Transcriptomics reveals different response mechanisms of Litopenaeus vannamei hemocytes to injection of Vibrio parahaemolyticus and WSSV.

As the most important cultural crustacean species worldwide, studies about Pacific white shrimp (Litopenaeus vannamei) have received more attention. It has been well-documented that various pathogens could infect L. vannamei, resulting in huge economic losses. The studies about the responding mechanism of L. vannamei to sole pathogens such as Vibrio parahaemolyticus and white spot virus (WSSV) have been extensively reported, while the studies about the differently responding mechanisms remain unclear. In the present study, we identified the differently expressed genes (DEGs) of L. vannamei hemocytes post V. parahaemolyticus and WSSV infection with RNA-seq technology and compared the DEGs between the two groups. The results showed 2672 DEGs post the V. parahaemolyticus challenge (1079 up-regulated and 1593 down-regulated genes), while 1146 DEGs post the WSSV challenge (1067 up-regulated and 513 down-regulated genes). In addition, we screened the genes that simultaneously respond to WSSV and V. parahaemolyticus (434), solely respond to WSSV (1146), and V. parahaemolyticus challenge (2238), respectively. Six DEGs involved in innate immunity were quantified to validate the RNA-seq results, and the results confirmed the high consistency of both methods. Furthermore, we found plenty of innate immunity-related genes that responded to V. parahaemolyticus and WSSV infection, including pattern recognition receptors (PRRs), the proPO activating system, antimicrobial peptides (AMPs), and other immunity-related proteins. The results revealed that they were differently expressed after different pathogen challenges, demonstrating the complex and specific recognition systems involved in defending against the invasion of different pathogens in the environment. The present study improved our understanding of the molecular response of hemocytes of L. vannamei to V. parahaemolyticus and WSSV stimulation.

Facile synthesis of mesoporous ZSM-5 aided by sonication and its application for VOCs capture.

Application of ultrasound power to the mother liquor is popular pretreatment for zeolite synthesis which offers a simple way of accelerating crystallization process and finetuning the properties of nanocrystalline zeolites. In this work, sonication-aided synthesis of mesoporous ZSM-5 at low temperature and ambient pressure was systematically studied, in an attempt to reach efficient and benign synthesis of zeolites with hierarchical pore structure, which has wide applications as catalysts and sorbents. The effects of sonication duration, power density, sonication temperature and seeding on the crystallization of ZSM-5 were investigated. The obtained samples were characterized by XRD, SEM, BET and VOCs capture. High quality mesoporous ZSM-5 can be obtained by a facile 5 d synthesis at 363 K, much faster than conventional hydrothermal synthesis. The reduced synthesis time was mainly attributed to the enhanced crystallization kinetics caused by the fragmentation of seeds and nuclei, while sonication radiation had little impact on the nucleation process. Compared with control sample, mesoporous ZSM-5 prepared by sonochemical method had higher surface area and mesoporosity which demonstrated improved adsorption performance for the capture of isopropanol.

Chlorella sp. transgenic with Scy-hepc enhancing the survival of Sparus macrocephalus and hybrid grouper challenged with Aeromonas hydrophila.

Two marine antimicrobial peptides (AMPs), PC-hepc from large yellow croaker (Pseudosciaena crocea) and scygonadin from mud crab (Scylla serrata), are potently active against specific bacteria and thus they could be used as substitutes for antibiotics in aquaculture. However, how to utilize the AMPs feasibly for marine cultured animals has been so far confused. In our study, a 510 bp of the Scy-hepc sequence was cloned into pMDC85 expression vector, which was then electroporated into Chlorella sp., and thus a transgenic Chlorella, in which the Scy-hepc gene was effectively expressed, was developed. The Scy-hepc fusion protein was successfully expressed in Chlorella sp. and it showed obvious bactericidal activity. In addition, the in vivo efficacy of the transgenic Chlorella was evaluated using Sparus macrocephalus and the hybrid Epinephelus fuscoguttatus (♀) × Epinephelus lanceolatus (♂). Results showed that the survival rate of S. macrocephalus fed with transgenic Chlorella (80 ± 10% after 72 h) was significantly higher than that of fish fed with the same dosage of wild-type Chlorella (33.33 ± 11.55% after 72 h). Similarly, results showed that the survival rate of the hybrid grouper fed with transgenic Chlorella (55 ± 5% after 36 h) was much higher than that of fish fed with the same dosage of wild-type Chlorella (25 ± 5% after 36 h). Therefore, in vitro and in vivo results indicated that the constructed transgenic Chlorella with the marine AMPs Scy-hepc could exert effective protection for fish against the Aeromonas hydrophila infection, providing an encouraging prospect for the expected use of transgenic Chlorella in aquaculture in future.

