Regulation of Anguillicola crassus (Nematoda) infrapopulations in their definitive host, the European eel, Anguilla anguilla

The parasitic nematode Anguillicola crassus was recently introduced into populations of the European eel, Anguilla anguilla. We investigated, under experimental conditions, the regulation of A. crassus infrapopulations. We tested the effects of (1) the resource-limited habitat of the parasite and (2) the coexistence of several developmental stages in its niche (the swim-bladder) on the composition of the infrapopulations. The results revealed that the respective effects of these factors differed substantially during the course of the infection. Third-stage larvae (L3s) establishment would not be constrained by the size of the swim-bladder. Their moult to fourth-stage larvae (L4s) would be accelerated as the number of L3s increased. The moulting time of L4s to adults would be reduced by males and would be constrained by the size of the swim-bladder. However, the moult of L4s to adults and their further development would be synchronized with those of the opposite sex. At the time of mating, the number of males and the body weight of adults would depend on the size of the swim-bladder. Soon after the laying of eggs, the developmental constraint on the late L3s would decrease. When adults die, constraints would cease and late larval stages would moult to become adults.     

Humoral immune response of European eel Anguilla anguilla experimentally infected with Anguillicola crassus

A humoral immune response of the European eel Anguilla anguilla elicited by an experimental infection was demonstrated for the first time against the swimbladder nematode Anguillicola crassus. Eels were experimentally infected once or repeatedly and the antibody response was observed over a period of 325 d. Specific antibodies against A. crassus in the peripheral blood of the eels were measured using an ELISA and the immunoblot technique. Anti-A. crassus antibodies were first observed 8 wk post infection, and appeared to be independent of both the number of infective third stage larvae (L3) administered and the frequency of administration. However, individual eels showed great differences in the course of the antibody response. The late appearance of antibodies in the peripheral blood supports the hypothesis that not the invading L3 but rather the adult parasites elicit the production of specific antibodies. A stage-specific antibody response against the L3 was not observed. Main antigens are located in the body wall, especially in the gelatinous outer cuticle, of adult A. crassus.     

Biased sex ratio in the European eel (Anguilla anguilla) swim-bladder parasite Anguillicola crassus, experimentally induced by 11-ketotestosterone

Parasites are intimately connected to the host in which they live, and some may be affected by the polluted environment of their host. The present study describes the effect of a steroid hormone (11-ketotestosterone) on the sex ratio of the invasive hematophagous nematode Anguillicola crassus Kuwahara, Niimi & Itagaki, 1974, when experimentally injected to European eels, Anguilla anguilla. Our results showed that this steroid induced a significant male-biased ratio in the nematode A. crassus infrapopulations, suggesting that the presence of endocrine disruptors in the environment may lead to skewed sex ratios among parasites.     

Histopathological changes in the swimbladder wall of the European eel Anguilla anguilla due to infections with Anguillicola crassus

The histopathological changes in swimbladders of European eels naturally and experimentally infected with Anguillicola crassus were studied using transmission and scanning electron microscopy. During the course of probably several infections swimbladders undergo characteristic changes. In addition to the thickening of the entire swimbladder wall, and to the folded internal surface of this organ, inflammation, migration of white blood cells, fibrosis and changes in the epithelial cells are frequently seen. Epithelial cells tend to proliferate heavily and form hyperplastic tissues; these processes are accompanied by changes in the internal structure of the cells. The normally cubic cells become spherical or columnar and form folds facing the lumen of the swimbladder. As a consequence, most of these cells lose contact with the blood vessels and show no strict polarity. In heavily affected swimbladders the basal labyrinth of the epithelial cells is reduced, i.e. becomes shorter and less densely packed. The lamina propria shows severe fibrosis with infiltration of white blood cells. Larvae of A. crassus, inhabiting the wall of the swimbladder, were found to be surrounded by cell debris, but this local necrosis does not affect the entire swimbladder in its overall structure. These histological findings can partly explain changes in the gas composition in eels infected with A. crassus.     

