Search

The testis-specific double sex and mab-3-related transcription factor 1 (DMRT1) has long been recognized as a crucial player in sex determination across vertebrates, and its essential role in gonadal development and the regulation of spermatogenesis is well established. Here, we report the cloning of the key spermatogenesis-related DMRT1 cDNA, named Tc-DMRT1, from the gonads of Tridacna crocea (T. crocea), with a molecular weight of 41.93 kDa and an isoelectric point of 7.83 (pI). Our hypothesis is that DMRT1 machinery governs spermatogenesis and regulates gonadogenesis. RNAi-mediated Tc-DMRT1 knockdown revealed its critical role in hindering spermatogenesis and reducing expression levels in boring giant clams. A histological analysis showed structural changes, with normal sperm cell counts in the control group (ds-EGFP) but significantly lower concentrations of sperm cells in the experimental group (ds-DMRT1). DMRT1 transcripts during embryogenesis exhibited a significantly high expression pattern (p < 0.05) during the early zygote stage, and whole-embryo in-situ hybridization confirmed its expression pattern throughout embryogenesis. A qRT-PCR analysis of various reproductive stages revealed an abundant expression of Tc-DMRT1 in the gonads during the male reproductive stage. In-situ hybridization showed tissue-specific expression of DMRT1, with a positive signal detected in male-stage gonadal tissues comprising sperm cells, while no signal was detected in other stages. Our study findings provide an initial understanding of the DMRT1 molecular machinery controlling spermatogenesis and its specificity in male-stage gonads of the key bivalve species, Tridacna crocea, and suggest that DMRT1 predominantly functions as a key regulator of spermatogenesis in giant clams. [ABSTRACT FROM AUTHOR]

Sex is an integral part of the evolution of species and often takes the form of intersex and sexual change in mollusks. As a major member of the deep-sea coral reef ecosystem, Tridacna crocea has complex gonadal development mechanisms and reproductive strategies, and the regulation model of spermatogenesis needs to be further explored. In this study, gonadal transcripts of T. crocea in resting, spermatogonia and sperm stage were evaluated. According to the statistics of bioinformatics analysis, a total of 10,260 DEGs were identified, including 3021 genes shared by three developmental stages. Through dynamic expression analysis, we identified the expression patterns and functional classification of genes that regulate spermatogenesis. These candidate genes were involved in regulatory mechanisms such as nucleotide binding, cell cycle, sexual reproduction and DNA replication. Subsequently, based on functional screening, PPI analysis was performed on the target gene cluster to learn the network relationships among candidate genes and other genes closely related to candidate genes. The results showed that in addition to candidate genes (zmcm6, gins2, mcm9, blm, mcm4, psmc3ip, haus, ccnb3, spice, rad1, rad17, ncapg, bub1b, lin9, tipin, and cks), several genes that have been shown to be associated with gonadal development were present in the network (such as fanci, aurka, kif20a and nek2), suggesting the reliability of the results. Finally, the expression levels of candidate genes between individuals at different developmental stages were compared with transcriptional expression profiles, which further proved that these candidate genes were related to T. crocea gonad development traits. These results make up for the lack of research on T. crocea gametogenesis, spermatogenesis and gonad development, and provide a reference for the research on Tridacna reproductive mechanism.

P-element-induced wimpy testis (Piwi) is a key gene involved in germ cell development in a diverse range of organisms. However, in giant clams, the function of Piwi remains unclear. In the present study, we isolated the full-length cDNA of Piwi ortholog (Tc-Piwi1) and analyzed its expression patterns in the gonads of adult and juvenile Tridacna crocea. The results of qPCR showed that the transcript of Tc-Piwi1 was mainly expressed in gonad tissue. In addition, the relative expression level of Tc-Piwi1 increased with the proliferation of male and female germ cells during the adult gonad development stage, suggesting that Tc-Piwi1 might be involved in gametogenesis. In situ hybridization revealed that Tc-Piwi1 RNA was located in female and male germ cells and strongly expressed in male germ cells in the early stage. Furthermore, immunohistochemical experiments further confirmed that Tc-Piwi1 was mainly located in primordial germ cells (PGCs), germ stem cells (GSCs), and female and male germ cells of early development, so it could be used as a marker gene of T. crocea germ cells. Whole-mount in situ hybridization suggested that Tc-Piwi1 was of maternal origin and located in two clusters of cells in the trochophore-larvae stage, implying that these cells might be putative PGCs during the embryo development. Finally, Tc-Piwi1 was used as a molecular marker to elucidate the gonadal formation, sex differentiation, and gonadal maturation process of juvenile T. crocea for the first time in the Tridacna family. Collectively, all these results revealed that Tc-Piwi1 was involved in germline formation and sex differentiation in T. crocea.

