Chih-Ching Chung

Identification of a High-Affinity Phosphate Transporter Gene in a Prasinophyte Alga, Tetraselmis chui, and Its Expression under Nutrient Limitation

ABSTRACT A high-affinity phosphate transporter gene, TcPHO, was isolated from a growth-dependent subtracted cDNA library of the marine unicellular alga Tetraselmis chui. The full-length cDNA of TcPHO obtained by 5′ and 3′ rapid amplification of cDNA ends was 1,993 bp long and encoded an open reading frame consisting of 610 amino acids. The deduced amino acid sequence of TcPHO exhibited 51.6 and 49.8% similarity to the amino acid sequences of PHO89 from Saccharomyces cerevisiae and PHO4 from Neurospora crassa, respectively. In addition, hydrophobicity and secondary structure analyses revealed 12 conserved transmembrane domains that were the same as those found in PHO89 and PHO4. The expression of TcPHO mRNA was dependent on phosphate availability. With a low-phosphate treatment, the TcPHO mRNA concentration increased sharply to 2.72 fmol μg of total RNA−1 from day 1 to day 2 and remained at this high level from days 2 to 4. Furthermore, rescue treatment with either phosphate or p-nitrophenyl phosphate effectively inhibited TcPHO mRNA expression. In contrast, TcPHO mRNA expression stayed at a low level (range, 0.25 to 0.28 fmol μg of total RNA−1) under low-nitrate conditions. The expression pattern suggests that TcPHO can be used as a molecular probe for monitoring phosphorus stress in T. chui.

Distribution patterns and phylogeny of marine stramenopiles in the north pacific ocean.

ABSTRACT Marine stramenopiles (MASTs) are a diverse suite of eukaryotic microbes found in marine environments. Several MAST lineages are thought to contain heterotrophic nanoflagellates. However, MASTs remain uncultured and data on distributions and trophic modes are limited. We investigated MASTs in provinces on the west and east sides of the North Pacific Subtropical Gyre, specifically the East China Sea (ECS) and the California Current system (CALC). For each province, DNA was sampled from three zones: coastal, mesotrophic transitional, and more oligotrophic euphotic waters. Along with diatoms, chrysophytes, and other stramenopiles, sequences were recovered from nine MAST lineages in the six ECS and four CALC 18S rRNA gene clone libraries. All but one of these libraries were from surface samples. MAST clusters 1, 3, 7, 8, and 11 were identified in both provinces, with MAST cluster 3 (MAST-3) being found the most frequently. Additionally, MAST-2 was detected in the ECS and MAST-4, -9, and -12 were detected in the CALC. Phylogenetic analysis indicated that some subclades within these lineages differ along latitudinal gradients. MAST-1A, -1B, and -1C and MAST-4 size and abundance estimates obtained using fluorescence in situ hybridization on 79 spring and summer ECS samples showed a negative correlation between size of MAST-1B and MAST-4 cells and temperature. MAST-1A was rarely detected, but MAST-1B and -1C and MAST-4 were abundant in summer and MAST-1C and MAST-4 were more so at the coast, with maximum abundances of 543 and 1,896 cells ml−1, respectively. MAST-4 and Synechococcus abundances were correlated, and experimental work showed that MAST-4 ingests Synechococcus. Together with previous studies, this study helps refine hypotheses on distribution and trophic modes of MAST lineages.

Cooccurrence of ScDSP Gene Expression, Cell Death, and DNA Fragmentation in a Marine Diatom, Skeletonema costatum

ABSTRACT A novel death-specific gene, ScDSP, was obtained from a death stage subtraction cDNA library of the diatom Skeletonema costatum. The full length of ScDSP cDNA was 921 bp in length, containing a 699-bp open reading frame encoding 232 amino acids and two stretches of 66 and 156 bp in the 5′ and 3′ untranslated regions, respectively. Analysis of the peptide structure revealed that ScDSP contained a signal peptide domain, a transmembrane domain, and a pair of EF-hand motifs. When S. costatum grew exponentially at a rate of 1.3 day−1, the ScDSP mRNA level was at 2 μmol · mole 18S rRNA−1. In contrast, when the culture entered the death phase with a growth rate decreasing to 0.5 day−1, ScDSP mRNA increased dramatically to 668 μmol · mole 18S rRNA−1, and a high degree of DNA fragmentation was simultaneously observed. Under the influence of a light-dark cycle, ScDSP expression in both exponential and stationary phases clearly showed a diel rhythm, but the daily mean mRNA level was significantly higher in the stationary phase. Our results suggest that ScDSP may play a role in the molecular mechanism of self-destructive autolysis in phytoplankton under stress.

Nitric oxide as a signaling factor to upregulate the death-specific protein in a marine diatom, Skeletonema costatum, during blockage of electron flow in photosynthesis.

ABSTRACT To determine the physiological functions of a novel death-specific protein gene, Skeletonema costatum DSP-1 (ScDSP-1) in a marine diatom, Skeletonema costatum, the mRNA abundance of ScDSP-1 was measured in cultures subjected to light manipulation and treatments with various chemicals. When cells were transferred to a dim light intensity of 15 μmol m−2 s−1, ScDSP-1 mRNA levels showed a transient increase of 1 to 17.2 μmol (mol 18S rRNA)−1 in 60 h. Furthermore, treatments with the photoinhibitors 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) resulted in high ScDSP-1 mRNA levels, which reached 943 and 72 μmol (mol 18S rRNA)−1, respectively. Treatment with the nitric oxide (NO) donor diethylamine nitric oxide also induced ScDSP-1 expression, and this inducible expression was inhibited by the NO scavenger hemoglobin. Additionally, the expression of ScDSP-1 mRNA elicited by DCMU and DBMIB was efficiently reduced when cultures were pretreated with the cell-penetrating NO scavenger 2-(4-carboxyphenyl)-4,5-dihydro-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide. In contrast, treatment with another photoinhibitor, paraquat, had no effect on ScDSP-1 expression. Our results indicated that NO is the crucial secondary messenger which signals the expression of ScDSP-1 when electron flow between photosystem II and photosystem I is blocked in S. costatum cells. In addition, the discovery of a similar gene, ScDSP-2, is briefly described.