Prevalence and abundance of Anguillicola crassus in the European eel (Anguilla anguilla) at a thermal discharge site on the Swedish coast

The accidental introduction and establishment of a population of the sanuivorous swimbladder nematode, Anguillicola crassus , in eels in an area of the Baltic Sea off the coast of Sweden that is affected by thermal water discharge was studied bi-annually over a 5-year period. In 1987, none of 387 eels examined was found to be affected. The first case of infection was recorded in 1988 and within 3 years the prevalence of infection had increased to about 60 %. By 1991, this was the most heavily infested area in Swedish waters. Whereas for yellow eels in the surrounding archipelago records have been kept only sporadicall since the infestation became established, in 1989–91 it was observed that both the prevalence and the abundance of infection were significantly higher in spring than during the second half of the year. When the frequency distributions of parasites for this period 1989–91 are compared, it is evident that this seasonality is connected with an autumn increase in the proportion of eels free from infestation, rather than a reduced proportion of those heavily infected. Furthermore, no seasonality was noted in the proportion of developmental stages of the parasite. Consequently, the observed seasonality is suggested mainly to be an effect of immigrating heathy eels.     

Evaluation of an ELISA and immunoblotting for studying the humoral immune response in Anguillicola crassus infected European eel Anguilla anguilla

The applicability of an enzyme-linked immunosorbent assay (ELISA) for the detection of anguillicolosis in feral eels was examined using a crude antigen preparation from the body wall of adult Anguillicola crassus. The screening consisted of samples from 100 feral European eels Anguilla anguilla. As a reference the actual status of infection was determined by dissection of the eels' swim-bladders. The ELISA results were compared with a background value calculated from the results obtained from 43 non-infected farm eels. The screened samples had a high prevalence of A. crassus (83 %); however, the specificity and the negative predictive value of the ELISA were low compared to the high positive predictive value. Nonetheless, the reproducibility (precision) of the test was satisfactory, and for the non-infected reference group specificity was 97.7 %. Although the ELISA, as used in the present study, is not applicable for diagnostic purposes, it represents a useful tool for the investigation of the specific humoral immune response of eels against A. crassus under controlled experimental conditions. Immunoblots using crude antigen preparations from different parts of adult A. crassus as well as a crude somatic third-stage (L3) antigen preparation illustrated that only antigens associated with the body wall of adult A. crassus are potentially suitable for diagnostic purposes. Despite the fact that antibodies against Raphidascaris acus cross-reacted with 3 body wall antigens of A. crassus, the most encouraging results were obtained with the antigen preparation from the outer cuticle of adult A. crassus which yielded a conspicuous, broad band at about 100 kDa.     

Distribution,prevalence and intensity of Anguillicola crassus (Nematoda) infection in Anguilla anguilla in the Republic of Ireland

This study is the first comprehensive documentation of the geographical range of Anguillicola crassus in its host, the European eel Anguilla anguilla, in the Republic of Ireland. The prevalence and intensity of infections across 234 sites and 93 river basins in Ireland comprising rivers, lakes and transitional waters (estuaries) were analysed. While only 32% of the river basins were affected by this nematode, they correspond to 74% of the total wetted area. Significant differences in infection levels among water body types were found with lakes and transitional waters yielding the highest values, which can be attributed to the proportions of juvenile (total length, L_T < 300 mm) A. anguilla caught. There were no significant differences in infection levels between water body types for adult A. anguilla or between sexes for any water body type. Prevalence was significantly lower in juvenile compared with adult A. anguilla captured in rivers and a positive correlation between infection levels and host size‐classes was found. Future efforts should focus on monitoring the spread of A. crassus infections and assessing the swimbladder health of A. anguilla in Ireland.     

Vaccination of eels (Anguilla japonica and Anguilla anguilla) against Anguillicola crassus with irradiated L3

The original host of the swimbladder nematode Anguillicola crassus, the Japanese eel (Anguilla japonica) and the recently colonized European eel (Anguilla anguilla) were immunized with 40 irradiated (500 Gy) 3rd-stage larvae (L3) of this parasite and challenged with an infection of 40 normal L3. The immunization induced a significant reduction of the number of adult worms developing from the challenge infection in A. japonica, but not in A. anguilla. The induced resistance (calculated using the relation of the number of adult worms in immunized eels and in non-immunized control eels) in A. japonica was 87.3%+/-30.4%. Following a single infection, the percentage of adult worms found in A. japonica was lower as compared to A. anguilla, and the few adult worms were much smaller, revealing a lower susceptibility of A. japonica to A. crassus in comparison to A. anguilla. Both eel species developed an antibody response against A. crassus, but the level of antibody responses was not positively correlated with the protection against infection, suggesting that the antibody response is not a key element in resistance of eels against A. crassus. This study suggests that the original host of A. crassus is able to mount efficient protective immune responses against its parasite, whereas the newly acquired host seems to lack this ability.     