Giant clams are marine bivalves that inhabit Indo-Pacific coral reefs. The boring giant clam, Tridacna crocea, exhibits bright and conspicuous mantle coloration based on the specialized cells (iridocytes) that generate structural colors. In order to illustrate the coloring mechanism of individual iridocytes, the reflection spectra curve of iridocytes was obtained by a micro-hyperspectral imager. TEM images were obtained to show the inner nanostructures of iridocytes. FDTD simulation was conducted to analyze the relationship between the color of iridocytes and the unique lamellar structure. We found that the laminae in the regular arrangement within cells govern the coloration of individual iridocytes. With the gradual increase of lamellar thickness and spacing, the color of the structure varies from bright violet to orange-red, forming a full visible spectrum. This study provides a new understanding of the various colors produced by individual iridocytes.

Serum amyloid protein (SAA) is known as an acute reactive protein of innate immunity in mammals. However, in invertebrates, the role of SAA in innate immunity is still unclear. In this study, a full-length cDNA of the SAA gene (named TcSAA) was cloned from Tridacna crocea , mollusca. The gene includes a 193 bp 5′ untranslated region (UTR) and a 129 bp 3' UTR sequence, and the open reading frame (ORF) with 393 bp nucleotides encodes a polypeptide of 130 amino acids. Tc SAA contains a typical signal peptide and an SAA functional domain. The mRNA expression of TcSAA was detected in all 12 selected tissues and 7 different developmental stages. Furthermore, the expression of TcSAA was increased quickly in hemocytes after challenge with V. coralliilyticus or LPS. Furthermore, rTc SAA could bind V. coralliilyticus and V. alginolyticus , and the protein could reduce the lethality rate of the clams from 80% to 55% which caused by V. coralliilyticus about 48 h after injection. In summary, these results indicate that Tc SAA may act as a marker for monitoring health and protecting T. crocea. • A full-length cDNA of serum amyloid gene (named as TcSAA) in T. crocea was cloned. • TcSAA is expressed increased radically after V. coralliilyticus and LPS chanllege. • rTc SAA was expressed and found to bind to V. coralliilyticus and V. alginolyticus. • rTc SAA could reduce the lethality rate of the T. crocea which caused by V. coralliilyticus. [ABSTRACT FROM AUTHOR]

The boring giant clam (Tridacna crocea) is the smallest species of the Tridacna genus, but they strongly express mantle color polymorphism. These color morphs represent potential genetic resources for aquaculture to supply the aquarium market, and the evaluation of their genetic and production characteristics is important. Wild yellow-green clams were consistently larger than blue clams in both the Huangyan Island and Zhaoshu Island populations in the Southern China Sea. To determine the relationship between mantle color and phenotypic traits, blue, yellow-green and control strains (various colors) were constructed and cultured for 1.5 years at an experimental station in Sanya, Hainan Island Province. All three strains had a high fertilization rate, but the survival to D larvae of the blue strain was significantly lower than the other two strains. Larval survival rate was over 85 % for all three strains, but the metamorphosis rate of the control strain was significantly lower than that of the other two strains. During the grow-out stage, the survival rate of the control strain from day 120 to day 480 was highest and that of the blue strain was lowest of the three groups. After metamorphosis, growth divergence of the three strains varied until day 120. Consistent growth trends appeared from day 120 to day 180, with the yellow-green strain largest, followed by the control strain. The process of mantle coloration was divided into three stages: the non-coloring period (day 0–60), the coloring period (day 60–180), and the stable expression period (day 180 onward). The yellow-green and blue mantle colors are genetically stable. This study provides a base for analysis of mantle color heredity and the genetic breeding of giant clams, and has great potential applications in the aquaculture industry, aquarium market and the coral reef tourism.

