Lab 6 - PHYLUM NEMATODA Nematodes are almost unbelievably abundant. Not only are there more than 15,000 known species of roundworms, but there are many thousands of individual nematodes in even a single handful of garden soil. Some species of roundworm may contain more than 27 million eggs at one time and lay more than 200,000 of them in a single day. Some scientists have estimated that there may be as many as half a million unknown species of roundworm yet to be discovered; an estimate based on the fact that many new species are still being discovered, that relatively few people are looking for more species, and that most roundworms look pretty much alike. If the estimated number of species is anywhere close to correct, it would mean that roundworms are the second most diverse group of animals, trailing behind only the arthropods. General Characteristics Nematodes are bilaterally symmetrical, worm-like organisms that are surrounded by a strong, flexible noncellular layer called a cuticle. The body plan is simple. The body of a nematode is long and narrow, resembling a tiny thread in many cases, and this is the origin of the group's name. The word "nematode" comes from a Greek word “nema” meaning "thread". The epidermis (skin) of a nematode is highly unusual; it is not composed of cells like other animals, but instead is a mass of cellular material and nuclei without separate membranes. This epidermis secretes a thick outer cuticle, which is both tough and flexible. Most are dioecious and show considerable sexual dimorphism, with the female usually larger and the tail of the male being more curled. In females, the reproductive tract opens separately from the digestive tract, while in males the digestive tract joins the reproductive tract forming a cloaca. Nematodes vary in size from less than 1 mm to over 1 meter in length. Juvenile nematodes require several molts before the adult stage is reached, and development may be either direct or may require an intermediate host. 2 Development All nematodes pass through 5 developmental stages separated by 4 molts. Pre-adult stages are referred to as larval or juvenile stages and differ from the adult stage in both size and in the extent of development of the reproductive system. M1 M2 M3 M4 Egg J1 J2 J3 J4 Adult Nematode Morphology Digestive System: The nematodes have a complete digestive tract. Find the mouth opening at the blunt end of the worm. The mouth opens into the characteristic pharynx. In most cases the pharynx is highly muscular -- it often has a terminal bulb where it joins the intestine. A nerve ring encircles a constriction of the pharynx, but it is seldom visible. The pharynx is continuous with the thin-walled intestine (which lacks any musculature). The digestive system terminates near the end of the worm at the anus. (In male nematodes the reproductive and digestive system join together forming a cloaca, which terminates at the anus.) Reproduction: Most female nematodes have a complex, double set of reproductive organs. In nematodes the reproductive system is in the form of a continuous tube, which is usually subdivided into ovary, oviduct, and uterus. The ovaries are separated from the double uteri by an oviduct in which fertilization typically occurs. Uteri flow into a common vagina that serves as a canal for expulsion of eggs and for movement of sperm upwards towards the oviducts. The vagina opens to the outside at the genital pore (also called the vulva). The uterus can be distinguished as that part of the tract in which young worms (larvae) can be seen moving about. Most female nematodes lay eggs, but in some like the vinegar eel, the trichina worm, and filarial worms, the eggs hatch before they leave the uterus. The adult males are distinctly smaller than the females. This is the case with most nematodes. They can be differentiated from females by their smaller size, by the presence of chitinous spicules, the accessory organ of copulation, near the cloaca, and by the lack of a genital pore (other than that of the anus). The spicule serves to guide sperm into the female tract during copulation. Male nematodes commonly have a single reproductive tract. The vas deferens (sperm duct) joins the digestive tract to form the cloaca, which leads to the anus. An enlarged portion of the sperm duct, the seminal vesicle, is often present anteriorly. The vas deferens is continuous anteriorly with the testis -- it is often difficult or impossible to differentiate these portions of the system. We have included a diagram of general nematode morphology in this hand out. Use this diagram to become familiar with the nematode. 