Contents of several inorganic substances in European eel infected and uninfected by Anguillicola crassus (Nematoda).

The content of 5 macroelements and 5 microelements were analyzed using the atomic absorption method in muscle samples of European eels infected and uninfected by Anguillicola crasus. The mean contents of these substances in infected eels were statistically highly significantly lower in Ca, P, Fe, Mn, but only statistically significantly lower in Na, Mg, Zn and Cu as compared to uninfected fishes. These differences are discussed in relation to hematophagus feeding and pathogenity of the parasite.     

Physiological responses to acute temperature increase in European eels Anguilla anguilla infected with Anguillicola crassus

The swimbladder parasite, Anguillicola crassus has infected, and spread rapidly, through European eel Anguilla anguilla (L.) populations over the past 20 to 25 yr. Our aim in the present studies was to elucidate whether the presence of A. crassus in these eels alters their rapid physiological responses to an acute temperature increase, compared to the response of uninfected fish. Both infected and uninfected fish showed significant increases in plasma cortisol after 2 h at a raised environmental temperature with increased plasma glucose after 6 h. However, infected eels exhibited a slight lag in glucose mobilisation, which may be due to the metabolic cost of harbouring a sanguiverous parasite. Both infected and uninfected fish showed a significant increase in haematocrit after 6 h of temperature elevation, but only uninfected fish exhibited a significant increase in haemoglobin at this point. However, there were no significant changes in mean erythrocyte haemoglobin concentration in either group. Our results suggest that acute temperature increase alone is unlikely to cause significant mortality of A. crassus-infected European eels; however, the effects of chronic increases in temperature in combination with other factors such as toxicants and hypoxia requires examination.     

Ichthyophthirius multifiliis (Ciliophora) exit from gill epithelium

The first change in the sequence of morphological events occurring as fully developed Ichthyophthirius multifiliis trophonts spontaneously left gill epithelium or as younger trophonts departed, following experimentally induced death of the fish, was the separation of parasites from overlying host cells. Discharge of contractile vacuoles may have played a role in this process. Spaces then appeared between host cells, and individual epithelial cells became vacuolated. Finally, the epithelium ruptured and the parasites swam free. In induced exit after three days residence in the host, departure of the trophont was evident only after autolysis of epithelium had occurred. Induced departure of trophonts after four days residence was more rapid, suggesting an active role for the parasite in exit. Changes in parasite and epithelium observed in induced exit were similar to those in spontaneous departure after five days residence.     

Pop up satellite tags impair swimming performance and energetics of the European eel (Anguilla anguilla)

Pop-up satellite archival tags (PSATs) have recently been applied in attempts to follow the oceanic spawning migration of the European eel. PSATs are quite large, and in all likelihood their hydraulic drag constitutes an additional cost during swimming, which remains to be quantified, as does the potential implication for successful migration. Silver eels (L_T = 598.6±29 mm SD, N = 9) were subjected to swimming trials in a Steffensen-type swim tunnel at increasing speeds of 0.3–0.9 body lengths s⁻¹, first without and subsequently with, a scaled down PSAT dummy attached. The tag significantly increased oxygen consumption (MO₂) during swimming and elevated minimum cost of transport (COT_min) by 26%. Standard (SMR) and active metabolic rate (AMR) as well as metabolic scope remained unaffected, suggesting that the observed effects were caused by increased drag. Optimal swimming speed (U _opt) was unchanged, whereas critical swimming speed (U _crit) decreased significantly. Swimming with a PSAT altered swimming kinematics as verified by significant changes to tail beat frequency (f), body wave speed (v) and Strouhal number (St). The results demonstrate that energy expenditure, swimming performance and efficiency all are significantly affected in migrating eels with external tags.     

