Development and intensity dependence of Ornithodiplostomum ptychocheilus metacercariae in fathead minnows (Pimephales promelas)

Intensity-dependent development of Ornithodiplostomum ptychocheilus metacercariae was studied in fathead minnows (Pimephales promelas) exposed to 0, 20, or 120 cercariae. Subsamples of hosts were necropsied at 2-wk intervals to monitor parasite recruitment, growth, and time to encystment. The complex development of metacercariae within the cranium of minnows involved growth, encystment, and consolidation phases, each of which were affected by intensity. At the end of the growth phase, metacercariae from low-dose fish were 20% longer than those from high-dose fish and the latter took 2-4 wk longer to encyst. At the end of the postencystment consolidation phase (6-8 wk postinfection), the size of metacercariae decreased by approximately 50%. The rate of consolidation was slower in high-dose fish. Our results show that time, intensity, and temperature affect development of O. ptychocheilus. Because metacercariae development and differentiation are linked to infectivity, events occurring in intermediate hosts can potentially impact the structure and size of trematode suprapopulations.     

High intensity and prevalence of two species of trematode metacercariae in the fathead minnow (Pimephales promelas) with no compromise of minnow anti-predator competence

Opportunity for parasites to manipulate host behavioral phenotype may be influenced by several factors, including the host ecology and the presence of cohabiting parasites in the same host. Metacercariae of Ornithodiplostomum ptychocheilus and "black spot" Crassiphiala bulboglossa have similar life cycles. Each parasite uses a littoral snail as a first intermediate host, fathead minnows as a second intermediate host, and a piscivorous bird as a final host. Metacercariae of black spot encyst in the dermal and epidermal tissues, while metacercariae of O. ptychocheilus encyst on the brain over a region that coordinates optomotor responses. Because of site differences within the host, we predicted that O. ptychocheilus metacercariae might manipulate the behavioral phenotype of minnows to facilitate transmission to the final host, but metacercariae of black spot would not. In our study population, prevalence was 100% for O. ptychocheilus , with an overall median intensity of 105 metacercariae per minnow. Prevalence of black spot was 60%, with a median abundance and intensity of 12 and 20 metacercariae per minnow for the overall sample and for infected fish, respectively. Minnows accumulated both parasites over time, producing significant correlations between intensity and minnow body length and between intensities of the 2 parasites. Minnows infected with black spot had on average twice as many O. ptychocheilus metacercariae as similar-sized minnows without any black spot cercariae. We found no correlation between body condition of minnows and intensity for either parasite. We measured 2 aspects of anti-predator competence to test for effects linked to parasite intensity. We found no correlation between intensity of either species of parasite and latency to behavioral response to attack from a mechanical model heron, nor was there any effect of parasite intensity on a measure of shoaling affinity. The absence of any detectable effect of metacercariae on anti-predator competence in minnows may reflect selection against parasite pathology from predation by non-hosts of the parasites and overwinter mortality due to low dissolved oxygen.     

Infectivity of Ornithodiplostomum ptychocheilus and Posthodiplostomum minimum (Trematoda: Diplostomidae) cercariae following exposure to cadmium

The effect of cadmium (Cd) exposure on infectivity of cercariae of 2 trematode species (Ornithodiplostomum ptychocheilus and Posthodiplostomum minimum) to their second intermediate fish host was evaluated. Individual fathead minnows (Pimephales promelas) were exposed to cercariae that had been treated with Cd solutions at concentrations of 0, 2, 20, or 200 microg/L for 2 hr. Two weeks later, the numbers of encysted metacercariae in the brain (O. ptychocheilus) and body cavity (P. minimum) of the fish were evaluated. ANOVA analyses indicated a strong negative effect of Cd concentration on cercaria infectivity. The species x Cd concentration interaction was not significant, indicating that the magnitude of Cd-induced reduction in infectivity was similar between O. ptychocheilus and P. minimum. The results show that short-term exposure to Cd, even at low concentrations, interferes with transmission processes that affect the recognition and penetration of cercariae, the migration and survival of metacercariae within the second intermediate host, or both.     

