Adaptive spatial utilization of host mussels by the Japanese rosy bitterling Rhodeus ocellatus kurumeus

The spatial pattern of resource utilization for oviposition in the Japanese rosy bitterling Rhodeus ocellatus kurumeus was investigated in two field experiments. The distribution of rosy bitterling eggs deposited in the four demibranchs of the gills of the test mussel species, Anodonta woodiana , differed with mussel reproductive state in pair spawnings, but not in group spawnings. In pair spawnings, female rosy bitterling may have had more time to select the site of oviposition in the gill in relation to the sex and reproductive state of the mussel, thereby maximizing embryo survival. Thus, the inner gill of female mussels may have been selected in preference to the outer gills to avoid mortalities of eggs due to the presence of the mussel's own embryos (glochidia) in the outer gill chambers. In male mussels, female rosy bitterling distributed their eggs equally among all parts of the mussel gill, thereby minimizing density-dependent mortality of embryos. During group spawnings, however, female rosy bitterling may have been more constrained in their decision making, ovipositing in the inner gills irrespective of mussel sex.     

The bitterling–mussel symbiosis: a model for host‐parasite adaptation

Bitterlings (Acheilognatinae) are a monophyletic group of cyprinid fishes that lay their eggs in the gill chamber of freshwater mussels. They have evolved many behavioural, morphological and physiological adaptations to the symbiosis. Female bitterling develop a long ovipositor that insert into the exhalant siphon of a mussel and males fertilize the eggs by releasing sperm over the inhalant siphon of the mussel. Embryos hatch within 2 days but develop inside the mussel for further 3 to 6 weeks. Embryos are adapted to the low oxygen environment in the mussel's gill chamber. Both males and females discriminate among mussels in relation to their quality as host for developing embryos. On the other hand, mussels used for oviposition have larvae that obligate ectoparasites on fish. Here I review current knowledge on the status of the symbiosis, developmental and behavioural adaptations by bitterling and mussel and summarize costs and benefits to both symbionts. Further, I use a recent well‐resolved bitterling phylogeny to emphasize the potential of this model system to study the evolution of this symbiosis, which is a part of the ongoing study.     

A proximate cue for oviposition site choice in the bitterling (Rhodeus sericeus)

1. We investigated two possible proximate cues used for oviposition site choice by females of the bitterling ( Rhodeus sericeus ), a freshwater fish that spawns on the gills of live unionid mussels. The two cues were the flow velocity and/or oxygen content of water emerging from the exhalant siphon of a mussel.
2. Field observations showed that female bitterling always inspected the exhalant siphons of mussels before they spawned in them. Siphon inspection was not always a prelude to spawning and it may serve as a means of assessing mussel quality. Female skimming behaviour, swimming over a mussel without spawning, may also be used to assess mussel quality, although the mechanism for this is unclear.
3. Measurements of the flow velocity of water emerging from the exhalant siphons of four mussel species ( Anodonta anatina, A. cygnea, Unio pictorum and U. tumidus ) showed a significant difference among species, with U. tumidus having the highest mean flow velocity and U. pictorum the lowest.
4. Measurements of the change in oxygen concentration of water entering a mussel inhalant siphon and leaving its exhalant siphon in field and laboratory studies showed a significant difference among the four mussel species, with A. cygnea exhibiting a significantly higher change in oxygen concentration than the other species.
5. The presence of bitterling embryos in the gills of a mussel significantly increased its oxygen consumption whereas larval glochidia had no significant effect. We discuss oxygen availability as a possible proximate cue for oviposition site choice in bitterling.     

