Carotenoid-based colour of acanthocephalan cystacanths plays no role in host manipulation

Manipulation by parasites is a catchy concept that has been applied to a large range of phenotypic alterations brought about by parasites in their hosts. It has, for instance, been suggested that the carotenoid-based colour of acanthocephalan cystacanths is adaptive through increasing the conspicuousness of infected intermediate hosts and, hence, their vulnerability to appropriate final hosts such as fish predators. We revisited the evidence in favour of adaptive coloration of acanthocephalan parasites in relation to increased trophic transmission using the crustacean amphipod Gammarus pulex and two species of acanthocephalans, Pomphorhynchus laevis and Polymorphus minutus. Both species show carotenoid-based colorations, but rely, respectively, on freshwater fish and aquatic bird species as final hosts. In addition, the two parasites differ in the type of behavioural alteration brought to their common intermediate host. Pomphorhynchus laevis reverses negative phototaxis in G. pulex, whereas P. minutus reverses positive geotaxis. In aquaria, trout showed selective predation for P. laevis-infected gammarids, whereas P. minutus-infected ones did not differ from uninfected controls in their vulnerability to predation. We tested for an effect of parasite coloration on increased trophic transmission by painting a yellow-orange spot on the cuticle of uninfected gammarids and by masking the yellow-orange spot of infected individuals with inconspicuous brown paint. To enhance realism, match of colour between painted mimics and true parasite was carefully checked using a spectrometer. We found no evidence for a role of parasite coloration in the increased vulnerability of gammarids to predation by trout. Painted mimics did not differ from control uninfected gammarids in their vulnerability to predation by trout. In addition, covering the place through which the parasite was visible did not reduce the vulnerability of infected gammarids to predation by trout. We discuss alternative evolutionary explanations for the origin and maintenance of carotenoid-based colorations in acanthocephalan parasites.     

Field evidence of host size-dependent parasitism in two manipulative parasites

The distribution of parasites within host natural populations has often been found to be host age-dependent. Host mortality induced by parasites is the commonest hypothesis proposed for explaining this pattern. Despite its potential importance in ecology, the parasitism intensity in relation with the host age has rarely been studied in the field. The 2 manipulative acanthocephalans, Polymorphus minutus and Pomphorhynchus laevis, use the amphipod Gammarus pulex as an intermediate host, and their infection intensity and incidence among G. pulex populations were examined by analyzing 2 large samples of hosts collected in eastern France. Both parasites had low prevalence in the host populations, but their mean abundances were highly related with gammarid age. For the 2 acanthocephalans, results reported a disappearance or an absence of heavily infected hosts in the older host age classes. These results suggested that parasites that alter intermediate host behavior for enhancing their transmission success to the definitive host reduce the survival of their intermediate host. In conclusion, manipulative parasites might act as a mechanism regulating the density of gammarid populations.     

Differential influence of Pomphorhynchus laevis (Acanthocephala) on brain serotonergic activity in two congeneric host species

The physiological mechanisms by which parasites with complex life cycles manipulate the behaviour of their intermediate hosts are still poorly understood. In Burgundy, eastern France, the acanthocephalan parasite Pomphorhynchus laevis inverses reaction to light in its amphipod host Gammarus pulex, but not in Gammarus roeseli, a recent invasive species. Here, we show that this difference in manipulation actually reflects a difference in the ability of the parasite to alter brain serotonergic (5-HT) activity of the two host species. Injection of 5-HT in uninfected individuals of both host species was sufficient to inverse reaction to light. However, a difference in brain 5-HT immunocytochemical staining levels between infected and uninfected individuals was observed only in G. pulex. Local adaptation of the parasite to the local host species might explain its inability to manipulate the behaviour and nervous system of the invasive species.     

Infection with an acanthocephalan manipulates an amphipod's reaction to a fish predator's odours

Many parasites with complex life cycles increase the chances of reaching a final host by adapting strategies to manipulate their intermediate host's appearance, condition or behaviour. The acanthocephalan parasite Pomphorhynchus laevis uses freshwater amphipods as intermediate hosts before reaching sexual maturity in predatory fish. We performed a series of choice experiments with infected and uninfected Gammarus pulex in order to distinguish between the effects of visual and olfactory predator cues on parasite-induced changes in host behaviour. When both visual and olfactory cues, as well as only olfactory cues were offered, infected and uninfected G. pulex showed significantly different preferences for the predator or the non-predator side. Uninfected individuals significantly avoided predator odours while infected individuals significantly preferred the side with predator odours. When only visual contact with a predator was allowed, infected and uninfected gammarids behaved similarly and had no significant preference. Thus, we believe we show for the first time that P. laevis increases its chance to reach a final host by olfactory-triggered manipulation of the anti-predator behaviour of its intermediate host.     