Histopathological and ultrastructural observations of metacercarial infections of Diplostomum phoxini (Digenea) in the brain of minnows Phoxinus phoxinus

The spatial distribution and histopathological changes induced by metacercariae of the digenean trematode Diplostomum phoxini (Faust, 1918) in the brains of European minnows Phoxinus phoxinus (L.) from the River Endrick, Scotland, were studied by light and electron microscopy. Post-mortem examination of a sample of 34 minnows revealed that 50% (n = 17) of the population was infected with 13.7 +/- 2.6 (mean +/- SE; range 1 to 38) metacercariae per infected host. Serial histological sections of the infected minnow brains revealed that the metacercariae were unevenly distributed throughout the brain, with aggregations occurring in the cerebellum, the medulla oblongata and the optic lobes. In fish with highest intensities of infection, over 40% of the cerebellar area and about 30% of the medulla oblongata area were occupied by larvae. Metacercariae disrupt the integrity of brain tissue, with individuals being found in small pockets surrounded by cellular debris. Metacercariae were rarely encountered on the surface of the brain. Electron microscopic examination of infection sites revealed that the granular layer surrounding metacercariae was necrotic, exhibited nuclear degradation and was marked by vacuolation of the cytoplasm. Rodlet cells, the only inflammatory cell types recorded in this study, were found only in parasitized brains and in close proximity to the teguments of metacercariae. It is hypothesised that secretions released from the teguments of metacercariae are a counter response to protect the metacercariae from the fish brain's cellular defence mechanisms.     

Studies on the infectivity of Diplostomum spathaceum in rainbow trout (Oncorhynchus mykiss)

The infectivity of Diplostomum spathaceum (Digenea: Trematoda) cercariae to rainbow trout and the efficacy of the diplostomule migration to the lens following different routes of administration was examined. The optimum age of infectivity for cercariae was between 0-5 h after liberation from the snail and for intraperitoneally injected diplostomules, 5 h post-transformation in vitro through fish skin. After exposure of the entire fish body or head to cercariae, metacercariae first appeared in the lens at 5 h and their numbers gradually increased until 22 h. Following exposure of the tail region of rainbow trout to cercariae, metacercariae first appeared in the lens at 14 h. Significantly more metacercariae established in the lens of fish following exposure of the fish head compared with the tail region; 40% of penetrating cercariae reached the lens of fish following exposure of the head or entire body, 20% of cercariae or diplostomules injected either intraperitoneally, intramuscularly or intracardially reached the lens while only 5% of cercariae established as metacercariae following exposure of the tail region.     

倪氏复口吸虫在稀有鮈鲫体内的移行及对其耳石微结构的影响

倪氏复口吸虫是我国淡水鱼类的一种常见的危害严重的寄生虫。利用稀有鮈鲫作为寄主,研究了倪氏复口吸虫在其体内的移行途径,并观察了复口吸虫的感染是否在寄主耳石上留下标记轮。倪氏复口吸虫尾蚴从稀有鮈鲫体表的各个部位侵入,但主要集中在头部,侵入体内的尾蚴通过肌肉和结缔组织直接进入眼球。感染倪氏复口吸虫的稀有鮈鲫耳石表面微结构发生变化,部分鱼耳石上出现明显的标记轮。     

Density and maturation of rodlet cells in brain tissue of fathead minnows (Pimephales promelas) exposed to trematode cercariae

Evidence for the presumed linkage between the enigmatic rodlet cells of fish and exposure to helminths is anecdotal and indirect. We evaluated the proliferation and development of rodlet cells in the optic lobes of fathead minnows exposed to cercariae of Ornithodiplostomum ptychocheilus. Mean rodlet cell densities (ca. 10/mm²) in the optic lobes were similar between unexposed controls and minnows with 1- and 2-week old infections. Rodlet cell densities increased at 4 weeks p.i., reaching maxima (ca. 200/mm²) at 6 weeks p.i., followed by a decline at 9 weeks. This temporal pattern of proliferation and maturation paralleled the development of metacercariae within the optic lobes. Unencysted metacercariae develop rapidly within tissues of the optic lobes for approximately 4 weeks after penetration by cercariae, then shift to the adjacent meninges to encyst. The former stage is associated with tissue damage, the latter with massive inflammation of the meninges. Thus, peak densities and maturation of rodlet cells correspond to the period when inflammation of the meninges caused by the large metacercarial cysts is at a maximum. Our results support recent contentions that rodlet cells comprise part of the host inflammatory defence response.     