The oxidative stress of Phanerochaete chrysosporium against lead toxicity.

Among the technologies for heavy metal remediation, bioremediation technology has gained extensive attention because of its low processing costs and high efficiency. The white-rot fungus Phanerochaete chrysosporium (P. chrysosporium) which has a good tolerance to heavy metals has been widely used in the heavy metal bioremediation. In order to figure out the molecular mechanisms involved in the oxidative stress of P. chrysosporium against metal toxicity, we examined the effect of Pb(2+) on the levels of reactive oxygen species and the production of malondialdehyde. Results showed that P. chrysosporium could adjust Pb-stressed condition by regulating the unique oxidation-antioxidation process in cells and kept a balance between oxidation and antioxidation when it was threatened by a different dose of Pb(2+). Investigations into the oxidative stress of P. chrysosporium to lead could not only provide a better understanding of the relationship between lead and oxidative stress in P. chrysosporium, but also offer important informations on the development of fungal-based remediation technologies to reduce the toxic effects of lead.

Effect of low-concentration rhamnolipid on adsorption of Pseudomonas aeruginosa ATCC 9027 on hydrophilic and hydrophobic surfaces.

The effects of low-concentration monorhamnolipid (monoRL) on the adsorption of Pseudomonas aeruginosa ATCC 9027 grown on glucose or hexadecane to glass beads with hydrophobic or hydrophilic surfaces was investigated using batch adsorption experiments. Results showed that adsorption isotherms of the cells on both types of glass beads fitted the Freundlich equation better than the Langmuir equation. The Kf of the Freundlich equation for adsorption of hexadecane-grown cell to glass beads with hydrophobic surface was remarkably higher than that for adsorption of hexadecane-grown cell to glass beads with hydrophilic surface, or glucose-grown cell to glass beads with either hydrophilic or hydrophobic surface. Furthermore, it decreased with the increasing monoRL concentration. For both groups of cells, the zeta potential was close to each other and stable with the increase of monoRL concentration. The surface hydrophobicity of hexadecane-grown cells, however, was significantly higher than that of the glucose-grown cells and it decreased with the increase of monoRL concentration. The results indicate the importance of hydrophobic interaction on adsorption of bacterial cells to surfaces and monoRL plays a role in reducing the bacterial adsorption by affecting cell surface hydrophobicity.

Highly sensitive electrochemical sensor using a MWCNTs/GNPs-modified electrode for lead (II) detection based on Pb(2+)-induced G-rich DNA conformation.

A sensitive electrochemical lead ion (Pb(2+)) sensor based on carboxylic acid group functionalized multi-walled carbon nanotubes (MWNTs-COOH) and direct electrodeposited gold nanoparticles (GNPs) was developed for Pb(2+) detection. The DNA capture probe was self-assembled onto the surface of the modified electrode for hybridizing with the guanine-rich (G-rich) aptamer probe and for forming the DNA double helix structure. When Pb(2+) was added in, the DNA duplex unwound and formed a stabilized G-quadruplex (G4) due to the Pb(2+)-induced G-rich DNA conformation. Also, methylene blue (MB) was selected as the G4-binding indicator. Compared with previous Pb(2+) sensors, the proposed sensor had better sensitivity, because the modified MWCNTs/GNPs could provide a large surface area and good charge-transport capacity to dramatically improve the DNA attachment quantity and sensor performance. The sensor could detect Pb(2+) in a range from 5.0 × 10(-11) to 1.0 × 10(-14) M, with a detection of 4.3 × 10(-15) M.

Role of low-concentration monorhamnolipid in cell surface hydrophobicity of Pseudomonas aeruginosa: adsorption or lipopolysaccharide content variation.

