Contrasting alterations in brain chemistry in a crustacean intermediate host of two acanthocephalan parasites.

The dynamic properties of neural systems throughout life can be hijacked by so-called manipulative parasites. This study investigated changes in the brain chemistry of the amphipod Gammarus fossarum in response to infection with two trophically-transmitted helminth parasites known to induce distinct behavioral alterations: the bird acanthocephalan Polymorphus minutus and the fish acanthocephalan Pomphorhynchus tereticollis. We quantified brain antioxidant capacity as a common marker of homeostasis and neuroprotection, and brain total protein, on 72 pools of six brains. We analyzed the concentration of serotonin (5HT), dopamine (DA) and tyramine in 52 pools of six brains, by using ultrafast high performance liquid chromatography with electrochemical detection (UHPLC-ECD). Brain total protein concentration scaled hypo-allometrically to dry body weight, and was increased in infected gammarids compared to uninfected ones. The brain of gammarids infected with P. minutus had significantly lower total antioxidant capacity relative to total proteins. Infection with P. tereticollis impacted DA level compared to uninfected ones, and in opposite direction between spring and summer. Brain 5HT level was higher in summer compared to spring independently of infection status, and was decreased by infection after correcting for brain total protein concentration estimated from dry whole-body weight. The potential implication of 5HT/DA balance in parasite manipulation, as a major modulator of the reward-punishment axis, is discussed. Taken together, these findings highlight the need to consider both brain homeostatic and/or structural changes (antioxidant and total protein content) together with neurotransmission balance and flexibility, in studies investigating the impact of parasites on brain and behavior.

Microbial artists: the role of parasite microbiomes in explaining colour polymorphism among amphipods and potential link to host manipulation.

Parasite infections are increasingly reported to change the microbiome of the parasitized hosts, while parasites bring their own microbes to what can be a multi-dimensional interaction. For instance, a recent hypothesis suggests that the microbial communities harboured by parasites may play a role in the well-documented ability of many parasites to manipulate host phenotype, and explain why the degree to which host phenotype is altered varies among conspecific parasites. Here, we explored whether the microbiomes of both hosts and parasites are associated with variation in host manipulation by parasites. Using colour quantification methods applied to digital images, we investigated colour variation among uninfected Transorchestia serrulata amphipods, as well as amphipods infected with Plagiorhynchus allisonae acanthocephalans and with a dilepidid cestode. We then characterized the bacteriota of amphipod hosts and of their parasites, looking for correlations between host phenotype and the bacterial taxa associated with hosts and parasites. We found large variation in amphipod colours, and weak support for a direct impact of parasites on the colour of their hosts. Conversely, and most interestingly, the parasite's bacteriota was more strongly correlated with colour variation among their amphipod hosts, with potential impact of amphipod-associated bacteria as well. Some bacterial taxa found associated with amphipods and parasites may have the ability to synthesize pigments, and we propose they may interact with colour determination in the amphipods. This study provides correlational support for an association between the parasite's microbiome and the evolution of host manipulation by parasites and host-parasite interactions more generally.

Subcutaneous occurrence of encysted Acanthocephala in an anuran amphibian from Brazil.

Acanthocephalans, in their adult stage, are obligatory parasites of many types of vertebrates, including anuran amphibians. Their complex life cycle always involves an arthropod intermediate host but may include non-obligatory strategies that could improve transmission success, such as paratenic infections. In paratenic hosts, these parasites are normally found loose in the body cavity or encysted in internal organs. Here, we present the first report of acanthocephalans found encysted under the skin of an amphibian (i.e., external to its body cavity). The specimen, a clay robber frog [Haddadus binotatus (Spix, 1824)], had been collected in an Atlantic Forest area in southeastern Brazil. Upon examination of the frog, we recovered two specimens of acanthocephalan (Order Echinorhynchida) encysted under the skin of its venter. Considering the host's relatively small size and its thin ventral musculature, we believe that the acanthocephalans may have accidentally trespassed the muscular tissue while attempting to encyst in the frog's internal body wall.

Patterns in parasite diversity and interactions with anurans from the Atlantic Forest.