According to the RGB law display, the polymorphism of the giant clam mantle color pattern is through four iridocytes. The boring giant clam (Tridacna crocea) exhibits diverse mantle colors, including blue, green, purple, gold, and orange. In order to evaluate the genetic laws driving these mantle color patterns, a complete diallel cross between two color strains [blue strain (only blue iridocyte) and the yellow-green strain (yellow and green iridocytes)] was performed. Using a single-to-single mating system, two intra-strain crosses (BB and YY) and two reciprocal inter-strain crosses (BY and YB) were produced in triplicates. Higher fertilization rate and hatching rate were observed in all experimental groups, suggesting that there was no sperm–egg recognition barrier between the two strains. In the grow-out stage, the size of the reciprocal hybrids was larger than that of the two pure strains with a degree of heterosis. In addition, compared with the two pure strains, the hybrids have higher larval metamorphosis rate and higher survival rate. At 1 year of age, the mantle color pattern of pure strains showed 100% stable inheritance, while the reciprocal hybrids exhibited colorful patterns (a combination of blue, yellow, and green), suggesting that there was a genetic recombination of the mantle colors during the stable expression period. These results provide a theoretical basis for the formation of the mantle color of giant clam and its genetic segregation law, as well as provide guidance for genetic breeding of giant clams.

Molluscan shells are among the most fascinating research objects because of their diverse morphologies and textures. The formation of these delicate biomineralized structures is a matrix-mediated process. A question that arises is what are the essential components required to build these exoskeletons. In order to understand the molecular mechanisms of molluscan shell formation, it is crucial to identify organic macromolecules in different shells from diverse taxa. In the case of bivalves, however, taxon sampling in previous shell proteomics studies are focused predominantly on representatives of the class Pteriomorphia such as pearl oysters, edible oysters and mussels. In this study, we have characterized the shell organic matrix from the crocus clam, Tridacna crocea, (Heterodonta) using various biochemical techniques, including SDS-PAGE, FT-IR, monosaccharide analysis, and enzyme-linked lectin assay (ELLA). Furthermore, we have identified a number of shell matrix proteins (SMPs) using a comprehensive proteomics approach combined to RNA-seq. The biochemical studies confirmed the presence of proteins, polysaccharides, and sulfates in the T. crocea shell organic matrix. Proteomics analysis revealed that the majority of the T. crocea SMPs are novel and dissimilar to known SMPs identified from the other bivalve species. Meanwhile, the SMP repertoire of the crocus clam also includes proteins with conserved functional domains such as chitin-binding domain, VWA domain, and protease inhibitor domain. We also identified BMSP (Blue Mussel Shell Protein, originally reported from Mytilus), which is widely distributed among molluscan shell matrix proteins. Tridacna SMPs also include low-complexity regions (LCRs) that are absent in the other molluscan genomes, indicating that these genes may have evolved in specific lineage. These results highlight the diversity of the organic molecules – in particular proteins – that are essential for molluscan shell formation.

Molluscan shells are among the most fascinating research objects because of their diverse morphologies and textures. The formation of these delicate biomineralized structures is a matrix-mediated process. A question that arises is what are the essential components required to build these exoskeletons. In order to understand the molecular mechanisms of molluscan shell formation, it is crucial to identify organic macromolecules in different shells from diverse taxa. In the case of bivalves, however, taxon sampling in previous shell proteomics studies are focused predominantly on representatives of the class Pteriomorphia such as pearl oysters, edible oysters and mussels. In this study, we have characterized the shell organic matrix from the crocus clam, Tridacna crocea , (Heterodonta) using various biochemical techniques, including SDS-PAGE, FT-IR, monosaccharide analysis, and enzyme-linked lectin assay (ELLA). Furthermore, we have identified a number of shell matrix proteins (SMPs) using a comprehensive proteomics approach combined to RNA-seq. The biochemical studies confirmed the presence of proteins, polysaccharides, and sulfates in the T. crocea shell organic matrix. Proteomics analysis revealed that the majority of the T. crocea SMPs are novel and dissimilar to known SMPs identified from the other bivalve species. Meanwhile, the SMP repertoire of the crocus clam also includes proteins with conserved functional domains such as chitin-binding domain, VWA domain, and protease inhibitor domain. We also identified BMSP (Blue Mussel Shell Protein, originally reported from Mytilus), which is widely distributed among molluscan shell matrix proteins. Tridacna SMPs also include low-complexity regions (LCRs) that are absent in the other molluscan genomes, indicating that these genes may have evolved in specific lineage. These results highlight the diversity of the organic molecules – in particular proteins – that are essential for molluscan shell formation. [ABSTRACT FROM AUTHOR]

