So Yun Park

Biochemical Characterization of Soluble Acid and Alkaline Invertases from Shoots of Etiolated Pea Seedlings

Soluble invertase was purified from pea (Pisum sativum L.) by sequential procedures entailing ammonium sulfate precipitation, DEAE-Sepharose column, Con-A- and Green 19-Sepharose affinity columns, hydroxyapatite column, ultra-filtration, and Sephacryl 300 gel filtration. The purified soluble acid (SAC) and alkaline (SALK) invertases had a pH optimum of 5.3 and 7.3, respectively. The temperature optimum of two invertases was 37 degrees C. The effects of various concentrations of Tris-HCl, HgCl(2), and CuSO(4) on the activities of the two purified enzymes were examined. Tris-HCl and HgCl(2) did not affect SAC activity, whereas 10 mM Tris-HCl and 0.05 mM HgCl(2) inhibited SALK activity by about 50%. SAC and SALK were inhibited by 4.8 mM and 0.6 mM CuSO(4) by 50%, respectively. The enzymes display typical hyperbolic saturation kinetics for sucrose hydrolysis. The Kms of SAC and SALK were determined to be 1.8 and 38.6 mM, respectively. The molecular masses of SAC shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting were 22 kDa and 45 kDa. The molecular mass of SALK was 30 kDa. Iso-electric points of the SAC and SALK were estimated to be about pH 7.0 and pH 5.7, respectively.

Seasonal Dynamics and Metagenomic Characterization of Marine Viruses in Goseong Bay, Korea

Viruses are the most abundant biological entities in the oceans, and account for a significant amount of the genetic diversity of marine ecosystems. However, there is little detailed information about the biodiversity of viruses in marine environments. Rapid advances in metagenomics have enabled the identification of previously unknown marine viruses. We performed metagenomic profiling of seawater samples collected at 6 sites in Goseong Bay (South Sea, Korea) during the spring, summer, autumn, and winter of 2014. The results indicated the presence of highly diverse virus communities. The DNA libraries from samples collected during four seasons were sequenced using Illumina HiSeq 2000. The number of viral reads was 136,850 during March, 70,651 during June, 66,165 during September, and 111,778 during December. Species identification indicated that Pelagibacter phage HTVC010P, Ostreococcus lucimarinus OIV5 and OIV1, and Roseobacter phage SIO1 were the most common species in all samples. For viruses with at least 10 reads, there were 204 species during March, 189 during June, 170 during September, and 173 during December. Analysis of virus families indicated that the Myoviridae was the most common during all four seasons, and viruses in the Polyomaviridae were only present during March. Viruses in the Iridoviridae were only present during three seasons. Additionally, viruses in the Iridoviridae, Herpesviridae, and Poxviridae, which may affect fish and marine animals, appeared during different seasons. These results suggest that seasonal changes in temperature contribute to the dynamic structure of the viral community in the study area. The information presented here will be useful for comparative analyses with other marine viral communities.

Differential gene expression patterns during embryonic development of sea urchin exposed to triclosan

Triclosan (TCS; 2,4,4′‐trichloro‐2′‐hydroxydiphenyl ether) is a broad‐spectrum antibacterial agent used in common industrial, personal care and household products which are eventually rinsed down the drain and discharged with wastewater effluent. It is therefore commonly found in the aquatic environment, leading to the continual exposure of aquatic organisms to TCS and the accumulation of the antimicrobial and its harmful degradation products in their bodies. Toxic effects of TCS on reproductive and developmental progression of some aquatic organisms have been suggested but the underlying molecular mechanisms have not been defined. We investigated the expression patterns of genes involved in the early development of TCS‐treated sea urchin Strongylocentrotus nudus using cDNA microarrays. We observed that the predominant consequence of TCS treatment in this model system was the widespread repression of TCS‐modulated genes. In particular, empty spiracles homeobox 1 (EMX‐1), bone morphogenic protein, and chromosomal binding protein genes showed a significant decrease in expression in response to TCS. These results suggest that TCS can induce abnormal development of sea urchin embryos through the concomitant suppression of a number of genes that are necessary for embryonic differentiation in the blastula stage. Our data provide new insight into the crucial role of genes associated with embryonic development in response to TCS.