Knowledge of parasite-host interactions is essential for understanding factors associated with the ecology and evolution of both groups. Some aspects, such as host size and phylogeny, as well as parasite specificity, are significant predictors that help unveil the parasite-host relationship. Thus, the goals of this study were: (1) to describe parasite diversity in regions of the Atlantic Forest; (2) to analyze which host characteristics can influence parasite richness of anuran's parasite component community; and (3) to investigate if the prevalence of parasite infection is related to specificity metrics (ecological and phylogenetic), number of infected hosts and parasite's abundance. We identified 49 parasite taxa, classified into three phyla: Nematoda, Acanthocephala, and Platyhelminthes. Supporting the existing literature, our findings corroborate the positive relationship between host size and parasite richness, further emphasizing the significance of this predictor. Parasite prevalence in the host community is related to the number of infected host species and parasite abundance, but not to phylogenetic and ecological specificity indices. This shows that parasite prevalence is strongly associated with infection opportunity, host sampling effort, and high parasite abundance.

Intermediate host patterns of acanthocephalans in the Weser river system: co-invasion vs host capture.

Anthropogenic interference is a major driver of ecological change in freshwater ecosystems. Pollution and the introduction of new species not only alter macrozoobenthic community structures, but can also affect their respective parasite communities. The ecology of the Weser river system experienced a drastic decline in biodiversity over the past century due to salinization caused by the local potash industry. As a response, the amphipod Gammarus tigrinus was released into the Werra in 1957. A few decades after the introduction and subsequent spread of this North American species, its natural acanthocephalan Paratenuisentis ambiguus was recorded in the Weser in 1988, where it had captured the European eel Anguilla anguilla as a novel host. To assess the recent ecological changes in the acanthocephalan parasite community, we investigated gammarids and eel in the Weser river system. In addition to P. ambiguus, 3 Pomphorhynchus species and Polymorphus cf. minutus were discovered. The introduced G. tigrinus serves as a novel intermediate host for the acanthocephalans Pomphorhynchus tereticollis and P. cf. minutus in the tributary Werra. Pomphorhynchus laevis is persistent in the tributary Fulda in its indigenous host Gammarus pulex. Pomphorhynchus bosniacus colonized the Weser with its Ponto-Caspian intermediate host Dikerogammarus villosus. This study highlights the anthropogenically driven changes in ecology and evolution in the Weser river system. Based on morphological and phylogenetic identification, the shifts in distribution and host usage described here for the first time contribute to the puzzling taxonomy of the genus Pomphorhynchus in times of ecological globalization.

Infection with an acanthocephalan helminth reduces anxiety-like behaviour in crustacean host.

Trophically transmitted heteroxenous parasites of diverse clades can decrease or reverse antipredator behaviours in their intermediate hosts, thereby increasing their chances of reaching their final hosts. Such behavioural alterations could result from compromised cognitive abilities affecting fear- or more generally stress-related neurophysiological pathways. We tested this hypothesis in a key model system in the study of parasitic manipulation, the fish acanthocephalan parasite Pomphorhynchus tereticollis and its intermediate crustacean host Gammarus fossarum, using the 'threat of electric shock' paradigm. We exposed uninfected and infected G. fossarum to chronic and/or acute electric shock programs at two different intensities (voltage), and then quantified their sheltering behaviour as a proxy for anxiety-like state. Infected gammarids did not express anxiety-like response to electric shocks, while uninfected gammarids hid more when exposed to acute treatments, and when exposed to the high intensity chronic treatment. Interestingly, the lack of response in infected gammarids depended on parasite developmental stage. Our results support the hypothesis that this acanthocephalan parasite impacts the general anxiety-like circuitry of their intermediate host. Further studies are needed to investigate whether it involves inappropriate processing of information, impaired integration, or altered activation of downstream pathways initiating behavioural action.

Narrower isotopic niche size in fish infected by the intestinal parasite Pomphorhynchus sp. compared to uninfected ones.

Examples of parasite-related effects on intermediate crustacean hosts are numerous but their ecological consequences on their vertebrate hosts are scarce. Here, we address the role of macroparasite infections on the trophic niche structure of definitive hosts and its potential physiological consequences using wild fish populations infected with an acantochephalan parasite Pomphorhynchus sp., a trophically transmitted intestinal worm. Infected and uninfected fish were sampled from six populations on the Marne River, France and the prevalence of intestinal parasites in the host populations ranged from 50% to 90%. Although the isotopic ratios (δ13 C and δ15 N) did not differ between infected and uninfected fish, we found a consistent pattern of isotopic niche size being considerably smaller in infected hosts when compared with noninfected ones. This was not explained by interindividual differences in intrinsic factors such as length/age or body condition between infected and uninfected fish. These results suggest a potential niche specialization of infected fish, which did not impair their energetic status.

