In the Asia-Pacific region, the COI DNA test revealed the divergence of the bivalve mollusc Mactra chinensis into three species; can these species be distinguished using shell coloration and sperm structure?

According to COI DNA barcoding testing, the marine bivalve mollusc Mactra chinensis, which is native to the Asia-Pacific region, diverged into three species. These species were preliminary characterized as M. chinensis COI clade I, M. chinensis COI clade II and M. chinensis COI clade III. To find out whether it is possible to morphologically distinguish samples representing genetic clades, we examined the color of the shells and the structure of the spermatozoa. It was found that the number of detected coloration types exceeds the number of detected species. In addition, it was shown that individuals belonging to the same genetic clade can have shells of different colors. Consequently, it is impossible to choose one type of shell coloration as a species-specific trait. For sperm, the sperm morphological patterns found in each of the three species are consistent with the M. chinensis sperm model described in previous reports. However, the single sperm variant is also not applicable to discriminate between species derived from M. chinensis, since heterogeneous variants of spermatozoa differing in the length of the acrosomal rod were found. We hypothesized that genetic divergence of species could cause a shift towards predominance of one of the sperm variants, and that species-specific sperm morphs could be quantitatively dominant in molluscs belonging to different clades. However, the dominant sperm morphs were the same in COI clade I and COI clade III. Thus, dominant sperm morphs are useless as species-specific traits. However, shell color and sperm parameters are specific to different geographic regions, and it seems that unique environmental factors can determine shell color and sperm morphology. As a result, both shells and spermatozoa can be used to distinguish the geographical forms of M. chinensis, regardless of the belonging of the forms to a particular genetic clade. Here we propose the introduction of geographic identifiers, in which the shell color and parameters of sperm sets are used as morphological criteria to determine the geographical origin of mollusc specimens belonging to the M. chinensis species complex.

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 < 0.05). The association analysis identified 20 SNPs in the exons of Mm-BCDO2, among which three loci (i.e., c.984A > C, c.1148C > T, and c.1187A > T) were remarkably related (P < 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 < 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.

A Genome-Wide Association Study Identifies Candidate Genes Associated With Shell Color in Bay Scallop Argopecten irradians irradians

Molluscan shell color has consistently drawn attention for its abundant diversity and commercial use in shellfish breeding projects. Recently, two new strains of bay scallop (Argopecten irradians irradians) with different shell colors as marked phenotypic traits have been artificially bred to improve their economic values; however, the inheritance mechanism of their shell pigmentation is still unclear. In this study, a genome-wide association study (GWAS) was conducted to determine the genetic basis of shell color in bay scallops utilizing 29,036 high-quality single-nucleotide polymorphisms (SNPs) derived from 80 purple-red (PP) and 80 black-brown (BP) shell color individuals. The result of the GWAS showed that 469 SNPs (p <1.72E−6) significantly associated with shell color were mainly distributed in chromosome 7. The top three SNPs (i.e., chr7-12764003, chr7-13213864, and chr7-11899306) are located in the genic region of G-protein-coupled receptor-like 101 (GRL101), polyketide synthase 1 (PKS1), and phosphoinositide phospholipase C (PLC1), which have been widely reported to be involved in pigmentation. Successfully, the top three SNPs were verified in another non-breeding bay scallop population. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses obtained 38 GO terms covering 297 genes and aggregating pathways involving 252 annotated genes. Specifically, the expression profiles of the top three identified candidate genes were detected in mantles of PP and BP individuals by real-time quantitative reverse transcription PCR. The significantly higher expression levels of GRL101 (6.43-fold) and PLC1 (6.48-fold) in PP, and PKS1 (12.02-fold) in BP implied that GRL101 and PLC1 potentially functioned in PP shell coloration, and black pigmentation in BP might be principally regulated by PKS1. Our data provide valuable information for deciphering the phenotype differences of shell color in the bay scallop.

Genotypic and Dietary Effects on Egg Quality of Local Chicken Breeds and Their Crosses Fed with Faba Beans

The quality of chicken eggs is an important criterion for food safety and the consumers’ choice at the point of sale. Several studies have shown that egg quality can be influenced by the chickens’ genotype and by the composition of the diet. The present study aimed to evaluate the effect of faba beans as a substitute for soybeans in the diet of chickens originating from traditional low-performance breeds in comparison with high-performing laying type hens and their crosses on egg quality parameters. Chickens of six different genotypes were fed either with a feed mix containing 20% faba beans with high or low vicin contents or, as a control, a feed mix containing soybeans. The genotypes studied were the local breeds Vorwerkhuhn and Bresse Gauloise, as well as commercial White Rock parent hens and their crosses. Yolk weight, Haugh units, yolk and shell color, the frequency of blood and meat spots and the composition of the eggs were significantly influenced by the genotype. The feeding of faba beans had an effect on yolk and shell color, Haugh units and shell portion, while there was no significant influence on the frequency of blood and meat spots.

Identification of shell-color-related microRNAs in the Manila clam Ruditapes philippinarum using high-throughput sequencing of small RNA transcriptomes

Shell-color polymorphism is a common phenomenon in several mollusk species and has been associated with thermal capacity, developmental stability, shell strength, and immunity. Shell-color polymorphism has been related to the differential expression of genes in several signal transduction pathways; however, the functions of micro-RNAs (miRNAs) in shell-color formation remain unclear. In the present study, we compared high-quality, small-RNA transcriptomes in three strains of the Manila clam Ruditapes philippinarum with specific shell-color patterns, artificially selected for six generations. Totals of 114 known and 208 novel miRNAs were identified by high-throughput sequencing, of which nine known and one novel miRNA were verified by stem-loop quantitative real time-polymerase chain reaction. Predicted miRNA targets were subjected to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. miR-137 and miR-216b and the Hedgehog signaling pathway and Wnt signaling pathway were identified as being potentially involved in pigment formation and regulation in R. philippinarum. These results may help to clarify the role of miRNAs in shell coloration and shed light on the mechanisms regulating color formation in bivalve shells.