Invertebrate Anatomy OnLine

Opisthorchis sinensis ©

Chinese Liver Fluke


Copyright 2001 by

Richard Fox

Lander University


            This is one of many exercises available from Invertebrate Anatomy OnLine , an Internet laboratory manual for courses in Invertebrate Zoology.   Additional exercises can be accessed by clicking on the links on the left.   A glossary and chapters on supplies and laboratory techniques are also available.   Terminology and phylogeny used in these exercises correspond to usage in the Invertebrate Zoology textbook by Ruppert, Fox, and Barnes (2004).   Hyphenated figure callouts refer to figures in the textbook.   Callouts that are not hyphenated refer to figures embedded in the exercise. The glossary includes terms from this textbook as well as the laboratory exercises.  


Bilateria, Protostomia, Platyhelminthes P, Neodermata, Trematoda C, Digenea sC, Opisthorchiida O, Opisthorchiata sO, Opisthorchoidea SF, Opisthorchidae F, (Fig 10-48)

Platyhelminthes P

            Flatworms, or platyhelminths, are bilaterally symmetrical metazoans with three tissue layers.   Unlike most triploblastic animals, they are compact and have no coelom (body cavity) surrounding the viscera and no hemal system.   The gut, if present, has a single opening to the exterior.   An anterior brain with associated concentration of sense organs is present, as expected of bilaterians.   Flatworms are complex animals with elaborate hermaphroditic reproductive systems.   Fertilization is internal with copulation.   They may be free-living or parasitic.  


            Neodermata includes the parasitic flatworms, most of which are flukes or tapeworms.   Most parasitic flatworms are endoparasites with complex life cycles requiring multiple hosts including a definitive host inhabited by the adult worm and one or more intermediate hosts inhabited by juvenile stages of the worm.   The defining neoderm characteristic is the neodermis, or tegument.   Neodermis is an epidermis specialized for living in a potentially hostile environment from which it must absorb food but reject toxins. The neodermis is a syncytium with its cell nuclei submerged below the basal lamina (Fig 10-33*, 10-36).    

Trematoda C

            Trematoda consists mostly of flukes belonging to Digenea.   Flukes are important animal parasites and several important human and livestock diseases are caused by them. The small taxon Aspidogastrea also belongs to Trematoda.

Digenea s C

            Digeneans are compact, bilaterally symmetrical, endoparasitic flatworms known as flukes.   A blind gut with mouth and pharynx is present but there is no anus.   Osmoregulation is accomplished via protonephridia.   The integument is an elaborate syncytial neodermis without a cuticle.   There are no respiratory or hemal systems and hermaphroditism is the rule.   About 11,000 species are known making this the second largest taxon (after nematodes) of parasitic worms. Size ranges from less than 1 mm to 6 cm.

            At least two hosts are required to complete the life cycle and this is the basis for the name “ digenea”.  The definitive host is always a vertebrate and the intermediate host is usually a gastropod mollusc (Fig 10-34B). An additional intermediate host, if present, is an arthropod or fish.  In the definitive host the parasite typically inhabits either the hemal system or the gut and its derivatives.


Laboratory Specimens

            The anatomy of digenetic trematodes is usually studied in introductory laboratories using commercially prepared wholemount slides.   The Chinese liver fluke, Opisthorchis (= Clonorchis) sinensis, or the sheep liver fluke, Fasciola hepatica, are the most frequently used species.   Due to its convenient small size and transparency Opisthorchis is an especially good subject for an introductory study.   This exercise is based on commercially prepared wholemount slides of Opisthorchis.   The adult inhabits the bile ducts of the liver of any of several mammals including humans, cats, and dogs. It is an important human parasite in the orient.  

            The quality of staining varies in commercially prepared slides and greatly affects the visibility of structures.   Few slides are perfectly stained and there will undoubtedly be structures you cannot find on your slide.   It will help to use more than one slide.  