Effects of Asian Dust Storms on Synechococcus Populations in the Subtropical Kuroshio Current

Asian dust storms (ADSs) are the major source of dust deposition in the Northwest Pacific Ocean. To gain a better understanding on how ADSs affect the ecology of picophytoplankton in this oligotrophic region, five oceanographic cruises were conducted between March 15 and April 15, 2006 on a segment of the Kuroshio Current near the shelf break of the East China Sea (25.05° N, 123.15° E). During the study period, three ADS events were recorded and increases in nutrient concentrations as well as mixing depths were observed. Most of the ADS events stimulated the growth of Synechococcus, but the abundance of Prochlorococcus either remained unaffected or showed mild declines. A more detailed study was conducted during the ADS event between March 16 and 19. Phylogenetic analysis of 16S ribosomal RNA nucleotide sequences revealed that most of the newly appeared Synechococcus belonged to the clade II lineage. Furthermore, messenger RNA (mRNA) levels of three nutrient deficiency indicators, including idiA (an iron deficiency indicator), ntcA (a nitrogen deficiency indicator), and pstS (a phosphorus deficiency indicator), were analyzed by real-time quantitative reverse-transcription polymerase chain reaction. As this ADS event proceeded, mRNA levels of all these indicators decreased from relatively high to non-detectable values. These results suggest that the contributions of iron, nitrogen, and phosphate by the dust deposition from ADSs promote the growth of Synechococcus in the Kuroshio Current.

Strains of the Morphospecies Ploeotia costata (Euglenozoa) Isolated from the Western North Pacific (Taiwan) Reveal Substantial Genetic Differences

Two phagotrophic euglenid strains (Strains Pac and Tam) were isolated from coastal locations in Taiwan. Ultrastructural characteristics of the strains included five pellicle strips joined at the posterior end. The strips were formed by major grooves with bifurcated edges. At the cell anterior, the feeding structure formed a lip. Underneath the lip was a comb composed of layers of microtubules. Farther back, two supporting rods tapered toward the posterior end, and a number of vanes with attached microtubules were present between the rods. The morphological characteristics agree with Ploeotia costata Strain CCAP 1265/1. However, the 18S rDNA sequences of Strains Pac/Tam lacked a group I intron and possessed three extra insertions of 116, 67, and 53 bp. Phylogenetic analysis indicated low sequence similarity between Strains Pac/Tam and CCAP 1265/1 (92%). The morphospecies P. costata apparently includes a substantial level of DNA sequence divergence, and likely represents multiple molecular species units.

Community Composition of Photosynthetic Picoeukaryotes in a Subtropical Coastal Ecosystem, with Particular Emphasis on Micromonas

Photosynthetic picoeukaryotes (PPEs) are important constituents in picoplankton communities in many marine ecosystems. However, little is known about their community composition in the subtropical coastal waters of the Northwestern Pacific Ocean. In order to study their taxonomic composition, this study constructed 18S rRNA gene libraries using flow cytometric sorting during the warm season. The results show that, after diatoms, prasinophyte clones are numerically dominant. Within prasinophytes, Micromonas produced the most common sequences, and included clades II, III, IV, and VI. We are establishing the new Micromonas clade VI based on our phylogenetic analysis. Sequences of this clade have previously been retrieved from the South China Sea and Red Sea, indicating a worldwide distribution, but this is the first study to detect clade VI in the coastal waters of Taiwan. The TSA‐FISH results indicated that Micromonas clade VI peaked in the summer (~4 × 102 cells/ml), accounting for one‐fifth of Micromonas abundance on average. Overall, Micromonas contributed half of Mamiellophyceae abundance, while Mamiellophyceae contributed 40% of PPE abundance. This study demonstrates the importance of Micromonas within the Mamiellophyceae in a subtropical coastal ecosystem.

Predator and prey biodiversity relationship and its consequences on marine ecosystem functioning—interplay between nanoflagellates and bacterioplankton

The importance of biodiversity effects on ecosystem functioning across trophic levels, especially via predatory–prey interactions, is receiving increased recognition. However, this topic has rarely been explored for marine microbes, even though microbial biodiversity contributes significantly to marine ecosystem function and energy flows. Here we examined diversity and biomass of bacteria (prey) and nanoflagellates (predators), as well as their effects on trophic transfer efficiency in the East China Sea. Specifically, we investigated: (i) predator diversity effects on prey biomass and trophic transfer efficiency (using the biomass ratio of predator/prey as a proxy), (ii) prey diversity effects on predator biomass and trophic transfer efficiency, and (iii) the relationship between predator and prey diversity. We found higher prey diversity enhanced both diversity and biomass of predators, as well as trophic transfer efficiency, which may arise from more balanced diet and/or enhanced niche complementarity owing to higher prey diversity. By contrast, no clear effect was detected for predator diversity on prey biomass and transfer efficiency. Notably, we found prey diversity effects on predator–prey interactions; whereas, we found no significant diversity effect on biomass within the same trophic level. Our findings highlight the importance of considering multi-trophic biodiversity effects on ecosystem functioning in natural ecosystems.