Digenea
Synonym
Flukes
Classification
Class of Platyhelminthes
General Information
The 6000 species of digenetic trematodes are very common and widespread parasites of all classes of vertebrates and may inhabit (as adult or juvenile worms) nearly every organ of their hosts (Table 1). Externally they are characterized by a sucker around the mouth and an additional ventral sucker or acetabulum that is involved both in the attachment to host surfaces and in locomotion. The shape and location of these suckers is species-specific. Digenean development occurs in at least two different hosts and involves several generations. The basic life-cycle pattern employed by digeneans and examples of their larval stages are displayed diagrammatically in Figs. 1–5.
System
The proposed classification of the class Digenea according to the origin of the protonephridial excretory system (Cyrtocyte, Platyhelminthes/Fig. 24) is, in outline (excluding several families), as follows:
Class: Digenea
Superorder: Anepitheliocystidia (in the adult worms the wall of the larval excretory bladder is retained)
Order: Strigeatida (cercariae fork-tailed; miracidia with two pairs of protonephridia)
Families: Diplostomatidae, Schistosomatidae, Spirorchidae, Bucephalidae, Strigeidae, Cyclocoelidae
Order: Echinostomatida (cercariae with simple tail, miracidia with a single pair of protonephridia)
Families: Echinostomatidae, Fasciolidae, Gastrodiscidae, Paramphistomatidae
Superorder: Epitheliocystidia (excretory bladder is newly formed by mesodermal cells; tail of cercariae unforked)
Order: Plagiorchiida (eggs operculated: oral stylet usually present in oral sucker of cercariae; tail of cercariae often lacks excretory vessels)
Families: Dicrocoeliidae, Plagiorchiidae, Prosthogonimidae, Troglotrematidae
Order: Opisthorchiida (eggs operculated; cercariae lack an oral stylet; tail of cercariae always contains excretory vessels)
Families: Opisthorchiidae, Heterophyidae
Important Species
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| Table 1. Important families and species of digenean trematodes
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Life Cycle
Except for some groups (e. g., the dioecious Schistosomatidae; Table 1), flukes are hermaphroditic, utilizing sexual reproduction (with cross-insemination) in the final host. The tanned eggs are produced after fertilization inside a complex system consisting of a central ootype (oogenotop), an ovary, vitelline glands, Mehlis's glands, and uterus (Fig. 6, Fig. 7). These eggs leave the host in feces, urine, or sputum, and the zygote within the eggs develops (or has already developed by this stage) into a ciliated larva (miracidium; Fig. 2) In general this stage infects a gastropod mollusk or a lamellibranch (by penetration or via oral uptake = Clonorchis) as first intermediate host, inside which a polyembryonic, mitotic reproduction occurs involving different developmental stages (sporocysts, rediae) and leading finally to the production of numerous motile and infective cercariae (Fig. 5). The latter leave the first intermediate host, often with a marked rhythm and, in some species, enter a second intermediate host (e.g. Clonorchis) or attach to the surface of plants, or in others directly penetrate the final host (Table 1, Fig. 2).
Inside the second intermediate host or on the surface of plants the cercariae encyst and develop into metacercariae; cyst walls are totally or partly produced by cystogenous glands in the cercarial apex.
The metacercariae grow and become mature adults when orally ingested by the final host. Inside the final host they feed on its fluids depending on their final habitat (Table 1). Nutrients are taken up and digested by means of their anus-less, branched intestine (Fig. 3, Fig. 4) and via the characteristic syncytial tegument (Platyhelminthes/Fig. 11, Platyhelminthes/Fig. 12, Platyhelminthes/Fig. 13).
Reproduction
While the formation of the digenean eggs and of the first larva (miracidium) is rather well understood (reproductive organs), the ontogeny of other larvae remains a problem which is very complex and far from being solved. Electron microscopic studies have shown that the light microscopically visible germ balls consist of mitotically dividing cells which give rise to embryos (polyembryony?) (Fig. 9; sporocysts, rediae, or cercariae) and to a line of new germ cells that become included in these embryonic stages. Since the absence of meiotic processes is not proven, the exact definition remains doubtful. The same is true for embryonic development in the monogenean genus Gyrodactylus. In any case, however, both processes are very successful means of increasing the biotic potential and producing numerous daughter organisms.
