The high mobility group A (HMGA) proteins have a close relation with chromatin remodeling and cancer transformation. They are widely expressed during embryogenesis, whereas their expression is absent or very low in adult tissues. The full open reading frame (ORF) of Bombyx mori high mobility group A (BmHMGA) was cloned with the cDNA of B. mori pupae, and the recombinant plasmid, pET-28a(+)-BmHMGA was constructed. The fusion protein His-BmHMGA was correctly expressed in BL21Star (DE3). New Zealand rabbits were immunized with fusion protein purified by Ni-NTA affinity chromatography and the polyclonal antibody was obtained. The antibody had a high specificity and a titer of over 1∶12 800. The results of real-time PCR (RT-PCR) showed that the transcription levels of the BmHMGA gene declined with developmental stage in the order egg, fifth instar larva, pupa and moth. In the fifth instar larva stage, the transcription levels of the BmHMGA gene varied between tissues, declining in the order head, epidermis, ovaries, testis, fat body, stigma, Malpighian tubules, silk glands and midgut. The results of Western blotting and semi-quantitative analysis by enzyme-linked immuno sorbent assay (ELISA) showed that the BmHMGA content was the highest in the egg and the lowest in the moth, and that in the fifth instar larva, the protein levels were obviously higher in the head than that in other tissues. No signals were detected in the midgut or Malpighian tubules. These results were identical to those of RT-PCR. The result of immunocytochemistry in BmN cells showed that the proteins were evenly distributed in the cytoplasm.

Gallibacterium infection was a new infectious disease observed in some areas of China, which was mainly harmful for laying hen, resulting in salpingitis, oophoritis, peritonitis, septicemia, hepatitis, enteritis, respiratory tract diseases, and so on. So far, most of the G. anatis were isolated from poultry, including chickens, ducks, pigeons, geese, parrots, quail and guinea fowl, etc. Isolation of Gallibacterium from pigeons was not reported in China. A contagious disease occurred in one breeding farm of pigeon in Sichuan Province in January of 2013, with the characteristic signs of depressed wasting, dyspnea, diarrhea, and so on. The morbidity was about 43%, and the mortality was about 18%. The disease was mainly found in female pigeons, and female pigeons of different ages could be infected, especially pigeons of 1 to 1.5 years old. Mild bleeding and mucus could be observed in diseased pigeons, and other gross lesions could also be observed including hemorrhage of lung, peritonitis, salpingitis, mild bleeding of duodenum, and so on. This disease had caused great economic losses to the pigeon farm. In order to propose a proper treatment program for this disease, representative samples were collected from disease pigeons for bacteria isolation and virus detection. The bacteria were isolated by blood agar and were identified by following test: morphological observation, biochemical identification, animal experiment, antibiotic sensitive experiment, 16S rRNA gene sequence analysis. Animal experiments were conducted by infecting mice and pigeons to observe its pathogenicity. For the virus detection, the nucleotide sequences of infectious bronchitis virus (IBV), avian influenza virus subtype (H9N2) and Newcastle disease virus (NDV) were detected by RT-PCR. The results showed that NDV was detected from the lung and trachea of some sick pigeons, and one bacterial strain was isolated from the livers and lungs of sick pigeons and named as SC01. The SC01 strain was gram-negative (G-), short rod-shaped bacteria. It was pathogenic to mice, and the median lethal dose to mice was 1.26×109 CFU/mL. Biochemical test showed that it can ferment most sugars and alcohols. Sensitive drug screen revealed that the SC01 strain was sensitive to ceftriaxone sodium, cefoperazone sodium, florfenicol, cephalexin, cefazolin, and resistant to neomycin, streptomycin, erythromycin, sulfafurazole, etc. The 16S rRNA gene sequence of the SC01 strain was blasted in GenBank, and the results showed that the nucleotide sequence homologies of SC01 and Gallibacterium, Pasteurella, Haemophilus, Actinobacillus were relatively high. The 16S rRNA nucleotide sequence homologies of SC01 and four G. genomo sp. 3 strains were 98.5%99.5%, and the highest homology (99.5%) came from the comparison between the sequences of SC01 and EU423996. Phylogenetic analysis showed that SC01 strain and nine Gallibacterium strains formed an independent big branch in phylogenetic tree. The SC01 strain and G. genomo sp. 3 fell into the same small branch. Meanwhile, Pasteurella, Haemophilus and Actinobacillus formed another big branch in phylogenetic tree. In conclusion, a G. genomo sp. 3 was isolated from diseased pigeons, and its biological character, pathogenicity, drug sensitivity and 16S rRNA sequence were also analyzed. The results provide an important reference for the epidemiological investigation, diagnosis and therapy of G. genomo sp. 3 infection in China. In addition, the appearance of G. genomo sp. 3 infection in pigeons may provide a new challenge for the prevention and therapy of pigeon disease in China.