The swimbladder nematode Anguillicola crassus in the European eel Anguilla anguilla and the Japanese eel Anguilla japonica: differences in susceptibility and immunity between a recently colonized host and the original host

The swimbladder nematode Anguillicola crassus originates from Asia where it is a parasite of the Japanese eel Anguilla japonica. After its introduction to Europe about 25 years ago, the parasite spread rapidly within the indigenous populations of the European eel Anguilla anguilla and subsequently the prevalence and mean intensity appeared to stabilize. Under experimental and aquaculture conditions the na?ve new host appears to be more susceptible to A. crassus compared to the original host. Both eel species develop a immune response against A. crassus. The antibody response is well characterized for the European eel, but poorly characterized for the Japanese eel. It remains unclear if antibodies have any protective function against A. crassus. Encapsulation of larvae of A. crassus can be observed in naturally infected European eels. However, encapsulation of larvae following experimental infection has not been detected in European eels, but only in Japanese eels. Reinfection experiments and intraperitoneal injection of A. crassus homogenates failed to demonstrate the development of acquired immunity in European eels. Immunization with irradiated third stage larvae provided preliminary evidence for acquired immunity against A. crassus in the Japanese eel, but not in the European eel.     

Ichthyophthirius multifiliis (Ciliophora) Invasion of Gill Epithelium1

The sequence of morphologic events associated with Ichthyophthirius rnultifliis invasion of gill epithelium began in the theront with differentiation of secretory mucocysts and the perforatorium. After escaping from the cyst the theront, which stained intensely with Mallory' stain, sought a host. As it approached the host epithelium, the contents of the mucocysts were discharged, enveloping the ciliate in sticky material, which made initial contact with the host epithelium. Rapid penetration by the theront caused disruption of one or more host cells and resulted in a focal necrosis associated with the anterior margin of the ciliate. Within five minutes postexposure, the parasite completed its invasion of the epithelial layer and stained less intensely. The remnants of host cells disrupted during its entry surrounded the trophont until they were ingested by the parasite. Within 40 min postexposure, synthetic activity of the parasite appeared to increase as evidenced by an abundance of organelles, particularly ribosomes and crystalline mucocysts. At this point, the overlying host epithelium appeared normal.     

A pseudocystic lipoma in the European eel (Anguilla anguilla)

This is the first report of lipoma in the European eel Anguilla anguilla. In a single eel that was obtained from a polyculture fish farm in northern Egypt, a yellowish swelling (10 mm in diameter) was observed near the tip of the lower lip. The tumor was composed of mature lipocytes that occasionally fused into an unlined cystic space.     

Ichthyophthirius multifiliis (Ciliophora) Exit from Gill Epithelium1

The first change in the sequence of morphological events occurring as fully developed Ichthyophthirius multifiliis trophonts spontaneously left gill epithelium or as younger trophonts departed, following experimentally induced death of the fish, was the separation of parasites from overlying host cells. Discharge of contractile vacuoles may have played a role in this process. Spaces then appeared between host cells, and individual epithelial cells became vacuolated. Finally, the epithelium ruptured and the parasites swam free. In induced exit after three days residence in the host, departure of the trophont was evident only after autolysis of epithelium had occurred. Induced departure of trophonts after four days residence was more rapid, suggesting an active role for the parasite in exit. Changes in parasite and epithelium observed in induced exit were similar to those in spontaneous departure after five days residence.     

Ichthyophthirius multifiliis (Ciliophora) Development in Gill Epithelium1

ABSTRACT. The development of Ichthyophthirius multifiliis trophonts in gill epithelium of channel catfish was studied on the first five days post-exposure (PE) by light and electron microscopy. Trophonts increased in average diameter from 48 μm at one day PE to 248 μm at five days PE. Although theronts invaded any part of the gill epithelium, at three days PE most parasites were found adjacent to major blood vessels, particularly the afferent vessel. From one to three days PE, 70–90% of trophonts were immediately adjacent to gill epithelium, but at four and five days PE only 50% of trophonts were closely apposed by host epithelial cells. Notable ultrastructural changes in the trophonts over the five-day period occurred in the mucocysts and lipid inclusions, both of which increased markedly in number. The few crystalline mucocysts present at one day PE were near the cell periphery but rarely attached to the plasma membrane. At three days PE, crystalline mucocysts were significantly more abundant, and at five days PE, they occurred in large numbers throughout the cytoplasm as well as at the periphery. At three days PE, secretory mucocysts were first observed. Contractile vacuoles were more prominent at five days PE than earlier in development. Development of mucocysts and lipid reserves is probably essential to survival and reproduction of the ciliate after it leaves the host.