Some digenean species belonging to the superfamilies Brachylaemoidea (e.g., Postharinostomum spp., Leucochloridium spp.) and Bucephaloidea (e.g., Bucephalus spp.) form branching sporocysts. By constrictions such branches may split off and finally grow to their former size. This occurs by mitotic divisions of undifferentiated cells which are found below the body wall (Platyhelminthes/Fig. 10).
Integument
The shape and the development of the peculiar body cover is described under Platyhelminthes/Integument. In addition it is noteworthy that all larval stages seem to have a surface coat containing proteoglycans (acid mucopolysaccharides). In the miracidium even the cilia are covered by a surface coat. Mother sporocysts of Fasciola hepatica and Schistosoma mansoni show an amplification of the surface area by a mixture of branching folds and microvilli. Both are covered by a fuzzy surface coat. Vesicles at the base of the microvilli suggest the occurrence of endocytosis. Rediae also have a surface coat. Although there is evidence that they are able to take up nutrients through the mouth into the small digestive system, absorption of nutrients by endocytosis through the tegument has been observed for glucose, a polysaccharide, and amino acids.
Host Finding
Miracidia
Miracidia which actively reach their aquatic snail hosts are infectious for only a few hours, and their behavior seems to center entirely around finding and invading the host snail. Their behavior patterns have functions in at least four phases of host-finding:
- Dispersal. After escaping from the egg, some miracidial species tend to disperse from the site of origin. They swim relatively fast and linearly, and even their orientation to light and gravity seems to be conductive to dispersal. Species which later infect ground-dwelling hosts can exhibit photopositive and geonegative orientation for approximately the first hour after emergence. During this phase some miracidia are not as responsive and infective to their host snails as they are in later phases of the host-finding process.
- Microhabitat selection. Most miracidia tend to accumulate in the microhabitats which are preferred by their host snails. This is achieved by responding to environmental stimuli such as light, gravity, temperature, and the magnetic field. The type of orientation may resemble very closely that of the host snail. For example, Schistosoma mansoni miracidia show a geonegative and photopositive orientation as do their host snails. The type of orientation may be modified by certain environmental conditions. For example, the negative photo-response of Schistosoma haematobium miracidia reverses with decreasing temperature and they were found to infect their host snails in summer preferentially at the bottom of the water and in winter near the water surface. Miracidia of Schistosoma japonicum reverse their photopositive orientation to a negative one with increasing temperature and/or light intensity, a behavior which also resembles that of the host snails.
The photopositive orientation of at least Schistosoma mansoni miracidia is a phototaxis, as the miracidia are able to detect the relative intensites of two separate sources of light. The wavelengths to which the organisms respond have been determined for miracidia of Fasciola hepatica, Schistosomatium douthitti, S. mansoni and Bunodera mediovitellata. They all respond maximally to blue-green light between 500 and 550 nm, which penetrates deepest into clear water. S. mansoni and B. mediovitellata miracidia prefer in addition red-brown light of 650 nm, which is typical for muddy waters.
Several miracidial species are able to perform geoorientation independently of their photoresponses, but the mechanisms involved in this type of orientation are not well understood. A special mechanism occurs in miracidia of Philophthalmus gralli. Their strong geopositive orientation seems to be brought about by a sensitive north-seeking magnetotaxis in the northern hemisphere.
- Host-directed orientation. When miracidia enter the host snail's active space, they approach the host by responding to snail-emitted chemical signals. The type of orientation is a chemokinesis in most species studied so far, e.g. in Schistosoma mansoni, Schistosoma haematobium, Trichobilharzia ocellata, Fasciola hepatica, Echinostoma caproni. The organisms increase their rate of change of direction (RCD) in increasing stimulus concentration gradients and they perform a sharp 180E turn when the concentration of the stimulus decreases (Fig. 10 A,B). For this type of orientation, the miracidia must only detect an increase or a decrease of the stimulus concentration. This is possible although the miracidia rotate along their long axis, which supports a straight path of movement. However, S. japonicum miracidia show a directed chemotaxis (Fig. 10C) and the rotating organisms seem to detect the direction of the concentration gradient in an unknown way.