Parkinsons disease (PD) is a common and age-related progressive neurodegenerative disorder, and its pathogenesis is not yet entirely clear. The present study is to establish a model easy to control and better simulate the clinical symptoms, processes and pathological changes of PD patients, expecting to provide a foundation and platform service for pathogenesis and treatment research of Parkinsons disease. Twelve healthy aging female rhesus monkeys, divided into experimental group (nine rhesus monkeys) and control group (three rhesus monkeys) randomly, were respectively daily injected a small dose (0.2 mg/(kg·d)) of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and the same dose of saline by intramuscular injection for 45 days repeatedly. The behavioral manifestations of all monkeys were evaluated before and after the injection for 30 min, respectively. After sacrificed, the expression and distribution of tyroxine hydroxylase (TH) and α-synuclein (α-syn) in substantia nigra were studied by immunohistochemical method, and the immunopositive total area and average optical density were used to validate the model. The results of behavioral manifestations indicated that, experimental animals showed three different degrees of clinical manifestations and could be divided into three subgroups (three animals each subgroup) according to the behavioral and clinical manifestations. The first subgroup showed typical behavioral manifestations of Parkinsons disease and one animal was in the moribund period. The activity, posture and bradypraxia of the third subgroup were slightly abnormal, and behavioral manifestations of the second subgroup were between the first and the third subgroup. However the control group did not show any abnormality.
The immunopositive productions of TH were mainly distributed at the cytoplasm and neurites. Compared with the control group, the positive cytoplasm and neurites of the first subgroup were severely reduced, even no positive production was observed in partial substantia nigra pars compacta; the neuron structure of the third subgroup was mild blurred, and neurites were slightly shorter and fewer; the positive productions and structure of the second subgroup were between the first
and the third subgroup. Also compared with the control group, the positive total area of each group decreased by 71.90%, 61.90% and 45.74%, showing statistical significance (P<0.01).
The immunopositive productions of α-syn mainly distributed at neuritis, and also distributed at the intercellular substance, and Lewy bodies were detected at the substantia nigra pars compacta of the moribund animal. Compared with the control group, positive total area of each group increased by 170.29%,137.82% and 47.88%, showing statistical significance (P<0.01). The above results suggest that the aging rhesus PD animal model can be established via small doses and repeated intramuscular injection of MPTP, and the PD model can better simulate the clinical behavioral manifestations, disease processes and pathological changes of PD patients, so the model is more reliable for etiology, pathogenesis, drug therapy and gene therapy of Parkinsons disease.

Phalaenopsis spp. is one of the most important orchids, which has a great economic value as potted plants in China. However, growth disorders like damage to membranes, generation of reactive oxygen species (ROS), protein denaturation, and accumulation of toxic compounds due to low temperature stress have become one of the key constraints for Phalaenopsis consumption during winter season in north subtropical area of China. There are a large number of reports about using salicylic acid (SA) to alleviate the negative effects of chilling stress on plants. However there is rare report about the changes of antioxidant systems under chilling stress with exogenous plant regulators application such as SA in Phalaenopsis. The objectives of this study are to investigate the alleviating effects of exogenous SA application on Phalaenopsis under low temperature stress and in recovery growth, to determine the changes of antioxidative enzyme activities and key indicators of photosynthetic system. Five different cultivars of Phalaenopsis (“Hunyan” “Dalajiao” “Tianjiao” “Tianxianggongzhu” and “V31”) were foliar sprayed with a series of SA concentrations (0,0.2,0.4,0.6,0.8 mmol/L). After foliar application, the plants were transferred to a growth chamber treated at low temperature (LT) of 11 ℃ /6 ℃ (day/night) for 3 days and then to recover growth at regular temperature 27 ℃ /22 ℃ (day/night) for 7 days. The changes of antioxidant system and photosynthetic system in Phalaenopsis leaves were tested. The results showed that the foliar spray of SA could obviously alleviate the negative effects of LT stress. In this study, the best concentration of SA was from 0.4 to 0.6 mmol/L. Appropriate concentrations of SA could significantly increase the chlorophyll content, Fv/Fm, superoxide dismutase (SOD) activity, ascorbic acid (AsA) and glutathione (GSH) contents and improve the chilling tolerance of Phalaenopsis. The activity of ascorbate peroxidase (APX) and soluble sugar content were decreased in most of LT stress treatments. After LT stress, exogenous SA could promote the plant growth. After 7 days recovery growth, application of 0.6 mmol/L SA increased SOD and APX activities, chlorophyll content and Fv/Fm, and decreased AsA and GSH contents. The treatment enhanced the photosynthetic and antioxidant capacity of the plants. These results indicated that the application of appropriate concentration of SA can improve the LT tolerance of Phalaenopsis. Compared the different varieties of Phalaenopsis, it is concluded that the variety “Huanyan” is of strong LT tolerance and more sensitive and effective to SA application.

