Pearl Mussel Research Articles

The revenant: rediscovery of Margaritifera homsensis from Orontes drainage with remarks on its taxonomic status and conservation (Bivalvia: Margaritiferidae)

Since Margaritifera marocana (Pallary, 1918) and M. laosensis (Lea, 1863) were rediscovered, M. homsensis (Lea, 1865) remains the only pearl mussel species known solely based on old shell samples from natural history museums. This is also the last pearl mussel species, which is absent in a phylogeny of the family. Here, we aimed to provide an integrative revision of the taxonomic status of M. homsensis from the Orontes Basin. Using a newly collected specimen from the River Karasu, Hatay Province, southern Turkey, five gene partitions were sequenced, the cytochrome c oxidase subunit I (COI), large ribosomal subunit rRNA (16S), large ribosomal subunit rDNA (28S) and its D3 expansion segment (D3), and small ribosomal subunit rDNA (18S). The multi-gene phylogeny indicates that M. homsensis is a sister taxon of M. auricularia, but both these species are closely related to M. marocana by nuclear genes. The main conchological features, i.e., the shell shape, teeth morphology, and mantle attachment scars, as well as Fourier shell shape analysis have not shown principal differences between M. homsensis and M. auricularia. Based on these data, we concluded that M. homsensis is a valid species that is most closely related to M. auricularia. Special conservation efforts for a population of M. homsensis discovered in Turkey, including the formation of a nature reserve, might contribute to the conservation of the species. Finally, an extensive search for surviving populations in Orontes drainage (southern Turkey, Lebanon, and Syria) and the Nahr-el-Kabir River (Lebanon and Syria) remains necessary to develop a transboundary conservation strategy for this unique taxon.

A qPCR MGB probe based eDNA assay for European freshwater pearl mussel (Margaritifera margaritifera L.)

Abstract1. Environmental (e)DNA assays are becoming increasingly used to detect rare or invasive aquatic species.2. The Critically Endangered freshwater pearl mussel Margaritifera margaritifera is undergoing range‐wide reduction in population numbers and distribution.3. An eDNA assay to detect the presence of M. margaritifera was developed, based on the mitochondrial cytochrome oxidase I gene, utilizing species‐specific primers, a minor groove binding (MGB) probe and quantitative (q)PCR approaches.4. The results from this pilot study demonstrated high sensitivity both in laboratory and field trials, and provide a valuable non‐invasive tool for detecting M. margaritifera.

Estimates of genetic parameters and genotype by environment interactions for shell nacre color and growth traits in the purple freshwater pearl mussel Hyriopsis cumingii

Hyriopsis cumingii, one of the most commercially important freshwater pearl mussel in China, produced most of white and orange cultured pearls. However, purple freshwater cultured pearls, with higher commercial value, are the most popular due to its rarity. For increasing the yields of purple pearl, one selection breeding program for improving shell nacre color were developed in H. cumingii. Genetic parameters and genotype by environment interactions for shell nacre color and growth traits were estimated from the fifth-generation purple selective strain of H. cumingii reared at two sites (Chongming and Jinhua). Four shell nacre color parameters and six shell growth traits were recorded by 1142 individuals at 12-month-old. Microsatellite-based DNA parentage analysis was used to assign the mussels to 13 paternal half-sib families, including 37 full-sib families. Genetic diversity was found to be high in the selected strain, with a mean expected heterozygosity at 6 microsatellite loci equal to 0.791. Heritability estimates of four shell nacre color parameters, lightness (L*), redness (a*), yellowness (b*), and total color change (dE) were 0.33 ± 0.19, 0.17 ± 0.10, 0.69 ± 0.10, and 0.33 ± 0.19, respectively. Heritability estimates were 0.20 ± 0.03 for shell length, 0.16 ± 0.06 for shell height, 0.17 ± 0.02 for shell width, 0.23 ± 0.01 for body weight, 0.15 ± 0.06 for mantle weight, and 0.31 ± 0.03 for shell weight. No significant phenotypic or genetic correlation was found between shell nacre color parameters and growth traits. All shell nacre color parameters and growth traits, except for shell nacre color parameter b*, exhibited may be significant genotype by environment interactions at the two sites. The results suggest that shell nacre color parameters may be treated as selective traits for further improving shell nacre color. And shell nacre color may be selected for independently of selection on growth performance. Some considering to the likely genotype by environment interactions for shell nacre color and growth traits should be taken in future selective breeding programs for large-scale culture practices.