Much research was performed using various methods to identify the attractive components of snail tissues, mucus and snail conditioned water (SCW). Most work dealed with miracidia of S. mansoni, but the results were very controversial. Small molecular compounds with low host specificity were described, such as amino acids, magnesium ions, decreasing calcium ion concentration, peptides, short-chain fatty acids, N-acetylneuraminic acid, ammonia, hydrogen ions, glutathione, glucose. However, recent studies identified macromolecular glycoconjugates as the exclusively attracting molecules of SCW in miracidia of S. mansoni, S. haematobium, T. ocellata, F. hepatica, and E. caproni, and there is some evidence that other species are also attracted by macromolecules. The molecules which attract S. mansoni, T. ocellata, and F. hepatica have very similar chemical characteristics, their saccharide chains are linked to a core protein via an O-glycosidic linkage probably between threonine or serine and N-acetylgalactosamine, and their identification signal is encoded in the carbohydrate moiety. At least the miracidia of F. hepatica, T. ocellata and an Egyptian strain of S. mansoni identify their respective host-snail species with a very high specificity and sensitivity (Fig. 11). The signaling macromolecules are effective at concentrations as low as 1 mg in 10 000 liters of water, and the signaling carbohydrate structures not analyzed so far may be recognized at an even considerably lower concentration.
- Host contact and invasion. After contact with solid surfaces miracidia show at least 12 different behavior patterns (MacInnis), the typical behavior at the suitable host snail's surface is “repeated investigation”. This response is stimulated by the same host-specific glycoconjugates which stimulate the preceding chemo-orientation, at least in S. mansoni, S. haematobium, Trichobilharzia ocellata, and F. hepatica. The releasing cues for the subsequent attachment and penetration behavior patterns are poorly studied. The attachment of F. hepatica miracidia seems to be supported by snail mucus, short chain fatty acids and mechanical interaction with snail epidermis structures.
A typical characteristic of digenean-snail interactions is a stringent degree of specificity. Each species of digenean is capable of developing only in very few of the available snails, often only a single species or geographic strain. The highly specific host finding behavior of miracidia is obviously fully adapted to this situation.The high specificity, which underlies miracidial host finding, may be easily overlooked in laboratory experiments, where the miracidia are exposed to the snails in a few milliliters of water. However, in the field the specific host finding behavior might be a determining factor of the parasite-snail compatibility. Field- or semifield-studies should now characterize the impact of miracidial host-finding on transmission success. The fact that the host snails signal their species specificity with such a precision despite 400 million years of coevolution with their digenean parasites suggests that the snails might need the signal molecules for other purposes. They might serve as pheromones for intraspecific communication of the snails, and the digeneans possibly just misuse them for safe host identification. Whether the attractants can be used for specific control methods still has to be investigated.
Cercariae
Cercariae of many digeneans find and recognize their hosts with complex behavior patterns and via very different, highly sensitive and specific receptors. They have evolved an enormous diversity of host-finding strategies and can achieve very high host specificities. This indicates that cercarial host finding plays a central role in the transmission of digeneans and that it is an important determinant in the evolution of their life cycles.
Cercariae of many digenean species invade their hosts directly within aquatic habitats. They are provided with limited accumulated energy reserves and must invade the hosts within their short life span of only 1–3 days. Most species support the transmission success by producing high numbers of cercariae (up to 500 000 cercariae per host snail daily). In addition to this, most cercariae show complex behavior patterns which seem entirely dedicated to finding hosts. The various cercarial behavior patterns have functions in the following host-finding processes:
- Departure from the snail host and dispersal. Immediately after leaving the snail hosts some species (such as Diplostomum spathaceum and Schistosoma haematobium) show a particularly high swimming activity which carries them away from the snails towards their preferred position within the habitats. Some cercarial species that infest snails as second intermediate hosts avoid super-infections of the first intermediate host snails that emit them with characteristic behavior. They show phases of converted orientation towards light and gravity in their immediate post-shedding period, and they search their second intermediate hosts later. E.g., cercariae of Hypoderaeum conoideum start chemotactic orientation towards snail hosts at the earliest 1 h after they have left their first intermediate host snails.
