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Embryonic development is often considered shielded from the effects of natural selection, being selected primarily for reliable development. However, embryos sometimes represent virulent parasites, triggering a coevolutionary "arms race" with their host. We have examined embryonic adaptations to a parasitic lifestyle in the bitterling fish. Bitterlings are brood parasites that lay their eggs in the gill chamber of host mussels. Bitterling eggs and embryos have adaptations to resist being flushed out by the mussel. These include a pair of projections from the yolk sac that act as an anchor. Furthermore, bitterling eggs all adopt a head-down position in the mussel gills which further increases their chances of survival. To examine these adaptations in detail, we have studied development in the rosy bitterling ( Rhodeus ocellatus ) using molecular markers, X-ray tomography, and time-lapse imaging. We describe a suite of developmental adaptations to brood parasitism in this species. We show that the mechanism underlying these adaptions is a modified pattern of blastokinesis-a process unique, among fish, to bitterlings. Tissue movements during blastokinesis cause the embryo to do an extraordinary "front-flip" on the yolk. We suggest that this movement determines the spatial orientation of the other developmental adaptations to parasitism, ensuring that they are optimally positioned to help resist the ejection of the embryo from the mussel. Our study supports the notion that natural selection can drive the evolution of a suite of adaptations, both embryonic and extra-embryonic, via modifications in early development., Competing Interests: Competing interests statement:The authors declare no competing interest.

Monogenean parasites are often co-introduced with their fish hosts into novel areas. This study confirmed co-introduction of two dactylogyrids, Dactylogyrus squameus Gusev, 1955 and Bivaginogyrus obscurus (Gusev, 1955), and a newly described gyrodactylid species, Gyrodactylus pseudorasborae n. sp. into Europe along with their fish host, the invasive topmouth gudgeon Pseudorasbora parva (Temminck & Schlegel) from East Asia. All three species were observed in the lower Dnieper and middle Danube basin regions and had slightly larger haptoral hard parts than the same parasites in their native range. While dactylogyrids occurred sporadically, we recorded regular infection by G. pseudorasborae n. sp. at relatively high prevalence and abundance. This latter species was observed in both the native and non-native range of topmouth gudgeon, and resembles Gyrodactylus parvae You et al., 2008 recently described from P. parva in China. Both species were distinguished based on genetic analysis of their ITS rDNA sequence (6.6% difference), and morphometric differences in the marginal hooks and male copulatory organ. Phylogenetic analysis of dactylogyrid monogeneans showed that B. obscurus clustered with Dactylogyrus species parasitising Gobionidae and Xenocyprididae, including D. squameus, supporting recent suggestions of a paraphyletic origin of the Dactylogyrus genus. In addition to co-introduced parasites, topmouth gudgeon was infected with a local generalist, G. prostae Ergens, 1964, increasing the number of monogeneans acquired in Europe to three species. Nevertheless, monogenean infections were generally lower in non-native host populations, potentially giving an advantage to invading topmouth gudgeon., (© M. Ondračková et al., published by EDP Sciences, 2023.)

Polymorphism of the major histocompatibility complex (MHC), DAB1 gene was characterized for the first time in the European bitterling (Rhodeus amarus), a freshwater fish employed in studies of host-parasite coevolution and mate choice, taking advantage of newly designed primers coupled with high-throughput amplicon sequencing. Across 221 genotyped individuals, we detected 1-4 variants per fish, with 28% individuals possessing 3-4 variants. We identified 36 DAB1 variants, and they showed high sequence diversity mostly located within predicted antigen-binding sites, and both global and codon-specific excess of non-synonymous mutations. Despite deep divergence between two major allelic lineages, functional diversity was surprisingly low (3 supertypes). Overall, these findings suggest the role of positive and balancing selection in promotion and long-time maintenance of DAB1 polymorphism. Further investigations will clarify the role of pathogen-mediated selection to drive the evolution of DAB1 variation., (© 2022. The Author(s).)