We report the development and characterization of nine microsatellite markers from the Tridacna crocea genome. In a wild population of 20 individuals from Ulugan Bay in the Republic of the Philippines, we observed an average of 9 alleles per locus and expected heterozygosity ranging from 0.41 to 0.92. Cross-species priming was tested in three additional Tridacna species with moderate success. These markers will be used in studies of fine-scale population genetic structure and gene flow in T. crocea. The markers may also be useful in a forensic context by identifying source populations of giant clams collected illegally for the live aquarium trade, while protecting legal exports from aquaculture facilities.

In this study, we first determined and described the mitochondrial genome of Tridacna crocea, a member of the subfamily Tridacninae. The existing mitogenome of T. crocea is 19,157 bp in length, containing 13 protein-coding genes, 2 rRNA genes, 24 tRNA genes, and 1 non-coding control region. Phylogenetic analysis revealed that T. crocea was closely related to T. squamosa.

Survival and growth of post-metamorphic Tridacna crocea juveniles were improved by inducing symbiosis with fresh or cloned zooxanthellae (Tg10) derived from T. gigas. Although clam growth was best with Tc4, survival was also poorest. Symbiosis with specific zooxanthellae was established at the pediveliger stage, with reinfection a few days after. It is suggested that while survival and growth may be easily monitored and may be used as indicators of good performance of a functional holobiont, other phenotypic traits such as resistance to disease, bleaching, etc. may also be considered in evaluating the effectivity of the selected zooxanthellae.

In May 2010, an expedition on the research vessel Akademik Oparin explored the populations of the clam Tridacna crocea on reefs of the Bay Canh and Cau islands of the Cön Dao Archipelago in the South China Sea. A retrospective approach using phototechniques revealed a great similarity of the vertical distribution, growth, size, and age composition of the tridacna at these islands. The maximum density of tridacna (25 ind/m) was characteristic of a depth of about 3 m in the area of reef flat transition to reef slope. The growth of clams can be approximated by a Bertalanffy-type equation: L = 133.8[1 − e]. The lack of discreteness in the size and age compositions of populations of T. crocea indicated that population recruitment was quite stable over the years. This tends to maintain the population in a relatively stable condition, which enabled us to consider it as a possible reserve for the artificial restoration of populations of the species in impoverished coastal areas of Vietnam. [ABSTRACT FROM AUTHOR]

The article presents a case study of an adult, hermaphroditic Tridacna crocea ornamental clam imported from Vietnam, which is found to be moribund in a home aquarium. The clam was subjected to necropsy and was found to exhibit visceral mass edema, emaciation, and a rare multifocal off white to light tan-colored gill nodules. The clam was diagnosed to have infected with the Perkinsus sp. pathogen.

The Coral Triangle is the global center of marine biodiversity; however, its coral reefs are critically threatened. Because of the bipartite life history of many marine species with sedentary adults and dispersive pelagic larvae, designing effective marine protected areas requires an understanding of patterns of larval dispersal and connectivity among geographically discrete populations. We used mtDNA sequence data to examine patterns of genetic connectivity in the boring giant clam (Tridacna crocea ) in an effort to guide conservation efforts within the Coral Triangle. We collected an approximately 485 base pair fragment of mtDNA cytochrome c oxidase 1 (CO1) from 414 individuals at 26 sites across Indonesia. Genetic structure was strong between regions (φST= 0.549, p < 0.00001) with 3 strongly supported clades: one restricted to western Sumatra, another distributed across central Indonesia, and a third limited to eastern Indonesia and Papua. Even within the single largest clade, small but significant genetic structure was documented (φST= 0.069, p < 0.00001), which indicates limited gene flow within and among phylogeographic regions. Significant patterns of isolation by distance indicated an average dispersal distance of only 25–50 km, which is far below dispersal predictions of 406–708 km derived from estimates of passive dispersal over 10 days via surface currents. The strong regional genetic structure we found indicates potent limits to genetic and demographic connectivity for this species throughout the Coral Triangle and provides a regional context for conservation planning. The recovery of 3 distinct evolutionarily significant units within a well-studied taxonomic group suggests that biodiversity in this region may be significantly underestimated and that Tridacna taxa may be more endangered than currently recognized. [ABSTRACT FROM AUTHOR]