            Use the compound microscope to examine a stained wholemount of Opisthorchis (Fig 1, 10-34A*, 10-37).  

External Features

            Note the size and shape of the worm.   This species reaches 25-30 mm in length and is strongly flattened dorsoventrally.   The anterior end of the worm narrows gradually to a pointed tip.  The wider posterior end is bluntly rounded.    At the extreme anterior end a conspicuous oral sucker (Fig 1, 10-34A) surrounds the mouth and is used to attach to the host and maintain its position.   The circular ventral sucker, or acetabulum, is located on the ventral surface a short distance posterior to the oral sucker.   It also attaches to the host but is not associated with the gut.

Body Wall

            The body wall consists of an outer syncytial neodermis whose nuclei are sunken below the basal lamina (Fig 10-36).   The true epidermis is lost during larval development. There is no cuticle.   Muscles underlie the neodermis and a mesenchymal parenchyma fills the interior of the animal.   Organs are embedded in the parenchyma.   Trematodes, like other flatworms, are compact (acoelomate) and have neither mesodermal body cavity nor mesothelium.  

Digestive System

            The mouth opens into a muscular, sucking pharynx (Fig 1, 10-16C), which is used to pump food into the gut.   Opisthorchis feeds on blood, epithelium, mucus, and fluids from the wall of the host’s bile duct.   Immediately posterior to the pharynx is a short, inconspicuous esophagus. The esophagus quickly bifurcates to form two long, unbranched intestinal ceca that extend along the sides to the posterior end of the worm.   Hydrolysis and absorption occur in these ceca.   They end blindly and there is no anus.    

Excretory/Osmoregulatory System

            Protonephridia are scattered through the mesenchyme but they are not visible in these preparations.  They drain into a pair of branching, lateral excretory canals, which likewise are usually not visible.   The lateral canals drain into a large median excretory bladder at the posterior end of the worm. The bladder, which is easily observed (Fig 1, 10-34A, 10-37), opens to the exterior by a large nephridiopore at the posterior end of the worm.  

Respiratory System

            Opisthorchis inhabits an anoxic environment and is a facultative anaerobe with no special respiratory surface.

Fluid Transport System

            Flatworms have no hemal system.   The gut itself distributes food to the tissues and is often called the gastrovascular cavity in recognition of its dual roles in digestion and transport. Flukes also absorb nutriment directly across the cuticle-free syncytial neodermis.  

Nervous System

            The nervous system is usually not visible in preparations of this type.   The brain is a bilobed cerebral ganglion dorsal to the esophagus.   Three pairs of longitudinal nerve cords (dorsal, lateral, and ventral) exit the brain and are connected with each other by numerous transverse commissures.

Reproductive System

            Opisthorchis, like other platyhelminths, is hermaphroditic and has a complex reproductive system with (nearly) independent male and female systems sharing only the common gonopore.  The reproductive system is large and well-developed, occupying a large proportion of the interior, as is often the case with parasites.   Many of the ducts mentioned in the following descriptions are difficult to find but if it is in bold type, it should be visible in most preparations.

Figure 1.   Ventral view of an adult Chinese liver fluke, Opisthorchis sinensis.   Fluke26L.gif

Figure 1

Male System

            The male system begins with two large, irregularly branched testes located in the posterior third of the worm (Fig 1, 10-34A, 10-17).   The testes are large, easily seen, and usually stain dark pink.   One is the anterior testis and the other the posterior testis, situated as their names suggest. Each has a central area from which extend wide, blunt, branched lobes reminiscent of the pseudopodia of Amoeba.   A slender, inconspicuous duct, the vas efferens, which you probably will not see, arises near the center of each testis.  

            The two vasa efferentia unite near the middle of the body to form the short vas deferens which is obscured by the voluminous uterus in the center of the worm.   The vas deferens quickly widens to become the muscular seminal vesicle.   It usually stains pink in contrast with the brown uterus. This meandering tube extends anteriorly and eventually joins the uterus and the two open together to the exterior via the common gonopore.   In ventral view the seminal vesicle is hidden by the uterus and may be difficult to observe.   The seminal vesicle stores autosperm produced by the testes.  

