MOLECULAR IDENTIFICATION OF JUVENILE NEOECHINORHYNCHUS SPP. (PHYLUM: ACANTHOCEPHALA) INFECTING OSTRACOD AND SNAIL HOSTS PROVIDES INSIGHT INTO ACANTHOCEPHALAN HOST USE.

The role of invertebrates in some acanthocephalan life cycles is unclear because juvenile acanthocephalans are difficult to identify to species using morphology. Most reports suggest acanthocephalans from turtle definitive hosts use ostracods as intermediate hosts and snails as paratenic hosts. However, laboratory studies of the life cycle suggest that ostracods and snails are both required hosts in the life cycle. To elucidate the role of ostracods and snails in acanthocephalan life cycles better, we collected 558 freshwater snails of 2 species, including Planorbella cf. Planorbella trivolvis and Physa acuta, from 23 wetlands in Oklahoma, U.S.A., and examined them for acanthocephalan infections. Additionally, we examined 37,208 ostracods of 4 species, Physocypria sp. (morphotype 1), Cypridopsis sp., Stenocypris sp., and Physocypria sp. (morphotype 2) for juvenile acanthocephalans from 2 wetlands in Oklahoma. Juvenile acanthocephalans were morphologically characterized, and the complete internal transcribed spacer (ITS) region of nuclear rDNA was sequenced from acanthocephalans infecting 11 ostracod and 13 snail hosts. We also sampled 10 red-eared slider turtles, Trachemys scripta elegans, and 1 common map turtle, Graptemys geographica, collected from Oklahoma, Arkansas, and Texas and recovered 1,854 adult acanthocephalans of 4 species. The ITS of 17 adult acanthocephalans of 4 species from turtle hosts were sequenced and compared to juvenile acanthocephalan sequences from ostracod and snail hosts from this study and GenBank to determine conspecificity. Of the 23 locations sampled for snails, 7 (30%) were positive for juvenile acanthocephalans in the genus Neoechinorhynchus. The overall prevalence and mean intensity of acanthocephalans in Planorbella cf. P. trivolvis and P. acuta were 20% and 2 (1-6) and 2% and 1 (1), respectively. In contrast, only 1 of 4 species of ostracods, Physocypria sp. (morphotype 1), was infected with larval/juvenile Neoechinorhynchus spp. with an overall prevalence of 0.1% and a mean intensity of 1 (1-2). Although 4 species of acanthocephalans infected turtle definitive hosts, including Neoechinorhynchus chrysemydis, Neoechinorhynchus emydis, Neoechinorhynchus emyditoides, and Neoechinorhynchus pseudemydis, all the ITS sequences from cystacanths infecting snail hosts were conspecific with N. emydis. In contrast, the ITS sequences from larval/juvenile acanthocephalans from ostracods were conspecific with 2 species of acanthocephalans from turtles (N. emydis and N. pseudemydis) and 1 species of acanthocephalan from fish (Neoechinorhynchus cylindratus). These results indicate that N. emydis infects freshwater snails, whereas other species of Neoechinorhynchus appear not to infect snail hosts. We document new ostracod and snail hosts for Neoechinorhynchus species, including the first report of an ostracod host for N. pseudemydis, and we provide novel molecular barcodes that can be used to determine larva, juvenile, and adult conspecificity of Neoechinorhynchus species.

Histopathology of lesions caused by Pseudocorynosoma constrictum (Acanthocephala: Polymorphidae) in the ileum of blue-winged teal.

Pseudocorynosoma constrictum (Van Cleave, 1918) is a polymorphid acanthocephalan that attaches to the digestive tract of waterfowl to complete its life cycle, causing severe histological damage to its definitive avian hosts. In the present study, we present a histopathological analysis of the lesions that P. constrictum induced in the layers of the ileum of the blue-winged teal Anas discors. The results revealed that worms insert the attachment structures into the inner gut muscular layer, which causes substantial swelling, haemorrhaging and necrosis in the tissue near the parasite's proboscis. We also observed that the number of parasites attached to the tissue can obstruct the intestinal lumen; in the most serious case, we observed more than 30 parasites penetrating completely the walls of the bird intestine.