Dysbiosis of fish gut microbiota is associated with helminths parasitism rather than exposure to PAHs at environmentally relevant concentrations.

Although parasite infection and pollution are common threats facing wild populations, the response of the gut microbiota to the joint impact of these stressors remains largely understudied. Here, we experimentally investigated the effects of exposure to Polycyclic Aromatic Hydrocarbons (PAHs) and infection by a common acanthocephalan intestinal parasite (Pomphorhynchus sp.) on the gut microbial flora of a freshwater fish, the European chub (Squalius cephalus). Naturally infected or uninfected individuals were exposed to PAHs at environmentally realistic concentrations over a five-week period. Characterization of the gut bacterial community through 16S rRNA gene amplicon sequencing revealed that parasitic infection was a more structuring factor of bacterial diversity and composition than PAH exposure. Specifically, chub infected by Pomphorhynchus sp. harbored significantly less evenly represented gut bacterial communities than the uninfected ones. In addition, substantial changes in sequence abundance were observed within the main bacterial phyla, including the Firmicutes, Fusobacteriota, Actinobacteriota, and Proteobacteria. Again, these compositional changes correlated with host infection with Pomphorhynchus sp., confirming its pivotal role in gut microbial assemblage. Overall, these results highlight the importance of defining the parasitic status of individuals when conducting microbial ecotoxicological analyses at the digestive tract level, as this should lead to better understanding of microbiota modulations and help to identify microbial markers specifically associated with chemicals.

Parasite infection influences the biomarker response and locomotor activity of Gammarus fossarum exposed to conventionally-treated wastewater.

Modern wastewater treatment plants cannot completely remove pollutants. Often, effluents entering the aquatic environment still contain micropollutants such as pharmaceuticals or pesticides, which may impose adverse effects on aquatic biota. At the same time, a large proportion of free-living aquatic species are known to be infected with parasites, which raises the question of interactions between environmental stressors (such as micropollutants) and parasite infection. We chose the freshwater amphipod Gammarus fossarum (Koch, 1835) as a test organism to investigate potential pollutant-parasite interactions. This gammarid is frequently used in ecotoxicological tests and is also commonly infected with larvae of the acanthocephalan parasite species Polymorphus minutus (Zeder, 1800) Lühe, 1911. We exposed infected and uninfected specimens of G. fossarum to conventionally-treated wastewater and river water in a 22-day flow channel experiment. The test organisms' response was measured as mortality rates, concentrations or activities of five biomarkers, and overall locomotor activity. No significant differences were found between mortality rates of different exposure conditions. Contrastingly, three biomarkers (phenoloxidase activity, glycogen, and lipid concentrations) showed a significant increase in infected gammarids, while the effect of the water type was insignificant. Infected gammarids also showed a significantly higher locomotor activity in both water types. Our results suggest that the response of G. fossarum during the exposure experiments was mainly driven by parasite infection. This implies that parasites may act as additional biotic stressors in multiple stressor scenarios, and therefore, might play an important role when measuring the response of organisms to chemical stressors. Future ecotoxicological studies and assessments thus should consider parasite infection as an additional test parameter.

STABLE ISOTOPE SIGNATURES OF AN ACANTHOCEPHALAN AND TREMATODE FROM THE HERBIVOROUS MARINE FISH KYPHOSUS BIGIBBUS (PERCIFORMES: KYPHOSIDAE).