            The common gonopore, or genital pore, is on the ventral surface immediately anterior to the ventral sucker.   It is small and obscure but usually appears as a pink spot on the anterior edge of the ventral sucker.   Opisthorchis, unlike most flukes, has no eversible copulatory cirrus (Fig 10-37) relying instead on the muscular seminal vesicle to transfer sperm.

Female System

            The female system is more complex, partly because trematodes produce ectolecithal eggs consisting of an oocyte surrounded by yolk cells.   Oocytes are produced by the ovary whereas yolk cells are produced by two independent vitellaria.   Hermaphroditism and internal fertilization also contribute to the complexity of the system.

            Begin with the easily located seminal receptacle, a large, ovoid organ where allosperm (from another worm) are stored (Fig 1, 10-34A, 10-37).   It is usually stained pink. The seminal receptacle is drained by the sperm duct which joins the oviduct.

            The single germarium (ovary) stains dark pink and lies on the midline immediately anterior to the seminal receptacle.   The germarium is smaller (and usually darker) than the seminal receptacle.   The ootype is a glandular chamber near the center of the worm but it usually cannot be distinguished in these preparations (Fig 10-37). The germarium connects with the ootype via the oviduct, neither of which is visible.  

            The two large vitellaria produce vitellocytes (= yolk cells).   They lie lateral to the intestinal ceca, between the ceca and the edge of the body.   They are large and are usually brownish-yellow in stained preparations.   Each is drained by a large, easily seen (usually) vitelline duct (= vitellarium duct).   The two ducts join each other to form a short common duct that delivers vitellocytes to the ootype.

            The large brown uterus exits the ootype and extends anteriorly to its union with the seminal vesicle at the common gonopore.   It is a wide convoluted tube packed with dark, yellow-brown embryonated eggs.   The uterus and seminal vesicle share the common gonopore.

Reproduction and Life Cycle

            Adults Opisthorchidae parasitize the branches of the bile duct of fish-eating mammals, including humans.   The life cycle requires two intermediate (snail and fish) and one definitive (mammal) host for completion.


            Sperm from the testes pass via the vasa efferentia and vas deferens to the seminal vesicle for storage.   During copulation sperm are forced by the muscular seminal vesicle from each worm into the uterus of the partner.   The sperm move up the uterus to the seminal receptacle where they are stored.  

            Later, oocytes leave the germarium and are fertilized by sperm from the seminal receptacle as they enter the ootype.   The fertilized oocytes, still in the ootype, are then enclosed in a layer of vitellocytes from the vitellaria.   Vacuoles in the yolk cells release liquid protein which hardens to form a tough eggshell.   Additional components of the eggshell may be secreted by the epithelium lining the ootype.   Shelled eggs are stored in the uterus where meiosis is completed, union of pronuclei accomplished, and development begins.   The so-called “eggs” in the uterus are actually shelled embryos, sometimes referred to as embryonated eggs to distinguish them from unicellular haploid ova.

Life Cycle

            Development in the uterus is rapid and by the time they are released from the common gonopore of the adult worm each "egg" contains a miracidium larva (Fig 10-34B, 10-35A). These embryonated “eggs” pass down the bile duct of the definitive host and into its gut. They exit the body of the host with its feces and some end up in freshwater habitats.  

           The Chinese liver fluke has three hosts, the last of which inhabits the bile ducts of humans and other mammals.   The first intermediate host is an aquatic snail, the second intermediate host is a fish, usually a cyprinid (e.g. carp), and the definitive host is a mammal (human, pig, dog, cat, rat, camel, and probably any other mammal that eats raw fish).  

