Identification and Expression Characterization of the Smad3 Gene and SNPs Associated with Growth Traits in the Hard Clam (Meretrix meretrix)

It has been demonstrated that the sekelsky mothers against decapentaplegic homolog 3 (Smad3) plays an important role in the growth and development of vertebrates. However, little is known about the association between the Smad3 gene and the growth traits of mollusks. In this study, Smad3 from the hard clam Meretrix meretrix (Mm-Smad3) was cloned, characterized, and screened for growth-related single nucleotide polymorphisms (SNPs) in its exons. The full-length cDNA of Mm-Smad3 was 1938 bp, encoding a protein with 428 amino acid residues. The protein sequence included an MH1 (27–135 aa) and MH2 domain (233–404 aa). Promoter analysis showed that the promoter sequence of Mm-Smad3 was 2548 bp, and the binding sites of Pit-1a, Antp, Hb, and other transcription factors are related to the growth and development of hard clams. The phylogenetic tree was divided into two major clusters, including mollusks and vertebrate. The expression level of Mm-Smad3 was predominantly detected in the mantle and foot, while extremely less expression was observed in the digestive gland. The low expression level of Mm-Smad3 was detected at the stages of unfertilized mature eggs, fertilized eggs, four-cell embryos, blastula, gastrulae, trochophore, and D-shaped larvae, whereas an opposite trend was observed regarding the highest expression at the umbo larvae stage (p < 0.05). In the mantle repair experiment, the time-course expression profiles showed that compared to the expression level at 0 h, Mm-Smad3 significantly decreased at 6 h (p < 0.05) but increased at 12 and 48 h. Further, the association analysis identified 11 SNPs in the exons of Mm-Smad3, of which three loci (c.597 C > T, c.660 C > T, c.792 A > T) were significantly related to the growth traits of clam (p < 0.05). Overall, our findings indicated that Mm-Smad3 is a growth-related gene and the detected SNP sites provide growth-related markers for molecular marker-assisted breeding of this species.

Sex ratio, gonadal and condition indexes of the Asiatic hard clam, Meretrix meretrix in Marudu Bay, Malaysia

Duisan L, Salim G, Ransangan J. 2021. Sex ratio, Gonadal and Condition indexes of the Asiatic Hard Clam, Meretrix meretrix in Marudu Bay, Malaysia. Biodiversitas 22: 4895-4904. Asiatic hard clam, Meretrix meretrix is one of the important shellfishery resources in Marudu Bay, Sabah, Malaysia. It is among the most popular clam species being widely traded in the local wet markets around Sabah, Malaysia. Unfortunately, the shellfishery management for this species has not been well established. In addition to overexploitation, habitat destruction is also one of the significant threats to this species due to the extensive land use of the coastal areas in Sabah. Hence, conservation and breeding efforts for this species are greatly required. Therefore, the current study was conducted to examine the sexual maturity of the clam with respect to shell length classes for artificial seed production purposes. For this study, a total of 86 clam specimens were randomly collected from mudflats in Marudu Bay. The specimens were utilized for gonad histological and condition analyses. The clams were grouped into three shell length classes; (3.00-4.99) cm, (5.00-6.99) cm, and (7.00-8.99) cm prior to the analyses. Results showed the natural stock of the Asiatic hard clams in Marudu Bay was dominated by females (1.39:1) over males with no hermaphroditism observed. The gonadal index was recorded higher among clams with shell lengths between 5.00 and 6.99 cm. The condition index analysis also recorded high (>4.0) for clams in all the shell length classes. The findings of this study suggest that the clams with shell lengths between 5.00 cm and 7.00 cm are already fully matured and can be utilized as a broodstock candidate for an artificial breeding program in the hatchery.

Stock Assessment of Hatchery-Released Clam Meretrix meretrix in an Estuary of China From the Perspectives of Population Ecology and Genetic Diversity

In this study, the release effects of the buried clam Meretrix meretrix in a northern estuary of China (Shuangtaizi Estuary) was assessed in terms of population ecology and genetic variability by using longer-term monitoring data from 2011 (used as the baseline data) to 2018. A total of 2.4 × 108 hatchery juveniles were released from 2012 to 2014, and follow-up monitoring survey of the growth, distribution, density, and biomass of M. meretrix population was carried out. Results showed a temporary spike in the population of M. meretrix, and the population underwent considerable year-to-year fluctuations in total abundance and biomass. Meanwhile, the age structure of the populations from 2011 to 2018 is still an unstable expanding type. Genetic analysis based on 10 polymorphic microsatellite markers revealed no significant genetic differentiation and weak genetic structures between years but higher genetic diversities in 2014–2018 than in the initial period (2013) of stock restoration. Our study suggests that effective policies for management and conservation to restore M. meretrix population by local governments are still needed in the long run.

Characterization and Function Analysis of β, β-carotene-9′, 10′-oxygenase 2 (BCDO2) Gene in Carotenoid Metabolism of the Red Shell Hard Clam (Meretrix meretrix)

The relationship between carotenoid and shellfish shell color has gained increasing attention. β, β-carotene-9′,10′-oxygenase 2 (BCDO2) is a key enzyme in animal carotenoid metabolism, and its accumulation affects the change in body color, as demonstrated in mammals, birds, and fish. However, it is unclear whether BCDO2 is involved in the formation of the red shell color of clam. To explore the molecular structure and biological function of BCDO2 gene in the process of carotenoids accumulation, in this study, the BCDO2 from hard clam Meretrix meretrix (designated as Mm-BCDO2) was cloned and characterized, and the single-nucleotide polymorphisms (SNPs) associated with shell color were detected. The results of qRT-PCR indicated that Mm-BCDO2 gene was expressed in all six tested tissues, and the expression of mantle was significantly higher than other tissues (P &lt; 0.05). The association analysis identified 20 SNPs in the exons of Mm-BCDO2, among which three loci (i.e., c.984A &gt; C, c.1148C &gt; T, and c.1187A &gt; T) were remarkably related (P &lt; 0.05) to the shell color of clam. The western blot analysis revealed that the expression level of Mm-BCDO2 in the mantle of red shell clams was stronger than that of white shell clams (P &lt; 0.05). Further, the immunofluorescence analysis indicated that the single-layer columnar cells at the edge of the mantle were the major sites for the Mm-BCDO2 secretion. This study explored the potential impacts of BCDO2 gene on the shell color of M. meretrix, which provided a theoretical basis for a better understanding of the important role of BCDO2 in carotenoid metabolism.