3 Schematic of Nematode Female Schematic of Nematode male Schematic of Nematode Male 4 CLASS ENOPLEA: Order Trichurida Nematodes in this Order are characterized by: • A body that is filiform anteriorly. • A mouth without lips. • An extremely long, thin, nonmuscular pharynx, called a stichosome. Within the stichosome is a row (or rows) of large cells, called stichocytes. • A single reproductive system in females, and the majority of species are oviparous. The vulva is commonly located near the base of the pharynx. • Males have a single spicule or none at all. • The life cycle is usually direct, with no intermediate host required. • Parasites of nearly all organs of all classes of vertebrates. You are responsible for two nematodes in the Order Trichurida; Trichuris trichiura and Trichinella spiralis. Trichuris tricuhura, the human whipworm, is a human parasite found throughout the world, but most prevalent in the tropics. The incidence of infection may reach 25% in parts of the southeastern United States. Typically, infections involve fewer than 100 worms and are relatively symptomless. However, in more intense infections, the mortality rate can be 1 in 1000. The worms are relatively large (30-50mm), with an extremely narrow anterior end and broader posterior ends, hence the name “whipworm”. These parasites occur in the large intestine where they embed their anterior ends beneath the mucosal surface and feed on cells in the lamina propria. The protruding posterior ends of the worms find each other for mating and females deposit eggs into the intestinal lumen so they pass out with the feces. Slide: Adult Trichuris trichiura, male and female (w.m.) Study the adult worm: • Distinguish the long filiform anterior end from the fusiform posterior. • Note the single row of large cells (stichocytes) surrounding the long, thin pharynx. The entire structure is referred to as the stichosome. Stichocytes secrete material that aid in digestion and that modulate the host reaction to the parasite. • The pharynx occupies about two- thirds of the body length. • The anterior end of the pharynx lacks stichocytes. • The anus is located near the tip of the tail. 5 In the male: • Note the coiled tail • The single spicule surrounded by a spiny spicule sheath. • The testis is singular, long, and convoluted, and gives the appearance of square- shaped compartments along its length. • Follow the testis forward from the region of the cloaca to a point near the termination of the stichosome. The tube now turns back on itself as a large uncoiled vas deferens. • Posterior, the vas deferens narrows, then widens again to form an ejaculatory duct that joins the intestine to form the cloaca. In the female • Note the bluntly rounded posterior end. • The vulva is located near the anterior end of the fusiform body region, near the junction of the pharynx and intestine. • A coiled vagina runs to the posterior from the vulva to its junction with a wider uterus. • The uterus runs to a coiled oviduct and sacculate ovary. • Eggs within the uterus are unembryonated, and have a characteristic barrel shape with a plug at each end. Slide: Trichuris eggs: Trichuris eggs are easily recognized by their prominent bipolar plugs and large size. Females produce 3000 to 20,000 eggs per day. Eggs embryonate within three weeks after leaving the host’s body and can remain infective for months if they are deposited in moist soil in the shade. Infection is acquired when a suitable host ingests embryonated eggs. Adults live for several years, so large numbers may accumulate in a person, even where the rate of new infection is low. 6 Trichinella spiralis, the trichina worm, is one of the most studied of all nematodes. It is the smallest nematode parasitic in humans, has one of the most unique life cycles, is one of the most widespread, and is one of the most medically important parasites in the world. Adult worms lie buried in the mucosa of the small intestine. Males die shortly after copulation. Females are viviparous, giving birth to living young in the tissues of the intestine. Juvenile nematodes are transported via the lymph or blood to all parts of the body. Further development only occurs in striated muscle, especially those muscles that are active. They penetrate individual muscle fibers, absorb nutrients from the muscle cell, and increase in length to about 1.0 mm in eight weeks, at which time they are infective. During this time they assume a spiral shape and become encysted by infiltrating leukocytes. They may remain viable for many years. Transmission is through ingestion of the larvae infected meat. Upon ingestion by a host, the cycle repeats; therefore, one animal serves as both definitive and intermediate host, with the juvenile and adult inhabiting different organs. Most mammals are susceptible to infection. Trichnella spp. generally have reduced host specificity, although some species tend to occur in different host associations. The life cycle depends on scavenging food chains: first stage larvae embedded in muscle ingested by a predator or scavenger and develop to adulthood and produce infective larvae in the muscles of the scavenger.