A role of rhamnolipid biosurfactant to enhance the biodegradation of hydrocarbons is known to be enhancing bacterial cell surface hydrophobicity (CSH) and adhesion of cells to hydrocarbons. Assumptions regarding the mechanism for rhamnolipid in changing CSH of Gram-negative bacteria are rhamnolipid-induced release of lipopolysaccharide (LPS) from the cell's outer membrane and adsorption/orientation of rhamnolipid on the cell surface. In this study, the relation between cell-wall LPS or rhamnolipid content and CSH of a Pseudomonas aeruginosa bacterium subjected to rhamnolipid treatment was investigated to add insights to the mechanism. Results showed that the initial CSH was determined by the type of substrate the cells grow on and the stage of growth. For glucose-grown cells with low initial CSH and high LPS content, rhamnolipid sorption in cell wall had no discernable effect on CSH. For cells grown on glycerol with medium initial CSH and low LPS content, rhamnolipid sorption increased CSH of exponential-phase cells but decreased that of stationary-phase cells. For hexadecane-grown cells with high initial CSH and high LPS content, rhamnolipid sorption decreased CSH of both exponential-phase and stationary-phase cells. The results indicated that CSH has a better correlation to the content of rhamnolipid in the cell wall than to the content of LPS in the presence of rhamnolipid treatment and that rhamnolipid adsorption may be an important mechanism for rhamnolipid to alter CSH of P. aeruginosa.

[Purification and characterization of the biosurfactant rhamnolipid].

Biosurfactant rhamnolipid is a metabolic intermediate produced by microorganisms under a certain condition. There are the polar hydrophilic group and the non-polar hydrophobic group in rhamnolipid molecule which always exhibits high surface or interfacial activity. A reliable separation and purification method as well as component identification technique is essential for success of production process. The rhamnolipid was produced by aerobic fermentation using Pseudomonas aeruginosa CCTCC AB93066 in this study. It was separated from the culture by acid precipitation and purified by column chromatography until two groups of monorhamnolipid and dirhamnolipid were obtained. High performance liquid chromatography with mass spectrometry (HPLC-MS) examination showed that either the monorhamnolipid or the dirhamnolipid contained three major species. They were RhaC10C10, RhaC10C12-H2, RhaC10C12 for monorhamnolipid and Rha2C10 C10, Rha2C10 C12-H2, Rha2 C10 C12 for dirhamnolipid. The results of the study suggested that Pseudomonas aeruginosa CCTCC AB93066 is a good strain for rhamnolipid production. Acid precipitation-column chromatography technique is good for purification of rhamnolipid. Meanwhile, HPLC-MS is a reliable method for identifying components of rhamnolipid with high sensitivity and accuracy.

Study on biodegradation process of lignin by FTIR and DSC.

The biodegradation process of lignin by Penicillium simplicissimum was studied to reveal the lignin biodegradation mechanisms. The biodegradation products of lignin were detected using Fourier transform infrared spectroscopy (FTIR), UV-Vis spectrophotometer, different scanning calorimeter (DSC), and stereoscopic microscope. The analysis of FTIR spectrum showed the cleavage of various ether linkages (1,365 and 1,110 cm(-1)), oxidation, and demethylation (2,847 cm(-1)) by comparing the different peak values in the corresponding curve of each sample. Moreover, the differences (Tm and ΔHm values) between the DSC curves indirectly verified the FTIR analysis of biodegradation process. In addition, the effects of adding hydrogen peroxide (H2O2) to lignin biodegradation process were analyzed, which indicated that H2O2 could accelerate the secretion of the MnP and LiP and improve the enzymes activity. What is more, lignin peroxidase and manganese peroxidase catalyzed the lignin degradation effectively only when H2O2 was presented.

Synthesis of gold-cellobiose nanocomposites for colorimetric measurement of cellobiase activity.

