heavily ulcerated colon becomes very delicate Sometimes a granulomatous mass called ameboma(阿米巴肿, forms in the wall of the intestine and may obstruct the bowel. It result of cellular responses to a chronic ulcer and often still contains active trophozoites. The condition is rare Symptoms The symptoms of amebiasis are far from clear cut and depend in large measure on the extent of tissue invasion and on whether the infection is confined to the intestinal tract or has spread to involve other organs. According WHO Report on Amebiasis, the symptoms of amoebiasis involve I Asymptomatic infections II Symptomatic infections A Intestinal amebiasis: (1) Dysenteric; (2)Nondysenteric clitis B. Extraintestinal amebiasis. (1)Hepatic: a Acute nonsuppurative; b liver abscess (2)Pulmonary (3). Other extraintestinal foci(very rare) E. histolytica infection in man is variable. In endemic area, nearly of individuals harbouring E. histolytica in their intestinal tract are asymptomatic cyst passers, while remainders have invasive intestinal amoebiasis or extra-intestinal invasive amoebiasis such as amoebic liver abscess Incubation period is usually long and often indefinite. It may be short in rare instances 1) Intestinal amoebiasis(肠阿米巴病): Intestinal amebiasis is the most common form of nfection and may be asymptomatic(无症状的, Certain patients with intestinal amebiasis have vague and nonspecific abdominal symptoms. Acute amoebiasis patients have more definite symptoms, such as diarrhea or dysentery(痢疾,腹泻) abdominal pain and cramping, flatulence (肠胃气涨), anorexIa(食欲减退), weight loss, and chronic fatigue. This term should be reserved or those who actually have dysentery Asymptomatic cyst passers: It is the common clinical form of intestinal amoebiasis Gastrointestinal manifestations are not specific. It consists of colicky lower abdominal pain and increased frequency of bowel movements with loose watery diarrhoea. The condition is intermittent and chronic in nature @Symptomatic intestinal amoebiosis Non-dysenteric colitis: It is a well recognised condition It is usually chronic Symptomatology consists of intermittent diarrhoea, abdominal pain, flatulence and loss of weight These cases have less colonic inflammation and small amoebic ulcers amoebae in their stool and associated antibodies in the serum he patient. They respond well to treatment with anti-amoebic drus Acute amoebic dysentery: It is the common form of invasive intestinal amoebiasis. The condition is characterised by the presence of blood and mucus in the stool, accompanied by abdominal pain, tenderness, tender hepatomegaly(肝脾肿大) and rectal tenesmus. Fever is
25 older lesions usually the amoebae, assisted by bacteria, may break through the muscularis mucosae, infiltrate the submucosa (粘膜下层), and even penetrate the muscle layers and serosa. This enables trophozoites to be carried by blood and lymph to ectopic sites throughout the body where secondary lesions then form. A high percentage of deaths result from perforated colons with concomitant peritonitis (腹膜炎). Surgical repair of perforation is difficult because a heavily ulcerated colon becomes very delicate. Sometimes a granulomatous mass called ameboma(阿米巴肿)，forms in the wall of the intestine and may obstruct the bowel. It result of cellular responses to a chronic ulcer and often still contains active trophozoites. The condition is rare. Symptoms The symptoms of amebiasis are far from clear cut and depend in large measure on the extent of tissue invasion and on whether the infection is confined to the intestinal tract or has spread to involve other organs. According WHO Report on Amebiasis, the symptoms of amoebiasis involve: ⅠAsymptomatic infections ⅡSymptomatic infections A. Intestinal amebiasis: (1) Dysenteric; (2) Nondysenteric clitis B. Extraintestinal amebiasis: (1) Hepatic: a. Acute nonsuppurative; b.liver abscess (2) Pulmonary (3).Other extraintestinal foci (very rare) E. histolytica infection in man is variable. In endemic area, nearly 90% of individuals harbouring E. histolytica in their intestinal tract are asymptomatic cyst passers, while remainders have invasive intestinal amoebiasis or extra-intestinal invasive amoebiasis such as amoebic liver abscess. Incubation period is usually long and often indefinite. It may be short in rare instances. 1) Intestinal amoebiasis(肠阿米巴病): Intestinal amebiasis is the most common form of infection and may be asymptomatic (无症状的). Certain patients with intestinal amebiasis have vague and nonspecific abdominal symptoms. Acute amoebiasis patients have more definite symptoms, such as diarrhea or dysentery (痢疾，腹泻), abdominal pain and cramping, flatulence (肠胃气涨), anorexia (食欲减退), weight loss, and chronic fatigue. This term should be reserved for those who actually have dysentery. ①Asymptomatic cyst passers: It is the common clinical form of intestinal amoebiasis. Gastrointestinal manifestations are not specific. It consists of colicky lower abdominal pain and increased frequency of bowel movements with loose watery diarrhoea. The condition is intermittent and chronic in nature. ②Symptomatic intestinal amoebiosis Non-dysenteric colitis: It is a well recognised condition. It is usually chronic. Symptomatology consists of intermittent diarrhoea, abdominal pain, flatulence and loss of weight. These cases have less colonic inflammation and small amoebic ulcers, amoebae in their stool and associated antibodies in the serum of the patient. They respond well to treatment with anti-amoebic drugs. Acute amoebic dysentery: It is the common form of invasive intestinal amoebiasis. The condition is characterised by the presence of blood and mucus in the stool, accompanied by abdominal pain, tenderness, tender hepatomegaly (肝脾肿大) and rectal tenesmus. Fever is