This study focused on the total mercury content in fish from seven drinking-water reservoirs located in the Morava River Basin: Bojkovice, Boskovice, Hubenov, Karolinka, Landstejn, Ludkovice and Nova Rise in the Czech Republic. A total of 308 fish were collected for the analysis. The content of total mercury was measured in the muscle tissue of bream, roach and perch using atomic absorption spectrometry and varied from 0.057±0.009 to 0.440 mg kg -1 in bream, from 0.030±0.005 to 0.393±0.138 mg kg -1 in roach and from 0.092±0.007 to 0.638±0.042 mg kg -1 in perch. The highest total mercury content was found in perch from Landstejn and the lowest was measured in roach from Ludkovice. A positive statistically significant relationship was found between fish weight and total mercury content in fish muscle for almost all species and all sampling sites, except for roach from Ludkovice. A total of 19 samples exceeded the maximum mercury level set by legislation on food contaminants-0.5 mg kg -1 in freshwater fish., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The genes of the major histocompatibility complex (MHC) are an essential component of the vertebrate immune system and MHC genotypes may determine individual susceptibility to parasite infection. In the wild, selection that favors MHC variability can create situations in which interspecies hybrids experience a survival advantage. In a wild system of two naturally hybridizing leuciscid fish, we assessed MHC IIB genetic variability and its potential relationships to hosts' ectoparasite communities. High proportions of MHC alleles and parasites were species-specific. Strong positive selection at specific MHC codons was detected in both species and hybrids. MHC allele expression in hybrids was slightly biased towards the maternal species. Controlling for a strong seasonal effect on parasite communities, we found no clear associations between host-specific parasites and MHC alleles or MHC supertypes. Hybrids shared more MHC alleles with the more MHC-diverse parental species, but expressed intermediate numbers of MHC alleles and positively selected sites. Hybrids carried significantly fewer ectoparasites than either parent species, suggesting a hybrid advantage via potential heterosis., (© 2021. The Author(s).)

Hybrid advantage, described as the superiority of hybrids in some traits over their parents and termed the "heterosis effect," is widely documented in the case of reciprocal crosses of parental species (i.e., hybrids representing the F1 generation). In fish, high survival, fast growth and better health status have been widely documented in F1 hybrids. Nonetheless, the effects of interspecific hybridization on vigour, physiology and immunity-related traits in fish are largely unknown, especially concerning native systems of coexisting parental and hybrid genomes in the same habitat. The present study examined the potential physiological and immune aspects of hybrid heterosis by comparing condition status (measured especially by indexes), haematological profile, glucose concentration and selected parameters of non-specific and specific immunity between the evolutionarily divergent non-congeneric cyprinoid species Abramis brama and Rutilus rutilus and their hybrids representing the F1 generation, all of them caught in nature. Clear differences were documented for vigour-related, physiological and immune parameters between the two divergent species. Hybrids generally tended to express intermediate characters of the measured traits, likely generated by the evolutionary divergence of the hybridizing species; nonetheless, for some traits, hybrids exhibited a character that was more similar to one parental species than to the other. This was interpreted as the heterozygote advantage for F1 hybrids. It is suggested that a maternally inherited genetic background may potentially influence the expression of some branches of non-specific immunity or other aspects related to the fish health status., (© 2020 Fisheries Society of the British Isles.)

Background: Host specificity is one of the outputs of the coevolution between parasites and their associated hosts. Several scenarios have been proposed to explain the pattern of parasite distribution in parental and hybrid genotypes ranging from hybrid resistance to hybrid susceptibility. We hypothesized that host-parasite co-adaptation limits the infection of host-specific parasites in hybrid genotypes even under the condition of the high frequency of hybrids. The experimental monogenean infection in pure breeds of Blicca bjoerkna and Abramis brama and cross-breeds (the F1 generation of hybrids) under the condition of similar frequencies of pure and hybrid genotypes was investigated. We also examined the potential effect of the maternal origin of hybrids (potential co-adaptation at the level of mitochondrial genes) on monogenean abundance., Methods: Pure breeds of two cyprinids and two cross-breeds (one with B. bjoerkna, the next with A. brama in the maternal positions) were exposed to infection by monogeneans naturally occurring in B. bjoerkna and A. brama. The experiment was run under similar frequencies of the four breed lines., Results: We showed similar levels of monogenean infection in B. bjoerkna and A. brama. However, each species harboured specific monogenean fauna. Hybrids harboured all monogenean species specifically infecting one or the other species. Monogenean infection levels, especially those of Dactylogyrus specific to A. brama, were lower in hybrids. For the majority of host-specific parasites, there was no effect of the maternal origin of hybrids on monogenean abundance. Asymmetry was found in the distribution of specific parasites in favour of specialists of B. bjoerkna in the monogenean communities of hybrids., Conclusions: Our results indicate that the maternal mtDNA of hybrids is not an important predictor of host-specific monogenean infection, which may suggest that mitochondrial genes are not strongly involved in the coadaptation between monogeneans and their associated hosts. The asymmetry of species-specific parasites suggests similarity between the molecular components of the immune mechanisms in hybrids and B. bjoerkna. Our results revealed a difference between the degree of host-parasite coadaptation in specific parasites of A. brama and the degree of host-parasite coadaptation in specific parasites of B. bjoerkna and their associated hosts.