Effects of two acanthocephalan parasites on the fecundity and pairing status of female Gammarus pulex (Crustacea: Amphipoda)

Acanthocephalan parasites are known to alter the reproductive biology and physiology of their hosts in various ways. In this study we investigated the influence of two acanthocephalan parasites, Pomphorhynchus laevis and Polymorphus minutus, on the fecundity and pairing success of female Gammarus pulex. The results show that P. laevis and P. minutus affect female intermediate host reproduction in different ways. Females infected with P. minutus were totally castrated, whereas those infected with P. laevis only showed reduced fecundity. The oocytes of P. laevis-infected females showed a similar structure to those of uninfected females, although infected females had a higher proportion of oocytes that had failed to reach complete maturity. In comparison, the oocytes of P. minutus-infected females demonstrate a clearly altered structure that suggests a major disruption to the process of vitellogenesis. In the field, males paired more frequently with uninfected females than with infected ones, and is a stronger effect for P. minutus-infected females than P. laevis-infected females. We suggest that the difference in pairing success of P. minutus-infected and P. laevis-infected females is a direct result of the different effects that the two parasites have on female fecundity.     

Infection with acanthocephalans increases the vulnerability of Gammarus pulex (Crustacea, Amphipoda) to non-host invertebrate predators

Phenotypic alterations induced by parasites in their intermediate hosts often result in enhanced trophic transmission to appropriate final hosts. However, such alterations may also increase the vulnerability of intermediate hosts to predation by non-host species. We studied the influence of both infection with 3 different acanthocephalan parasites (Pomphorhynchus laevis, P. tereticollis, and Polymorphus minutus) and the availability of refuges on the susceptibility of the amphipod Gammarus pulex to predation by 2 non-host predators in microcosms. Only infection with P. laevis increased the vulnerability of amphipods to predation by crayfish, Orconectes limosus. In contrast, in the absence of refuges, the selectivity of water scorpions, Nepa cinerea, for infected prey was significant and did not differ according to parasite species. When a refuge was available for infected prey, however, water scorpion selectivity for infected prey differed between parasite species. Both P. tereticollis- and P. laevis-infected gammarids were more vulnerable than uninfected ones, whereas the reverse was true of P. minutus-infected gammarids. These results suggest that the true consequences of phenotypic changes associated with parasitic infection in terms of increased trophic transmission of parasites deserve further assessment.     

Do distantly related parasites rely on the same proximate factors to alter the behaviour of their hosts?

Phylogenetically unrelated parasites often increase the chances of their transmission by inducing similar phenotypic changes in their hosts. However, it is not known whether these convergent strategies rely on the same biochemical precursors. In this paper, we explored such aspects by studying two gammarid species (Gammarus insensibilis and Gammarus pulex; Crustacea: Amphipoda: Gammaridae) serving as intermediate hosts in the life cycle of two distantly related parasites: the trematode, Microphallus papillorobustus and the acanthocephalan, Polymorphus minutus. Both these parasite species are known to manipulate the behaviour of their amphipod hosts, bringing them towards the water surface, where they are preferentially eaten by aquatic birds (definitive hosts). By studying and comparing the brains of infected G. insensibilis and G. pulex with proteomics tools, we have elucidated some of the proximate causes involved in the parasite-induced alterations of host behaviour for each system. Protein identifications suggest that altered physiological compartments in hosts can be similar (e.g. immunoneural connexions) or different (e.g. vision process), and hence specific to the host-parasite association considered. Moreover, proteins required to alter the same physiological compartment can be specific or conversely common in both systems, illustrating in the latter case a molecular convergence in the proximate mechanisms of manipulation.     