- Habitat selection. Most cercariae tend to disperse in the habitats normally frequented by their potential hosts. For example, cercariae of human schistosomes accumulate in the uppermost water layers, whereas parasites of ground-dwelling invertebrates orientate towards the bottom of waters, and some of them select microhabitats there with a defined light intensity. This habitat selection is achieved with different types of spontaneous swimming behavior and by responses to environmental stimuli, such as gravity, light and shadow, water currents, temperature and physical boundaries. The different species show an enormous diversity of mechanisms in responding to environmental cues. For example, some species achieve geo-orientation exclusively by gravity-determined swimming movements, other species respond only to the direction of light radiation, and still others adjust their position in the water column by responding only to light intensity, independent of the direction of the light source.
- Localization in host time. Even the time of day the cercariae occupy the microhabitats is correlated with the habits of the subsequent hosts. This is in part achieved by the pattern of cercarial emergence from the host snail. For example, the human schistosomes leave their host snails in the middle of the day, Schistosomatium douthitti at night when their rodent hosts are active, and S. margrebowiei shows two emission peaks, one at dawn and one at dusk, i.e. when their antelope hosts water. Even an intraspecific polymorphism in the shedding patterns was found: S. mansoni cercariae left their snails in foci with humans as hosts mainly at 11–12 h, but in nearby foci with rats as hosts at 16–17 h. The emergence behavior is hereditary and it is synchronized in schistosomes mainly by the photoperiod, but the thermoperiod and other factors also have an effect.
- Host-directed orientation. In many cercariae a first contact with the host seems to occur by chance. However, some species respond to stimuli emanating from their hosts in such a way, that the chances of encountering the host increase. Three types of stimulating host cues have been studied in more detail: chemical cues, water turbulence and touch, and dark stimuli (Table 2).
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| Table 2. Host cues supposed to facilitate approach of swimming cercariae toward the host
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Chemical cues: Chemo-orientation towards the host has been found in echinostome cercariae, which invade slowly moving aquatic snails (Table 2). Three species show chemokinetic orientation, they simply turn back when the concentrations of snail-emitted amino acids, urea and ammonia decrease. However, one species can swim by being directed chemotactically along increasing concentration gradients of snail-emitted peptides. A prerequisite for this chemotaxis is that the cercariae can detect the direction of the concentration gradient. Most other cercariae, which invade faster moving hosts, do not seem to use long-distance chemo-orientation. Only S. mansoni is able to orientate in an unknown way along artificially high concentration gradients of linoleic acid.
Water turbulence and touch: The schistosome cercariae respond only poorly or not at all to water currents. This may help to avoid energy-consuming responses to the frequent encounters with water-inhabitating non-host organisms. The cercariae of the duck parasite T. ocellata react to water currents with forward swimming and a readiness to respond to host cues with attachment. This response is inhibited when the parasites save energy by clinging to the water surface (Fig. 12). All fish-invading cercariae of Table 2 respond to water currents by starting to swim, which may increase the chance of a contact with the moving hosts. At least in D. spathaceum and Opisthorchis viverrini water turbulence not only triggers long swimming movements but also evokes attachments, when the current-excited cercariae swim against solid substrates.
Dark stimuli: All fish parasites of Table 2 respond to shadow stimuli with the start of swimming movements. Whether this behavior may support an encounter with the fish hosts is not clear, as the shadow stimulus simultaneously inhibits the triggered swimming bursts and as it does not stimulate attachments. Among the Schistosoma species of Table 2 only S. haematobium responds significantly to shadows. The cercariae shift to a high swimming activity, which leads them upward in the water column, and this may increase the chance for an encounter with their human hosts. In the duck parasite Trichobilharzia ocellata a shadow response plays a dominant role in host finding (Fig. 3). These cercariae prefer an energy-saving position clinging to the water surface, where they do not respond to the frequent water current stimuli. But when a shadow falls on them they swim away from its source, usually downward. This may increase the chances of encountering the feet of their duck hosts. The shadow simultaneously triggers a readiness to respond to thermal and chemical duck foot cues with attachment behavior. Wavelengths to which cercariae respond maximally are 500 nm in Trichobilharzia ocellata, 490 nm in S. mansoni, and 550 nm in Bunodera mediovitellata.
