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The moon jellyfish Aurelia coerulea is a nuisance species around the world, and is considered highly tolerant of a wide range of environmental conditions. The recruitment success during the early life stages of scyphomedusae can have a major effect on the abundance of the adult medusa population and can contribute to jellyfish blooms. The environmental stress factors of elevated copper concentrations and reduced salinity often act simultaneously during the summer time. In this study, we investigated the combined effects of three concentrations of copper (0, 10 and 25 μg L−1) and reduced salinity (from ambient 31 psu to 22 psu) on the early life stages of A. coerulea. We found that the swimming speed of the planula larvae of A. coerulea was significantly affected by the copper concentration. Planula larvae of A. coerulea from the highest copper concentration (25 μg L−1) moved slower at than at lower copper concentrations. The results showed significant interactive effects between copper concentrations and salinity on the settlement of A. coerulea planulae. High copper concentration (25 μg L−1) and reduced salinity (22 psu) significantly increased the settlement of A. coerulea planulae. Additionally, copper concentration had a significant effect on the asexual reproduction rate of A. coerulea polyps, which was significantly higher at the moderate copper concentration (10 μg L−1). These results suggest that the current concentration of copper was not a stress factor for the early life stages of A. coerulea and may potentially stimulate population increases of A. coerulea in Chinese coastal waters.

The dinoflagellate Karlodinium veneficum that is usually present at relatively low cell abundances is a globally-distributed harmful algal bloom-forming species, which negatively affects marine ecosystems, fisheries, and human health. Hence, an efficient detection platform for the rapid and sensitive identification of K. veneficum is highly demanded. In this study, a method referred to as recombinase polymerase amplification coupled with lateral flow dipstick (RPA-LFD) was developed for the rapid detection of K. veneficum. The primers for RPA and the detection probe for LFD were designed to specially target the internal transcribed spacer of K. veneficum by molecular cloning and multiple alignments of the related sequences. The developed RPA can gain an approximately 300 bp specific band from K. veneficum. Successful amplification for RPA could be achieved at a temperature range of 35 °C–45 °C. RPA for 30 min could produce enough products that could generate clearly visible electrophoresis bands and were adequate for subsequent LFD analysis. The RPA products can be visually detected by the naked eyes through an LFD after an automatic chromatography for 5 min at room temperature. The developed RPA-LFD was exclusively specific for K. veneficum and displayed no cross-reactivity with other algal species that are commonly distributed along the Chinese coast. In addition, the lowest detection limit of RPA-LFD was 10 ng μL−1 of genomic DNA and 0.1 cell mL−1, which was 100-fold sensitive than conventional PCR. In conclusion, the developed RPA-LFD assay in this study can be used as a rapid and sensitive method to monitor K. veneficum in the future.

Harmful algal blooms (HABs) have adverse effects on the marine ecological environment, public health, and marine economy. Thus, methods for the accurate and rapid identification of harmful algal species are urgently needed for the effective monitoring of the occurrence of HABS. A method for the parallel detection of harmful algal species must be established because various HAB-forming algal species coexist in the marine environment. This work developed a multiplex PCR (mPCR) method that can simultaneously detect six common HAB-forming microalgal species distributed along the coast of China: Karlodinium veneficum (Kv), Chattonella marina (Cm), Skeletonema spp., Scrippsiella trochoidea (St), Karenia mikimotoi (Km), and Prorocentrum donghaiense (Pd). Specific mPCR primers were designed from the internal transcribed spacer rDNA or large subunit rDNA gene of the target algal species. The mPCR conditions were optimized. Each mPCR primer was subjected to a cross-reactivity test with other microalgae to confirm the specificity of the developed mPCR system. The results of the system stability test indicated that the background concentration of DNA tested did not affect the performance of the established mPCR system. The results of the sensitivity test showed that the detection limit of the proposed mPCR system for Kv, Cm, Km, and Pd was 0.6 ng μL-1 and that for Skeletonema spp. and St was 0.06 ng μL-1. Additional mPCR analysis with spiked field samples revealed that the detection limit of the mPCR system for Km, Pd, and Kv was 60 cells, whereas that for Cm, Skeletonema spp., and St was 6 cells. The convenience and accuracy of the established mPCR assay were further validated through tests with field samples. The proposed mPCR assay is characterized by parallel analysis, strong specificity, and stability and can be used to supplement morphology-based detection methods for algal species.