Tomato gray mold is a common disease which is caused by Botrytis cinerea and does harm to tomato seriously. Chemical control of tomato gray mold is easy to produce negative effects, such as drug resistance, pesticide residues and ecological imbalance. Biological control has advantages of high efficiency, no pollution, no drug resistance and combining prevention with treatment. Bio-organic fertilizer has both superiorities of bio-fertilizer and organic fertilizer, which can not only increase production, improve product quality, but also regulate and improve the composition of soil microorganisms, and then reduce the danger of crop diseases to some extent. Screening effective antagonistic strains of tomato gray mold and then preparing antagonistic agent and bio-organic fertilizer are important biotechnology ways to prevent and control tomato gray mold. The studies on solid fermentation of antagonistic strains are less involved at home and abroad. This study aimed at screening an effective antagonistic strain of tomato gray mold, exploring the solid fermentation conditions, preparing biological organic fertilizer and studying its bio-control effect on tomato gray mold. The strains of soil samples were isolated by the method of dilution coated tablet and streak plate, and then screened by tablet confrontation. Furthermore, they were identified by morphological and ITS sequence analysis. The influence factors on the conidial concentration of this strain were investigated by single factor and orthogonal experiments, including solid medium, external carbon, external nitrogen, external phosphate, the ratio of solid material to water, amount of bottled solid material, inoculums percentage, illumination, temperature and time of fermentation. The antagonistic strain was added to the rotten organic fertilizer to ferment, and then it was applied to pot experiment. After isolating, primary screening and rescreening, a strain of number Z47 was obtained eventually, which had the best inhibition effect and its inhibition rate reached 74.80% on B. cinerea. The strain was identified as Penicillium oxalicum. Fermentation condition optimization results for the P. oxalicum Z47 were as follows. The orthogonal optimization results for powdered rice hulls∶bran, maltose, ammonium sulfate, and dipotassium hydrogen phosphate in medium formula were 8∶2, 1.5%, 2% and 0.15%, respectively. The orthogonal optimization results for the ratio of solid material to water, amount of bottled solid material, inoculums percentage, illumination conditions and fermentation temperature in medium formula were 1∶0.6 (g/mL), 15 g, 12.5% (mL/g), dark totally and fermentation at a fluctuant temperature of 25—20—28 ℃ for 2 days, respectively. P. oxalicum Z47 had the highest spore number when cultured for 12 days under the optimal solid medium and culture conditions. The result of the pot experiment showed that the control effect of tomato gray mold by applying biological organic fertilizer which was rich in antagonistic strain Z47 was up to 73.67%, which was significantly higher than the treatment of antagonistic strain Z47 liquid. In sum, under the optimization of solid fermentation, the spore numbers of P. oxalicum Z47 were up to 5×109 CFU/g. The fermentation of biological organic fertilizer has good potential for prevention of tomato gray mold.

Stropharia rugoso-annulata is an edible fungi with strong physiological activity and resistance, which can be cultivated by using rice straw, water bamboo leaves, wheat straw, and other agricultural waste resources.
Zizania latifolia is the largest in the planting area of aquatic vegetables in Zhejiang Province, which was developed rapidly in recent years, and the area was more than 2 667 million square meters. Consequently, a large area of water bamboo planting has produced large amounts of water bamboo leaves, which is a waste of resources and environmental pollution by dumping and incineration, resulting in ecological and economic pressure. At present, the use of rice straw in planting S. rugoso-annulata has been already very mature. In this study, the best cultivation scheme and formula was determined through the comparison of applying different proportions of water bamboo leaves in the culture of S. rugoso-annulata strains, with the rice straw as the control, in order to provide scientific basis for production of S. rugoso-annulata. In order to improve the cultivation of S. rugoso-annulata and reduce the pollution caused by Z. latifolia leaves, three different S. rugoso-annulata strains were cultivated on seven mediums containing different proportions of Z. latifolia leaves to select the best strain and suitable cultivation medium. The results indicated that strain A1 showed the best mycelial growth and yield, followed by strain A2 and A3. Z. latifolia leaves affected the growth of S. rugoso-annulata in a certain degree, especially for mycelial growth, but no significant effect (P<0.05) was observed when the proportion was less than 60%. Therefore, the strain A1 cultivated on substrates with no more than 60% of Z. latifolia leaves would be an appropriate cultivation pattern, which can make full use of the wasted Z. latifolia leaves without significantly affected yield of S. rugoso-annulata. It is suggested that the leaves of Z. latifolia are not a good resources for S. rugoso-annulata cultivation, but it can reduce pollution caused by wasted Z. latifolia leaves.