Within-river variation in growth and survival of juvenile freshwater pearl mussels assessed by in situ exposure methods

The early post-parasitic phase is considered to be the most vulnerable life stage of the endangered freshwater pearl mussel (Margaritifera margaritifera), yet little is known about the spatial variability in juvenile performance at a river scale. The main aim of this study was to determine within-river variation in habitat suitability for juvenile M. margaritifera. The growth and survival rates of juveniles (one and two year old) were measured in the Vltava River (Czech Republic) in relation to several physical and chemical variables using a total of 166 bioindication units (individual mesh cages and sandy boxes) during two thermally different vegetation seasons. Three spatial scales were studied: a longitudinal river profile, bottom habitat types, and hyporheic microhabitats. Both the survival rate and growth of the exposed juveniles varied considerably within the studied river. The effect of the longitudinal river profile and the roles of temperature, oxygenation within microhabitats, episodic pollution, and exposure method were demonstrated. This study represents the first example of a hierarchized approach to the assessment of a riverine mussel biotope with the use of bioindication methods. The results demonstrate the importance of the scale and bioindication method used in understanding the suitability of a river environment to juvenile freshwater mussels.

Atlantic salmon (Salmo salar) and brown trout (Salmo trutta) differ in their suitability as hosts for the endangered freshwater pearl mussel (Margaritifera margaritifera) in northern Fennoscandian rivers

Abstract European populations of the freshwater pearl mussel (FPM, Margaritifera margaritifera) have collapsed across much of the species’ geographic range and, despite many types of conservation intervention, the number of successful restoration efforts has been low. The goal of this study was to determine whether there were population‐specific differences in the suitability of Atlantic salmon (Salmo salar) and brown trout (S. trutta) as hosts for the parasitic glochidium larvae of FPM. We predicted that such differences would depend on the historical occurrence of these salmonid species in FPM habitats. We studied the potential host specificity both in the field and in laboratory by exposing salmonid fish to FPM glochidia originating from nine populations in different types of streams and rivers within three large river basins in northern Fennoscandia. The exposures showed remarkable population‐specific differences in the suitability of host species. In large main channels, previously colonised by Atlantic salmon but mostly now dammed for hydropower production, the occurrence of FPM glochidia was always highest on salmon. Moreover, glochidia on salmon were often larger than the conspecifics on brown trout. Conversely, in small tributaries with no salmon history, brown trout was generally the best, or the only suitable, host for FPM. The adaptation of certain FPM populations to salmon is a highly significant finding, which offers — together with the hydropower dam construction and the salmonid stocking practices that often favour brown trout — an explanation for the collapse of FPM populations living in former Atlantic salmon rivers. This study illustrates the indirect but substantial effects that dams and changes in fish communities may induce, and emphasises the need to determine the most suitable host species for each FPM population in order to inform conservation and management actions.

Molecular characterization of two distinct Smads gene and their roles in the response to bacteria change and wound healing from Hyriopsis cumingii

The proteins of Smad family are critical components of the TGF-β superfamily signal pathway. In this paper, we cloned two intracellular mediators of TGF-β signaling, Smad3 and Smad5, from the pearl mussel Hyriopsis cumingii. The full length cDNA of HcSmad3 and HcSmad5 were 2052 bp and 1908 bp and encoded two polypeptides of 418 and 461amino acid residues, respectively. The deduced amino acid of HcSmad3 and HcSmad5 possessed two putative conserved domains, MH1 and MH2, a conserved phosphorylation motif SSXS at the carboxyl-terminal. The two Smad genes were detected muscle, mantle, hepatopancreas and gill, but with a very low level in heamocytes. The transcripts of Smad3 and Smad5 were up-regulated in hemocytes and hepatopancreas after A. hydrophila and PGN stimulation. However, the expression of Smad3 and Smad5 were only up-regulated in hepatopancreas after A. hydrophila stimulation. The transcripts of Smad3 and Smad5 had a slight change in hepatopancreas after PGN stimulation. The transcripts of HcSmad3 showed very little increase and HcSmad5 mRNA significantly up-regulated after wounding.