Behavioral responses of Poecilia vivipara (Osteichthyies: Cyprinodontiformes) to experimental infections of Acanthocollaritrema umbilicatum (Digenea: Cryptogonimidae)

The swimming behavior of Poecilia vivipara was evaluated using an image analysis system comparing laboratory-reared uninfected fish before and after experimental infection with different intensities of cercariae of the trematode Acanthocollaritrema umbilicatum. Two experiments were performed, each with 30 fish which were individually exposed to 30 and 50 cercariae, respectively, shed from experimentally infected molluscs, Heleobia australis. Before and after (17–27 days) infection, the behavior of each fish was monitored in terms of Distance travelled, Ambulatory time, Stereotypic time, Resting time and Average speed. At the end of the experiments, the fish were dissected to count the number of metacercariae recovered. In the experiment with 30 cercariae, fish with 2–10 metacercariae did not exhibit any significant differences in their swimming activity, but those with 11–22 metacercariae had a significantly enhanced Stereotypic time and a reduced Time Resting. In the experiment with 50 cercariae, fish with 5–22 metacercariae had an enhanced Distance travelled and a reduced Average speed; highly significant differences occurred with regard to all behavioral parameters when considering the subgroup 23–36 metacercariae: Distance travelled, Stereotypic time, Resting time, Ambulatory time and Average speed. The swimming behavior of P. vivipara changed influenced by an intensity-dependence on metacercariae of A. umbilicatum, supporting the prediction that parasites are able to alter the behavior of their hosts.     

Clonorchis sinensis metacercarial infection in the pond smelt Hypomesus olidus and the minnow Zacco platypus collected from the Soyang and Daechung Lakes

The pond smelt Hypomesus olidus and minnow Zacco platypus were collected from the Soyang and Daechung Lakes in January 2003, and their metacercarial infections was examined by the muscle compression and artificial digestion techniques. In the Soyang Lake, 161 metacercariae of Clonorchis sinensis (0.35 per fish) were harvested from 459 pond smelts examined. Also, 13 metacercariae of C. sinensis (0.43 per fish), 1 of Metagonimus sp., 4 of Echinostoma sp., 148 of Centrocestus armatus and 44 unidentified species were collected from 30 minnows. In the Daechung Lake, 369 metacercariae of C. sinensis (3.69 per fish) and 51 unidentified species were recovered from 100 pond smelts. The metacercariae of C. sinensis were fed to experimental rats, in which the adult flukes were identified. The pond smelts and minnows collected from the Soyang and Daechung Lakes were verified to be the second intermediate hosts and the sources of human C. sinensis infection.     

Population dynamics of Ornithodiplostomum ptychocheilus metacercariae in fathead minnows (Pimephales promelas) from four northern-Alberta lakes

Annual, seasonal, and interlake variation in prevalence and intensity of Ornithodiplostomum ptychocheilus (Faust) metacercariae was assessed in populations of fathead minnows (Pimephales promelas) collected from 4 lakes in north-central Alberta. Mean metacercariae intensity in young-of-the-year minnows varied extensively (5-123 metacercariae/host) among year, month, and lakes. In 2 of the lakes, prevalence always reached 100%, and mean intensity always peaked in September or October. The high spatial and annual variation in metacercarial recruitment was partly attributable to variation in host size. but variation in water depth, temperature, snail densities, and bird visitation likely also played a role. A laboratory experiment demonstrated that host and metacercariae survival was intensity-independent during a period of simulated winter. Thus, metacercariae recruited in the fall survive until the following spring.     