Tea polyphenols (TPs), as one kind of effective components in tea, are composed of catechins, anthocyanins, flavonoids, etc. Catechins take up 70%80% of TPs. The epigallocatechin gallate (EGCG) is the most abundant and important bioactivity among TPs. Various studies have demonstrated that EGCG possesses a higher efficient bioactivity compared with other components of tea polyphenols, especially in inhibiting carcinogenesis and preventing metastasis in established tumors. Therefore, it was studied widely to develop new methods or techniques to enhance the efficiency of extraction and purification of TPs (including EGCG). Now column chromatography is the main method in purification industry of TPs. However, this technique involves of organic solvents and complex operational processes, which leads to an environmental pollution and increases the additional costs virtually. The demand for an environment-friendly and cheaper technique in TPs industry is growing rapidly while more people become to pay attention to ambient bio-environment, so seeking for some new methods or new materials is of great significance. The hyperbranched poly (amine-ester) (HPAE) is a new sort of materials, having internal cellular structure and a large amount of hydroxyl terminals. HPAE has been widely used as surface modifiers, drug carriers and adsorbents for heavy metal ions on account of its high adsorption properties and complex ability. To date, there is no research on the adsorption of tea polyphenols by HPAE. The objective of this study is to investigate the characteristics of HPAE as a new environment-friendly and low-energy adsorbent for the adsorption of tea polyphenols from aqueous solutions. The adsorbent HPAE was prepared by one step method with 1-(bis (2-hydroxyethyl) amino) pentan-3-one (as a AB2 monomer) and 2-ethyl-2-(hydroxymethyl)-1,3-propanediol (as a core molecule) in our laboratory. The kinetic experiments of adsorption were done with 10 mL HPAE (1%, w/w) and 40 mL 98% TPs solution (3.5 g/L) under the condition of 298 K and pH 7.0. The TPs contents in supernatant were determined by HPLC methods. The experiments about effects of different pH values (from 4.0 to 8.0) on adsorption also were done with similar steps with 40 mL TPs solution (1.25 g/L). Finally, the comparison of the adsorption capacity was made among HPAE and 16 kinds of traditional macroporous resins under the same condition of 298 K, pH=7. The results indicated that the adsorption of tea polyphenols by HPAE increased with time and reached equilibrium time in six hours of contact. The experimental value of the equilibrium adsorption capacity Qe (Exp) was 279.69 mg/g. The R2 values were 0.614 8, 0.997 7 and 0.744 5 for the pseudo-first-order kinetic model, pseudo-second-order kinetic model and intraparticle diffusion kinetic model, respectively. The pseudo-second-order kinetic model due to its higher R2 value and closest Qe,2 (Cal), was considered to be the best choice to describe the adsorption behavior of tea polyphenols by HPAE, suggesting that the adsorption mechanism might be a physical adsorption process. The increase in pH from 4 to 8 resulted in the increase in the TPs adsorption capacity, indicating that pH had an obvious effect on the TPs adsorption capacity onto HPAE and adsorbed tea polyphenols could be desorbed by controlling pH value in acid medium. The maximum adsorption capacity of HPAE was 194.90 mg/g at the present experiment condition, and far greater than the adsorption capacity of macroporous resins studied widely in TPs industry, such as LSA-8, LSA-20, XDA-1, HPD600, NKA, AB-8, D3520, D4020, PVPP and PA, etc. In conclusion, the new adsorbent HPAE has excellent adsorption and desorption properties for tea polyphenols. Its adsorption capacity could achieve 194.90 mg/g, and adsorption of TPs could be desorbed easily by adjusting pH instead of using organic solvents. HPAE shows a great potential as an environment-friendly and high efficiency

