Tiezhu Mi

Diversity, Abundance, and Spatial Distribution of Ammonia-Oxidizing β-Proteobacteria in Sediments from Changjiang Estuary and Its Adjacent Area in East China Sea

Changjiang Estuary, the largest estuary in China, encompasses a wide range of nutrient loading and trophic levels from the rivers to the sea, providing an ideal natural environment to explore relationships between functional diversity, physical/chemical complexity, and ecosystem function. In this study, molecular biological techniques were used to analyze the community structure and diversity of ammonia-oxidizing bacteria (AOB) in the sediments of Changjiang Estuary and its adjacent waters in East China Sea. The amoA gene (encoding ammonia monooxygenase subunit A) libraries analysis revealed extensive diversity within the β-Proteobacteria group of AOB, which were grouped into Nitrosospira-like and Nitrosomonas-like lineages. The majority of amoA gene sequences fell within Nitrosospira-like clade, and only a few sequences were clustered with the Nitrosomonas-like clade, indicating that Nitrosospira-like lineage may be more adaptable than Nitrosomonas-like lineage in this area. Multivariate statistical analysis indicated that the spatial distribution of the sedimentary β-Proteobacterial amoA genotype assemblages correlated significantly with nitrate, nitrite, and salinity. The vertical profile of amoA gene copies in gravity cores showed that intense sediment resuspension led to a deeper mixing layer. The horizontal distribution pattern of amoA gene copies was nearly correlated with the clayey mud belt in Changjiang Estuary and its adjacent area in East China Sea, where higher β-Proteobacteria phylogenetic diversity was observed. Meanwhile, those areas with high amoA copies in the surface sediments nearly matched those with low concentrations of dissolved oxygen and ammonium in the bottom water.

Seasonal Variation and Sources of Dissolved Nutrients in the Yellow River, China

The rapid growth of the economy in China has caused dramatic growth in the industrial and agricultural development in the Yellow River (YR) watershed. The hydrology of the YR has changed dramatically due to the climate changes and water management practices, which have resulted in a great variation in the fluxes of riverine nutrients carried by the YR. To study these changes dissolved nutrients in the YR were measured monthly at Lijin station in the downstream region of the YR from 2002 to 2004. This study provides detailed information on the nutrient status for the relevant studies in the lower YR and the Bohai Sea. The YR was enriched in nitrate (average 314 μmol·L−1) with a lower concentration of dissolved silicate (average 131 μmol·L−1) and relatively low dissolved phosphate (average 0.35 μmol·L−1). Nutrient concentrations exhibited substantial seasonal and yearly variations. The annual fluxes of dissolved inorganic nitrogen, phosphate, and silicate in 2004 were 5.3, 2.5, and 4.2 times those in 2002, respectively, primarily due to the increase in river discharge. The relative contributions of nutrient inputs to nitrogen in the YR were: wastewater > fertilizer > atmospheric deposition > soil; while to phosphorus were: wastewater > fertilizer > soil > atmospheric deposition. The ratios of N, P and Si suggest that the YR at Lijin is strongly P-limited with respect to potential phytoplankton growth.

Molecular characterisation, evolution and expression of hypoxia-inducible factor in Aurelia sp.1.

The maintenance of physiological oxygen homeostasis is mediated by hypoxia-inducible factor (HIF), a key transcriptional factor of the PHD-HIF system in all metazoans. However, the molecular evolutionary origin of this central physiological regulatory system is not well characterized. As the earliest eumetazoans, Cnidarians can be served as an interesting model for exploring the HIF system from an evolutionary perspective. We identified the complete cDNA sequence of HIF-1α (ASHIF) from the Aurelia sp.1, and the predicted HIF-1α protein (pASHIF) was comprised of 674 amino acids originating from 2,025 bp nucleotides. A Pairwise comparison revealed that pASHIF not only possessed conserved basic helix-loop-helix (bHLH) and Per-Arnt-Sim (PAS) domains but also contained the oxygen dependent degradation (ODD) and the C-terminal transactivation domains (C-TAD), the key domains for hypoxia regulation. As indicated by sequence analysis, the ASHIF gene contains 8 exons interrupted by 7 introns. Western blot analysis indicated that pASHIF that existed in the polyps and medusa of Aurelia. sp.1 was more stable for a hypoxic response than normoxia.