Altered host behaviour and brain serotonergic activity caused by acanthocephalans: evidence for specificity

Manipulative parasites can alter the phenotype of intermediate hosts in various ways. However, it is unclear whether such changes are just by-products of infection or adaptive and enhance transmission to the final host. Here, we show that the alteration of serotonergic activity is functionally linked to the alteration of specific behaviour in the amphipod Gammarus pulex infected with acanthocephalan parasites. Pomphorhynchus laevis and, to a lesser extent, Pomphorhynchus tereticollis altered phototactism, but not geotactism, in G. pulex, whereas the reverse was true for Polymorphus minutus. Serotonin (5-hydroxytryptamine, 5-HT) injected to uninfected G. pulex mimicked the altered phototactism, but had no effect on geotactism. Photophilic G. pulex infected with P. laevis or P. tereticollis showed a 40% increase in brain 5-HT immunoreactivity compared to photophobic, uninfected individuals. In contrast, brain 5-HT immunoreactivity did not differ between P. minutus-infected and uninfected G. pulex. Finally, brain 5-HT immunoreactivity differed significantly among P. tereticollis-infected individuals in accordance with their degree of manipulation. Our results demonstrate that altered 5-HT activity is not the mere consequence of infection by acanthocephalans but is specifically linked to the disruption of host photophobic behaviour, whereas the alteration of other behaviours such as geotactism may rely on distinct physiological routes.     

The effect of the acanthocephalan parasite Pomphorhynchus laevis on the lipid and glycogen content of its intermediate host Gammarus pulex

Besides conspicuous changes in behaviour, manipulative parasites may also induce subtle physiological effects in the host that may also be favourable to the parasite. In particular, parasites may be able to influence the re-allocation of resources in their own favour. We studied the association between the presence of the acanthocephalan parasite, Pomphorhynchus laevis, and inter-individual variation in the lipid and glycogen content of its crustacean host, Gammarus pulex (Amphipoda). Infected gravid females had significantly lower lipid contents than uninfected females, but there was no difference in the lipid contents of non-gravid females and males that were infected with P. laevis. In contrast, we found that all individuals that were parasitised by P. laevis had significantly increased glycogen contents, independent of their sex and reproductive status. We discuss our results in relation to sex-related reproductive strategies of hosts, and the influence they may have on the level of conflict over energy allocation between the host and the parasite.     

Is the host or the parasite the most locally adapted in an amphipod-acanthocephalan relationship? A case study in a biological invasion context

Manipulative endoparasites with complex life cycles can alter their intermediate host immunity and behaviour in ways that increase survival probability within the host body cavity and enhance successful transmission to the definitive host. These parasitic manipulations are variable among and within parasite species and may result from co-evolutionary processes, in which the parasite is constrained for adaptation to the local intermediate host. Hence, arrival of a new host species in a local host population may promote local parasite maladaptation. This study tested the occurrence of local adaptation in two distantly located populations of the acanthocephalan parasite Pomphorhynchus laevis and its effect on the immunity and behaviour of its gammarid intermediate host Gammarus roeseli. This was done in France (an area for which G. roeseli is a recent invader) and Hungary (an area from which G. roeseli was believed to be native). As expected, we found no alteration in G. roeseli's immune defence and behaviour associated with infection by P. laevis in localities, where the gammarid is invasive. Unexpectedly, we found similar results in Hungarian populations, where the parasite was even more exposed to the host immune response. Whilst these results suggest maladaptation of the parasite to the gammarid in both countries, they also suggest that the gammarid host might be locally adapted to the parasite. Genetic analyses were performed on both the parasite and the host and the results suggest that the two subsets of populations we studied harbour rather isolated host-parasite systems, both probably deriving from a common ancestral population. We propose that G. roeseli is also of recent acquisition in Hungary, and that a recent co-evolutionary history between P. laevis and G. roeseli in association with a long generation time in the parasite has constrained parasite adaptations in Europe or even favoured host adaptation to the parasite.     

Parasitic infection and developmental stability: fluctuating asymmetry in Gammarus pulex infected with two acanthocephalan species

Several studies have reported a negative association between developmental stability and parasitic infection. However, the host-parasite associations examined so far consist only of a limited number of parasite taxa, and developmental stability was appraised on definitive hosts. The present study examines the association between infection by 2 acanthocephalan parasites. Pomphorhynchus laevis and Polymorphus minutus, and the developmental stability of their common intermediate host Gammarus pulex. Developmental stability was estimated from the fluctuating asymmetry (FA) levels of 6 morphological traits. A positive association was found between FA and infection. Infected gammarids tended to be more asymmetrical than the noninfected ones for an index generated by combining FA scores from 2 characters out of the 6 studied, even though no significant relationships were found between FA levels and parasitic loads. The simultaneous presence of both acanthocephalan species in the same host seems to be associated with increased FA levels of gammarids, but this trend was not statistically significant. For the same characters, males exhibited higher levels of FA than females.     