Austropeplea ollula (Pulmonata: Lymnaeidae): a new molluscan intermediate host of a human intestinal fluke, Echinostoma cinetorchis (Trematoda: Echinostomatidae) in Korea

Three freshwater snail species of the family Lymnaeidae have been reported from Korea, Radix auricularia coreana, Austropeplea ollula and Fossaria truncatula. Out of 3 lymnaeid snail species, A. ollula was naturally infected with the Echinostoma cinetorchis cercariae (infection rate = 0.7%). In the experiments with the laboratory-bred snails, F. truncatula as well as A. ollula was also susceptible to the E. cinetorchis miracidia with infection rates of 25% and 40%, respectively. All of three lymnaeid snail species exposed to the E. cinetorchis cercariae were infected with the E. cinetorchis metacercariae. It is evident that A. ollula acts as the first molluscan intermediate host of E. cinetorchis in Korea, and F. truncatula may be a possible candidate for the first intermediate host of this intestinal fluke. Also, three lymnaeid snail species targeted were experimentally infected with E. cinetorchis metacercariae.     

Changes in minnow, Pimephales promelas Rafinesque, schooling behaviour associated with infections of brainencysted larvae of the fluke, Ornithodiplostomum ptychocheilus

The schooling behaviour of unparasitized fathead minnows, Pimephales promelus Rafinesque, was compared to that of minnows infected with brain-dwelling metacercariae of the fluke, Ornithodiplostomum ptychocheilus (Faust). Laboratory results showed that schools of infected fish divided more frequently, were less compact, and occupied positions closer to the water surface than control schools. These results are discussed with reference to the possibility that this larval parasite manipulates minnow behaviour to increase host vulnerability to predation.     

Relationship between Snail Population Density and Infection Status of Snails and Fish with Zoonotic Trematodes in Vietnamese Carp Nurseries

Background

Fish-borne zoonotic trematodes (FZT) are a food safety and health concern in Vietnam. Humans and other final hosts acquire these parasites from eating raw or under-cooked fish with FZT metacercariae. Fish raised in ponds are exposed to cercariae shed by snail hosts that are common in fish farm ponds. Previous risk assessment on FZT transmission in the Red River Delta of Vietnam identified carp nursery ponds as major sites of transmission. In this study, we analyzed the association between snail population density and heterophyid trematode infection in snails with the rate of FZT transmission to juvenile fish raised in carp nurseries.

Methodology/Principal Findings

Snail population density and prevalence of trematode (Heterophyidae) infections were determined in 48 carp nurseries producing Rohu juveniles, (Labeo rohita) in the Red River Delta area. Fish samples were examined at 3, 6 and 9 weeks after the juvenile fish were introduced into the ponds. There was a significant positive correlation between prevalence of FZT metacercariae in juvenile fish and density of infected snails. Thus, the odds of infection in juvenile fish were 4.36 and 11.32 times higher for ponds with medium and high density of snails, respectively, compared to ponds where no infected snails were found. Further, the intensity of fish FZT infections increased with the density of infected snails. Interestingly, however, some ponds with no or few infected snails were collected also had high prevalence and intensity of FZT in juvenile fish. This may be due to immigration of cercariae into the pond from external water sources.

Conclusions/Significance

The total number and density of potential host snails and density of host snails infected with heterophyid trematodes in the aquaculture pond is a useful predictor for infections in juvenile fish, although infection levels in juvenile fish can occur despite low density or absence infected snails. This suggests that intervention programs to control FZT infection of fish should include not only intra-pond snail control, but also include water sources of allochthonous cercariae, i.e. canals supplying water to ponds as well as snail habitats outside the pond such as rice fields and surrounding ponds.     