Fifteen polymorphic microsatellite markers were developed for Tridacna maxima in order to assess self-recruitment and larval dispersal within and among MPAs in New Caledonia and provide baseline data for conservation management of this species. Number of alleles varied from 5 to 25 per locus, observed and expected heterozygosities ranged from 0.171 to 0.860 and 0.591 to 0.935 respectively. Significant deviations from HWE were detected in eight loci. Cross-amplifications were tested in five other species of Tridacnidae ( T. crocea, T. squamosa, T. derasa, T. noae and Hippopus hippopus). [ABSTRACT FROM AUTHOR]

In this study, we first determined and described the mitochondrial genome of Tridacna crocea, a member of the subfamily Tridacninae. The existing mitogenome of T. crocea is 19,157 bp in length, containing 13 protein-coding genes, 2 rRNA genes, 24 tRNA genes, and 1 non-coding control region. Phylogenetic analysis revealed that T. crocea was closely related to T. squamosa. [ABSTRACT FROM AUTHOR]

Giant clams are able to grow in oligotrophic tropical marine environments with the aid of photosynthates from its symbiotic zooxanthellae. Therefore, light intensity affects the survival and growth of giant clams, and is one of the most important environmental factors in giant clam cultures. In this study, two size classes of Tridacna crocea (small: ~50 mm shell length, large: ~85 mm shell length) were cultured at three light intensities (5000, 10,000, and 15,000 lx) for 16 weeks, to evaluate the effects of light intensity on the survival, growth performance and biochemical composition of different size clams. T. crocea under 15,000 lx light intensity showed a significantly higher survival rate and relative growth rate (RGR) (P <.0001). The RGR in the small group was significantly higher than the large group under 10,000 and 15,000 lx light intensity (P <.0001), while higher RGR was observed in the large group under 5000 lx light intensity (P =.0009). The content of eight amino acids in the mantle was significantly affected by light intensity (P <.05), while only one amino acid (glycine) was found in the adductor muscle (P =.0005). Similarly, there were significant differences in ten mantle amino acids between small and large size groups (P <.05), while three amino acids were found in the adductor muscle (P <.05). C22:6n3 (DHA) was the major polyunsaturated fatty acid (PUFA) in T. crocea. In the adductor muscle, the C22:6n3 profile significantly increased with increasing light intensity and clam size (P <.0001), while no significant differences of C22:6n3 profiles were found among different light intensities in the mantle (P =.3345). Our results demonstrate that a light intensity of 15,000 lx is suitable for T. crocea (45–90 mm in shell length) rearing, and that large size T. crocea has higher tolerance to weak light intensity than small individuals. • The optimal light intensity of T. crocea (shell length 45–90 mm) was 15,000 lx. • The survival and growth of T. crocea were affected by both light intensity and clam size. • The amino acid contents in the mantle were affected by both light intensity and clam size. • The fatty acid profiles of T. crocea were strongly affected by both light intensity and clam size. [ABSTRACT FROM AUTHOR]