Hydrogenated diamond-like carbon (DLC) film was prepared on the stainless steel by plasma enhanced chemical vapor deposition technique and its tribocorrosion behaviors under different lubricating conditions were investigated. The results show that the hydrogenated DLC film exhibits superior tribocorrosion resistance than the steel substrate in water. The addition of graphene oxide (GO) in water can further improve the tribocorrosion resistance of the DLC film. The excellent tribocorrosion resistance of the DLC film lubricated with the GO dispersed in water is attributed to the high hardness and chemical inertness of the DLC film and the deposited film of GO nanosheets on the wear surface that has lubrication function for reducing wear and acts as barrier layer against corrosion during rubbing process.

Harmful algal blooms (HAB) involving toxic microalgae have posed a serious threat to the marine industry and environment in the past several decades. Efficient techniques are required to monitor the marine environment to provide an effective warning of imminent HAB. Sequenced the partial large subunit rDNA (D1-D2) sequences of eight toxic harmful algae that are commonly distributed along the Chinese coast were cloned. Specific padlock probes (PLP) that contain linker regions composed of universal primer binding sites and Zip sequences were designed from the obtained target DNA. Taxonomic probes complementary to the Zip sequences were tailed and spotted onto a nylon membrane to prepare a DNA array. An optimized multiplex hyperbranched rolling circle amplification (MHRCA) was used to produce biotin-labeled amplified products. Heat-denatured MHRCA products were used to hybridize with DNA array, followed by dot coloration. An MHRCA-based membrane DNA array assay (MHBMDAA) for detecting toxic microalgae was developed. The specificity of the MHBMDAA was confirmed by double cross-reactivity tests of PLP and taxonomic probes. The MHBMDAA was competent for detecting the simulated samples with 103 to 10-1 cells mL-1, which is 10-fold more sensitive than a multiplex PCR-based membrane DNA array. The effectiveness of the MHBMDAA was also validated by testing with natural samples from the East China Sea. Results indicated that the MHBMDAA provides a valuable tool for the sensitive and reliable detection of toxic microalgae for early warning and research purposes.

In recent years, the scale and frequency of outbreaks of harmful algal blooms (HABs) have increased year by year due to the intensification of seawater eutrophication and global climate change. HABs have become a global marine ecological and environmental problem, which poses a serious threat to human health, marine ecological security, and economic development. The establishment of detection technology for harmful microalgae is fundamental to the early warning and prevention of HABs. To date, several detection methods have been developed for harmful microalgae, they however lack a unified classification standard. It is difficult to use a reasonable mix of all the developed methods to improve the accuracy of detection results. Here, all of the established detection methods for harmful microalgae were reviewed, including morphological structure-based detection methods, cytochrome-based detection techniques, immunoassays, and nucleic acid-based detection methods. The principles, advantages, and weaknesses of these methods were highlighted. Their application in the detection of harmful microalgae was summarized. Overall, different detection methods are suitable for different purposes. Further development of more accurate, cost-effective, efficient, and rapid detection technology is required in the future. This review is expected to provide a reference for research related to the monitoring of marine environment, early warning of HABs, and the molecular identification of harmful microalgae.

Prorocentrum donghaiense is an important dinoflagellate as it frequently forms harmful algal blooms that cause serious damage to marine ecosystems and fisheries in the coast of East China Sea. Previous studies showed that phosphorus acquisition (especially inorganic phosphorus) was the limiting factor for P. donghaiense growth. However, the responsive mechanism of this microalga under dissolved inorganic phosphorus (DIP) limitation is poorly understood. In this work, the physiological parameters and differentially expressed genes in P. donghaiense response to DIP limitation were comparatively analyzed. DIP-depleted P. donghaiense displayed decreased growth rate, enlarged cell size, decreased cellular phosphorus content, and high AP activities. A forward suppression subtractive hybridization (SSH) library representing differentially upregulated genes in P. donghaiense under DIP-depleted conditions was constructed, and 134 ESTs were finally identified, with a significant identity (E values

In this work, a rapid, simple and sensitive colorimetric sensor for simultaneous (or respective) detection of Cr3 + and Pb2 + using tyrosine functionalized gold nanoparticles (AuNPsTyr) has been developed. Tyrosine, a natural and zwitterionic amino acid, could be as a reducing and capping agent to synthesise AuNPs and allow for the simultaneous and selective detection of Cr3 + and Pb2 +. Upon the addition of Cr3 + or Pb2 + (a combination of them), the color of AuNPsTyr solution changes from red to blue grey and the characteristic surface plasmon resonance (SPR) band is red-shifted to 580 nm due to the aggregation of AuNPs. Interestingly, the aggregated AuNPsTyr can be regnerated and recycled by removing Pb2 + and Cr3 +. Even after 3 rounds, AuNPsTyr show almost the same A580 nm / A520 nm value for the assays of Pb2 + and Cr3 +, indicating the good recyclability of the colorimetric sensor. The responding time (within 1 min) and sensitivity of the colorimetric sensor are largely improved after the addition of 0.1 M NaCl. Moreover, the AuNPsTyr aggregated by Cr3 + or Pb2 + (a combination of them) show excellent selectivity compared to other metal ions (Cr3 +, Pb2 +, Fe2 +,Cu2 +,Zn2 +,Cr6 +,Ni2 +,Co2 +,Hg2 +,Mn2 +,Mg2 +,Ca2 +,Cd2 +). More importantly, the developed sensor manifests good stability at room temperature for 3 months, which has been successfully used to determine Cr3 + and Pb2 + in the real water samples with a high sensitivity.