Metagenomic characterization of viral communities in Goseong Bay, Korea

In this study, seawater samples were collected from Goseong Bay, Korea in March 2014 and viral populations were examined by metagenomics assembly. Enrichment of marine viral particles using FeCl3 followed by next-generation sequencing produced numerous sequences. De novo assembly and BLAST search showed that most of the obtained contigs were unknown sequences and only 0.74% of sequences were associated with known viruses. As a result, 138 viruses, including bacteriophages (87%), viruses infecting algae and others (13%) were identified. The identified 138 viruses were divided into 11 orders, 14 families, 34 genera, and 133 species. The dominant viruses were Pelagibacter phage HTVC010P and Roseobacter phage SIO1. The viruses infecting algae, including the Ostreococcus species, accounted for 9.4% of total identified viruses. In addition, we identified pathogenic herpes viruses infecting fishes and giant viruses infecting parasitic acanthamoeba species. This is a comprehensive study to reveal the viral populations in the Goseong Bay using metagenomics. The information associated with the marine viral community in Goseong Bay, Korea will be useful for comparative analysis in other marine viral communities.

Hypoxia-modulated gene expression profiling in sea urchin (Strongylocentrotus nudus) immune cells

Hypoxia is an issue that affects ocean coastal waters worldwide. It has severe consequences for marine organisms, including death and rapid adaptive changes in metabolic organization. Although some aquatic animals are routinely exposed and resistant to severe environmental hypoxia, others such as sea urchins (Strongylocentrotus nudus) have a limited capacity to withstand this stress. In this study, hypoxia induced a significant increase in the number of red spherule cells among coelomocytes, which function as immune cells. This suggests that sea urchin immune cells could be used as a biological indicator of hypoxic stress. In the current study, we used cDNA microarrays to investigate the differential expression patterns of hypoxia-regulated genes to better understand the molecular mechanisms underlying the response of immune cells to hypoxia. Surprisingly, the predominant major effect of hypoxia was the widespread suppression of gene expression. In particular, the expression of RNA helicase and GATA-4/5/6 was decreased significantly in response to hypoxia, even in field conditions, suggesting that they could be utilized as sensitive bioindicators of hypoxic stress in the sea urchin.

Development of DNA chip for verification of 25 microalgae collected from southern coastal region in Korea

Countless species occur in the marine microalgal domain. Some are used as health functional foods or medical products but many species are harmful such as those that cause the red tide. Therefore, it is necessary to conduct prompt and accurate identification of microalgal species. As it is quite difficult to accurately distinguish all species in terms of morphology, we performed DNA barcoding analysis using molecular markers for more accurate and rapid screening. DNA barcoding analysis, i.e., DNA chip technology, is a powerful method for studies on microalgal taxonomy and biodiversity. We used the mitochondrial cytochrome c oxidase subunit I (mtCOI) as a barcoding gene to identify microalgal species. In this study, the diversity and phylogenetic differences among different microalgae were analyzed. Additionally, a microalgal species-specific probe was screened by 21–23 bp and the result was printed on silylated slide for use in a robotic microarrayer. As a result, we performed a DNA chip assay for each of 25 microalgal species and determined that the COI barcode gene was suitable as a marker gene, as it could identify various microalgae from the Korean South Sea by species.

Monitoring the seasonal dynamics of microalgae in the South Sea of Korea by use of a cytochrome c oxidase I DNA barcode

Microalgae are important primary producers in the marine environment, and are also important sources of health foods and medical products. Rapidly growing microalgae have potential use for the production of commercial products, but can also cause harmful microalgal blooms in natural ecosystems. There are many available techniques for the identification of microalgae in natural ecosystems. We used the mitochondrial cytochrome c oxidase subunit I gene as DNA barcode to identify 14 species of microalgae from the South Sea of Korea and to discriminate among similar biogeographic subgroups within species. In addition, we designed species-specific primers targeting the cytochrome c oxidase subunit I gene to evaluate monthly changes of microalgae throughout the year. Chaetoceros brevis, Asterionellopsis glacialis and Stephanopyxis turris were present during all seasons, whereas Skeletonema japonicum, Nitzschia improvisa, Ditylum brightwellii and Chaetoceros diadema were only detected during winter and spring. Our results indicate that species-specific polymerase chain reaction of the cytochrome c oxidase subunit I gene can be used to monitor the seasonal dynamics of microalgae in the South Sea of Korea. This polymerase chain reaction detection method successfully identified the 14 most common species of microalgae with the same polymerase chain reaction condition near Tongyeong, in the South Sea of Korea.