- Host contact and invasion. After contact with the host's surface, the cercariae recognize and invade the host with a series of behavior patterns which can be carried out spontaneously, or in response to host signals of quite different host specificity: (1) Attachment to the host; (2) enduring contact with (remaining on) the host; (3) creeping to suitable entry sites; (4) penetration into the skin with elements such as penetration movements, shedding of the tail, secretion of acetabular gland contents, and tegument transformation for immune evasion. Each of these behavior patterns may be stimulated independently by separate host signals. Some species achieve a very narrow host specificity employing strategies including: (1) A chronological succession of responses to different stimuli, which themselves may be of low specificity; (2) responses to a combination of different stimuli; (3) sensitive responses to individual stimuli with a high host specificity. A typical characteristic of cercarial host finding is a high diversity of strategies. Each species studied so far recognizes its host with responses to host signals that differ considerably from those of other species, even when these invade the same host genera. This may reflect adaptations to maximal transmission successes within differing ecological conditions. The stimulating cues for cercarial behavior after host contact have been analyzed for species invading mammals, birds, and fish (Table 3).
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| Table 3. Host signals as stimuli for cercarial host-recognition and invasion phases
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Schistosomes invading mammalian hosts (Table 3) find their host in a characteristic manner, either by responses to different host signals or by an individual sensitivity of the responses. For example, S. mansoni cercariae respond in each of the host-finding phases with a particular sensitivity to chemical host cues. It was speculated that this might represent an adaptation to clear water habitats or to an infection near the water surface, where the detection of chemical cues is not disturbed by mud components. In contrast to that, S. haematobium cercariae respond most sensitively to thermal stimuli. They attach maximally to substrates when their temperature exceeds ambient temperature by only 1° C (S. mansoni 11–13° C), and they migrate along temperature gradients as low as 0.03° C/mm (S. mansoni 0.15° C/mm). This specialization on thermal host cues was interpreted as adaptation to invading the hosts in cooler habitats, where warm host surfaces are especially contrasting. A very low specificity occurs in the host finding of S. japonicum cercariae. They attach to and remain on the host purely by chance and their penetration behavior and tegument transformation can be stimulated by warmth alone. This bears the risk of cercarial losses by penetration attempts into warm non-host substrates. However, this behavior enables the S. japonicum cercariae to invade a broad spectrum of mammals including species with a low content of free fatty acids in their skin surface. Their poor response to chemical host cues was interpreted as an adaptation to the muddy habitats in which they actually invade their hosts, i.e. habitats where chemical components of mud could interfere with chemoreception.
A common characteristic of all schistosomes studied so far is that they respond very sensitively to certain free fatty acids of mammalian skin surface with penetration behavior and transformation of their tegument for immune evasion. There is evidence that the fatty acids act via chemoreceptors and neural mechanisms. However, they may also have other functions in the invasion processes. S. mansoni and T. ocellata cercariae synthesize various eicosanoids when they are incubated in essential fatty acids, and it was suggested that these might directly act in the tegument transformation and immune evasion processes.
Host recognition of bird-invading schistosomatids has been studied only in 3 species in detail (Table 3). The host cues to which they respond are also characteristic for mammalian skin and this suggests, that they are not adapted to avoiding unsuccessful penetrations into mammals. In fact, several bird-invading schistosomatids are known to cause cercarial dermatitis in humans. Each of the species studied so far responds to ceramides or cholesterol as host cues. These lipids also occur in mammalian skin surface, but they are lacking in the birds' uropygial gland secretions which are distributed on the feathers. Therefore, the response to these lipids may help to avoid useless attachments or penetration attempts to bird feathers.
Detailed data are available only for 3 fish-invading species (Table 3). The attachment response of D. spathaceum is stimulated by host-derived carbon dioxide, and the cercariae attach to nearly all animals. The disadvantage of the low specificity may be overcome by a high sensitivity to carbon dioxide, which allows a very fast attachment. The fish specificity is then achieved during the enduring contact and penetration phases, where the D. spathaceum cercariae respond to macromolecules. Each of the 3 fish-invading species achieves its fish specificity by responses to at least 2 different types of macromolecules. How the specificity is encoded in the signaling proteins and glycoconjugates is poorly understood. There is some evidence that terminally positioned sialic acids in the mucus are used to distinguish a host fish from mucus covered aquatic invertebrates.