The bitterling–mussel interaction as a test case for co‐evolution

The European bitterling Rhodeus sericeus (Cyprinidae) spawns in the gills of freshwater mussels (Unionidae) and shows some obvious adaptations to this type of spawning, such as the development of an ovipositor. Furthermore, recent studies have shown that the fish avoid species of mussels that have a high likelihood of ejecting their eggs prematurely. This leads to the question of whether the interaction between bitterling and mussels could represent a case of co‐evolution, involving evolutionary responses by both species to selection imposed by the other. The evidence for and against co‐evolution is reviewed, incorporating new results from two sets of experiments designed to test for adaptive choices by bitterling according to the mussels' sex and reproductive state, as well as a preliminary study of potential benefits for mussels from exposure to bitterling. Host preferences by bitterling, both among and within mussel species, may indeed have evolved in response to differences in benefits for offspring survival. There is no evidence yet for any benefits to mussels from receiving eggs, whereas there are costs due to reduced ventilation rates when the gills contain bitterling eggs. While there are differences among mussel species and individuals in their tendency to reject bitterling embryos, these differences do not provide strong evidence for co‐evolution. For example, they may reflect differences in host physiology such as ventilation rate and generalized responses to expelling objects from their gills. Therefore, while bitterling are well adapted for their obligate spawning relationship with mussels, it has been much more difficult to find evidence for adaptations by mussels for dealing with bitterling. This suggests that any co‐evolutionary dynamics between bitterling and mussels may be asymmetric, with stronger responses to selection by the fish than by mussels.     

Reproductive ecology of the striped bitterling Acheilognathus cyanostigma (Cyprinidae: Acheilognathinae)

The reproductive ecology of the striped bitterling Acheilognathus cyanostigma was investigated in a small pond in Mie Prefecture, central Honshu, Japan. A. cyanostigma was the only species of bitterling in the study pond, where only a single species of mussel Anodonta woodiana was present. Spawning of A. cyanostigma was recorded between early April and early July, peaking between late April and mid-June in 2003. Ovipositor length during the spawning period ranged from 19.2 to 42.8 mm, and was positively correlated with female body length, but did not show significant seasonal variation. The eggs are elliptical with a volume of approximately 1.5 mm³. Egg size correlates positively with female body size, and both egg shape and volume changed significantly with season. The embryos were located on host mussel gills approximately 30 mm from the exhalant siphon and were found more frequently on the inner rather than the outer mussel gills. The possible ecological significance of these observations is discussed in the context of the adaptations of A. cyanostigma for utilizing mussels for oviposition.     

Histological structure of the female gonads and ovipositor of the European bitterling,Rhodeus amarus (Bloch, 1782) (Cyprinidae: Acheilognathinae)

We investigated the histological structure of the female gonads and ovipositor of the European bitterling, Rhodeus amarus. The base of the ovipositor was formed by the conical organ or ‘eminence’. Based on the structure of the conical organ, including a special distribution of collagenous and muscular fibrils, a well‐developed vascular system and numerous scyphoid mucous cells in the internal epithelium, this organ was used for temporary storage of oocytes during the spawning period and for their movement along the ovipositor. An extensive network of blood vessels, muscular fibrils and numerous collagenous fibrils in the connective tissue of the ovipositor may make a functional contribution to the ovipositor by making it firmer during egg laying. Mucous cells were detected in the medial and distal regions of the ovipositor, which may play a role in facilitating insertion of the ovipositor into the exhalant siphon of a mussel during oviposition. European bitterling are batch spawners, and the female spawns eggs in clutches at intervals during the breeding season, which were visible as three distinct cohorts of oocytes in the ovary.     

Factors affecting seasonal mortality of rosy bitterling (Rhodeus ocellatus kurumeus) embryos on the gills of their host mussel

I investigated the seasonal change in factors affecting embryonic mortality in the rosy bitterling, Rhodeus ocellatus kurumeus, a freshwater fish that spawns on the gills of living unionid mussels. Research was conducted in a small pond during 1999 and 2001 in which bitterling were provided with Anodonta sp. mussels for spawning. Bitterling spawned between April and July, peaking mid–late May. Seasonal survival rate of bitterling embryos in their mussel hosts was unimodal, with a peak between late April and mid May (about 70% of total spawnings). In mid April, survival was about 50%. The lowest survival was from late May to July (0%). Losses of bitterling embryos from mussels were identified by ejections from the mussel host. Ejections were categorized as either ejections of live embryos, or ejections of embryos that died in the mussel and were subsequently expelled from the mussel. Ejection rates of live embryos were higher in the earlier part of the spawning period (early–mid April) and dead embryo ejections in the later period (after June). The ejection rate of live embryos was higher among younger embryos earlier in the season, probably because of the incomplete development of morphological and behavioural traits associated with maintaining the embryo inside the mussel gill chambers, and as a consequence of a more protracted developmental period at low temperatures making them more susceptible to ejection. The ejection rate of dead embryos was higher in older embryos later in the season, and in larger mussels and at high embryo densities. The survival of embryos in mussels was probably related to oxygen availability, with mortalities probably caused by asphyxiation. Increased embryo mortalities may arise through competition among embryos, between embryos and mussel, and ambient dissolved oxygen levels. The optimal period for bitterling to spawn may represent a balance between two opposing factors; with positive and negative effects of a seasonal rise in temperature directly affecting embryonic growth rate and oxygen availability.An erratum to this article can be found at This revised version was published online January 2005 with the correction of the authors name.     