A highly sensitive and visible-light-driven photoelectrochemical (PEC) sensor has been constructed for the quantitative assay of organophosphorus pesticides like chlorpyrifos (Chlo) using hollow Co9S8@CdS heterostructures. The Co9S8@CdS heterostructures were created by growing CdS onto Co9S8 surface via one-step oil-bath route, of which Co9S8 was fabricated with hollow tubular structures through the Co(CO3)0.35Cl0.2(OH)1.1-templated solvothermal method. It was discovered that the so-obtained Co9S8@CdS with direct Z-scheme heterojunctions could exhibit larger photocurrents than the Co9S8 or CdS because of the greatly enhanced separation efficiency of photo-generated carriers. What’s more, the photocurrent signal of Co9S8@CdS-based PEC sensor could be selectively inhibited with the addition of Chlo through the specific chelation between the N and S atoms of Chlo with C N and P S groups and Co sites of Co9S8@CdS so as to suppress the carriers transferring. The developed PEC sensor could exhibit a linear response for Chlo ranging from 0.050 to 1000 ppb, with the detection limit value down to 0.015 ppb, which was far below than those of currently reported PEC sensors. Besides, the proposed routes may provide guidelines for designing various direct Z-scheme heterojunction-based PEC analysis platforms for determining environmental pollutants like Chlo.

Graphene/GaN nanocomposites have important applications in GaN-based LEDs. Since the electrical transport characteristics of this composite have been widely studied, the thermal transport properties have not been systematically studied. Herein, the thermal transport properties of suspended graphene and graphene supported by GaN including length, substrate effect and temperature dependence are carefully studied by molecular dynamics (MD) simulation. When the length is 250 nm, the thermal conductivity of GaN-supported graphene is 1567.5±46.4 W/mK, which is about 85% of that for the suspended graphene at room temperature. More interesting, a theoretical model is used to describe substrate effect by modifying the phonon dispersion and density of states, and the conclusion of this model is consistent with our MD simulations. Moreover, the underlying mechanism of substrate effect and temperature dependence is revealed by spectral energy density (SED) analysis. Our study reports the finite suppression of GaN on the thermal conductivity of graphene, which provides insights for thermal management of GaN-based LEDs.

A near-infrared (NIR) light-driven photoelectrochemical (PEC) sensing platform was established for the quantitative assay of Hg2+ using Ag@Ag2S nanocomposites. Ag2S quantum dots (QDs) with the bead-chain-like nanostructure were fabricated simply via one-step oil-bath route to be further deposited with metallic Ag through the photo-reduction method to yield Ag@Ag2S nanocomposites. It was found that Ag@Ag2S-based sensor could present more than twice larger PEC responses than the Ag2S-based one, due to the synergetic combination of metallic Ag with the surface plasmon resonance (SPR) effects and Ag2S QDs with the NIR-light-irradiated PEC properties. More importantly, the developed PEC sensor could display the photocurrent responses that are selectively inhibited in the presence of Hg2+ by forming Ag-Hg amalgam to accelerate the recombination of photo-induced electron-hole pairs. Under NIR light, it can detect Hg2+ in the concentrations linearly ranging from 0.0010 to 5.0 nM, with a limit of detection of about 0.50 pM. This work may guide a promising combination of Ag2S of NIR QDs with metallic Ag of SPR effects for designing various PEC sensing platforms for the complex environmental monitoring of different heavy-metal ions like Hg2+.