Parasitic infections elicit host defences that pose energetic trade-offs with other fitness-related traits. Bitterling fishes and unionid mussels are involved in a two-way parasitic interaction. Bitterling exploit mussels by ovipositing into their gills. In turn, mussel larvae (glochidia) develop on the epidermis and gills of fish. Hosts have evolved behavioural responses to reduce parasite load, suggesting that glochidia and bitterling parasitism are costly. We examined the energetic cost of parasitism on both sides of this relationship. We used intermittent flow-through respirometry to measure (1) standard metabolic rate (SMR) of individual duck mussels Anodonta anatina (a common bitterling host) before and during infection by embryos of the European bitterling Rhodeus amarus, and (2) SMR and maximum oxygen uptake (MO 2 max) of individual R. amarus before and during infection with glochidia of the Chinese pond mussel Sinanodonta woodiana (a mussel species that successfully infects bitterling). As predicted, we observed an increase in mussel SMR when infected by bitterling embryos and an increased SMR in glochidia-infected bitterling, though this was significantly mediated by the time post-infection. Contrary to our predictions, glochidia infection did not impair MO 2 max and the number of glochidia attached to gills positively (rather than negatively) correlated with MO 2 max. The results suggest that tolerance is the prevailing coping mechanism for both fish and mussels when infected, while resistance mechanisms appear to be confined to the behavioural level.

South-east Europe, along with the adjacent region of south-west Asia, is an important biodiversity hotspot with high local endemism largely contributed by contemporary continental lineages that retreated to southern refugia during colder Quaternary periods. We investigated the genetic diversity of the European bitterling fish (Rhodeus amarus) species complex (Cyprinidae) across its range in the western Palearctic, but with a particular emphasis in the region of Balkan, Pontic and Caspian refugia. We genotyped 12 polymorphic microsatellite loci and a partial sequence of mitochondrial gene cytochrome b (CYTB) for a set of 1,038 individuals from 60 populations. We used mtDNA sequences to infer phylogenetic relationships and historical demography, and microsatellite markers to describe fine-scale genetic variability and structure. Our mtDNA analysis revealed six well-supported lineages, with limited local co-occurrence. Two lineages are distributed throughout central and western Europe (lineages "A" and "B"), with two zones of secondary contact. Another two lineages were restricted to the Ponto-Aegean region of Greece (lineages "C" and "D") and the final two lineages were restricted south of the Caucasus mountains (lineage "E" from the Black Sea watershed and lineage "F" from the Caspian watershed). A signal of recent expansion was revealed in the two widespread lineages and the Ponto-Aegean lineage "C". The geographic distribution of clusters detected by nuclear microsatellites corresponded well with mitochondrial lineages and demonstrated finely sub-structured populations. A profound population structure suggested a significant role of genetic drift in differentiation among lineages. Lineage divergence in the Ponto-Aegean and Caspian regions are substantial, supporting the validity of two described endemic species (Rhodeus meridionalis as lineage "D" and Rhodeus colchicus as lineage "E") and invite taxonomic evaluation of the other two southern lineages (Thracean "C" and Caspian "F")., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Carassius gibelio is an extraordinary cyprinid species exhibiting both sexual and asexual reproduction. We hypothesized that parasitism selection is one of the potential mechanisms contributing to the coexistence of the two reproductive forms of C. gibelio living in the same habitat. We performed a four-year study to investigate the dynamics of parasite infection in C. gibelio . According to the Red Queen prediction, the asexual form is a target of parasite adaptation due to its low genetic variability. Both sexual and gynogenetic forms of C. gibelio exhibited similar levels of prevalence, with monogeneans being the most frequently observed parasite group. We observed the temporal dynamics of parasite infection in the last year of investigation, when both forms were more strongly parasitized. The sexual form was more parasitized by ectoparasites in the first and last years and less parasitized by nematodes in the last year when compared to the gynogenetic form. We found no trend of high parasite infection in gynogenetic mtDNA haplotypes. We conclude that Red Queen dynamics is not the mechanism driving parasite infection in sexual-gynogenetic C. gibelio over a long time scale. Alternatively, we suggest that the dynamics of parasite infection in this complex may be generated by multiple mechanisms.

The aim of this study was to evaluate the influence of preservation method on the results of parasite community studies. Two host species, European perch Perca fluviatilis and European bitterling Rhodeus amarus, were examined for parasites after having been subjected to 4 different storage treatments: freezing, preservation in 4% formaldehyde or 70% ethanol and transportation of live (fresh) fish as a control. Preservation prior to dissection resulted in a loss of information, leading to incomplete quantitative data (all preservation treatments), qualitative data (ethanol and formaldehyde preservation) and a lowered ability to determine parasites to species level based on morphology compared to dissecting fresh fish. Of the more abundant taxa, only crustaceans and acanthocephalans provided relatively even results between treatments. We conclude that preservation media, such as ethanol or formaldehyde, significantly affects the ability to obtain precise parasite community data; hence, we recommend the use of freshly sacrificed fish for parasite community studies whenever possible. Alternatively, freezing may prove acceptable for evaluating parasite community taxonomic composition.