A possible evolutionary lag in the relationship between freshwater mussels and European bitterling

The coevolutionary dynamics between European bitterling Rhodeus amarus and freshwater unionid mussels, which the former parasitize by laying eggs on their gills, were tested. In a series of experiments fish preferences and mussel responses were compared in parasites and hosts of recent (Europe) and ancient (Asia) sympatry. Rhodeus amarus readily oviposited on the gills of all mussel species tested. Fish that laid their eggs on the gills of Asian Anodonta woodiana, however, suffered a dramatic reduction in reproductive success compared to fish that oviposited on the gills of European mussels: Unio pictorum , Unio tumidus , Anodonta anatina and Anodonta cygnea . This difference was the result of egg ejection behaviour by mussels rather than the unsuitability of the internal gill environment for European bitterling embryo development. The ejection response of mussels with a long sympatry with European bitterling was considerably more pronounced than that of mussels with a substantially shorter sympatry. The data support a coevolutionary arms race between bitterling and mussels and point to an evolutionary lag in the relationship between R. amarus and its European mussel hosts.     

The European bitterling: a model for oviposition decision

For oviparous species, oviposition decisions can have significant fitness consequences. The European bitterling ( Rhodeus sericeus ) is a cyprinid fish that lays its eggs on the gills of freshwater mussels. Because bitterling use a discrete spawning site that can be readily manipulated, they are unusually amenable to field and laboratory studies aimed at understanding the adaptiveness and consequences of oviposition decisions. Here I present data demonstrating the adaptiveness of oviposition decisions by female bitterling, and link these decisions, using game‐theory models, with bitterling population dynamics. I explain the proximate cues used by females in making oviposition choices, and how these respond to environmental variables. I also show how the oviposition decisions of males relate to the risk of sperm competition in mussels, and propose that oviposition choices may represent an inter‐sexual conflict in bitterling. Finally, I present behavioural and genetic data from mesocosm experiments to show the implications of global and local male densities for female oviposition opportunities, and consider the consequences for bitterling population dynamics.     

Bayesian inference supports the host selection hypothesis in explaining adaptive host specificity by European bitterling

Generalist parasites have the capacity to infect multiple hosts. The temporal pattern of host specificity by generalist parasites is rarely studied, but is critical to understanding what variables underpin infection and thereby the impact of parasites on host species and the way they impose selection on hosts. Here, the temporal dynamics of infection of four species of freshwater mussel by European bitterling fish (Rhodeus amarus) was investigated over three spawning seasons. Bitterling lay their eggs in the gills of freshwater mussels, which suffer reduced growth, oxygen stress, gill damage and elevated mortality as a result of parasitism. The temporal pattern of infection of mussels by European bitterling in multiple populations was examined. Using a Bernoulli Generalized Additive Mixed Model with Bayesian inference it was demonstrated that one mussel species, Unio pictorum, was exploited over the entire bitterling spawning season. As the season progressed, bitterling showed a preference for other mussel species, which were inferior hosts. Temporal changes in host use reflected elevated density-dependent mortality in preferred hosts that were already infected. Plasticity in host specificity by bitterling conformed with the predictions of the host selection hypothesis. The relationship between bitterling and their host mussels differs qualitatively from that of avian brood parasites.