Mapping distribution of cysts of recent dinoflagellate and Cochlodinium polykrikoides using next-generation sequencing and morphological approaches in South Sea, Korea

The total dinoflagellate cyst community and the cysts of Cochlodinium polykrikoides in the surface sediments of South Sea (Tongyeong coast), South Korea, were analysed using next-generation sequencing (NGS) and morphological approaches. Dinoflagellate cysts can be highly abundant (111–4,087 cysts g−1 dry weight) and have diverse species composition. A total of 35 taxa of dinoflagellate cysts representing 16 genera, 21 species (including four unconfirmed species), and 14 complex species were identified by NGS analysis. Cysts of Scrippsiella spp (mostly Scrippsiella trochoidea) were the most dominant and Polykrikos schwartzii, Pentapharsodinium dalei, Ensiculifera carinata, and Alexandrium catenella/tamarense were common. Thus, a combination of NGS and morphological analysis is effective for studying the cyst communities present in a given environment. Although C. polykrikoides developed massive blooms during 2013–2014, microscopy revealed low density of their cysts, whereas no cysts were detected by NGS. However, the vegetative C. polykrikoides not appeared during 2015–2017 in spite of the observation of C. polykrikoides cysts. This suggests that the C. polykrikoides blooms were not due to development of their cysts but to other factors such as currents transporting them to a marine environment suitable for their growth.

High diversity and potential translocation of DNA viruses in ballast water

Ballast water is a common vector for the transport of invasive species to new marine and aquatic environments. We used a metagenomics approach to examine the composition and diversity of viral communities in ballast water from ships originating in Mexico, Saudi Arabia, New York, and Panama, and in water from the port of their destination in Busan, Korea. Myoviridae was the most abundant virus family in ballast water, followed Podoviridae and Siphoviridae. We also identified viruses that infect invertebrates, amoebas, and algae in ballast water and in the Busan port water. Interestingly, there were several viruses that infect humans or other animals (Swinepox virus, Raccoonpox virus, Suid herpesvirus, and Human endogenous retrovirus) in the samples from New York and Panama. In addition, there were giant viruses in all the ballast water samples, especially, identified Megavirus chilensis in New York and Panama, and Pandoravirus salinus in Mexico and Saudi Arabia. These results provide detailed descriptions of the characteristics of the viruses present in ballast water, document significant viral diversity, and indicate the potential translocation of viruses via ballast water.

Integration of the nuclease protection assay with sandwich hybridization (NPA-SH) for sensitive detection of Heterocapsa triquetra

Microalgae are photosynthetic microorganisms that function as primary producers in aquatic ecosystems. Some species of microalgae undergo rapid growth and cause harmful blooms in marine ecosystems. Heterocapsa triquetra is one of the most common bloom-forming species in estuarine and coastal waters worldwide. Although this species does not produce toxins, unlike some other Heterocapsa species, the high density of its blooms can cause significant ecological damage. We developed a H. triquetra species-specific nuclease protection assay sandwich hybridization (NPA-SH) probe that targets the large subunit of ribosomal RNA (LSU rRNA). We tested probe specificity and sensitivity with five other dinoflagellates that also cause red tides. Our assay detected H. triquetra at a concentration of 1.5×104 cells/mL, more sensitive than required for a red-tide guidance warning by the Korea Ministry of Oceans and Fisheries in 2015 (3.0×104 cells/mL). We also used the NPA-SH assay to monitor H. triquetra in the Tongyeong region of the southern sea area of Korea during 2014. This method could detect H. triquetra cells within 3 h. Our assay is useful for monitoring H. triquetra under field conditions.