Stable isotope analyses of carbon and nitrogen (δ13C and δ15N) are useful for elucidating consumer relationships of free-living organisms, as carbon isotopes indicate dietary carbon sources and incremental increases in nitrogen isotopic enrichment are correlated with increases in trophic position. However, host-parasite relationships are more difficult to interpret using isotopes, as data from different host-parasite systems rarely show any consistent pattern. This inconsistency of pattern reflects the complexity of host-parasite relationships, but also the scarcity of data from a diverse assemblage of host-parasite systems. We present stable isotope data from a host-parasite system including 2 ecologically contrasting helminths, an acanthocephalan (Filisoma filiformis) and a digenetic trematode (Enenterum sp.), which co-occur in the intestine of the same marine fish (Kyphosus bigibbus), the diet of which consists almost exclusively of macroalgae. We obtained δ13C and δ15N data from K. bigibbus muscle, stomach contents, and pooled infrapopulations of Enenterum sp. and F. filiformis. Consistent with other isotope studies including acanthocephalans, F. filiformis was depleted in δ13C and δ15N relative to K. bigibbus. Although Enenterum sp. exhibited values for δ13C similar to those for F. filiformis, they were enriched in δ15N relative to the acanthocephalan, with a signature similar to that of K. bigibbus. These findings are discussed within a host-ecosystem context, highlighting the importance of considering species-specific biology when interpreting host-parasite relationships using stable isotopes. Our study adds to the growing body of literature indicating that absorptive feeders, such as acanthocephalans, are typically depleted in δ13C and δ15N relative to their hosts, whereas trematodes, with a greater diversity of feeding opportunities, exhibit a wide variety of isotopic signatures across life stage and different host-parasite systems.

Conspecific migration and environmental setting determine parasite infracommunities of non-migratory individual fish.

Parasite infracommunities tend to be stochastic in nature, although environmental characteristics such as the type of water source in streams and host traits can have an effect on the biotic assemblages and by extension the parasite fauna. We examined the effect of water source and the rate of adult fish migration on the metazoan parasite infracommunities of conspecific juvenile brown trout, Salmo trutta L. among streams flowing into Lake Lucerne (Switzerland). Juvenile (1 to 2-year old) fish harboured higher parasite species richness in groundwater-fed than in surface water-fed streams, whereas the rate of fish migration did not affect infracommunity richness. Heteroxenous species were more common in groundwater-fed streams with high and medium rates of trout migration, whereas infracommunities in surface water-fed streams and streams with low rates of fish migration were dominated by one monoxenous parasite or lacked infections. Similarity in the parasite infracommunity composition of juvenile trout across streams was explained by the interaction between type of water source and adult migration rates. Our conclusions support that similarity in the parasite composition of resident freshwater conspecifics can be predicted by the local environmental settings and host migratory behaviour, whereas parasite richness is mainly influenced by the environmental characteristics.

The phylogeographic puzzle of Pseudoacanthocephalus toshimai, an amphibian acanthocephalan in northern Japan.

The amphibian acanthocephalan, Pseudoacanthocephalus toshimai, was considered to be an island-endemic species in Hokkaido, Japan. However, the parasite was found from Rana ornativentris, Rana tagoi, Zhangixalus arboreus, and Bufo japonicus formosus in northern Honshu (Aomori and Iwate Prefectures), which is separated from Hokkaido by the Tsugaru Strait. The mitochondrial DNA-based phylogenetic and population genetic analyses of P. toshimai showed that the northern Honshu isolates are far distantly related to the Hokkaido isolates, and that a demographic population expansion occurred in Hokkaido during the recent geological past. The rich genetic diversity of P. toshimai in northern Honshu suggests a scenario that anuran hosts invaded Hokkaido together with P. toshimai via the land bridge of the Tsugaru Strait. However, the evolutionary history of Rana pirica, a main definitive host for P. toshimai in Hokkaido, is contradictory to the introduction scenario inferred from the parasite. The finding of several geographically mismatched isolates of P. toshimai from both northern Honshu and Hokkaido suggests a possibility that the migration of the parasite infrequently occurred between the two areas even after the land bridge disappeared. More detailed information on the evolutionary history of anurans is needed to resolve the biogeographical enigma of P. toshimai.

Trade-Offs with Growth Limit Host Range in Complex Life-Cycle Helminths.

AbstractParasitic worms with complex life cycles have several developmental stages, with each stage creating opportunities to infect additional host species. Using a data set for 973 species of trophically transmitted acanthocephalans, cestodes, and nematodes, we confirmed that worms with longer life cycles (i.e., more successive hosts) infect a greater diversity of host species and taxa (after controlling for study effort). Generalism at the stage level was highest for middle life stages, the second and third intermediate hosts of long life cycles. By simulating life cycles in real food webs, we found that middle stages had more potential host species to infect, suggesting that opportunity constrains generalism. However, parasites usually infected fewer host species than expected from simulated cycles, suggesting that generalism has costs. There was no trade-off in generalism from one stage to the next, but worms spent less time growing and developing in stages where they infected more taxonomically diverse hosts. Our results demonstrate that life-cycle complexity favors high generalism and that host use across life stages is determined by both ecological opportunity and life-history trade-offs.