Hybrids and their parasite diversity represent interesting models for evolutionary ecology. The modified immune response, shifted ecology, inheritance, and maternal ancestry of hybrid host fish are supposed to affect the diversity of their parasite communities. The pattern of metazoan parasite distribution in non-congeneric cyprinids - common bream (Abramis brama) and roach (Rutilus rutilus) (species with different morphology and ecology, and harbouring different specific parasites) - and their hybrids was analysed. Four static alternative scenarios based on parasite infection levels in hybrids and parental taxa are known. The hybrid resistance scenario predicts that hybrids are more resistant than parental taxa, resulting in low parasite infection in hybrids. This scenario is principally consistent with hybrid heterosis advantage. In accordance with this prediction, metazoan parasite abundance and prevalence were higher in parental species when compared with their hybrids. Alternatively, the dynamic Red Queen scenario of infection in hybridising systems predicts parasite adaptation to common hosts. Temporal (six sampling events) and spatial (two sampling sites) aspects as possible factors influencing parasite distribution were analysed. We found no support for this hypothesis, i.e. no changes in the frequency of hybrids or their parental species and no changes in parasite infection in parental species or hybrids were found in the different time periods. The effect of maternal ancestry on infection level was evident; hybrids exhibiting common bream mtDNA were more strongly parasitized by digeneans and crustaceans than hybrids exhibiting roach mtDNA. Hybrids harboured a majority of the specific parasites of both parental species; however, the level of infection of common bream-specific parasites (especially monogeneans) in hybrids was low. Such an asymmetrical distribution of parental species-specific parasites in hybrids may suggest the limited inheritance of protective immunological mechanisms from one parental species and reveal stronger coadaptation between common bream and its specific parasites., (Copyright © 2017 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

In this study, we assessed the impact of sampling method on the results of fish ectoparasite studies. Common roach Rutilus rutilus were sampled from the same gravel pit in the River Dyje flood plain (Czech Republic) using 3 different sampling methods, i.e. electrofishing, beach seining and gill-netting, and were examined for ectoparasites. Not only did fish caught by electrofishing have more of the most abundant parasites (Trichodina spp., Gyrodactylus spp.) than those caught by beach seining or gill-netting, they also had relatively rich parasite infracommunities, resulting in a significantly different assemblage composition, presumably as parasites were lost through handling and 'manipulation' in the net. Based on this, we recommend electrofishing as the most suitable method to sample fish for parasite community studies, as data from fish caught with gill-nets and beach seines will provide a biased picture of the ectoparasite community, underestimating ectoparasite abundance and infracommunity species richness.

The phylogenetic relationships and taxonomy of the spirlins in the genus Alburnoides are examined by comparative sequencing analysis of mitochondrial and nuclear markers. Molecular analyses revealed 17 Eurasian lineages divided into two main clades, termed the Ponto-Caspian and European in accordance with the lineage distribution. The indel diagnostics of β-actin and S7 markers and translation of cyt b to the amino acid chain were evaluated as a reliable identifying tool for most of the recognised lineages. Lineage richness is closely connected with the existence of known glacial refugia in most cases. The underestimation of species richness in the genus Alburnoides is confirmed: the genetic analyses support the validity of 11 morphologically accepted species; apart from them, four phylogenetic lineages requiring descriptions as separate species were revealed. The distribution area of the nominotypical species A. bipunctatus s. stricto is newly defined. Two diverging phylogenetic lineages, A. ohridanus, and A. prespensis complex, were observed in the Southeast Adriatic Freshwater Ecoregion, confirmed as a hotspot of endemic biodiversity. A. ohridanus demonstrates high divergence from the A. prespensis complex, represented by three similar mitochondrial lineages with the same nuclear haplotypes and sympatric occurrence. The range restricted endemism was confirmed for at least seven species. The Albanian river systems, as well as the wider Ponto-Caspian basin exhibit complications among definite species delineations and gaps in understanding of microevolutionary processes; these areas require further investigations., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Predicting the impacts of non-native species remains a challenge. As populations of a species are genetically and phenotypically variable, the impact of non-native species on local taxa could crucially depend on population-specific traits and adaptations of both native and non-native species. Bitterling fishes are brood parasites of unionid mussels and unionid mussels produce larvae that parasitize fishes. We used common garden experiments to measure three key elements in the bitterling-mussel association among two populations of an invasive mussel (Anodonta woodiana) and four populations of European bitterling (Rhodeus amarus). The impact of the invasive mussel varied between geographically distinct R. amarus lineages and between local populations within lineages. The capacity of parasitic larvae of the invasive mussel to exploit R. amarus was higher in a Danubian than in a Baltic R. amarus lineage and in allopatric than in sympatric R. amarus populations. Maladaptive oviposition by R. amarus into A. woodiana varied among populations, with significant population-specific consequences for R. amarus recruitment. We suggest that variation in coevolutionary states may predispose different populations to divergent responses. Given that coevolutionary relationships are ubiquitous, population-specific attributes of invasive and native populations may play a critical role in the outcome of invasion. We argue for a shift from a species-centred to population-centred perspective of the impacts of invasions., (© 2015 The Author(s).)