Acanthocephalan infection patterns in amphipods: a reappraisal in the light of recently discovered host cryptic diversity.

Amphipods are model species in studies of pervasive biological patterns such as sexual selection, size assortative pairing and parasite infection patterns. Cryptic diversity (i.e. morphologically identical but genetically divergent lineages) has recently been detected in several species. Potential effects of such hidden diversity on biological patterns remain unclear, but potentially significant, and beg the question of whether we have missed part of the picture by involuntarily overlooking the occurrence and effects of cryptic diversity on biological patterns documented by previous studies. Here we tested for potential effects of cryptic diversity on parasite infection patterns in amphipod populations and discuss the implications of our results in the context of previously documented host-parasite infection patterns, especially amphipod-acanthocephalan associations. We assessed infection levels (prevalence and abundance) of 3 acanthocephalan species (Pomphorhynchus laevis, P. tereticollis and Polymorphus minutus) among cryptic lineages of the Gammarus pulex/G. fossarum species complex and G. roeseli from sampling sites where they occur in sympatry. We also evaluated potential differences in parasite-induced mortality among host molecular operational taxonomic units (MOTUs)-parasite species combinations. Acanthocephalan prevalence, abundance and parasite-induced mortality varied widely among cryptic MOTUs and parasite species; infection patterns were more variable among MOTUs than sampling sites. Overall, cryptic diversity in amphipods strongly influenced apparent infection levels and parasite-induced mortality. Future research on species with cryptic diversity should account for potential effects on documented biological patterns. Results from previous studies may also need to be reassessed in light of cryptic diversity and its pervasive effects.

Helminthes in the Small Intestine of the Common Eider (Somateria mollissima) from the Eastern Murman: Impact on the Host Digestive Activity and Physiological State.

In this ecological and physiological study of the common eider (Somateria mollissima) nesting on the coast of Eastern Murman, the species composition of the bird helminth fauna, as well as the infection quantitative parameters, were determined. The common eider small intestine proved to be infected with trematodes of the genus Microphallus; three species of cestodes, namely, Lateriporus teres (Cestoda: Dilepididae), Fimbriarioides intermedia (Cestoda: Hymenolepididae), and Microsomacanthus diorchis (Cestoda: Hymenolepididae); and one species of acanthocephalan, Polymorphus phippsi (Palaeacanthocephala: Polymorphidae). At the sites of F. intermedia and M. diorchis locations within the intestine, the protease activity was reduced while in the foci infected with acanthocephalan P. phippsi, it was, on the contrary, increased. Glycosidase activity in the intestinal mucosa was reduced as compared to the control in birds infected by the cestodes M. diorchis. Hematological indices of the infected individuals were higher than the control parameters.

Distribution of the acanthocephalan Neoechinorhynchus buttnerae and semiquantitative analysis of histopathological damage in the intestine of tambaqui (Colossoma macropomum).

In this paper, we have described for the first time a semiquantitative method to evaluate histopathological damage, taking the degree of Neoechinorhynchus buttnerae attachment to the intestinal wall of the tambaqui (Colossoma macropomum), an important species in Brazilian aquaculture, into account. Twelve specimens of tambaqui were collected from a fish farm. Their bowels were removed and divided into seven morphologically distinct portions according to density and distribution of the parasite studies. Fragments from each fraction were histologically processed and analyzed. There was a clear preference on the part of N. buttnerae for the intermediate regions of the intestinal tube, where the highest densities were recorded. The intensity of damage to the host, estimated by calculating the Histopathological Alteration Index (HAI), showed severe and irreversible changes only where the parasite had its proboscis penetrated into the intestine wall.

Mass Mortality Attributed to Acanthocephaliasis at a Gull-billed Tern (Gelochelidon nilotica) Colony in Coastal California, USA.