Gold-cellobiose nanocomposites (GCNCs) were synthesized by reducing gold salt with a polysaccharide, cellobiose. Here, cellobiose acted as a controller of nucleation or stabilizer in the formation of gold nanoparticles. The obtained GCNCs were characterized with UV-visible spectroscopy; Zetasizer and Fourier transform infrared (FT-IR) spectrophotometer. Moreover, 6-Mercapto-1-hexanol (MCH) was modified on GCNCs, and the MCH-GCNCs were used to determine the cellobiase activity in compost extracts based on the surface plasmon resonance (SPR) property of MCH-GCNCs. The degradation of cellobiose on MCH-GCNCs by cellobiase could induce the aggregation, and the SPR absorption wavelength of MCH-GCNCs correspondingly red shifted. Thus, the absorbance ratio of treated MCH-GCNCs (A650/A520) could be used to estimate the cellobiase activity, and the probe exhibited highly sensitive and selective detection of the cellobiase activity with a wide linear from 3.0 to 100.0U L(-1) within 20 min. Meanwhile, a good linear relationship with correlation coefficient of R2=0.9976 was obtained. This approach successfully showed the suitability of gold nanocomposites as a colorimetric sensor for the sensitive and specific enzyme activity detection.

Heavy metal-induced glutathione accumulation and its role in heavy metal detoxification in Phanerochaete chrysosporium.

Phanerochaete chrysosporium are known to be vital hyperaccumulation species for heavy metal removal with admirable intracellular bioaccumulation capacity. This study analyzes the heavy metal-induced glutathione (GSH) accumulation and the regulation at the intracellular heavy metal level in P. chrysosporium. P. chrysosporium accumulated high levels of GSH, accompanied with high intracellular concentrations of Pb and Cd. Pb bioaccumulation lead to a narrow range of fluctuation in GSH accumulation (0.72-0.84 µmol), while GSH plummeted under Cd exposure at the maximum value of 0.37 µmol. Good correlations between time-course GSH depletion and Cd bioaccumulation were determined (R (2) > 0.87), while no significant correlations have been found between GSH variation and Pb bioaccumulation (R (2) < 0.38). Significantly, concentration-dependent molar ratios of Pb/GSH ranging from 0.10 to 0.18 were observed, while molar ratios of Cd/GSH were at the scope of 1.53-3.32, confirming the dominant role of GSH in Cd chelation. The study also demonstrated that P. chrysosporium showed considerable hypertolerance to Pb ions, accompanied with demand-driven stimulation in GSH synthesis and unconspicuous generation of reactive oxygen stress. GSH plummeted dramatically response to Cd exposure, due to the strong affinity of GSH to Cd and the involvement of GSH in Cd detoxification mechanism mainly as Cd chelators. Investigations into GSH metabolism and its role in ameliorating metal toxicity can offer important information on the application of the microorganism for wastewater treatment.

Diversity of two-domain laccase-like multicopper oxidase genes in Streptomyces spp.: identification of genes potentially involved in extracellular activities and lignocellulose degradation during composting of agricultural waste.

Traditional three-domain fungal and bacterial laccases have been extensively studied for their significance in various biotechnological applications. Growing molecular evidence points to a wide occurrence of more recently recognized two-domain laccase-like multicopper oxidase (LMCO) genes in Streptomyces spp. However, the current knowledge about their ecological role and distribution in natural or artificial ecosystems is insufficient. The aim of this study was to investigate the diversity and composition of Streptomyces two-domain LMCO genes in agricultural waste composting, which will contribute to the understanding of the ecological function of Streptomyces two-domain LMCOs with potential extracellular activity and ligninolytic capacity. A new specific PCR primer pair was designed to target the two conserved copper binding regions of Streptomyces two-domain LMCO genes. The obtained sequences mainly clustered with Streptomyces coelicolor, Streptomyces violaceusniger, and Streptomyces griseus. Gene libraries retrieved from six composting samples revealed high diversity and a rapid succession of Streptomyces two-domain LMCO genes during composting. The obtained sequence types cluster in 8 distinct clades, most of which are homologous with Streptomyces two-domain LMCO genes, but the sequences of clades III and VIII do not match with any reference sequence of known streptomycetes. Both lignocellulose degradation rates and phenol oxidase activity at pH 8.0 in the composting process were found to be positively associated with the abundance of Streptomyces two-domain LMCO genes. These observations provide important clues that Streptomyces two-domain LMCOs are potentially involved in bacterial extracellular phenol oxidase activities and lignocellulose breakdown during agricultural waste composting.