MALACOFAUNA OF THE KIVACH STRICT NATURE RESERVE

The malacofauna of the Kivach Strict Nature Reserve was studied. In total, 25 species of mollusks were found. This number included terrestrial gastropods 9 species, aquatic pulmonate gastropods 9 species, prosobranch gastropods 2 species, bivalves 5 species. For eight species the nature reserve's territory is close to the northern limit of their distribution in European Russia, and for two species it is close to the southern limit. Among the terrestrial gastropods one species (Arion subfuscus) prefers forest habitats; one species (Succinea putris) prefers living on wet land, but is also found in water on the trunks and branches of submerged trees; one species (Deroceras reticulatum) is synanthropic, found only in a residential area; and the rest prefer open sites. All the aquatic gastropods found in the nature reserve live in stagnant or slow-flowing water bodies. All the observed species of bivalves belong to the superfamily Unionacea. Three species live in lakes and river pools with slow stream. Two bivalve species (Unio crassus and Margaritifera margaritifera) inhabit rapids in the Suna River. The latter two species are red-listed. In fact, the freshwater pearl mussel Margaritifera margaritifera used to inhabit the Suna River in the past but went extinct as a result of human activities. The present-day population of M. margaritifera includes 400 individuals. These mussels were translocated from the Nemina River within the program for restoration of this species. A list of species of mollusks described from Karelia but not found in Kivach nature reserve is presented. It is quite likely, however, that some of these species do inhabit the territory of the reserve. This is especially so for small bivalves of the family Sphaeriidae. The resultant species list can serve as a starting point for further investigations of the reserve's malacofauna.

The effect of four microbial products on production performance and water quality in integrated culture of freshwater pearl mussel and fishes

An 80-day mesocosm experiment was conducted to evaluate the effect of four commercial microbial products on production performance and water quality in integrated culture of freshwater pearl mussel Hyriopsis cumingii, grass carp, gibel carp, silver carp and bighead carp. Five treatments were tested. One treatment with non-supplementation of microbial products served as control (C). In the other four treatments, Novozymes Pond Protect (NO), Bio-Form BZT-Water Reform (WR), Bacillus natto (BN) and Effective Microbes (EM) were added at the intervals of two weeks, respectively. Mussel yield declined in the tanks with supplementation of the microbial products. No significant differences were found in fish yield and chemical water quality among the treatments except total nitrogen (TN) was higher in tanks EM than in tanks C. Biomass of phytoplankton and Cyanophyta was higher in tanks NO, WR, BN and EM than in tanks C. Supplementation of the microbial products resulted in change in bacterial community in which Actinobacteria, Chlorobi, Verrucomicrobia and Proteobacteria dominated. Bacterial community in the tanks was significantly affected by TN, total phosphorus, chemical oxygen demand and Cyanophyta biomass. This study reveals that the function of these microbial products as probiotics is limited in H. cumingii farming.

Structural characteristics of gonadal development and hermaphroditic phenomenon in freshwater pearl mussel, Sinohyriopisis schlegelii (Hyriopisis schlegelii).

The period of gonads development was first studied from one to five years in the freshwater pearl mussel Hyriopisis schlegelii. It lasted for 36 months and was divided into three main stages: initiation of gonad formation, a stable growth phase, and a reproductive cell development phase. Each reproductive cycle consisted of five stages: proliferative stage (from late January to late February), growth stage (from late February to late March), maturation stage, spawning stage (from early April to late October) and recovery stage (from early November to late January). Interestingly, a hermaphroditic phenomenon was observed in this mussel for the first time, which appears during the development stage from 26 to 32 months. Male and female follicular tissues coexisted in hermaphrodite individuals with the male follicular tissue accounting for more than 90% of the whole gonad tissue. No hermaphroditic phenomenon was observed in matured gonad. We thus speculate that self-fertilization does not exist in H. schlegelii.