The ability to attract mates, acquire resources for reproduction, and successfully outcompete rivals for fertilizations may make demands on cognitive traits--the mechanisms by which an animal acquires, processes, stores and acts upon information from its environment. Consequently, cognitive traits potentially undergo sexual selection in some mating systems. We investigated the role of cognitive traits on the reproductive performance of male rose bitterling (Rhodeus ocellatus), a freshwater fish with a complex mating system and alternative mating tactics. We quantified the learning accuracy of males and females in a spatial learning task and scored them for learning accuracy. Males were subsequently allowed to play the roles of a guarder and a sneaker in competitive mating trials, with reproductive success measured using paternity analysis. We detected a significant interaction between male mating role and learning accuracy on reproductive success, with the best-performing males in maze trials showing greater reproductive success in a sneaker role than as a guarder. Using a cross-classified breeding design, learning accuracy was demonstrated to be heritable, with significant additive maternal and paternal effects. Our results imply that male cognitive traits may undergo intra-sexual selection., (© 2015 The Author(s) Published by the Royal Society. All rights reserved.)

Bitterlings are relatively small cypriniform species and extremely interesting evolutionarily due to their unusual reproductive behaviors and their coevolutionary relationships with freshwater mussels. As a group, they have attracted a great deal of attention in biological studies. Understanding the origin and evolution of their mating system demands a well-corroborated hypothesis of their evolutionary relationships. In this study, we provide the most comprehensive phylogenetic reconstruction of species relationships of the group based on partitioned maximum likelihood and Bayesian methods using DNA sequence variation of nuclear and mitochondrial genes on 41 species, several subspecies and three undescribed species. Our findings support the monophyly of the Acheilognathidae. Two of the three currently recognized genera are not monophyletic and the family can be subdivided into six clades. These clades are further regarded as genera based on both their phylogenetic relationships and a reappraisal of morphological characters. We present a revised classification for the Acheilognathidae with five genera/lineages: Rhodeus, Acheilognathus (new constitution), Tanakia (new constitution), Paratanakia gen. nov., and Pseudorhodeus gen. nov. and an unnamed clade containing five species currently referred to as "Acheilognathus". Gene trees of several bitterling species indicate that the taxa are not monophyletic. This result highlights a potentially dramatic underestimation of species diversity in this family. Using our new phylogenetic framework, we discuss the evolution of the Acheilognathidae relative to classification, taxonomy and biogeography., (Copyright © 2014 Elsevier Inc. All rights reserved.)

In some taxa, males perform multiple ejaculations, which may function in sperm competition or in maintaining a baseline density of spermatozoa in the female reproductive tract to ensure fertilization, a process that has been termed 'topping up'. We investigated the function of multiple ejaculations in two species of bitterling, the European bitterling (Rhodeus amarus) and Chinese rose bitterling (Rhodeus ocellatus). Bitterling oviposit in living freshwater mussels, with fertilization taking place within the mussel gill cavity. Thus, although fertilization is external, the mussel is analogous to the female reproductive tract in an internally fertilizing species. We measured the frequency of ejaculations and mussel inspections by individual males of two bitterling species in 28 replicated mesocosms and examined focal male responses to rival ejaculations and the presence of females in spawning condition. We used a model of ejaculatory behaviour to simulate the temporal abundance of spermatozoa in mussels. Male R. amarus exhibited high rates of ejaculation and inspection of the siphons of mussels and increased their ejaculation rate in response to the presence of females in spawning condition. Rhodeus ocellatus showed lower overall rates of ejaculation, but significantly elevated ejaculation rate in response to rival ejaculations. The ejaculatory strategy of R. amarus is one that maintains a minimum level of spermatozoa in mussels, which is elevated when the probability of oviposition increases. In contrast, R. ocellatus engages more directly in sperm competition with rivals. We discuss these results in the context of the function of multiple ejaculations and male mating tactics., (© 2014 The Authors. Journal of Evolutionary Biology © 2014 European Society For Evolutionary Biology.)

In this study, the effects of the eye fluke Diplostomum pseudospathaceum (Trematoda) infection on over-winter survival of young-of-the-year (YOY) European bitterling Rhodeus amarus (Cyprinidae) were examined between September 2010 and April 2011. The fish were reared in semi-natural conditions to ensure that results were not confounded by other parasite infections. The cumulative mortality of R. amarus from November until April was significantly higher in D. pseudospathaceum-infected fish (57·3%) compared to controls (42·1%). Infection of the parental generation did not have any effect on the mortality of juveniles. The results indicate that D. pseudospathaceum infection increases over-winter mortality of YOY R. amarus. The possible mechanisms causing mortality are discussed., (© 2014 The Fisheries Society of the British Isles.)