In this study, a novel detection method by PCR-based dot chromatography strip (PDCS) is proposed. To investigate the application of PDCS in the detection of harmful microalgae, the internal transcribed spacer sequence of Karlodinium veneficum, one of the most common bloom-forming species, was cloned and sequenced to design and screen specific primers with tag sequences and probes, including gold nanoparticle probe, test probe, and control probe. The PDCS was prepared manually, and PCR amplicons prepared from the genomic DNA of K. veneficum using tagged specific primers were analyzed by PDCS for visual detection of the target species. The resulting test strip showed red spots at the predicted test and control points visible to the naked eyes, showing the successful development of PDCS. This detection technique is independent of expensive experimental equipment (except a DNA thermal cycler for PCR) but requires an aliquot of PCR amplicons mixed with development buffer to apply to the sample pad of PDCS for approximately 10 min to visualize the analytical results. Cross-reactivity test with 21 microalgae, including K. veneficum, showed that the established PDCS technique has excellent specificity. The detection limit of PDCS was 9.13 × 10−2 ng μL−1 for genomic DNA and 5.3 × 105 cells L − 1 for crude DNA extracts of the target alga. In summary, the PDCS with high sensitivity and specificity can be prepared by hand, which is less expensive than traditional strip, thus providing a promising alternative to the detection of K. veneficum in natural samples.

Scallop Chlamys farreri is an important aquaculture species in northern China. However, its mass mortality caused by several pathogens can result in great economic loss and negative impacts to the sustainable development of the scallop industry. Thus, improving the overall understanding of immune response mechanisms involved in host-pathogen interactions is necessary. Ferritins are conserved molecules in organisms that are involved in diverse biological processes, such as mediating host-pathogen responses. In this study, we report a novel ferritin gene from C. farreri (denoted as CfFER). The full length of CfFER is 848 bp and contains a 5'-UTR of 113 bp, a 3'-UTR of 219 bp, and a complete open reading frame (ORF) of 516 bp. The ORF encodes a polypeptide of 171 amino acid residues with a molecular weight of approximately 19.95 kDa and an isoelectric point of 5.07. The CfFER protein exhibited typical ferritin structures, namely, a ferroxidase diiron center, a ferrihydrite nucleation center, and an iron-binding response signature. Phylogenetic analysis revealed that CfFER was closely related to other mollusk ferritin proteins. Expression of CfFER in different tissues was analyzed by quantitative real-time PCR, and results showed that CfFER was ubiquitously expressed in all examined tissues. The highest and lowest expression levels of CfFER were measured in the muscle and hemocyte, respectively. The relative mRNA expression of CfFER in response to bacterial (Vibrio anguillarum) and viral (acute viral necrobiotic virus) challenges sharply increased by ca. 5-fold about12 h post-infection (hpi) and then normalized at 48 hpi. Western blot analysis with polyclonal antibodies generated from the recombinant product of CfFER also demonstrated the presence of ferritin protein in hemocytes. These findings strongly suggest that CfFER is involved in the immune response of C. farreri and protection against pathogen challenge.

The magnetization behaviors in Sm–Ni–Fe–Cu alloys at low temperatures have been investigated. It was found that the hysteresis loops show wasp-waisted character at low temperatures, which has been proved to be related to the existence of multi-phases, the Fe/Ni soft magnetic phases and the CaCu 5 -type hard magnetic phase. A smooth-jump behavior of the magnetization is observed at T >5 K, whereas a step-like magnetization process appears at T 5 -type phase is responsible for such abnormal magnetization behavior. The magnetic moment reversal model with thermal activation is used to explain the relation of the critical magnetic field ( H cm ) to the temperature ( T >5 K). The reversal of the moment direction has to cross over an energy barrier of about 6.6×10 −15 erg. The step-like jumps of the magnetization below 5 K is proposed to be resulted from a sharp increase of the sample temperature under the heat released by the irreversible domain wall motion.

We study a firm that makes new products in the first period and collects used products through trade-in, along with new product sale, in the second period. To conduct a convincing analysis, we initially evaluate the problem in a duopoly situation in which one firm (firm A) implements trade-in and the other one (firm B) does not. We subsequently introduce the competitive environment in a two-period planning horizon to identify thresholds that determine the trade-in operations, and then derive the equilibrium decisions of the resulting scenarios. We characterize the optimal production quantities that are associated with parameter b (the sum of used product salvage value and government subsidy) in the Nash equilibrium. Results indicate that adopting trade-in could bring competitive advantage for firm A in terms of market share and profit. If the new product sale is comparatively profitable, then the trade-in firm may forgo some of the collection margin by raising the trade-in rebate and selling additional units to increase new product sale in the second period. Moreover, the total collection quantity does not always increase with government subsidy. We consequently expand the model to the case where both firms compete in trade-in and derive the corresponding decision space of the duopoly firms. Finally, we explore the effect of adopting trade-in on consumer surplus and compare it in the two models.