Encystment of parasitic freshwater pearl mussel (Margaritifera margaritifera) larvae coincides with increased metabolic rate and haematocrit in juvenile brown trout (Salmo trutta)

Gill parasites on fish are likely to negatively influence their host by inhibiting respiration, oxygen transport capacity and overall fitness. The glochidia larvae of the endangered freshwater pearl mussel (FPM, Margaritifera margaritifera (Linnaeus, 1758)) are obligate parasites on the gills of juvenile salmonid fish. We investigated the effects of FPM glochidia encystment on the metabolism and haematology of brown trout (Salmo trutta Linnaeus, 1758). Specifically, we measured whole-animal oxygen uptake rates at rest and following an exhaustive exercise protocol using intermittent flow-through respirometry, as well as haematocrit, in infested and uninfested trout. Glochidia encystment significantly affected whole-animal metabolic rate, as infested trout exhibited higher standard and maximum metabolic rates. Furthermore, glochidia-infested trout also had elevated levels of haematocrit. The combination of an increased metabolism and haematocrit in infested fish indicates that glochidia encystment has a physiological effect on the trout, perhaps as a compensatory response to the potential respiratory stress caused by the glochidia. When relating glochidia load to metabolism and haematocrit, fish with low numbers of encysted glochidia were the ones with particularly elevated metabolism and haematocrit. Standard metabolic rate decreased with substantial glochidia loads towards levels similar to those of uninfested fish. This suggests that initial effects visible at low levels of encystment may be countered by additional physiological effects at high loads, e.g. potential changes in energy utilization, and also that high numbers of glochidia may restrict oxygen uptake by the gills.

Short-term breeding: releasing post-parasitic juvenile Margaritifera into ideal small-scale receptor sites: a new technique for the augmentation of declining populations

The aim of this research was to assess the potential for augmentation of freshwater pearl mussel populations Margaritifera margaritifera through short-term captive breeding to early juvenile stage and thus avoiding the growing-on stage of juveniles in captivity, which can have a high mortality rate. Fish encysted with Margaritifera glochidia were transferred to a tank lined with a thin layer of river gravels. Juvenile mussels were left to drop-down into these gravels and were then transferred, with the gravels, into suitable habitat in their native river. Redox potential measurements and near bed flow velocities were measured in the tops and tails of riffles to determine the best potential areas to transfer the juveniles. Following a year in the wild, juvenile mussels were found to be living in the areas in which they had been placed. This technique is recommended for rivers with populations of Margaritifera that have lost habitat quality, but where preferential flows maintain some areas that are still capable of juvenile survival.

HcToll3 was involved in anti-Vibrio defense in freshwater pearl mussel, Hyriopsis cumingii

Toll-like receptors (TLRs) play an important role in the activation of innate immune response but their functions in bivalves remain largely unknown. In this study, we identified a TLR from the freshwater pearl mussel Hyriopsis cumingii (HcToll3) and investigated its functions in immunity. The full-length cDNA of HcToll3 is 3852 bp and includes an open reading frame (ORF) of 3228 bp that encodes a polypeptide of 1075 amino acids. The predicted HcToll3 protein shares similar structural characteristics with other known Toll family proteins. Quantitative real-time PCR analysis revealed that HcToll3 mRNA is broadly expressed in all of the examined tissues; its transcript level was significantly up-regulated by challenge with gram-negative bacteria Vibrio parahaemolyticus or lipopolysaccharide, but not gram-positive Staphylococcus aureus or peptidoglycan. RNA interference by siRNA results showed that HcToll3 regulated expression of whey acidic protein (HcWAP) and lysozymes (HcLyso1 and HcLyso2) in vivo and knockdown of HcToll3 suppressed the elimination of V. parahaemolyticus. These findings suggest that HcToll3 might be involved in anti-Vibrio defense in H. cumingii.