From 12 May 2013 to 29 May 2013, the Gull-billed Tern (Gelochelidon nilotica) colony at the San Diego Bay National Wildlife Refuge, California, US, experienced a mass die-off of at least 92 adults, representing 71-92% of the breeding population on the US west coast. Cause of death was determined to be peritonitis due to perforations of the intestine by a large quantity of acanthocephala (Profilicollis [=Polymorphus] altmani). This is a unique report of P. altmani infecting G. nilotica, and a report of a great impact to a tern population in southern California. Mole crabs (Emerita analoga), the intermediate host for P. altmani and a major component of the Gull-billed Tern diet in San Diego, were found in the stomachs of necropsied terns along with cystacanths, and are the presumed source of the parasite infection. The tern's dietary reliance upon mole crabs likely amplified parasite transmission and infection. We suggest additional research to determine factors that influence parasite infection of intermediate and definitive hosts, particularly mole crabs, given that they are a vital resource for migrating birds within the coastal zone.

Temperature rise and parasitic infection interact to increase the impact of an invasive species.

Invasive species often detrimentally impact native biota, e.g. through predation, but predicting such impacts is difficult due to multiple and perhaps interacting abiotic and biotic context dependencies. Higher mean and peak temperatures, together with parasites, might influence the impact of predatory invasive host species additively, synergistically or antagonistically. Here, we apply the comparative functional response methodology (relationship between resource consumption rate and resource supply) in one experiment and conduct a second scaled-up mesocosm experiment to assess any differential predatory impacts of the freshwater invasive amphipod Gammarus pulex, when uninfected and infected with the acanthocephalan Echinorhynchus truttae, at three temperatures representative of current and future climate. Individual G. pulex showed Type II predatory functional responses. In both experiments, infection was associated with higher maximum feeding rates, which also increased with increasing temperatures. Additionally, infection interacted with higher temperatures to synergistically elevate functional responses and feeding rates. Parasitic infection also generally increased Q10 values. We thus suggest that the differential metabolic responses of the host and parasite to increasing temperatures drives the synergy between infection and temperature, elevating feeding rates and thus enhancing the ecological impact of the invader.

[Infection of the pacific saury Cololabis saira by acanthocephalans in the Kuril Islands area].

The Pacific saury Cololabis saira (Brevoort, 1856) is one of the important target species of commercial fisheries. Food manufacturers and consumers encounter problems due to the infection of the saury by acanthocephalans, which are quite difficult to clean out completely during on-board catch processing. Infection of C. saira was not studied on a regular basis, therefore, our knowledge about the parasites of saury is fragmentary. This paper contains infection indices (only acanthocephalans) of the Pacific saury caught in the Kuril Islands area (Russian Exclusive Economic Zone) in 2015.

Host condition and accumulation of metals by acanthocephalan parasite Echinorhynchus gadi in cod Gadus morhua from the southern Baltic Sea.

In this study, we analyzed the relationship between concentration of metals in the host-parasite system (cod - acanthocephalan Echinorhynchus gadi) and Fulton's condition factor (FCF) of the host. The relationship between metal (Ca, Cd, Cr, Cu, Fe, Hg, K, Mg, Mn, Na, Pb, Sr, Zn) concentrations in E. gadi and cod tissues was expressed as a bioconcentration factor (BCF), the ratio of the concentration in the parasite tissue to that in host tissues. Acanthocephalans accumulated mainly toxic metals (Cd, Pb), as well as Sr, Ca, Na. Cadmium showed the highest bioconcentration in parasites (BCF >200) compared to fish muscle. Significant negative correlation was detected between FCF and the concentration of Cd and Hg in cod liver. In contrast, FCF was positively correlated with the concentration of Hg in acanthocephalan tissues.

Development and Validation of a Biodynamic Model for Mechanistically Predicting Metal Accumulation in Fish-Parasite Systems.