Trophic relationship between the Patinga hybrid (Piaractus mesopotamicus x Piaractus brachypomus) and Echinorhynchus gomesi Machado Filho, 1948 in fish farms.

A total of 122 Patinga specimens were collected from fish farms (P1, P2 and P3), and only those from fish farm P1 were shown to be infected with Echinorhynchus gomesi. In addition, fish in this study were shown to have diets that consisted of 21 different food items, and Notodiaptomus sp. (Copepoda: Calanoida) was identified as a potential intermediate host for E. gomesi.

Gastrointestinal helminths of little blue penguins, Eudyptula novaehollandiae (Stephens), from Otago, New Zealand.

Data regarding helminth communities can provide insights into health, feeding interactions, behaviour and evolution of their host organisms. Penguins (Spheniscidae) are important components of marine food webs and tracking their helminth communities can be indicative of ecosystem health. New Zealand is home to 5 of the world's 19 penguin species and little is known about their gastrointestinal helminths. Here, we provide the first study on the gastrointestinal helminths of little blue penguins from south-eastern South Island, New Zealand. The helminth community consisted of two species of tapeworm; Tetrabothrius lutzi and Tetrabothrius sp.; three nematode species, Contracaecum eudyptulae, Capillaria sp. and Stegophorus macronectes; two acanthocephalans, Andracantha sigma and Bolbosoma balaenae; and one trematode, Galactosomum otepotiense. The most prevalent parasites were T. lutzi, A. sigma, and C. eudyptulae. This work includes three new host records and five new geographic records. This is the first report of B. balaenae occurring in a host other than a marine mammal. This study adds to our knowledge about the helminth community of New Zealand little blue penguins, and includes new genetic data on helminth species, providing a baseline against which future studies may be compared.

Taxonomic resolution affects host-parasite association model performance.

Identifying the factors that structure host­parasite interactions is fundamental to understand the drivers of species distributions and to predict novel cross-species transmission events. More phylogenetically related host species tend to have more similar parasite associations, but parasite specificity may vary as a function of transmission mode, parasite taxonomy or life history. Accordingly, analyses that attempt to infer host−parasite associations using combined data on different parasite groups may perform quite differently relative to analyses on each parasite subset. In essence, are more data always better when predicting host−parasite associations, or does parasite taxonomic resolution matter? Here, we explore how taxonomic resolution affects predictive models of host−parasite associations using the London Natural History Museum's database of host­helminth interactions. Using boosted regression trees, we demonstrate that taxon-specific models (i.e. of Acanthocephalans, Nematodes and Platyhelminthes) consistently outperform full models in predicting mammal-helminth associations. At finer spatial resolutions, full and taxon-specific model performance does not vary, suggesting tradeoffs between phylogenetic and spatial scales of analysis. Although all models identify similar host and parasite covariates as important to such patterns, our results emphasize the importance of phylogenetic scale in the study of host­parasite interactions and suggest that using taxonomic subsets of data may improve predictions of parasite distributions and cross-species transmission. Predictive models of host­pathogen interactions should thus attempt to encompass the spatial resolution and phylogenetic scale desired for inference and prediction and potentially use model averaging or ensemble models to combine predictions from separately trained models.

Vicariance in a generalist fish parasite driven by climate and salinity tolerance of hosts.

Acanthocephalans are parasites with complex lifecycles that are important components of aquatic systems and are often model species for parasite-mediated host manipulation. Genetic characterization has recently resurrected Pomphorhynchus tereticollis as a distinct species from Pomphorhynchus laevis, with potential implications for fisheries management and host manipulation research. Morphological and molecular examinations of parasites from 7 English rivers across 9 fish species revealed that P. tereticollis was the only Pomphorhynchus parasite present in Britain, rather than P. laevis as previously recorded. Molecular analyses included two non-overlapping regions of the mitochondrial gene - cytochrome oxidase and generated 62 sequences for the shorter fragment (295 bp) and 74 for the larger fragment (583 bp). These were combined with 61 and 13 sequences respectively, from Genbank. A phylogenetic analysis using the two genetic regions and all the DNA sequences available for P. tereticollis identified two distinct genetic lineages in Britain. One lineage, possibly associated with cold water tolerant fish, potentially spread to the northern parts of Britain from the Baltic region via a northern route across the estuarine area of what is now the North Sea during the last Glaciation. The other lineage, associated with temperate freshwater fish, may have arrived later via the Rhine/Thames fluvial connection during the last glaciation or early Holocene when sea levels were low. These results raise important questions on this generalist parasite and its variously environmentally adapted hosts, and especially in relation to the consequences for parasite vicariance.