Three STATs are involved in the regulation of the expression of antimicrobial peptides in the triangle sail mussel, Hyriopsis cumingii

Janus kinase (Jak) and signal transducers and activators of transcription (STAT) signaling pathway is associated in antiviral and antibacterial immune response. Previous studies primarily investigated the function of STATs in mammals. For most invertebrates, only one STAT was found in each species, such as STAT92E was found in Drosophila melanogaster. The studies, which focus on the functional difference between various STATs in the same species of invertebrate, are limited. In the present study, three STATs (HcSTAT1, HcSTAT2 and HcSTAT3) were identified in triangle shell pearl mussel, Hyriopsis cumingii. Phylogenetic analysis showed that HcSTAT1 and HcSTAT3 were clustered with Homo sapiens STAT5, and HcSTAT2 was clustered with Pinctada fucata STAT and Crassostea gigas STAT6. All three STATs could be detected in all tested tissues (hemocytes, hepatopancreas, gill, mantle and foot), and were induced expression when challenged with Staphylococcus aureus or Aeromonas hydrophilia in hemocytes and hepatopancreas. HcSTAT1 regulated the expression of HcDef, HcWAP, HcThe and HcTNF. The expression of HcWAP and HcTNF was down-regulated in HcSTAT2-RNAi mussel. And HcSTAT3 affected the expression of HcTNF. The study is the first report of different functions in antibacterial immune responses between STATs in mollusks.

Interaction between the endangered freshwater pearl mussel Margaritifera margaritifera, the duck mussel Anodonta anatina and the fish host (Salmo): acquired and cross-immunity

The common duck mussel Anodonta anatina can live in sympatry with—and use the same host, brown trout (Salmo trutta)—as the endangered freshwater pearl mussel Margaritifera margaritifera. Since the glochidia release of A. anatina takes place seasonally earlier than that of M. margaritifera, brown trout can be sequentially exposed first to A. anatina and then to M. margaritifera. Cross-immunity, an immune reaction induced in fish host against glochidia after the infection with glochidia of another mussel species, is possible. Thus, it was studied experimentally if brown trout can be cross immunized against M. margaritifera by earlier infection with A. anatina. In addition, the hypothesis that consecutive exposures of same glochidial species in different years in the same host may create acquired immunity was tested in brown trout against M. margaritifera. Furthermore, the dose dependence of acquired immunity against M. margaritifera glochidia in the Atlantic salmon (S. salar) was also studied. Cross-immunity was not found; suggesting that occurrence of A. anatina does not pose a threat to M. margaritifera. Instead, acquired immunity and its dose dependence were evident, emphasizing the significance of availability of 0+ age group immunologically naive Atlantic salmon/brown trout for efficient conservation of M. margaritifera.

Duration of the parasitic phase determines subsequent performance in juvenile freshwater pearl mussels (Margaritifera margaritifera).

Host–parasite systems have been useful in understanding coevolutionary patterns in sympatric species. Based on the exceptional interaction of the long‐lived and highly host‐specific freshwater pearl mussel (FPM; Margaritifera margaritifera) with its much shorter‐lived host fish (Salmo trutta or Salmo salar), we tested the hypotheses that a longer duration of the parasitic phase increases fitness‐related performance of mussels in their subsequent post parasitic phase, and that temperature is the main factor governing the duration of the parasitic phase. We collected juvenile mussels from naturally and artificially infested fish from eight rivers in Norway. Excysted juvenile mussels were maintained separately for each collection day, under similar temperature and food regimes, for up to 56 days. We recorded size at excystment, post excystment growth, and survival as indicators of juvenile fitness in relation to the duration of the parasitic phase. We also recorded the daily average temperatures for the entire excystment period. We observed strong positive relationships between the length of the parasitic phase and the post parasitic growth rate, size at excystment and post parasitic survival. Temperature was identified as an important factor governing excystment, with higher temperatures decreasing the duration of the parasitic phase. Our results indicate that juvenile mussels with the longest parasitic phase have better resources (larger size and better growth rate) to start their benthic developmental phase and therefore to survive their first winter. Consequently, the parasitic phase is crucial in determining subsequent survival. The temperature dependence of this interaction suggests that climate change may affect the sensitive relationship between endangered FPMs and their fish hosts.