Because of different reported effects of parasitism on the accumulation of metals in fish, it is important to consider parasites while interpreting bioaccumulation data from biomonitoring programmes. Accordingly, the first step is to take parasitism into consideration when simulating metal bioaccumulation in the fish host under laboratory conditions. In the present study, the accumulation of metals in fish-parasite systems was simulated by a one-compartment toxicokinetic model and compared to uninfected conspecifics. As such, metal accumulation in fish was assumed to result from a balance of different uptake and loss processes depending on the infection status. The uptake by parasites was considered an efflux from the fish host, similar to elimination. Physiological rate constants for the uninfected fish were parameterised based on the covalent index and the species weight while the parameterisation for the infected fish was carried out based on the reported effects of parasites on the uptake kinetics of the fish host. The model was then validated for the system of the chub Squalius cephalus and the acanthocephalan Pomphorhynchus tereticollis following 36-day exposure to waterborne Pb. The dissolved concentration of Pb in the exposure tank water fluctuated during the exposure, ranging from 40 to 120 µg/L. Generally, the present study shows that the one-compartment model can be an effective method for simulating the accumulation of metals in fish, taking into account effects of parasitism. In particular, the predicted concentrations of Cu, Fe, Zn, and Pb in the uninfected chub as well as in the infected chub and the acanthocephalans were within one order of magnitude of the measurements. The variation in the absorption efficiency and the elimination rate constant of the uninfected chub resulted in variations of about one order of magnitude in the predicted concentrations of Pb. Inclusion of further assumptions for simulating metal accumulation in the infected chub led to variations of around two orders of magnitude in the predictions. Therefore, further research is required to reduce uncertainty while characterising and parameterising the model for infected fish.

Histopathology of Catostomus commersoni (White Sucker) Infected with the Acanthocephalan Pomphorhynchus bulbocolli.

This study was undertaken to assess the damage caused by Pomphorhynchus bulbocolli to Catostomus commersoni (white sucker). Three specimens of C. commersoni were collected during early September 2014 via gill net from Canadarago Lake (Otsego County, New York), then dissected and examined for intestinal parasites. One C. commersoni , collected from a tributary of Otsego Lake (Otsego County, New York), was used as a control in this study because it was not infected with intestinal helminths. Upon dissection, damage to the fish intestine was macroscopically visible, with the intestine perforated when infected with P. bulbocolli. Intestines observed to be infected with P. bulbocolli were opened with a longitudinal incision and fixed in neutral buffered formalin with the acanthocephalans remaining attached. Histological sections of intestine with P. bulbocolli attached were compared with histological sections of intestine in which no worms were present. Examination of sections revealed full penetration of the intestinal wall and tissue destruction to the mucosa, submucosa, stratum compactum, and circular and longitudinal muscle layers, as well as an extensive host immune response in the form of proliferation of cells at the sites of wounds. While these results were consistent with previous histopathological studies on this host and parasite species, the occurrence of pockets of hyaline degeneration in the muscularis reported here is a new finding for this host-parasite system, and it appears to be quite rare in the parasitological literature. It is hypothesized that the presence of hyaline degeneration may be related to secretion of trypsinlike proteins from the presoma of the acanthocephalan, a phenomenon suggested previously for the congener Pomphorhynchus laevis . The host-parasite interaction involving physical damage, secretion of enzymes, and an extensive host immune response may be the cause of the damage, but further research is needed to investigate the nature of these interactions.

Parasite-induced alteration of plastic response to predation threat: increased refuge use but lower food intake in Gammarus pulex infected with the acanothocephalan Pomphorhynchus laevis.

Larvae of many trophically-transmitted parasites alter the behaviour of their intermediate host in ways that increase their probability of transmission to the next host in their life cycle. Before reaching a stage that is infective to the next host, parasite larvae may develop through several larval stages in the intermediate host that are not infective to the definitive host. Early predation at these stages results in parasite death, and it has recently been shown that non-infective larvae of some helminths decrease such risk by enhancing the anti-predator defences of the host, including decreased activity and increased sheltering. However, these behavioural changes may divert infected hosts from an optimal balance between survival and foraging (either seeking food or a mate). In this study, this hypothesis was tested using the intermediate host of the acanthocephalan parasite Pomphorhynchus laevis, the freshwater amphipod Gammarus pulex. We compared activity, refuge use, food foraging and food intake of hosts experimentally infected with the non-infective stage (acanthella), with that of uninfected gammarids. Behavioural assays were conducted in four situations varying in predation risk and in food accessibility. Acanthella-infected amphipods showed an increase in refuge use and a general reduction in activity and food intake. There was no effect of parasite intensity on these traits. Uninfected individuals showed plastic responses to water-borne cues from fish by adjusting refuge use, activity and food intake. They also foraged more when the food was placed outside the refuge. At the intra-individual level, refuge use and food intake were positively correlated in infected gammarids only. Overall, our findings suggest that uninfected gammarids exhibit risk-sensitive behaviour including increased food intake under predation risk, whereas gammarids infected with the non-infective larvae of P. laevis exhibit a lower motivation to feed, irrespective of predation risk and food accessibility.

