Isolation and Characterization of Polysaccharides from the Ascidian Styela clava

Styela clava is an edible sea squirt farmed in Korea that has gradually invaded other seas, negatively impacting the ecology and economy of coastal areas. Extracts from S. clava have shown wide bioactivities, and ascidians have the unique capability among animals of biosynthesizing cellulose. Thus, S. clava is a relevant candidate for valorization. Herein, we aimed at surveying and characterizing polysaccharides in both tunic and flesh of this ascidian. To this end, we enzymatically hydrolyzed both tissues, recovering crystalline cellulose from the tunic with high aspect ratios, based on results from microscopy, X-ray diffraction, and infrared spectroscopy analyses. Alkaline hydroalcoholic precipitation was applied to isolate the polysaccharide fraction that was characterized by gel permeation chromatography (with light scattering detection) and NMR. These techniques allowed the identification of glycogen in the flesh with an estimated Mw of 7 MDa. Tunic polysaccharides consisted of two fractions of different Mw. Application of Diffusion-Ordered NMR allowed spectroscopically separating the low-molecular-weight fraction to analyze the major component of an estimated Mw of 40–66 kDa. We identified six different sugar residues, although its complexity prevented the determination of the complete structure and connectivities of the residues. The two more abundant residues were N-acetylated and possibly components of the glycosaminoglycan-like (GAG-like) family, showing the remaining similarities to sulfated galactans. Therefore, Styela clava appears as a source of nanocrystalline cellulose and GAG-like polysaccharides.

A new range-extending record of the invasive sea squirt S. clava in the north of Scotland

A new record of an individual of the invasive sea squirt, Styela clava, is reported from Scapa Flow in Orkney, Scotland. This represents a 1.54° latitudinal extension (168 km) from the previous northern-most record in UK waters. Diver surveys of hard substrates in the immediate area, part of local biosecurity protocols, did not find any additional individuals. Possible transport vectors, likelihood of an established reproductively active population, and implications to local biodiversity are discussed.

Analysis of the Microbial Community Structure in Coastal Sediment of an Ascidian Farm in South Korea through 16S rRNA Gene Amplicon Sequencing

Aquaculture places contamination pressure on the coastal environment. We investigated the microbial community structure changes in sediment in an ascidian Styela clava farm. Data profiling of the 16S rRNA gene amplicon sequence shows that the microbial diversity of sediment in the Styela clava farm is dominated by Proteobacteria phyla (relative abundance, 95.34 to 97.85%).

Genome-Wide Identification, Comparison, and Expression Analysis of Transcription Factors in Ascidian Styela clava

Tunicates include diverse species, as they are model animals for evolutionary developmental biology study. The embryonic development of tunicates is known to be extensively regulated by transcription factors (TFs). Styela clava, the globally distributed invasive tunicate, exhibits a strong capacity for environmental adaptation. However, the TFs were not systematically identified and analyzed. In this study, we reported 553 TFs categorized into 60 families from S. clava, based on the whole genome data. Comparison of TFs analysis among the tunicate species revealed that the gene number in the zinc finger superfamily displayed the most significant discrepancy, indicating this family was under the highly evolutionary selection and might be related to species differentiation and environmental adaptation. The greatest number of TFs was discovered in the Cys2His2-type zinc finger protein (zf-C2H2) family in S. clava. From the point of temporal view, more than half the TFs were expressed at the early embryonic stage. The expression correlation analysis revealed the existence of a transition for TFs expression from early embryogenesis to the later larval development in S. clava. Eight Hox genes were identified to be located on one chromosome, exhibiting different arrangement and expression patterns, compared to Ciona robusta (C. intestinalis type A). In addition, a total of 23 forkhead box (fox) genes were identified in S. clava, and their expression profiles referred to their potential roles in neurodevelopment and sensory organ development. Our data, thus, provides crucial clues to the potential functions of TFs in development and environmental adaptation in the leathery sea squirt.

Phylogenetic Analysis and Screening of Antimicrobial and Antiproliferative Activities of Culturable Bacteria Associated with the Ascidian Styela clava from the Yellow Sea, China

Over 1,000 compounds, including ecteinascidin-743 and didemnin B, have been isolated from ascidians, with most having bioactive properties such as antimicrobial, antitumor, and enzyme-inhibiting activities. In recent years, direct and indirect evidence has shown that some bioactive compounds isolated from ascidians are not produced by ascidians themselves but by their symbiotic microorganisms. Isolated culturable bacteria associated with ascidians and investigating their potential bioactivity are an important approach for discovering novel compounds. In this study, a total of 269 bacteria were isolated from the ascidian Styela clava collected from the coast of Weihai in the north of the Yellow Sea, China. Phylogenetic relationships among 183 isolates were determined using their 16S rRNA gene sequences. Isolates were tested for antimicrobial activity against seven indicator strains, and an antiproliferative activity assay was performed to test for inhibition of human hepatocellular carcinoma Bel 7402 and human cervical carcinoma HeLa cell proliferation. Our results showed that the isolates belonged to 26 genera from 18 families in four phyla (Firmicutes, Actinobacteria, Proteobacteria, and Bacteroidetes). Bacillus and Streptomyces were the most dominant genera; 146 strains had potent antimicrobial activities and inhibited at least one of the indicator strains. Crude extracts from 29 strains showed antiproliferative activity against Bel 7402 cells with IC50 values below 500 μg·mL−1, and 53 strains showed antiproliferative activity against HeLa cells, with IC50 values less than 500 μg·mL−1. Our results suggest that culturable bacteria associated with the ascidian Styela clava may be a promising source of novel bioactive compounds.