Overfishing in the Baltic Sea basin in Russia, its impact on the pearl mussel, and possibilities for the conservation of riverine ecosystems in conditions of high anthropogenic pressure

In recent studies nine populations of the freshwater pearl mussel have been described in the Baltic Sea basin in Russia. They are very scarce, although the condition of their habitats seems to be rather good. Overfishing of the host fish is a limiting factor for them. The number of salmon has decreased by at least 100 times over the past 200 years. Such a scale of decline tends to be hidden over time, and estimation of the normal conditions of the salmon–pearl mussel ecosystem becomes problematic. A significant increase in the number of salmon is necessary to prevent extinction of pearl mussels. Effective protected areas appear to be the only possibility for conservation of the pearl mussels and its host fish species.

The populations of the freshwater pearl mussel, Margaritifera margaritifera (Linnaeus, 1758), and the thick shelled river mussel, Unio crassus Philipsson, 1788, in Latvia

The populations of two endangered species—the freshwater pearl mussel Margaritifera margaritifera and the thick shelled river mussel Unio crassus in Latvia were studied. The specimens were counted, measured, population density and age structure were calculated. The possible host fish presence was found.

Comparative characteristics of the lipid status of gills of juvenile Atlantic salmon infected with glochidia of the freshwater pearl mussel living in rivers of the European North

A comparative study of the lipid status of gills of juvenile Atlantic salmon infested with glochidia of the freshwater pearl mussel inhabiting the Vuokinyoki (White Sea basin) and Syuskyuyanyoki (basin of Lake Ladoga) rivers in the fall is carried out. The most infested gills of juvenile salmon and the lower water temperature (1.3°C) in the Vuokinyoki River reveal higher cholesterol levels and cholesterol/phospholipids ratios, which are due to the influence of infection factors; thereby they slow down biochemical processes. It is noted that variations in the lipid composition in juvenile salmon the gills of which are infected with glochidia reflect the development of adaptive responses that maintain homeostasis of juveniles under invasion.

Variation in the COI gene of the freshwater pearl mussel Margaritifera margaritifera from River Vuokkijoki

The freshwater pearl mussel Margaritifera margaritifera L. is one of the most endangered freshwater mussels in the world. Effective conservation of threatened species requires not only ecological, but also genetic information from the target species and populations. Since low genetic diversity can reduce the ability of a species to adapt to environmental changes, maintaining genetic diversity has been identified as one of the key elements in successful conservation programs. We examined genetic variation of the freshwater pearl mussel from the River Vuokkijoki, Karelia, Russia. We sequenced a fragment of the cytochrome c oxidase subunit I gene (COI) from 22 individuals and compared the data to 32 previously published COI sequences available in GenBank. We identified 10 different COI haplotypes in the sequenced samples, three of which had not been previously reported. Our results show that the River Vuokkijoki has high genetic diversity and suggest that the colonization of this northern freshwater pearl mussel population might have occurred from multiple and even distant refugia. Therefore, the freshwater pearl mussel population of the River Vuokkijoki is valuable for the conservation of the whole species.

The relationship between the freshwater pearl mussel (Margaritifera margaritifera) and its hosts

Endangered freshwater pearl mussels Margaritifera margaritifera reveal a complex life cycle with an obligate host-dependent phase. Only two species, Atlantic salmon Salmo salar and brown trout S. trutta, are important hosts in Europe, indicating a high degree of specialization. Whilst freshwater pearl mussels with their filtering activity can provide important ecosystem services and indirectly improve the habitat quality for their salmonid hosts, their direct effects on physiological stress, reduced swimming performance, and increased mortality at high rates of infestation all support a parasitic character of the mussel during its hostdependent phase. From an evolutionary perspective, both the much greater generation time of the parasite compared to the host, as well as the great distribution ranges of M. margaritifera and its hosts should favour local adaptation patterns. The variable suitability of different salmonid strains and species as hosts for M. margaritifera and the resulting differences in the performance of larvae during the host-dependent phase indicate that host-management strategies should focus on maintaining high quality hosts at a regional scale to avoid selection or genetic drift effects which could erode the genetic and evolutionary potential for adaptation to changing environmental conditions.