Visible-light-driven and ultra-stable CoSx/CdS photocatalysts were controllably fabricated by the ZIF-67-templated one-pot solvothermal route for highly efficient photocatalytic Cr(VI) reduction. It was discovered that the resulting CoSx/CdS composites could present different photocatalytic activities controllably depending on the CoSx-to-CdS molar ratios used. The one with 1/16 M ratio could display the highest photocatalytic reduction performance for Cr(VI) with the efficiency up to 100% within 30 min, which is 5.6-fold and 2.1-fold higher than that of pristine CoSx and CdS, respectively. Herein, the components of CoSx and CdS with well-matched energy-band structures might serve separately as the electrons donor and the holes quencher to prolong the lifetime of photogenerated electrons in CoSx/CdS composites. A direct Z-scheme heterojunction could be thus constructed for promoting the charge carrier transferring with the improved redox abilities, leading to the robust photocatalysis of CoSx/CdS photocatalyst. Moreover, the developed CoSx/CdS photocatalyst could present superior stability with the structure and activity well survived after five photocatalytic cycles. This work may pave the way towards the controllable construction of a direct Z-scheme photocatalytic system by synergistically combining two kinds of components with matched energy-band structures, achieving the enhanced photocatalysis for the wide applications in the environmental remediation field.

A visible-light-driven photoelectrochemical (PEC) sensor has been developed for probing Hg2+ with “turn-on” signal output using hollow CoSx@CdS polyhedron as direct Z-scheme photocatalyst and Hg2+-recognition probe. The CoSx@CdS heterojunction photocatalysts were fabricated by growing CdS nanoparticles on the surface of cobalt sulfide (CoSx) via one-pot solvothermal route using metal-organic frameworks (MOFs) of ZIF-67 polyhedrons as the sacrificial templates and cobalt precursors. It was discovered that the photocurrent responses of the CoSx@CdS-modified ITO electrodes could be specifically turned on by Hg2+, in contrast to these of the CoSx or CdS-modified ones showing no significant Hg2+-induced photocurrent. Under visible light irradiation, herein, the synergetic combination of CoSx and CdS components could improve the carriers transferring of PEC system. More importantly, the photocurrents of the PEC sensor could be greatly enhanced in the presence of Hg2+ through the selective ion-exchange reaction to trigger the in-situ formation of a new Z-scheme heterojunction of CoSx@CdS/HgS photocatalysts, achieving the further promoted charge transferring and suppressed recombination of photogenerated electron-hole pairs. The developed double heterojunctions-based PEC sensors can detect Hg2+ with the concentrations ranging from 0.010 to 1000 nM, which performances are much better than those of the current PEC analysis methods. Besides, such a MOF-templated one-pot construction route may pave the way toward the design of various direct Z-scheme heterojunction photocatalysts of PEC probes of great interest.

Chattonella marina is one of the main algae that could cause harmful algal blooms. It has killed a large number of cultured fish in coastal areas of many countries, causing serious economic losses. Therefore, it is necessary to establish a method that can specifically detect C. marina at pre-bloom abundance, so that timely measures can be taken before this alga causes harm. In this study, a long probe, a short probe and a pair of amplification primers were first designed by using the internal transcribed spacer (ITS) sequence of C. marina as the target gene and using the CD74 gene of a distant species Gallus gallus as the base sequence. The double probes rolling circle amplification (dpRCA) system was then established with the designed probes and amplification primers. A novel detection protocol referred to as dpRCA-LFD by combining the dpRCA products and lateral flow dipstick (LFD) was finally established, which can make the detection results visible to the naked eyes. The reaction conditions of dpRCA were optimized and the optimal conditions were as follows: cycle number of ligation reaction, 12; ligation temperature, 58 °C; amplification temperature, 60 °C; and amplification time, 60 min. The specificity test that was performed using the optimized dpRCA conditions indicated that dpRCA-LFD was exclusively specific for the target alga. The tests with the genomic DNA of C. marina and the recombinant plasmid containing the ITS sequence of C. marina showed that the sensitivity of dpRCA-LFD was 100 times higher than that of conventional PCR. The detection limit (DL) for the genomic DNA was 8.3 × 10−3 ng µL−1 (8.3 × 10−3 ng per reaction), and the DL for the recombinant plasmid DNA was 7.8 copies µL−1 (7.8 copies per reaction). The practicality of the developed dpRCA-LFD was further validated by test with the spiked samples containing C. marina and field samples. The simulative test showed that the dpRCA-LFD has a DL of 10 cells mL−1. The dpRCA-LFD could successfully recognize the target cells from the field samples. In summary, the dpRCA-LFD established in this study has advantages of good specificity, high sensitivity, and easily visible detection results, and therefore is promising for the analysis of C. marina in field samples.