We examined the relationship between muscle and scale mercury (Hg) and lead (Pb) concentrations in chub Squalius cephalus L. from six riverine sites in the Czech Republic in order to determine whether scale analysis alone could provide a nonlethal and convenient method for prediction of heavy metal concentration in muscle tissue. Our results confirm tissue-specific heavy metal accumulation in chub, with Hg tending to accumulate primarily in muscle tissue and Pb in scales. We found no significant relationship, however, for concentrations of either Pb or Hg between muscle tissue and scales of chub. Our results indicate that scales cannot be used for predicting heavy metal contamination in muscle of chub and we recommend, therefore, that muscle biopsy methods continue as the preferred method of analysis.

The impact of multiple invading species can be magnified owing to mutual facilitation--termed 'invasional meltdown'--but invasive species can also be adversely affected by their interactions with other invaders. Using a unique reciprocal host-parasite relationship between a bitterling fish (Rhodeus amarus) and unionid mussels, we show that an invasive mussel reverses the roles in the relationship. Bitterling lay their eggs into mussel gills, and mussel larvae parasitize fish. Bitterling recently colonized Europe and parasitize all sympatric European mussels, but are unable to use a recently invasive mussel, Anodonta woodiana. The parasitic larvae of A. woodiana successfully develop on R. amarus, whereas larvae of European mussels are rejected by bitterling. This demonstrates that invading species may temporarily benefit from a coevolutionary lag by exploiting evolutionarily naive hosts, but the resulting relaxed selection may facilitate its exploitation by subsequent invading species, leading to unexpected consequences for established interspecific relationships.

The role of genetic benefits in female mate choice remains a controversial aspect of sexual selection theory. In contrast to "good allele" models of sexual selection, "compatible allele" models of mate choice predict that females prefer mates with alleles complementary to their own rather than conferring additive effects. While correlative results suggest complementary genetic effects to be plausible, direct experimental evidence is scarce. A previous study on the Chinese rose bitterling (Rhodeus ocellatus) demonstrated a positive correlation between female mate choice, offspring growth and survival, and the functional dissimilarity between the Major Histocompatibility Complex (MHC) alleles of males and females. Here we directly tested whether females used cues associated with MHC genes to select genetically compatible males in an experimental framework. By sequentially pairing females with MHC similar and dissimilar males, based on a priori known MHC profiles, we showed that females discriminated between similar and dissimilar males and deposited significantly more eggs with MHC dissimilar males. Notably, the degree of dissimilarity was an important factor for female decision to mate, possibly indicating a potential threshold value of dissimilarity for decision making, or of an indirect effect of the MHC.

Coevolutionary relationships between parasites and hosts can elevate the rate of evolutionary changes owing to reciprocal adaptations between coevolving partners. Such relationships can result in the evolution of host specificity. Recent methodological advances have permitted the recognition of cryptic lineages, with important consequences for our understanding of biological diversity. We used the European bitterling (Rhodeus amarus), a freshwater fish that parasitizes unionid mussels, to investigate host specialization across regions of recent and ancient sympatry between coevolving partners. We combined genetic data (12 microsatellite and 2 mitochondrial markers) from five populations with experimental data for possible mechanisms of host species recognition (imprinting and conditioning). We found no strong evidence for the existence of cryptic lineages in R. amarus, though a small proportion of variation among individuals in an area of recent bitterling-mussel association was statistically significant in explaining host specificity. No other measures supported the existence of host-specific lineages. Behavioural data revealed a weak effect of conditioning that biased behavioural preferences towards specific host species. Host imprinting had no effect on oviposition behaviour. Overall, we established that populations of R. amarus show limited potential for specialization, manifested as weak effects of host conditioning and genetic within-population structure. Rhodeus amarus is the only species of mussel-parasitizing fish in Europe, which contrasts with the species-rich communities of bitterling in eastern Asia where several host-specific bitterling occur. We discuss costs and constraints on the evolution of host-specific lineages in our study system and more generally., (© 2011 Blackwell Publishing Ltd.)

Susceptibility to parasite infection was examined in a field experiment for four populations of 0+ juvenile European bitterling (Rhodeus amarus): one sympatric to local parasite fauna, one allopatric, and two hybrid populations. Significantly higher parasite abundance was recorded in the allopatric bitterling population, suggesting a maladaptation of parasites to their sympatric host. Type of parasite life cycle played an important role in host-parasite interactions. While the abundance of allogenic species between populations was comparable, a significant difference was found in abundance of autogenic parasite species between fish populations, with the allopatric population more infected. These results correspond with a prediction of higher dispersion probability and higher gene flow among geographically distant populations of allogenic species as compared to autogenic species. Increased susceptibility to parasites that do not occur within the natural host's geographical distribution was found in the allopatric host, but only for autogenic species. A difference in infection susceptibility was detected among populations of early-hatched bitterling exposed to infection during a period of high parasite abundance and richness in the environment. Differences in parasite abundance and species diversity among populations diminished, however, with increasing time of exposure. No difference was found within late-hatched populations, probably due to a lower probability of infection in late-hatched cohorts.

