Press a small piece of freshwater fish flesh between two slides, examine for encysted metacercaria of C. sinensis under lower-lens microscope 3. Isolation of encysted metacercaria Take the 2nd intermediate host-freshwater fish and smash, digest with 250 ml digestive solution per 10 g flesh for 4 h"12 h(Digestive solution: pepsin 9.8g, I N HCI 164 mL NaCl 17 g add water to 2 000 ml). Discard rough matter by sieve filtration. After several times sedimentation of the filtrated solution, take the sediment and examine under microscop 4. Examination adult from infected animal Dissect the guinea pig(or white rat) which were infected with the aid of stomach tube approximately 30 C. sinensis cysts 40 days ago. Expose the common bile duct, gal bladder and search for the adult worms in the billiary tree
5 Press a small piece of freshwater fish flesh between two slides, examine for encysted metacercaria of C. sinensis under lower-lens microscope. 3. Isolation of encysted metacercaria Take the 2nd intermediate host-freshwater fish and smash, digest with 250 ml digestive solution per 10 g flesh for 4 h～12 h (Digestive solution: pepsin 9.8 g, 1 N HCl 164 ml, NaCl 17 g, add water to 2 000 ml). Discard rough matter by sieve filtration. After several times sedimentation of the filtrated solution, take the sediment and examine under microscope. 4. Examination adult from infected animal Dissect the guinea pig (or white rat) which were infected with the aid of stomach tube approximately 30 C. sinensis cysts 40 days ago. Expose the common bile duct, gall bladder and search for the adult worms in the billiary tree
PARAGONIMUS FLUKES Objectives and requirements 1. To study the life cycles of P. westermani, to learn more about the life history of trematodes 2. To study the main parasitized sites and major pathogenisis 3. To study the laboratory diagnosis of these flukes Observation and Experiment 1.P A. See the preserved specimen(Demonstration) B. Study the internal structures of stained specimen(Manipulation) Note the position of the suckers. Are they equal in size? The branched testes situated in posterior of the body, side by side, and posterior to uterus and ovary lobulated ovary is in the opposite side of uterus(fully filled with ova). Are these two female reproductive organs situated anterior or posterior to the worms? C. See the SEM photographs of Paragonimus spines (2)Larva and intermediate hosts(Demonstration) A. See the first and second intermediate hosts- Melania snail and crayfish. B. See the stained cercaria C. See the live cysts D Isolation of the encysted metacercaria from infected crabs(Students work in groups) Crushed a crab in a mortar. Add some 0.45% Nacl solution or tap water into motar and filter. Allow the filtrate stand and settle, examine sediments for encysted metacercaria under microscop (3)Egg A Morphology of egg(Manipulation) Measures 80118 W mX48-60 u m. Note its ellipsoidal shape, uneven thickness of eggshell, slightly oblique operculum and yellowish brown in color. A germinal cell with more than ten yolk cells inside the egg Materials examined; sputum, feces Examining methods: direct sputum smear, sputum concentration and direct fecal smear B. See the SEM photograph of egg (4)Pathology A. See the preserved infected lungs(Demonstration)
6 PARAGONIMUS FLUKES Objectives and Requirements 1. To study the life cycles of P. westermani, to learn more about the life history of trematodes. 2. To study the main parasitized sites and major pathogenisis. 3. To study the laboratory diagnosis of these flukes. Observation and Experiment 1. P. westermani (1) Adult worm A. See the preserved specimen (Demonstration). B. Study the internal structures of stained specimen (Manipulation). Note the position of the suckers. Are they equal in size? The branched testes are situated in posterior of the body, side by side, and posterior to uterus and ovary. The lobulated ovary is in the opposite side of uterus (fully filled with ova). Are these two female reproductive organs situated anterior or posterior to the worms? C. See the SEM photographs of Paragonimus spines. (2) Larva and intermediate hosts (Demonstration) A. See the first and second intermediate hosts-Melania snail and crayfish. B. See the stained cercaria. C. See the live cysts. D. Isolation of the encysted metacercaria from infected crabs (Students work in groups). Crushed a crab in a mortar. Add some 0.45% NaCl solution or tap water into motar and filter. Allow the filtrate stand and settle, examine sediments for encysted metacercaria under microscope. (3) Egg A. Morphology of egg (Manipulation). Measures 80～118μm×48～60μm. Note its ellipsoidal shape, uneven thickness of eggshell, slightly oblique operculum and yellowish brown in color. A germinal cell with more than ten yolk cells inside the egg. Materials examined: sputum, feces. Examining methods: direct sputum smear, sputum concentration and direct fecal smear. B. See the SEM photograph of egg. (4) Pathology A. See the preserved infected lungs (Demonstration)