A visible-light-driven photoelectrochemical (PEC) sensor has been developed for the "signal-on" analysis of Hg2+ by the synergetic combination of low-bandgap Ag2S and wide-bandgap ZnS quantum dots (QDs). Ag2S QDs were synthesized with bead-chain-like structure by the self-assembly route and further covalently bound with ZnS QDs to be coated onto the indium tin oxide (ITO) electrodes. It was discovered that the ZnS@Ag2S-modified electrodes could display the visible-light-driven PEC behavior, of which Ag2S and ZnS QDs could act as the PEC sensitizer and Hg2+-recognition probe, respectively. More importantly, the photocurrent responses of the developed electrodes could be specifically turned on in the presence of Hg2+ under the visible-light irradiation, presumably due to that Hg2+ might conduct a Zn-to-Hg exchange on ZnS QDs to trigger the formation of HgS/ZnS@Ag2S heterojunction towards the enhanced electron-hole separation. The as-prepared PEC sensor could facilitate the detection of Hg2+ with concentrations ranging from 0.010-1000 nM, with a detection limit of about 1.0 pM. Besides, the feasibility of practical applications of the developed PEC analysis strategy was verified by probing Hg2+ in environmental water samples. Such a visible-light-driven PEC detection platform with the unique "turn-on" signal output may promise for the extensive applications for Hg2+ evaluation.

Harmful algal blooms caused by Chattonella marina recently have caused severe negative effect on coastal economy worldwide, with increased occurrence frequency and scale. It is therefore vital to establish new methods for rapid detection of this alga. In this study, the internal transcribed spacer (ITS) sequence was used as the target gene for molecular detection of C. marina. First, four loop-mediated isothermal amplification (LAMP) primers were designed based on the six regions of ITS, and the LAMP reaction system was established using these primers. Next, a probe was designed to detect the LAMP products by lateral-flow dipstick (LFD). Finally, a new method for rapid and sensitive detection of C. marina that is referred to as LAMP-LFD was established. The LAMP reaction system, amplification time, and amplification temperature were particularly optimized. The optimal parameters are as follows: Mg2+ concentration, 10 mM；dNTP concentration, 1.2 mM；ratio of internal primer concentration to outer primer concentration, 8:1；reaction time, 60 min；and reaction temperature, 60 °C. Both specificity and sensitivity were tested using the optimized LAMP reaction system in combination with LFD (LAMP-LFD). The established LAMP-LFD displayed good specificity and no cross reaction was detected with non-target algal species. The detection limit of LAMP-LFD was 3.4 × 10−4 ng μL−1 (3.4 × 10−4 ng per reaction) for the genomic DNA of target algae, and 1.3 copies μL−1 (1.3 copies per reaction) for the plasmid DNA containing the target ITS. Sensitivity tests using genomic DNA and plasmid DNA as templates consistently revealed that LAMP-LFD is 100 times more sensitive than regular PCR. The established LAMP-LFD was applied to analyze the simulated samples and the results showed that the detection limit of LAMP-LFD could reach 1 cell mL−1. LAMP-LFD also demonstrated good specificity and sensitivity in the analysis of natural samples. The whole procedure of LAMP-LFD could be completed within 1.5 h. Taken together, the LAMP-LFD assay developed here is characterized by simplicity, high specificity and sensitivity, and rapidity and therefore is promising for rapid detection of C. marina.

The scallop Chlamys farreri is an important aquaculture species in northern China. However, the sustainable development of the scallop industry is currently threatened by several pathogens that cause mass mortality of this mollusk. Therefore, a complete understanding of the immune response mechanisms involved in host-virus interactions is necessary. This study reports a novel QM gene from C. farreri. This gene was first identified as a putative tumor suppressor gene from human and then confirmed to participate in several functions, including immune response. The QM gene from C. farreri (CfQM) was identified by suppression subtractive hybridization, and its full-length (763 bp) cDNA was obtained through rapid amplification of cDNA ends. The cDNA of CfQM contained a short 5'-UTR of 22 bp and a 3'-UTR of 84 bp. Its ORF comprised 657 nucleotides that encode 218 amino acids with a molecular weight of approximately 28.3 kDa and an isoelectric point of 10.06. The deduced amino acid sequence of CfQM contained a series of conserved functional motifs that belong to the QM family. Phylogenetic analysis revealed that CfQM was closely related to other mollusk QM proteins, and altogether they form a mollusk QM protein subfamily that displays evolutionary conservation from yeast to human. The tissue-specific expression and transcriptional regulation of CfQM were investigated by quantitative real-time PCR in response to bacterial (Vibrio anguillarum) and viral (acute viral necrobiotic virus) challenges. The transcript level of CfQM was high in all of the examined tissues in a constitutive manner. The highest and lowest expression levels of CfQM were measured in the hepatopancreas and hemocyte, respectively. Upon bacterial and viral challenges, the relative mRNA expression of CfQM sharply increased at 6 h post-infection (hpi) and then normalized at 48 hpi. These findings suggest that CfQM can respond to and protect against pathogen challenge. To the best of our knowledge, this study is the first report of the QM gene from scallop. The results presented herein provided new insights into the molecular basis of host-pathogen interactions in C. farreri.