Increased susceptibility to predation and altered anti-predator behaviour in an acanthocephalan-infected amphipod

According to the 'parasitic manipulation hypothesis', phenotypic changes induced by parasites in their intermediate hosts are effective means of increasing trophic transmission to final hosts. One obvious prediction, although seldom tested, is that increased vulnerability of infected prey to an appropriate predator should be achieved by the parasite altering the anti-predator behaviour of its intermediate host. In this study, we tested this prediction using the fish acanthocephalan Pomphorhynchus tereticollis and the freshwater amphipod Gammarus pulex. Firstly, we estimated the relative vulnerability of infected and uninfected gammarids to predation by the bullhead Cottus gobio in the field. Second, we investigated under experimental conditions how two common anti-predator behaviours of aquatic invertebrates, refuge use and short-distance reaction to predator chemical cues, were affected by infection status. We found that the prevalence of infection in the field was 10 times higher among gammarids collected from the stomach contents of bullheads compared with free-ranging individuals collected in the same river. In a microcosm uninfected gammarids, but not infected ones, increased the use of refuge in the presence of a bullhead. Finally, a behavioural experiment using an Y-maze olfactometer showed opposite reactions to predator odour. Whereas uninfected gammarids were significantly repulsed by the chemical cues originating from bullheads, infected ones were significantly attracted to the odour of the predator. Taken together, our results suggest that the alteration of anti-predator behaviour in infected G. pulex might enhance predation by bullheads in the field. Reversing anti-predator behaviour might thus be an efficient device by which parasites with complex life-cycles increase their trophic transmission to final hosts. Further studies should pay more attention to both the increased vulnerability of infected prey to an appropriate predator in the field and the influence of parasitic infection on the anti-predator behaviour of intermediate hosts.     

Pairing success and sperm reserve of male Gammarus pulex infected by Cyathocephalus truncatus (Cestoda: Spathebothriidea)

Manipulative parasites with complex life cycles are known to induce behavioural and physiological changes in their intermediate hosts. Cyathocephalus truncatus is a manipulative parasite which infects Gammarus pulex as intermediate host. G. pulex males display pre-copulatory mate guarding as a response to male-male competition for access to receptive females. In this paper, we tested the influence that C. truncatus-infection might have on male G. pulex sperm number and pairing success. We considered 3 classes of G. pulex males in our experiments: (i) uninfected males found paired in the field, (ii) uninfected males found unpaired in the field, or (iii) infected males found unpaired in the field. Both infected males and uninfected unpaired males paired less with a new female than uninfected paired males did. Furthermore, infected males appear to be at a strong disadvantage when directly competing for females with a healthy rival male, and had fewer sperm in their testes. We discuss the potential effect of male and female mating strategies on such male host mating alteration.     

Observations on the distribution, specificity and pathogenicity of the acanthocephalan Pomphorhynchus laevis (Müller)

The distribution, specificity and pathogenicity of the acanthocephalan Pomphorhynchus laevis (Müller) were studied in the River Avon, Hampshire. The parasite occurred in every fish species, but three categories of hosts could be distinguished. Using growth and maturation as criteria of specificity, only chub Leuciscus cephalus (L.) and barbel Barbus barbus (L.) were recognized as its preferred hosts. Parasites occasionally matured in trout Salmo trutta (L.) and dace Leuciscus leuciscus (L.), but none grew or matured in other host species. Changes in the abundance of P. laevis along the river were related to changes in the abundance of both the intermediate host, Gammarus pulex , and the preferred hosts, and its occurrence in fish to the importance of G. pulex in their diet. In the upper reaches of the River Avon and in other localities populations could be maintained at a low level by parasites maturing in trout, and presence and abundance at any site depended upon the presence and abundance of both intermediate and definitive hosts especially and upon the dietary preferences of the latter. The absence of P. laevis from many parts of Britain is nevertheless still inexplicable. The parasite caused local damage to the intestinal wall of fish, the extent of which varied from species to species, but did not affect host growth rate or cause direct mortality and P. laevis cannot be regarded as an important pathogen in the River Avon or any other British River.     