Note the capsules and their surrounding damaged tissues B. See the infected liver( Demonstration) 2. Pagumogonimus skrjabini (1)Adult worm(Demonstration) A. See the preserved specimen B. See the stained specimen (2)Intermediate host(Demonstration) ee the first intermediate host-snail (Tricula humida) Exercise 1. Draw P. westermani egg in detail and label 2. Label the adult worm of p westermani 3. Write the result report of cyst isolation, to have an analysis of epidemiology of P. 4. Draw a life cycle draft of P westermani Thinking 1. How to explain paragonimiasis is a natural epidemic source disease in mountainous egion and forest? and what are the epidemic characteristics of paragonimiasis? R eference 1. Detection of sputum for egg of P westermani Take a clean slide, place a drop of saline in the center of it, pick up a small amount of the sputum and mix it with the drop of saline, examine under microscope (2)Egg concentration Collect the sputum of 24 h, pour into a measuring glass and add equal quantity of 10% NaoH, mix, sediment naturally for 6 h8 h, remove supernatant, take the sediment and examine under microscop 2. Survey biological hosts of P westerman in epidemiology Choose one or more brooks, capture Melania snail and stream crab or crayfish (1)Break up Melania snail and place the internal organs on a slide, open them and put a drop of 0.45% saline, find sporocyst, redia and cercaria under microscop (2)Dissect stream crab or crayfish and find encysted metacercaria 3. Infection of p westermani for dog Isolation of cysts from crayfish and mixing with meat, feed dogs, each one with 100-200 cysts according to the sizes of dogs. Kill the infected dogs 2-3 months later, open chest and expose the lungs, on which the nodular cysts can be observed
7 Note the capsules and their surrounding damaged tissues. B. See the infected liver (Demonstration). 2. Pagumogonimus skrjabini (1) Adult worm (Demonstration). A. See the preserved specimen. B. See the stained specimen. (2) Intermediate host (Demonstration) See the first intermediate host-snail (Tricula humida). Exercise 1. Draw P. westermani egg in detail and label. 2. Label the adult worm of P. westermani. 3. Write the result report of cyst isolation, to have an analysis of epidemiology of P. westermani. 4. Draw a life cycle draft of P. westermani. Thinking 1. How to explain paragonimiasis is a natural epidemic source disease in mountainous region and forest? And what are the epidemic characteristics of paragonimiasis? Reference 1. Detection of sputum for egg of P. westermani (1) Sputum smear Take a clean slide, place a drop of saline in the center of it, pick up a small amount of the sputum and mix it with the drop of saline, examine under microscope. (2) Egg concentration Collect the sputum of 24 h, pour into a measuring glass and add equal quantity of 10% NaOH, mix, sediment naturally for 6 h～8 h, remove supernatant, take the sediment and examine under microscope. 2. Survey biological hosts of P. westerman in epidemiology Choose one or more brooks, capture Melania snail and stream crab or crayfish . (1) Break up Melania snail and place the internal organs on a slide, open them and put a drop of 0.45% saline, find sporocyst, redia and cercaria under microscope. (2) Dissect stream crab or crayfish and find encysted metacercaria. 3. Infection of P. Westermani for dog Isolation of cysts from crayfish and mixing with meat, feed dogs, each one with 100～200 cysts according to the sizes of dogs. Kill the infected dogs 2～3 months later, open chest and expose the lungs, on which the nodular cysts can be observed
dissect them to find adult worms FASCIOLOPSIS BUSKI Objectives and requirements 1. To study the life history and feature in morphology of F buski 2. To study the laboratory diagnosis of this fluke 3. To gain some knowledge of plant vectors and the importance in prophylax Observation and experiment 1. Adult worm (1)See the preserved specimens(Demonstration) Note the shape, size, color and the positions of oral and ventral suckers and their sizes. large and thick (1)Study the stained specimen(Manipulation First with naked eye, then under low power microscope, note the adhesive organ, ceca and reproductive system of both sexes. The intestine is composed of two blind tracts without side branching, and the ventral sucker is very large 2. Larva, intermediate host and vector(demonstration) (1)See the snail intermediate host(Planorbis (2)See the cercaria (3)See the stained specimen of cyst (4)See the vector: aquatic plants(caltrop, water chestnut etc.) 3. Ovum (1)See the preserved specimens(Manipulation) Note its large size, 130-140 u mX80-85 H m, oval shape, pale yellow in color, thin shell, small operculum, an germinal cell surrounded by yolk granules Materials examined: feces (2)See the SEM photograph of ovum Exercise Draw and label F. buski egg in detail. Thinking 1. Draw a life cycle draft of f. buski