High quality of coastal water is critical to marine ecosystems, marine fisheries, public health, and aquatic environment. Specially, bio-toxin derived from toxic microalgae is currently threatening many coastal countries. Therefore, development of rapid and sensitive methods for the detection of toxin-producing microalgae is necessary for warning of water quality. In this paper, we established a novel method for rapid and sensitive detection of Amphidinium carterae by hyperbranched rolling circle amplification (HRCA). The partial large subunit rDNA (LSU D1–D2) of A. carterae was sequenced to design species-specific padlock probe (PLP). The PLP-coupled with two amplification primers were employed for HRCA. The optimized HRCA conditions were as follows: padlock concentration, 20 pM; ligation temperature, 65 °C; ligation time, 15 min; amplification temperature, 61 °C; and amplification time, 15 min. The developed HRCA was confirmed to be specific for A. carterae by tests with other algae. The sensitivity of HRCA was 100-fold higher than regular PCR, exhibiting a detection limit of 1 fg/μL representing 283 copies for the recombinant plasmid containing the target LSU D1–D2, and 1 cell for target species. Finally, a simplified protocol was applied to the simulated field and environmental materials, and exhibited a good performance. The whole detection could be completed within 1.5 h, displaying a repeated detection limit of 1 cell. The positive HRCA results could be visualized through coloration reaction by adding the fluorescent dye SYBR Green I to the amplification products. The HRCA provides a useful tool to quickly screen large sample sets for A. carterae, as well as other toxic species.

Aberrant Notch signaling is observed in human hepatocellular carcinoma (HCC) and has been associated with the modulation of cell growth. However, the role of Notch signaling in HCC and its underlying mechanism remain elusive. RBP-J-interacting and tubulin-associated (RITA) mediates the nuclear export of RBP-J to tubulin fibers and downregulates Notch-mediated transcription. In this study, we found that RITA overexpression increased protein expression of p53 and Fbxw7 and downregulated the expression of cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. These changes led to growth inhibition and induced G0/G1 cell cycle arrest and apoptosis in SMMC7721 and HepG2 cells. Our findings indicate that RITA exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis and suggest a therapeutic application of RITA in HCC.

The sustainable development of the scallop Chlamys farreri industry in China is hindered by mass mortality mainly caused by a novel pathogen known as acute viral necrosis virus (AVNV). A better understanding of host-virus interactions, especially those at the molecular level, may facilitate the prevention and cure of AVNV infections. MicroRNAs (miRNAs) represent a class of small RNA molecules involved in several biological processes, including mediating host-pathogen responses. In this study, two hemocyte small RNA libraries were constructed from control (control library, CL) and AVNV-infected (infection library, IL) C. farreri for high throughput sequencing using Solexa technology. Acquired data were further used to identify conserved and novel miRNAs, screen differentially expressed miRNAs, and predict their target genes through bioinformatics analysis. Solexa sequencing produced 19,485,719 and 20,594,513 clean reads representing 2,248,814 and 1,510,256 unique small RNAs from CL and IL, respectively. A total of 57 conserved miRNAs were identified in both libraries, among which only two were unique to IL. Novel miRNA prediction using the Crassostrea gigas genome as a reference revealed 11 candidate miRNAs, 10 of which were validated by RT-PCR. Differential expression (p < 0.001) between libraries was observed in 37 miRNAs, among which 30 and 7 miRNAs were up- and downregulated, respectively. Expression differences were further confirmed by qRT-PCR. A sequence homology search against available C. farreri ESTs showed that these differentially expressed miRNAs may target 177 genes involved in a broad range of biological processes including immune defense and stress response. This study is the first to characterize C. farreri miRNAs and miRNA expression profiles in response to AVNV infection by deep sequencing. The results presented here will deepen our understanding of the pathogenesis of AVNV at the molecular level and provide new insights into the molecular basis of host-pathogen interactions in C. farreri.