            >a. If slides of fluke larval stages are available in the laboratory, examine them with 400X of the compound microscope.  All the larval stages have protonephridia but they are not usually visible.   Rediae, cercariae, and metacercariae have guts.

            The embryonated egg consists of a zygote, embryo, or larva enclosed in a capsule, or eggshell. A door, or operculum, is located at one end of the eggshell.

            The first intermediate host (snail) ingests an “egg” with its fully developed miracidium (Fig 10-35A).   The miracidium, now in the snail’s gut, emerges from the eggshell and migrates to the snail’s digestive cecum, which is a diverticulum of the gut.  In miracidium slides you should be able to see cilia on the surface and abundant germ cells inside.

            In the snail’s digestive cecum, the miracidium metamorphoses into a saclike sporocyst soon after ingestion (Fig 10-35B).   It lacks mouth or gut. Within the sporocyst numerous asexually produced germinal cells each develop into a multicellular but undifferentiated germ balls which then develop into larvae, known as rediae, which escape from the sporocyst when the sporocyst ruptures after about two weeks.

            The rediae (Fig 10-35C) remain in the snail.   Each has a gut consisting of mouth, muscular pharynx, and a short, unbranched intestine.   Depending on the quality of your slide, you may be able to see the oral sucker surrounding the mouth and the developing gut. Undifferentiated gem balls and immature cercariae can be seen with no difficulty.  The oldest cercariae have tails and should be recognizable.

            Within each of the hundreds of redia, more germ cells mature into another larval stage, the cercaria (Fig 10-35D,E).   This clonal reproduction, known as polyembryony, by sporocyst and redia greatly increases the number of larvae in the first intermediate host even though only one miracidium may have entered the host. The cercaria looks like a little fluke with a tail, oral sucker, ventral sucker, gut, and protonephridia but no reproductive system yet (Fig 10-35D,E).   The tail and suckers are easy to see.   Sometimes a bilobed gut with pharynx and ceca can be made out.  It may have eyes to help it locate a passing fish.   Most species have simple tails but in some it is forked.

            The cercariae are released into the water where they attach to the skin of a passing fish (the second intermediate host) and bore through the epidermis to encyst under a scale or in muscle as a metacercaria (Fig 10-35F).   The intermediate host is usually a member of the fish family Cyprinidae, which includes carp, an important food fish in the orient.   On a good slide you may be able to see the two suckers, pharynx, and bilobed intestine.  

            The definitive host is infected when it eats raw or poorly cooked fish with metacercariae in the muscles.   With the help of the host’s digestive enzymes young flukes excyst in the duodenum of the definitive host.   Although the route is uncertain, it is thought that young flukes simply migrate up the bile duct to the liver from the duodenum.  

            The worms mature in the liver, mate and produce the first eggs after about 3 months. The life cycle is best broken by preventing the contamination of water with human feces and by thoroughly cooking freshwater fishes before they are eaten.

*Hyphenated call-outs, such as this one, refer to figures in Ruppert, Fox, and Barnes (2004).   Those without hyphenation refer to figures embedded in this exercise.


            Brown FA . (ed)   1950.   Selected Invertebrate Types.   Wiley, New York.   597p.  

            Cameron TWM .   1944.   The morphology, taxonomy, and life history of Metorchis conjunctus (Cobb, 1860).   Can. J. Res., Sect. D., 22:6-16.

            Schmidt GO, Roberts LS .   1989.   Foundations of Parasitology, 4th ed.   Times Mirror/Mosby, New York.   750p.

Ruppert EE, Fox RS, Barnes RB.   2004. Invertebrate Zoology, A functional evolutionary approach, 7 th ed. Brooks Cole Thomson, Belmont CA. 963 pp.  


Compound microscope

Prepared wholemount of Opisthorchis



Opisthorchis sinensis wholemounts

Triarch, Carolina, Ward’s

Trematode egg, miracidium, redia, cercaria, and metacercaria slides


Rediae and cercariae

Carolina ,