The role of light and gravity in the experimental transmission of Echinostoma caproni (Digenea: Echinostomatidae) cercariae to the second intermediate host, Biomphalaria glabrata (Gastropoda: Pulmonata)

Trematode cercariae inhabit predictable environments and respond to trigger cues with genetically fixed releaser responses when foraging for the upstream host. The effect of light and gravity on the transmission of Echinostoma caproni cercariae to Biomphalaria glabrata was investigated experimentally. Transmission chambers were constructed of clear polyvinyl chloride pipe. Snails were constrained within the chamber to prevent movement, while permitting the cercariae to swim freely. A trial consisted of 2 infected B. glabrata shedding E. caproni cercariae placed at the center of the chamber, with 5 uninfected B. glabrata placed 10 cm on either side (or above and below) of the shedding snails as sentinels. There was no significant difference in the prevalence of infection sentinel snails in either experiment (light vs. dark or top vs. bottom); however, mean intensity was significantly higher in sentinel snails in the dark portion of the chamber (42.5 vs. 10.4; P = 0.001) and the top of the transmission chamber (66.1 vs. 38.0; P = 0.0003). There was a high correlation between the number of metacercariae collected from sentinel snails and the total number of infective units (metacercariae + unsuccessful cercariae): r = 0.992 (light vs. dark) and r = 0.957 (top vs. bottom), respectively, at cercariae densities estimated from 22 to 3,304/L. The results suggest that cercariae of E. caproni exhibit negative photo- and geotaxis in searching for a second intermediate host. Stereotypical releaser responses to environmental trigger cues (light and gravity) allow E. caproni cercariae to exploit flexible strategies for completing the life cycle consistent with the broad range second intermediate and definitive hosts used by E. caproni cercariae and adults, respectively.     

Infection by Paragonimus miyazakii cercariae of their crab hosts, Geothelphusa dehaani, by percutaneous penetration

Identification of the transmission routes of the trematode parasite Paragonimus miyazakii into different intermediate hosts would help to explain the natural distribution of the parasite. The behavior of P. miyazakii cercariae released from snails into water and in the presence of a living host or a whole crab leg was observed by stereoscopic or light microscopy at various times after exposure started. On encountering a crab leg or cheliped, the cercariae became entangled with the host via mucoid strands arising from the cercariae. Within 3 hr, most cercariae were attached to the host; cuticular penetration took between 5 and 6 hr, after which cercariae were found in the cavity of the leg. Crabs examined 102-149 days after exposure to the cercariae contained fully developed metacercariae. The metacercariae were fed to 2 rats, and the rats were killed 83 or 111 days later. Some of the metacercariae had reached maturity in the rats. That the cercariae were not ingested by the crabs but penetrated the crabs percutaneously (through hard as well as soft tissue) means that transmission can occur even in areas in which crabs and the host snails do not coexist, as they would if the usual route were oral (when the crabs ate infected snails).     

Distribution of metacercariae of Centrocestus formosanus (Trematoda: Heterophyidae) on the gills of Aplocheilus panchax

The metacercariae of Centrocestus formosanus (Nishigori 1924) occur on the gills of freshwater fishes in association with blood vessels. The microecology of the metacercariae on the gills of the cyprinid fish Aplocheilus panchax (Ham. & Buch.) was investigated. The cysts showed a preference for anterior gills and for the dorsal section of each gill. Variations in the density of infection, season and size of the fish do not affect the spatial distribution of metacercariae. It is presumed that ventilation water currents spread the cercariae onto the anterior gills and then the cercariae actively select the preferred habitat.     

Long-term stability in the population levels of the eyefluke Tylodelphys podicipina (Digenea: Diplostomatidae) in perch

Annual changes in the population size of the metacercariae of Tylodelphys podicipina in the eyes of perch in a small eutrophic lake were studied over ten years by following changes in prevalence, abundance and overdispersion of parasites throughout the life of each year class of fish. The population increased rapidly in the first two years after its introduction to the lake, but for the next six years fluctuated within very narrowly constrained limits before declining as a result of a catastrophic decline in the perch population. No evidence of parasite-induced host mortality was found: the decline in parasite abundance throughout the life of each year class was due to density-independent parasite mortality within the fish as a result of natural variation in the life span of the metacercariae. Levels of infection in the first year of a year class were determined principally by transmission processes, as transmission could only occur over a very brief period when release of cercariae from parent generation snails coincided with the presence of perch fry. Transmission appeared to be density-independent, and infection levels in perch to be determined by the dimensions of the transmission' window' and subsequently modified by parasite mortality. No evidence of host responses or any other regulatory factor of fish origin was found, and the infrapopulations in perch appear to be non-equilibrial and unstable. Although the possibility of regulatory processes acting on infrapopulations of the parasite in other hosts remains, the importance of transmission windows in determining infrapopulation levels in fish is emphasized.     