Prorocentrum donghaiense is the most common bloom-forming species in the East China Sea, causing serious damage to regional marine ecosystems, marine fisheries, public health, and aquatic environment. To minimize fisheries losses caused by this harmful alga, a simple and accurate detection method need to be developed to provide adequate early warning for P. donghaiense blooms. In this study, we report the development and application of reverse transcription-coupled loop-mediated isothermal amplification (RT-LAMP) in the field detection of P. donghaiense. The partial large subunit rDNA (LSU D1-D2), small subunit rDNA, and internal transcribed spacers of P. donghaiense were first sequenced to design species-specific amplification primers. Primer set screen showed that the LSU D1-D2 was the best target for LAMP in terms of sensitivity and stability. The amplification conditions including the dNTP and betaine concentrations, the inner primer to outer primer concentration ratio, reaction time, and temperature, were optimized. The specificity of RT-LAMP for P. donghaiense was also confirmed through tests using a few common harmful algae. RT-LAMP targeting RNA exhibited a detection limit of 0.6 cells, which is more sensitive than LAMP and PCR targeting DNA and reverse transcription-polymerase chain reaction (RT-PCR) targeting RNA. Finally, an improved protocol for natural samples was applied to the field material. The optimized detection protocol could be completed within 1 h. In addition, positive RT-LAMP results could be confirmed through the production of white magnesium pyrophosphate precipitate or through mixing the fluorescent dye GeneFinder (TM) with the amplification products. In summary, the established RT-LAMP is specific, sensitive, and rapid method that is promising for the field detection of P. donghaiense. (c) 2013 Elsevier B.V. All rights reserved.

Acute viral necrosis virus (AVNV) was newly reported as one causative agent responsible for mass mortality of adult Chinese scallop Chlamys farreri , which is widely cultured on northern China coast. Unfortunately, the interaction between virus and host is largely unknown. According to these, this study was undertaken to deeply explore the immune response of haemocyte against AVNV. Two-dimensional gel electrophoresis (2-DE) was introduced to produce protein expression profiles from samples taken at 24 h post-infection (hpi) from the haemocytes of C. farreri that were either specific pathogen free or else infected with AVNV. Forty-eight protein spots, which consistently showed either a marked change (≥1.5-fold difference) in accumulated levels or else were highly expressed in haemocytes, were selected for further investigation. In-gel trypsin digestion was conducted followed by matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF-MS). Matching search was subsequently performed throughout bioinformatics databases. A total of 42 proteins were identified, all of which were classified into eight categories according to their Gene Ontology annotations of biological processes and molecular functions, i.e. cytoskeleton proteins, proteins involved in metabolism, proteins related to calcium homeostasis, chaperone, proteins involved in immunity, proteins involved in transcriptional regulation, proteins related to signal transduction, and ungrouped proteins. The possible biological significance of some observed proteins in the host response to AVNV was discussed. These studies could be served as the first global analysis of differentially expressed proteins in haemocytes from AVNV-infected C. farreri , and in addition to increasing our understanding of the pathogenesis of this virus-associated scallop disease, the results presented here should be useful both for potential biomarkers identification and anti-virus approaches development as well.

Pulses of 177 fs and 1035 nm, with average power of 1.2 mW, have been generated directly from a passively mode-locked Yb-doped figure-of-eight fiber laser, with a nonlinear optical loop mirror for mode-locking and pairs of diffraction gratings for intracavity dispersion compensation. To our knowledge, these are the shortest pulses ever to come from a passively mode-locked Yb-doped figure-of-eight fiber laser. This represents a 5-fold reduction in pulse duration compared with that of previously reported passively mode-locked Yb-doped figure-of-eight fiber lasers. Stable pulse trains are produced at the fundamental repetition rate of the resonator, 24.0 MHz.

A simple folded-cavity used for intracavity frequency-doubled Nd:YVO4 laser was analyzed by transmission matrix and numerical calculation. We have selected a set of proper cavity parameters which can be used readily by operating at high power levels with low threshold, high e:ciency, and wide dynamic operating range. 5.6-W TEM 00 green laser has been obtained at a 22-W pumping power and with an optical–optical conversion e:ciency of 25.5%. ? 2002 Elsevier Science Ltd. All rights reserved.

The mechanism of pulse shaping in Kerr-lens mode-locking Ti : sapphire lasers is investigated. Considering the balanced interaction between self-phase modulation, introduced by Kerr effect in Ti : sapphire, and group-velocity dispersion provided by prism-pair, pulse-splitting reflected in the intensity autocorrelation trace and pulse-compression display that the pulses outputting from different positions in laser cavity have different durations. We have concluded that the solitonlike pulse shaping that results from the competition between intracavity self-phase modulation within the Ti : S and negative group-velocity dispersion play dominant role in pulse evolution in Kerr-lens mode-locking lasers.