Interspecific antagonism and virulence in hosts exposed to two parasite species

Co-infection of host organisms by multiple parasite species has evolutionary consequences for all participants in the symbiosis. In this study, we co-exposed aquatic-snails (Biomphalaria glabrata) to two of their trematode parasites, Schistosoma mansoni and Echinostoma caproni. In co-exposed snails, E. caproni prevalence was 63% compared to only 23% for S. mansoni. Co-exposed E. caproni-infected snails exhibited reduced fecundity, higher mortality, and higher parasite reproduction (higher virulence) compared to hosts exposed to echinostomes alone. Conversely, co-exposed S. mansoni-infected snails released fewer parasites and produced greater numbers of eggs compared to hosts exposed to S. mansoni alone. These results suggest that co-exposure not only influences the establishment (presence or absence) of particular parasite species, but also impacts host life history, parasite reproduction, and the virulence of the interaction.     

Host factors limiting monogenean infections: a case study

Comprehensive field data on polystomatid monogeneans record low prevalence and intensity of infection and suggest that worm burdens in this group are strongly regulated: thus, in the majority of Polystoma species infecting anuran amphibians mean abundance is typically less than one parasite/host. There is circumstantial evidence that the dominant control is attributable to host factors which over-ride variations in transmission success. This review provides a brief summary of information on Pseudodiplorchis americanus, a parasite of the desert toad, Scaphiopus couchii, and then focuses in detail on the spectrum of factors regulating infrapopulations of Protopolystoma xenopodis, a parasite of the aquatic Xenopus laevis. Infection levels of adult worms and their contribution to transmission are regulated by external environmental factors (especially temperature), by host factors (including behaviour and population density), and by a range of parasite factors including intra- and inter-specific competitive interactions and variations in intrinsic characters, especially survivorship and reproductive output. In addition to these factors whose primary effect is to modulate transmission rates, there is a major attrition in parasite numbers between invasion and maturity (3 months post-infection). Long-term laboratory experiments on the Xenopus laevis/Protopolystoma xenopodis interaction demonstrate a powerful acquired immune response. Primary infection is characterised by a high prevalence of established adult worms but the success of subsequent challenge infection is greatly reduced, leading to low prevalence and extended pre-patent period. In the small proportion of hosts supporting a second infection of adult parasites, surviving burdens are small (one to two worms/host) and show reduced egg production. These results provide an explanation for the low burdens encountered in field studies: a majority of adult X. laevis in natural populations are likely to exhibit strong, relatively long-term, post-infection immunity after the loss of a previous infection.     

The status of brown and rainbow trout, Salmo trutta and S. gairdneri as hosts of the acanthocephalan, Pomphorhynchus laevis

The status of brown and rainbow trout as hosts of Pomphorhynchus laevis was studied in the field and by means of laboratory investigations. Field data indicated that rainbow trout might belong to the group of preferred hosts of P. laevis , whereas brown trout belonged to the group in which the parasite achieved less than optimal growth and maturation. This was confirmed by laboratory infections. In rainbow trout P. laevis attained up to three times the growth rate in brown trout and maturation occurred whereas in brown trout establishment was lower, growth slower and no parasites matured. Changes in the behaviour of infected Gammarus pulex induced by the presence of P. laevis cystacanths were such as to render the shrimps more vulnerable to predation by trout and other surface and mid-water feeding fish, and selective predation upon infeged G. pulex by fish was demonstrated. nvestigations into the stimuli necessary for eversion of cystacanths of P. laevis revealed that the most important factor was a non-specific component of bile, and it was concluded that cystacanths were likely to evert in any species of fish. Recognition of the different status of brown and rainbow trout as hosts of P. laevis still fails to explain some peculiarities in the distribution of the parasite in the British Isles, where in Britain it occurs in trout in only one river but in Ireland in all rivers throughout the country. It is suggested that the Irish parasites may constitute a different strain of P. laevis , since they use a different species of intermediate host and are better able to survive in brown trout.     