Characteristics of eukaryotic microalgal community and its abiotic influencing factors during brown tide blooms near Qinhuangdao, China

The brown tides caused by the picoplanktonic pelagophyte Aureococcus anophagefferens in the coastal waters of Qinhuangdao, China have occurred periodically since 2009 and exerted deleterious effects on scallop cultivation in the area. In this study, clone libraries were constructed to determine the characteristics of the local eukaryotic microalgae community, and a real-time fluorescent quantitative PCR assay was performed to analyze the temporal and spatial variations in the 18S rDNA copies of A. anophagefferens samples collected in 2012. The results showed that A. anophagefferens was the dominant species in the local eukaryotic microalgae community during the brown tide in June 2012 and accounted for a large fraction of the community. A redundancy analysis (RDA) showed that the decreasing concentration of dissolved inorganic nitrogen (DIN), increasing amount of human aquaculture activities and suitability of spring/summer temperatures for the growth of A. anophagefferens may be the primary causes of the brown tide outbreaks in the Qinghuangdao scallop culture area.

Phosphorus speciation, transformation and retention in the Three Gorges Reservoir, China

Damming of river systems allowing the transformation of former rivers into artificial lakes will increase the transformation and retention of dissolved and sediment-associated phosphorus (P). The reservoir is therefore a ‘filter’ or ‘converter’, reducing and delaying the transport of nutrients to marine systems. Our study of the Three Gorges Reservoir (TGR) found that no stratification of phosphorus occurred, and the high particulate phosphorus (PP) concentrations upstream decreased gradually in the reservoir. Detrital P was found in greater concentrations in the surface sediment, accounting for 39% of PP; exchangeable P was rare and contributed very little to the total P budget. P forms and their concentrations in the suspended particulate matter varied throughout the TGR, with a significant increase of bioavailable P in the <8-μm particle fraction from 27% of PP in Fuling to 60% in Yichang, and decreasing detrital P and authigenic P in each grain size class. The TGR acted as a ‘converter’ for the dissolved reactive phosphorus, and it therefore plays a minor role in trapping incoming total dissolved phosphorus; whereas the TGR behaved as a ‘filter’ for the PP, especially for the coarse fraction, which resulted in the retention of 70% of the non-bioavailable PP. The controlling mechanism of P species and retention in the reservoir is particulate settling and its associated effects.

Quantitative PCR analysis of the spatiotemporal dynamics of Aureococcus anophagefferens and Minutocellus polymorphus and the relationship between brown tides and nutrients in the coastal waters of Qinhuangdao, China

Brown tides caused by the excessive growth of the pelagophyte Aureococcus anophagefferens have occurred frequently in Qinhuangdao, China, since 2009. In this study, the temporal and spatial variations of A. anophagefferens and Minutocellus polymorphus were analyzed using real-time fluorescent quantitative PCR. The relationship between nutrient contents and brown tide events as well as the impacts of the brown tides on the composition of phytoplankton is discussed. The results showed that A. anophagefferens reached peak densities in July. Brown tides occurred in the coastal waters of Funing and Changli, and the main affected areas shifted from south to north throughout the experiment. The A. anophagefferens brown tide event was not monospecific, as M. polymorphus and Prorocentrum minimum were also observed. A redundancy analysis (RDA) indicated that decreasing concentrations of dissolved inorganic nitrogen (DIN) and increasing concentrations of the available organic nitrogen compounds (urea, dissolved free amino acids (DFAA), and dissolved combined amino acids (DCAA)) from March to June may have resulted in the outbreak of the brown bloom of A. anophagefferens, coinciding with a reduction in the community diversity of phytoplankton in the coastal waters of Qinhuangdao.

Metacaspases and programmed cell death in Skeletonema marinoi in response to silicate limitation

Diatoms comprise up to 40% of annual productivity in the modern ocean, and some stress factors may alter their physiology and survival. Here, we describe the morphological, biochemical and molecular responses that occur during metacaspase-mediated programmed cell death (PCD) in response to silicate limitation in the ubiquitous diatom Skeletonema marinoi. Transmission electron microscopy revealed that vacuolization occurred while the plasma membranes remained intact. Caspase 3-like activity, antioxidant gene expression, phosphatidylserine externalization and activated caspase-like expression were measured using in vivo staining. The results show that each was significantly increased under silicate limitation conditions. Using quantitative reverse transcription-PCR, we identified six distinct putative metacaspase proteins in S. marinoi, and we show that each contains a conserved caspase-like domain and is differentially expressed and that some of their expression profiles were correlated with PCD. Silicate limitation could trigger PCD in S. marinoi as well as an increase of antioxidant genes. PCD could be employed as a strategy to decrease silicate demand of the whole population and probably regulate the fate of S. marinoi.