[Dependence of the structure of the capsule surrounding the acanthocephalan Corynosoma strumosum on the species of its natural paratenic host].

The fine and ultrafine structure of the capsules surrounding the acanthocephalan Corynosoma strumosum in natural paratenic hosts of three species--the whitespotted greenling Hexagrammos stelleri, Steller's sculpin Myoxocephalus stelleri, and the saffron cod Eleginus gracilis--was studied. The results of this study, together with earlier data, support the hypothesis that the structure of the capsule depends on the paratenic host species. Three types of capsules differing in the ratio of fibroblasts and inflammatory cells have been identified: fibroblastic (in Japanese smelt, rainbow smelt, and saffron cod), intermediate (in whitespotted greenling), and leukocytic (in Steller's sculpin and the yellowfin sole). It was assumed that the structure of the capsule is determined by the degree of mutual adaptation of organisms in a given host-parasite system.

Acanthocephalan parasites: help or burden in gammarid amphipods exposed to cadmium?

We investigated the influence of the acanthocephalan parasite Polymorphus minutus on the mortality of its intermediate host, Gammarus roeseli, exposed to cadmium, by the measure of LC(50-96h) values as well as the bioaccumulation of cadmium both in the host and in its parasite. LC(50) results revealed that infected G. roeseli males died less under cadmium stress than uninfected ones; while the converse has been observed in females. Cadmium resistance of infected males could be explained by a weaker bioconcentration factor (BCF) than in females. The lower BCF in infected individuals was closely related with an uptake of cadmium by P. minutus in its host. Nevertheless, although infected females had both weaker BCF and cadmium concentration in their body, the presence of P. minutus did not induce lower mortality than uninfected females. On the contrary, their sensitivity to cadmium was increased by the presence of P. minutus. We discuss the hypothesis that differences of mortality between uninfected and infected gammarids could be explained by a difference of cadmium bioconcentration in host, and by the cadmium bioaccumulation in the parasite. Indeed, results suggested that P. minutus could help G. roeseli to face with stress, what contributed to keep the host alive and favour the parasite transmission.

Assessing host-parasite specificity through coprological analysis: a case study with species of Corynosoma (Acanthocephala: Polymorphidae) from marine mammals.

In this paper we report an investigation of the utility of coprological analysis as an alternative technique to study parasite specificity whenever host sampling is problematic; acanthocephalans from marine mammals were used as a model. A total of 252 scats from the South American sea lion, Otaria flavescens, and rectal faeces from 43 franciscanas, Pontoporia blainvillei, from Buenos Aires Province, were examined for acanthocephalans. Specimens of two species, i.e. Corynosoma australe and C. cetaceum, were collected from both host species. In sea lions, 78 out of 145 (37.9%) females of C. australe were gravid and the sex ratio was strongly female-biased. However, none of the 168 females of C. cetaceum collected was gravid and the sex ratio was not female-biased. Conversely, in franciscanas, 14 out of 17 (82.4%) females of C. cetaceum were gravid, but none of 139 females of C. australe was, and the sex ratio of C. cetaceum, but not that of C. australe, was female-biased. In putative non-hosts, the size of worms was similar to that from specimens collected from prey. Results suggest that both acanthocephalans contact sea lions and franciscanas regularly. However, C. australe and C. cetaceum cannot apparently reproduce, nor even grow, in franciscanas and sea lions, respectively. Coprological analysis may represent a useful supplementary method to investigate parasite specificity, particularly when host carcasses are difficult to obtain.

Structure of capsule around acanthocephalan Corynosoma strumosum from uncommon paratenic hosts-lizards of two species.