8 dissect them to find adult worms. FASCIOLOPSIS BUSKI Objectives and Requirements 1. To study the life history and feature in morphology of F. buski. 2. To study the laboratory diagnosis of this fluke. 3. To gain some knowledge of plant vectors and the importance in prophylaxis. Observation and Experiment 1. Adult worm (1) See the preserved specimens (Demonstration). Note the shape, size, color and the positions of oral and ventral suckers and their sizes, large and thick. (1) Study the stained specimen (Manipulation). First with naked eye, then under low power microscope, note the adhesive organ, ceca and reproductive system of both sexes. The intestine is composed of two blind tracts without side branching, and the ventral sucker is very large. 2. Larva, intermediate host and vector (Demonstration) (1) See the snail intermediate host (Planorbis). (2) See the cercaria. (3) See the stained specimen of cyst. (4) See the vector: aquatic plants (caltrop, water chestnut etc.). 3. Ovum (1) See the preserved specimens (Manipulation). Note its large size, 130～140μm×80～85μm, oval shape, pale yellow in color, thin shell, small operculum, an germinal cell surrounded by yolk granules. Materials examined: feces. (2) See the SEM photograph of ovum. Exercise 1.Draw and label F. buski egg in detail. Thinking 1.Draw a life cycle draft of F. buski
SCHIASTOSOMA JAPONICUM Objectives and Requirements 1. To understand characteristics of life cycle and morphology of its developmental 2. To observe the pathological changes of schistosomiasis in infected animal 3. To study morphology of ova, laboratory diagnostic methods and their princi 4. To gain fundamental knowledge of infective stage, infection mode, methods and significance of immunologic tests commonly used Observation and Experiment L Adult worm (1)See the live specimen(Demonstration) (1)See the preserved specimen(Demonstration) Adults are dioecious. a male and a female live together (3)Study stained specimen(Manipulation Note the shape, size and color of both sexes, oral and ventral suckers and gynecophoral canal of male, the number, shape, size and arrangement of testes, ovary and uterus of female. Females are long and slender, the posterior half of the body is a little thick and dark brown in color owing to the vitelline gland and intestine. Near the anterior end of the body, there are two suckers. The part of the male body behind the ventral sucker is flattened, and rolled ventrally along the length to form a groove, canal is gynecophonus, into which the female worm fits What is the meaning of dioecious? (2)See the seM photograph of adult worm( Demonstration) 2. Larva and intermediate host(Demonstration) (1)See the live miracidia Note the feature of movement in water (2)Study the structure of stained specimen of miracidia. (3)See the SEM photograph of miracidia (4)Identify the intermediate host Oncomelania snail (5)See the live cercariae Note the posture of the body resting beneath water surface and its movement Study stained specimen, note the internal structures and the forked tail (6)See the SEM photograph of cercariae 3. ovu (1) Place a small drop n containing S. japonicum ova on a slide
9 SCHISTOSOMA JAPONICUM Objectives and Requirements 1. To understand characteristics of life cycle and morphology of its developmental stages. 2. To observe the pathological changes of schistosomiasis in infected animal. 3. To study morphology of ova, laboratory diagnostic methods and their principle. 4. To gain fundamental knowledge of infective stage, infection mode, methods and significance of immunologic tests commonly used. Observation and Experiment l. Adult worm (1) See the live specimen (Demonstration). (1) See the preserved specimen (Demonstration). Adults are dioecious, a male and a female live together. (3) Study stained specimen (Manipulation). Note the shape, size and color of both sexes, oral and ventral suckers and gynecophoral canal of male, the number, shape, size and arrangement of testes, ovary and uterus of female. Females are long and slender, the posterior half of the body is a little thick and dark brown in color owing to the vitelline gland and intestine. Near the anterior end of the body, there are two suckers. The part of the male body behind the ventral sucker is flattened, and rolled ventrally along the length to form a groove, canalis gynecophorus, into which the female worm fits. What is the meaning of dioecious? (2) See the SEM photograph of adult worm (Demonstration). 2. Larva and intermediate host (Demonstration) (1) See the live miracidia. Note the feature of movement in water. (2) Study the structure of stained specimen of miracidia. (3) See the SEM photograph of miracidia. (4) Identify the intermediate host Oncomelania snail. (5) See the live cercariae. Note the posture of the body resting beneath water surface and its movement. Study stained specimen, note the internal structures and the forked tail. (6) See the SEM photograph of cercariae. 3. Ovum (1) Place a small drop of suspension containing S. japonicum ova on a slide