The seasonal dynamics of biological traits in a population of European bitterling Rhodeus amarus from a small river in central Europe (River Kyjovka, Czech Republic) were investigated from April 2007 to March 2008. The reproductive season lasted from early April to mid-June, with a peak in late April when > 50% of females possessed ripe eggs in their ovaries. The sex ratio was female-biased, with a higher bias following the peak in reproduction, suggesting higher mortality of males than females over the reproductive period. The population was effectively annual, with a major decrease in adult fish abundance at the end of the reproductive period. Highest relative lipid content and condition factor were observed in February, both decreasing with an increase in investment into gonads. Seasonal dynamics in somatic condition, relative lipid content and gonad mass, and their comparison with other R. amarus populations, suggest that the study population allocated a disproportionally high level of resources to reproduction., (© 2011 The Authors. Journal of Fish Biology © 2011 The Fisheries Society of the British Isles.)

An understanding of recent evolutionary processes is essential for the successful conservation and management of contemporary populations, especially where they concern the introduction or invasion of species outside their natural range. However, the potentially negative implications of intraspecific introductions and invasions have attracted less attention, although they also represent a potential threat to biodiversity, and are commonly facilitated through human activities. The European bitterling (Rhodeus amarus) is a small cyprinid fish that decreased greatly in its distribution during the 1970s and 1980s and was subsequently included on many European conservation lists. This decline appears to have reversed, and the extent of its distribution now exceeds its former range. We used a combination of 12 microsatellite markers and cytochrome b sequences on a large data set (693 individuals) across the current range of the European bitterling to investigate possible scenarios for its colonization of Europe. We show that the inferred history of colonization of Europe was largely congruent between mitochondrial and nuclear markers. The most divergent mtDNA lineages occur in the Aegean region but probably are not reproductively isolated as the Aegean populations also displayed mtDNA haplotypes from other lineages and nuclear data indicated their close relationship to Danubian populations. Much of Europe is currently populated by descendants of two main lineages that came to natural secondary contact in western Europe. An approximate Bayesian computation analysis indicates different dates for admixture events among western and central European populations ranging from the last deglaciation (natural) to the last few centuries (human-assisted translocations)., (© 2010 Blackwell Publishing Ltd.)

Multiple stressor scenarios, as they are relevant in many watersheds, call for approaches extending beyond conventional chemical-focused approaches. The present study, investigated the fish population, represented by chub (Leuciscus cephalus), in the Bílina River (Czech Republic), which is impacted by various pollution sources and might pose a risk on the fish population. To confirm or reject this hypothesis it was examined whether there exists an association between abundance of chub and exposure to toxic chemicals as well as natural stressors, represented by parasites, and whether health-related suborganismal traits, namely, organ indices, tissue histopathology, and immune parameters, would help in revealing relationships between stressor impact and population status. Toxic pressure was assessed by the toxic unit approach, which gives an integrative estimate of toxic effect concentrations and by measuring the biomarkers cytochrome P4501A and vitellogenin, which indicate exposure to bioavailable arylhydrocarbon- or estrogen receptor ligands. Parasite pressure was estimated by determining abundance and species composition of ecto- and endoparasites of chub. Chub abundance was high upstream in the Bílina, low to zero in the middle stretches, and increased again downstream. Toxic pressure increased in the downstream direction, while parasite intensity decreased in this direction. Health status of chub did not differ clearly between up-, middle-, and downstream sites. Thus, it appears that neither toxic pressure nor parasite pressure nor their combination translates into a change of chub health status. By using varied assessment tools, this study provides evidence against a presumed causative role of toxicants impairing the fish ecological status of the Bílina River., (Copyright 2009 SETAC.)

Morphological (meristic and morphometric traits), karyological and molecular (microsatellites, cytochrome b) analyses were performed to characterize a hybrid of the roach Rutilus rutilus and nase Chondrostoma nasus. Meristic and morphometric traits were different between hybrid and both parental species. The number of chromosomes found in hybrid specimen indicated that this individual represents the post-F(1) generation of hybrids and the microsatellite analysis of the hybrid showed the presence of variants typical for R. rutilus and C. nasus.

A 15 year data set revealed that the abundance of 0+ year Rhodeus amarus in a regulated river was negatively associated with mean river discharge, while mean standard length was positively related to the cumulative number of degree-days > or =10 degrees C. Results are discussed in the context of the recent invasion success of R. amarus.