Molecular characterization of sulfate-reducing bacteria community in surface sediments from the adjacent area of Changjiang Estuary

Sulfate-reducing bacteria (SRB), which obtain energy from dissimilatory sulfate reduction, play a vital role in the carbon and sulfur cycles. The dissimilatory sulfite reductase (Dsr), catalyzing the last step in the sulfate reduction pathway, has been found in all known SRB that have been tested so far. In this study, the diversity of SRB was investigated in the surface sediments from the adjacent area of Changjiang Estuary by PCR amplification, cloning and sequencing of the dissimilatory sulfite reductase beta subunit gene (dsrB). Based on dsrB clone libraries constructed in this study, diversified SRB were found, represented by 173 unique OTUs. Certain cloned sequences were associated with Desulfobacteraceae, Desulfobulbaceae, and a large fraction (60%) of novel sequences that have deeply branched groups in the dsrB tree, indicating that novel SRB inhabit the surface sediments. In addition, correlations of the SRB assemblages with environmental factors were analyzed by the linear model-based redundancy analysis (RDA). The result revealed that temperature, salinity and the content of TOC were most closely correlated with the SRB communities. More information on SRB community was obtained by applying the utility of UniFrac to published dsrB gene sequences from this study and other 9 different kinds of marine environments. The results demonstrated that there were highly similar SRB genotypes in the marine and estuarine sediments, and that geographic positions and environmental factors influenced the SRB community distribution.

Seasonal and spatial distribution of ammonia-oxidizing microorganism communities in surface sediments from the East China Sea

Ammonia oxidation plays a significant role in the nitrogen cycle in marine sediments. Seasonal and spatial distribution of ammonia-oxidizing archaea (AOA) and betaproteobacteria (β-AOB) in surface sediments from the East China Sea (ECS) were investigated using ammonia monooxygenase α subunit (amoA) gene. In order to characterize the community of AOA and β-AOB, real-time quantitative polymerase chain reaction (qPCR) was carried out in this study, along with environmental parameters. The abundance of β-AOB amoA gene (2.17×106–4.54×107 copy numbers per gram wet weight sediment) was always greater than that of AOA amoA gene (2.18×105–9.89×106 copy numbers per gram wet weight sediment) in all sampling stations. The qPCR results were correlated with environmental parameters. AOA amoA gene copy numbers in April were positively related to temperature and nitrite concentration (ρ<0.05). β-AOB amoA gene copy numbers in August correlated negatively with salinity (ρ<0.01), and correlated positively with ammonium concentration (ρ<0.05). With the increase of salinity, the amoA gene copy ratio of AOB to AOA had a tendency to decrease, which suggested β-AOB dominated in the area of high level ammonium and AOA preferred high salinity area.

Transformation and source of nutrients in the Changjiang Estuary

The concentrations of phosphate (PO43−), ammonium, nitrite, nitrate, silicate, dissolved organic nitrogen (DON), dissolved organic phosphorus (DOP), particulate phosphorus (PP) and particulate nitrogen (PN) along the salinity gradient were measured in the Changjiang Estuary in April 2007. The behavior of nutrient species along the continuum from the freshwater to the coastal zone is discussed. In the mixing zone between the riverine and marine waters, nitrate and phosphate behave non-conservatively, while silicate behaves conservatively. Nutrient import was quantified from the river load. Nutrient export to the sea was quantified from river discharge and from the salinity-nutrient gradient in the outer estuary. Using these data, a nitrogen and phosphorus budget was made. The internal estuarine fluxes played an important role in the nutrient estuarine fluxes, which accounted for approximately 41% of the nitrogen flux and 45% of the phosphorus flux. The mixing experiments in the laboratory generally reproduced well the inorganic process affecting nutrient dynamics in the Changjiang Estuary, indicating that the primary P and N transformation processes were phosphate and nitrate desorption along the salinity gradient.