Molecular identification of developmental stages in Opecoelidae (Digenea)

Nuclear ribosomal DNA sequences represent a useful tool for distinction of poorly differentiated developmental stages, such as trematode cercariae or metacercariae. Here, the complete internal transcribed spacer region of the ribosomal DNA (ITS 1 + 5.8S + ITS 2) was sequenced for 29 specimens of the digenean family Opecoelidae, including 16 adult specimens and 13 undescribed larval stages (nine cercariae and four metacercariae) occurring in various marine host organisms. Six cercariae and three metacercariae were found to match their corresponding adult form. This work also revealed that cercariae of the same species are able to infect more than one gastropod host species, suggesting that the specificity for the first intermediate host within the Digenea may be lower than previously thought.     

Patterns of fish deformities and their association with trematode cysts in the Willamette River,Oregon

Synopsis We described skeletal deformities in Willamette River fishes from larval and juvenile specimens collected in 2002 and 2003. Deformities were found in most taxa examined but were more frequent in native broadcast spawners, especially minnows and suckers, than in native or exotic nest builders. Caudal deformities were uniformly distributed throughout the river, but precaudal deformities were more localized near the towns of Newberg and Salem (Wheatland Ferry). In northern pikeminnow, Ptychocheilus oregonensis, deformities were dependent on relative hatch date, with early season fish having about three times the deformity load as late season fish. In a subsample of northern pikeminnow and chiselmouth, Acrocheilus alutaceus, number of deformities was directly related to number of trematode metacercariae and precaudal deformities were twice as likely as caudal deformities to be associated with metacercariae. Based on a logistic regression, the probability of a precaudal deformity was dependent on number of metacercariae and geographic area with the area effect disappearing as the number of cysts increased. A separate analysis showed that some types of deformities were unlikely to be associated with metacercariae. However, even in cases where metacercariae were unlikely to be associated with deformities, metacercariae were usually present elsewhere in the fish and an indirect effect could not be dismissed. The taxonomic, spatial and temporal patterns of skeletal deformities in Willamette River fishes may be due to differences in intermediate host (snail or fish) resistance or susceptibility to parasites, to differences in life history ecology, or to anthropogenic effects that are manifest in increased precaudal deformities near Newberg and Wheatland Ferry or decreased rates elsewhere.     

Intensity-dependent alteration of minnow (Pimephales promelas) behavior by a brain-encysting trematode

We examined the relationship between the numbers of brain-encysting trematodes (Ornithodiplostomum ptychocheilus) and the magnitude of altered behaviors in fathead minnows (Pimephales promelas). Because cysts develop within a brain region that integrates visual stimuli with motor response. we evaluated the standard optomotor response (OMR). Monitoring this task involved recording the time minnows spent following a spinning drum, on which alternating black and white stripes had been painted. Minnows were exposed to 0, 5, 20, 120, and 300 cercariae and then their OMR was evaluated at 2-wk postinfection. Surprisingly, only minnows that had high numbers of parasites (155 +/- 31 worms/fish) or low numbers of parasites (3 +/- 3 worms/ fish) differed significantly in their optomotor performance compared with controls. Reduced OMR of heavily infected minnows was positively correlated with reduction in minnow activity. In contrast, reduced OMR in lightly infected minnows was independent of host activity and was likely associated with the rapid development of parasite larvae within the optic tecta. The nonlinear relationship between parasite intensity and effect on host behavior was consistent with an earlier study, but the underlying mechanisms producing this pattern are unknown.