The phylogenetic relationships among gudgeons that represent most nominal taxa within Gobio gobio sensu lato were examined by mitochondrial and nuclear genome sequencing. The molecular analyses confirmed the separate generic status of Gobio as a monophyletic group and revealed 15 Eurasian lineages divided into two main clades, the Northern European and the Ponto-Caspian. The validity of eleven nominal taxa as distinct species was confirmed, gudgeons from the Volga River basin were described as a new species G. volgensis, and three revealed phylogenetic lineages were submitted for a comprehensive revision as "species-in-waiting". The species G. gobio showed a wide range extending from the British Isles to the Black Sea coast and overlapped the areas of several other species. Four pure lineages were detected in the middle Danube River basin. The Crimean Peninsula was found to be a region with the occurrence of individuals of hybrid origin. This region will require special investigation to define species participating in hybridization events, and to establish further steps for the conservation of endemic native gudgeon species. A simple diagnostic method, based on different lengths of the PCR products, called "S7indel diagnostics" is presented for further taxonomic investigations in the genus Gobio.

Environmental and population parameters that influence the strength of sexual selection may vary considerably over the course of the reproductive season. However, the potential for sexual selection frequently fails to translate into variation in reproductive success among individuals. We investigated seasonal changes in variation in reproductive success, measured as the opportunity for sexual selection, using parentage analysis in 20 experimental populations of the European bitterling (Rhodeus amarus, Cyprinidae), a small freshwater fish with a promiscuous, resource-based mating system. We showed that although the largest males sired most offspring over the entire reproductive season, variation in reproductive success and hence the opportunity for sexual selection was low at the start of the season but increased significantly at its end. This seasonal difference probably arose from the superior competitive endurance of large males and from a higher temporal clustering of reproductively active females at the start of the breeding season than later in the season. The spatial distribution of oviposition sites had a negligible effect on the variation in reproductive success. We discuss the potential implications of our results for the importance and strength of sexual selection in natural populations.

Interspecific symbiotic relationships involve a complex network of interactions, and understanding their outcome requires quantification of the costs and benefits to both partners. We experimentally investigated the costs and benefits in the relationship between European bitterling fish (Rhodeus sericeus) and freshwater mussels that are used by R. sericeus for oviposition. This relationship has hitherto been thought mutualistic, on the premise that R. sericeus use mussels as foster parents of their embryos while mussels use R. sericeus as hosts for their larvae. We demonstrate that R. sericeus is a parasite of European mussels, because it (i) avoids the cost of infection by mussel larvae and (ii) imposes a direct cost on mussels. Our experiments also indicate a potential coevolutionary arms race between bitterling fishes and their mussel hosts; the outcome of this relationship may differ between Asia, the centre of distribution of bitterling fishes, and Europe where they have recently invaded.

Sexual selection involves two main mechanisms: intrasexual competition for mates and intersexual mate choice. We experimentally separated intrasexual (male-male interference competition) and intersexual (female choice) components of sexual selection in a freshwater fish, the European bitterling (Rhodeus sericeus). We compared the roles of multiple morphological and behavioural traits in male success in both components of sexual competition, and their relation to male reproductive success, measured as paternity of offspring. Body size was important for both female choice and male-male competition, though females also preferred males that courted more vigorously. However, dominant males often monopolized females regardless of female preference. Subordinate males were not excluded from reproduction and sired some offspring, possibly through sneaked ejaculations. Male dominance and a greater intensity of carotenoid-based red colouration in their iris were the best predictors of male reproductive success. The extent of red iris colouration and parasite load did not have significant effects on female choice, male dominance or male reproductive success. No effect of parasite load on the expression of red eye colouration was detected, though this may have been due to low parasite prevalence in males overall. In conclusion, we showed that even though larger body size was favoured in both intersexual and intrasexual selection, male-male interference competition reduced opportunities for female choice. Females, despite being choosy, had limited control over the paternity of their offspring. Our study highlights the need for reliable measures of male reproductive success in studies of sexual selection.

The reproductive success of alternative mating behaviours may vary within and among populations in relation to environmental factors and demographic parameters. We used behavioural and genetic data to investigate how male density affects reproductive success of territoriality and sneaking in the European bitterling (Rhodeus sericeus, Cyprinidae), a freshwater fish that spawns on the gills of living freshwater mussels. Keeping the number of spawning sites constant, we manipulated male densities in laboratory and mesocosm experiments. We showed that sneaked fertilizations were common in R. sericeus, and that they increased significantly with male density. Territorial mating was almost 17 times more successful than sneaking at the lowest male density treatment, and still 2-3 times more successful at intermediate densities. However, both behaviours conferred the same fitness pay-off at the highest male density. While the success of territorial males declined with male density, the success of individual sneaking males remained constant across densities. Notably, the capacity of territorial males to outcompete sneakers by preoviposition sperm loading was the best predictor of male reproductive success, rather than aggression, body size or postoviposition ejaculation.