Increased temperature has no consequence for behavioral manipulation despite effects on both partners in the interaction between a crustacean host and a manipulative parasite.

Parasites alter many traits of their hosts. In particular, parasites known as "manipulative" may increase their probability of transmission by inducing phenotypic alterations in their intermediate hosts. Although parasitic-induced alterations can modify species' ecological roles, the proximate factors modulating this phenomenon remain poorly known. As temperature is known to affect host-parasite associations, understanding its precise impact has become a major challenge in a context of global warming. Gammarids are ecologically important freshwater crustaceans and serve as intermediate hosts for several acanthocephalan species. These parasites induce multiple effects on gammarids, including alterations of their behavior, ultimately leading to modifications in their functional role. Here, experimental infections were used to assess the effect of two temperatures on several traits of the association between Gammarus pulex and its acanthocephalan parasite Pomphorhynchus laevis. Elevated temperature affected hosts and parasites in multiple ways (decreased host survival, increased gammarids activity, faster parasites development and proboscis eversion). However, behavioral manipulation was unaffected by temperature. These results suggest that predicted change in temperature may have little consequences on the trophic transmission of parasites through changes in manipulation, although it may modify it through increased infection success and faster parasites development.

Temporal variation in infection levels and reproductive traits of the acanthocephalan Pseudoleptorhynchoides lamothei in the blue sea catfish Ariopsis guatemalensis (Günther, 1864).

Current data on reproductive biology and population dynamics of the acanthocephalans are scarce mainly in regions from the tropical Pacific. An analysis was done to identify possible factors that influence variation in infection levels of the acanthocephalan Pseudoleptorhynchoides lamothei Salgado-Maldonado, 1976 in its final host, the blue sea catfish Ariopsis guatemalensis (Günther, 1864), and describe its main reproductive traits. A total of 1,094 A. guatemalensis were collected from Tres Palos Lagoon from August 2014 to December 2015. Prevalence of P. lamothei varied from 1.47% to 38.33%, and mean abundance from 0.03 to 4.44 helminths per examined host. In female P. lamothei relative fecundity increased with total length. Temporal variations in P. lamothei infection levels were attributed mainly to changes in host feeding and reproductive behaviour in response to local environmental factors as climatic season, and variations in water temperature.

Parasitism in Gambusia affinis: Fitness Effects in an Incipient Matrotroph.

Gambusia affinis (western mosquitofish) serves as a host for a variety of larval and adult parasites. Gambusia affinis is also an incipient matrotroph, exhibiting adjustments in post-fertilization provisioning to some offspring within a brood using recently acquired resources. Nutrient transfer to embryos is expected to limit the loss of embryo mass during development resulting in larger offspring. Since larger offspring are more likely to survive, maternal contributions are expected to increase fitness. The presence of parasites, particularly intestinal helminths, potentially reduces body condition and resources available for developing offspring, thereby reducing host fitness. The effects of parasitism on the fitness of G. affinis were investigated in the present study. Fish were collected from 3 sites monthly from June 2015 through August 2016. All helminth parasites were collected during necropsy and identified. Brood size and embryo developmental stage were recorded for each female fish. Additionally, 10 ova/embryos of each developmental stage from each female fish collected from May through August 2016 were haphazardly selected and individually weighed. From 429 female mosquitofish, 5,072 helminths were collected. Brood size varied among collection sites and was positively influenced by maternal body condition, the number of daylight hours, water temperature, and the intensity of both plerocercoid and adult Schyzocotyle acheilognathi. However, brood size was negatively related to the intensity of Neoechinorhynchus cylindratus cystacanth and an increasing number of days between collection and dissection. Embryo weight increased with the presence of either Camallanidae or Contracaecum multipapulatum, embryo developmental stage, and relative host density. These results indicate that some parasitic helminth species negatively affect the fitness of G. affinis, while some positively affect fitness, and that effect can vary with intensity.