Inbreeding depression is common, especially in hermaphroditic animals, and causes a decline in biological fitness. To evaluate the effects of inbreeding on giant clams, two by two factorial cross experiments between individuals were conducted on three species, Tridacna deras a, T. squamosa , T. crocea. Thus, self-fertilized progeny (f = 0.5) and cross-fertilized progeny (f = 0) were produced for each species. The effects of inbreeding depression on fertilization rate, hatching rate, progeny growth and survival were slight for all three species. However, heavy inbreeding depression was observed on the larval metamorphosis rates of T. squamosa and T. crocea , and very few self-fertilized juveniles from these two species were successfully obtained. Surprisingly, inbreeding depression of performance traits was not found in T. derasa. Our results show that medium (T. squamosa) and small type (T. crocea) giant clam are prone to serious inbreeding depression (δ>0.99) at the larval metamorphic stage, while little inbreeding depression occurred in the large type giant clam (T. derasa). Two-factor ANOVA of survival rates for the whole experimental period showed that progeny survival of medium and small type giant clams was always affected by the mating strategy. Almost no inbreeding depression on growth traits was found these species, which may be due to the nutritional strategy of these autotrophic species. Our study proposes a theory regarding the magnitude of inbreeding depression in different giant clam species based on the evolution of the wild populations and provides a practical account of a mating strategy suitable for seed production for giant clam aquaculture. • Inbreeding depression of phenotypic traits for three giant clams were firstly evaluated. • Heavy inbreeding depression occurred metamorphosis stage for the medium (T. squamosa) and small type (T. crocea) giant clams. • Hardly no inbreeding depression was found on the larger type giant clam (T. derasa). • These results provides a practical account of a mating strategy suitable for seed production for aquaculture. [ABSTRACT FROM AUTHOR]

The boring giant clam, Tridacna crocea , is distributed along coasts in the South China Sea, and has formed several geographic populations. These populations represent potentially different genetic resources for aquaculture, and the evaluation of their genetic and production characteristics is important. To determine these performance characteristics, complete diallel crosses between two geographical populations with about 600 km distance, Zhaoshu Island and Huangyan Island, were conducted using pooled gametes at an experimental station in Sanya, Hainan Island Province. Two intra-population crosses (ZZ and HH) and two reciprocal inter-population crosses (ZH and HZ) were carried out successfully three times. High fertilization rates and D larvae rate were observed in all the experimental groups, suggesting that there was no sperm-egg recognition barrier between the two geographic populations. Reciprocal hybrids had a higher survival rate than the pure populations, but with the exception of metamorphosis rate there were no significant differences between the inter- and intra-population crosses. Growth heterosis was evident in both the spat and adult stages, and was mainly influenced by egg origin and mating strategy in the adult stage. In view of the growth differences between the two pure populations' progeny, the Huangyan Island population's progeny was more suitable for giant clam aquaculture than the Zhaoshu Island population's progeny. Our results demonstrate that crossbreeding between the Huangyan Island and Zhaoshu Island populations of boring giant clams can produce considerable heterosis, which has promising prospects for application in the giant clam aquaculture industry in the South China Sea. • Two population's progeny exhibited obvious difference in phenotypic traits as well as their parents. • Crossbreeding between two populations produced positive heterosis without any fertilization barrier. • Performance traits of reciprocal hybrids is asymmetric. • HH and HZ progeny would be excellent lines as choices for the giant clam aquarium market and aquaculture industry. [ABSTRACT FROM AUTHOR]

Interspecific hybridization has been used as an important tool for genetic manipulation in mollusks, with the aim to produce a new species with desirable traits from both the parental species. A two by two factorial cross between sympatric giant clam species, the fluted giant clam, Tridacna squamosa (S) and the boring clam, Tridacan crocea (C), yielded hybrids that were fast growing (characteristic of the S) and had brightly colored mantles (characteristic of C). Reciprocal hybrids (SC and CS) were obtained by the acceptable level of fertilization and hatching rate compared to these of two parental species, and the survival rate of the hybrids was significantly higher than that of the parental species. Yearling hybrids exhibited fully developed gonads and normal gametogenesis when sexually mature. Relative to CC, CS and SC also showed significant growth advantages in shell length and wet weight. The shell morphology of the hybrids was mainly dictated by maternal effect, while mantle coloration was dictated by paternal effect for SC progeny and parental effect for CS progeny. Our results revealed that reciprocal hybrids exhibited great growth heterosis and survival advantage (compared to C), as well as better mantle colors. Thus, these hybrids have great potential for application in the aquarium market and aquaculture industries across the globe. • A two by two factorial cross between two giant clams, Tridacna squamosa and T. crocea was successfully conducted. • Reciprocal hybrids exhibited larger growth heterosis and survival advantage, as well as a beautiful mantle coloration. • These hybrids showed great potential for application in the aquarium market and giant clam aquaculture industry. [ABSTRACT FROM AUTHOR]