Host manipulation of a freshwater crustacean (Gammarus roeseli) by an acanthocephalan parasite (Polymorphus minutus) in a biological invasion context

Several gammarid species serve as intermediate hosts for the acanthocephalan parasite Polymorphus minutus. This parasite influences gammarid behaviour in order to favour transmission to its ultimate host, generally a bird. We investigated this host manipulation in Gammarus roeseli, a gammarid species introduced in France 150 years ago which now coexists with several exotic species from different origins. In the field, vertical distribution of G. roeseli revealed a higher proportion of infected individuals close to the water's surface and the size distribution of infected gammarids revealed predation pressure on infected individuals. However, under laboratory conditions both infected and non-infected individuals remained benthic. The addition of a second gammarid, Dikerogammarus villosus, to the experimental device involved a vertical displacement of infected G. roeseli towards the water's surface. Dikerogammarus villosus, originating from the Ponto-Caspian basin, can be considered as an aggressive predator. The substitution of D. villosus with Atyaephyra desmarestii, a planktivore decapod, did not alter the gammarids' distribution, with both infected and uninfected G. roeseli staying benthic. Thus, biotic interactions between D. villosus and G. roeseli represent selective pressure encouraging the expression of manipulated behaviour in infected amphipods. Through manipulation, P. minutus was found to increase the survival of infected G. roeseli when faced with non-host predators and to make it more vulnerable to predation by the parasite's definitive host.     

Larval helminths in intermediate hosts: does competition early in life determine the fitness of adult parasites?

Density-dependent effects on parasite fitness have been documented from adult helminths in their definitive hosts. There have, however, been no studies on the cost of sharing an intermediate host with other parasites in terms of reduced adult parasite fecundity. Even if larval parasites suffer a reduction in size, caused by crowding, virtually nothing is known about longer-lasting effects after transmission to the definitive host. This study is the first to use in vitro cultivation with feeding of adult trematodes to investigate how numbers of parasites in the intermediate host affect the size and fecundity of adult parasites. For this purpose, we examined two different infracommunities of parasites in crustacean hosts. Firstly, we used experimental infections of Maritrema novaezealandensis in the amphipod, Paracalliope novizealandiae, to investigate potential density-dependent effects in single-species infections. Secondly, we used the crab, Macrophthalmus hirtipes (Ocypodidae), naturally infected by the trematodes, M. novaezealandensis and Levinseniella sp., the acanthocephalan, Profilicollis spp., and an acuariid nematode. These four helminths all develop and grow in their crustacean host before transmission to their bird definitive host by predation. In experimental infections, we found an intensity-dependent establishment success, with a decrease in the success rate of cercariae developing into infective metacercariae with an increasing dose of cercariae applied to each amphipod. In natural infections, we found that M. novaezealandensis-metacercariae achieved a smaller volume, on average, when infrapopulations of this parasite were large. Small metacercariae produced small in vitro-adult worms, which in turn produced fewer eggs. Crowding effects in the intermediate host thus were expressed at the adult stage in spite of the worms being cultured in a nutrient-rich medium. Furthermore, excystment success and egg-production in M. novaezealandensis in naturally infected crabs were influenced by the number of co-occurring Profilicollis cystacanths, indicating interspecific interactions between the two species. Our results thus indicate that the infracommunity of larval helminths in their intermediate host is interactive and that any density-dependent effect in the intermediate host may have lasting effects on individual parasite fitness.     

Parasitic infection manipulates sodium regulation in the freshwater amphipod Gammarus pulex (L.)

The acanthocephalan parasite Polymorphus minutus induces both physiological and behavioural effects in its intermediate host, Gammarus pulex. The net effect of parasite infection is to increase the likelihood of transmission to the definitive host. Osmoregulation is an energetically expensive mechanism that allows G. pulex to survive in dilute media. Any factor influencing osmoregulation is thus likely to affect the allocation of resources to other areas. This study investigated whether P. minutus infection alters sodium regulation in G. pulex. Haemolymph sodium concentration, water permeability and sodium fluxes were measured over the salinity acclimation range of G. pulex. Water permeability was unaltered by either acclimation salinity or parasite infection. Acclimation to 12‰ significantly raised the haemolymph sodium concentration, reduced the sodium influx, and increased the sodium efflux, to the same extent in both uninfected and infected G. pulex. However, parasite infection induced a significant increase in haemolymph sodium concentration in G. pulex acclimated to 6‰, which was not observed in uninfected G. pulex acclimated to the same salinity. Also, both sodium influx and sodium efflux were significantly lower in parasitized G. pulex acclimated to 6‰, when compared to uninfected G. pulex acclimated to the same salinity. It was concluded that the parasite induced disturbances to sodium regulation in G. pulex acclimated to 6‰ were a functional consequence of the manipulative strategy employed to alter behaviour, rather than a primary target.