Micromorphology and ultrastructure of capsule forming around acanthocephalan Corynosoma strumosum in uncommon paratenic hosts-lizards Lacerta agilis and Lacerta viridis-have been studied. Experimental infestation of the lizards by acanthocephalans obtained from naturally infested sea fishes showed that only small amount of parasites occurred in the intestine of the host was able to migrate into body cavity and to be encapsulated. Micromorphology of capsules of different ages from different species of lizards and micromorphology and ultrastructure of capsules at the age of 1.5 and 10 days appeared to be similar. In the capsule's structure cells of inflammatory rank were prevailing: mononuclear and multinuclear macrophages, eosinophils, and basophils. Fibroblasts were not numerous and were located only in the outer part of a capsule; exocellular collagen fibers were absent. Inflammatory character of capsule confirms the idea that lizards are unsuitable paratenic hosts for corynosomes.

Intestinal immune response of Silurus glanis and Barbus barbus naturally infected with Pomphorhynchus laevis (Acanthocephala).

Immunopathological and ultrastructural studies were conducted on the intestine of barbel Barbus barbus and sheatfish Silurus glanis that were naturally infected with the acanthocephalan Pomphorhynchus laevis. Enteric helminths often cause inflammation of the digestive tract, inducing the recruitment of different types of immune cells at the site of infection. The results of our study clearly demonstrated that mast cells (MC) were the dominant immune cells which occur at the site of inflammation in both hosts. MC were associated with fibroblasts and were found in close proximity to, and inside, the capillaries of the intestine, thus, migration of mast cells via the bloodstream was suggested. Significant degranulation of MC was present. Immunohistochemical staining revealed met-enkephalin and serotonin (5-HT) in intestinal MC of both uninfected and infected barbel and the absence of the antimicrobial peptides piscidin 3 and piscidin 4 in both species. Data are discussed with respect to host immune response to an intestinal helminth and compared with other host-parasite systems.

Co-variation between the intensity of behavioural manipulation and parasite development time in an acanthocephalan-amphipod system.

Pomphorhynchus laevis, a fish acanthocephalan parasite, manipulates the behaviour of its gammarid intermediate host to increase its trophic transmission to the definitive host. However, the intensity of behavioural manipulation is variable between individual gammarids and between parasite populations. To elucidate causes of this variability, we compared the level of phototaxis alteration induced by different parasite sibships from one population, using experimental infections of Gammarus pulex by P. laevis. We used a naive gammarid population, and we carried out our experiments in two steps, during spring and winter. Moreover, we also investigated co-variation between phototaxis (at different stages of infection, 'young' and 'old cystacanth stage') and two other fitness-related traits, infectivity and development time. Three main parameters could explain the parasite intra-population variation in behavioural manipulation. The genetic variation, suggested by the differences between parasite families, was lower than the variation owing to an (unidentified) environmental factor. Moreover, a correlation was found between development rate and the intensity of behavioural change, the fastest growing parasites being unable to induce rapid phototaxis reversal. This suggests that parasites cannot optimize at the same time these two important parameters of their fitness, and this could explain a part of the variation observed in the wild.

Lueheia inscripta (Westrumb, 1821) (Acanthocephala: Plagiorhynchidae) in anurans (Leptodactylidae: Bufonidae) from Mexico.

Juveniles of Lueheia inscripta (Westrumb, 1821 Travassos, 1919 (Acanthocephala: Plagiorhynchidae), an acanthocephalan with six lemnisci, are reported and described from mesenteries of frogs Leptodactylus fragilis Brochi, 1877 and a toad Bufo marinus (Linnaeus, 1758) from Morelos state, Mexico. These are new host records extending the known geographical distribution of this species from Brazil and Puerto Rico to Mexico.

Solearhynchus kostylewi (Meyer, 1932) comb. nov. (acanthocephala: Echinorhynchidae), a rare parasite of Solea solea (pisces: soleidae) in the Gemlik Bay, Sea of Marmara.

Paracanthocephaloides kostylewi (Meyer, 1932), a rare parasite of the common sole Solea solea, previously placed in the Arhythmacanthidae, is transfered to the genus Solearhynchus de Buron & Maillard, 1985 of the family Echinorhynchidae as Solearhynchus kostylewi (Meyer, 1932) new comb., because its proboscis is armed by a single type of hooks, the posterior hook in each row is pinlike and with feebly developed root. The species is redescribed on the basis of newly collected material from the Gemlik Bay, Sea of Marmara. A key to the species of Solearhynchus is provided.