Soil organic matter has an important role in soil fertility and agricultural productivity. Many practices have been used to increase organic carbon in soil, particularly nitrogen fertilization, zero-tillage agriculture, and the addition of large amounts of manure to soil. These practices do not sequester significant quantities of C into soil because most of the organic matter is not stable and is mineralized very fast. Many factors contribute to soil organic C stabilization, including soil clay content, rainfall, and temperature regimes. In recent years, there has been considerable interest in the use of biochar from pyrolysis of renewable biomass to sequester C and improve soil productivity. Much of the stimulus for this interest comes from research on the soils of the Amazon basin, known as Terra Preta de Indio, that contain variable quantities of organic black carbon considered to be of anthropogenic origin. Biochar can improve nutrient availability, cation exchange capacity, bulk density, and water-holding capacity, but these effects depend on the feedstock, prolysis conditions. It is important to evaluate the effects of biochar on soil fertility under different soil and climatic regimes to increase our understanding of potential interactions before widespread use of biochar in agricultural systems. Although biochar has been shown to increase soil fertility and productivity in the tropics, there is limited information about influences of biochar on transformation and chemical forms of C, N and P in soils. Therefore, an incubation experiment was conducted to study the effects of biochar application on transformation and chemical forms of C, N and P in soils with different pH. The experiment included four treatments, i.e., control without application of any chemical fertilizers and biochar, conventional fertilization with application of chemical fertilizers, biochar treatment with application of biochar but without any chemical fertilizers, and conventional fertilization + biochar treatment with application of both biochar and chemical fertilizers. The treated soils were incubated at temperature of 2035 ℃ for 12 months, and the incubated soils were characterized for different forms of C, N, and P and potential capacities of N leaching and volatilization loss. The results showed that application of biochar increased soil pH, particularly for acidic soil. Application of biochar increased significantly the accumulation of organic C, microbial biomass C and humic/fulvic acids （
H/F） of the soils, and decreased water soluble C in both acidic and neutral soils, and proportion of total organic C in easily oxidized organic carbon. Addition of biochar could promote the transformation of rapidly available N into slowly available N, and thus decrease potential of N leaching loss. However, application of biochar increased the volatilization loss of NH3 in the soils, and the effect on volatilization loss was less in the acidic soil than that in both neutral and calcareous soils. The application of biochar increased total P in the soils. The application of biochar increased proportion of water soluble P for the soils without addition of chemical fertilizers, but it decreased the proportion for the soils with addition of chemical fertilizers. Application of biochar increased proportions of bio-available P such as NaHCO3-IP and NaHCO3-OP in the soils, and decreased the proportions of NaOH-IP and residual P. Proportion of HCl-P increased when biochar was applied. It is concluded that application of biochar is benefit to increasing fixation of organic carbon and modifying availability of N and P in soils. It is recommended that biochar could be preferentially applied in acidic soil.

Groundwater is not only an important component of the hydrologic cycle but also an important drinking water source. Excessive nitrogen fertilizer and organic manure application has caused severe nitrate pollution in groundwater, which threats human health. Bioremediation of groundwater nitrate using denitrifying forms of bacteria found in groundwater is one of the effective methods for nitrogen removal. Carbon source is one of the necessities for heterotrophic denitrification, and water temperature and hydraulic retention time can also significantly influence the nitrate removal efficiency. The main objectives of this study are to: 1) investigate the performance of denitrification using different single carbon sources; 2) evaluate the effects of temperature, hydraulic retention time (HRT) and pH on denitrification performance and screen the best solid carbon source for nitrate removal. This research focused on nitrate removal using commonly available agricultural solid organic residues as the carbon sources. Particularly, corncob, rice straw, and newspaper were employed as the carbon sources for nitrate removal at different temperatures and hydraulic retention times (HRT) in the laboratory. The results indicated that when newspaper was used as carbon source, nitrate concentration was reduced from 30 to around 15 mg/L in 24 hours, and the nitrite concentration was much higher than the standard of the drinking water quality. On the other hand, use of corncob and rice straw as carbon sources could almost completely remove the nitrate within 24 hours. It was observed that small amount of nitrite was accumulated in the denitrification process when corncob and rice straw were used as the carbon sources. Denitrification was markedly affected by temperature. When rice straw and corncob were used as carbon sources, almost 100% nitrate removal was achieved within 60 hours at 20, 25 or 30 ℃, while only 50% nitrate removal could be achieved at 15 ℃. Nitrite concentration increased at first, then decreased to below 0.1 mg/L within 24, 40 and 24 hours, respectively, at 20, 25 and 30 ℃ when corncob or rice straw was used as carbon source. High nitrate removal efficiency could be achieved at the four different treatments of HRT by using corncob or rice straw as carbon source. But the time required for the near-complete removal of nitrate (stable operation) increased with the decrease of HRT. This study indicated that complete nitrate removal can be achieved readily using rice straw or corncob as carbon source by adjusting HRT or temperature in water, which can be potentially implemented in environmental engineering practices.

Malachite green (MG) is a kind of triphenylmethanedyes, which had ever been widely used around the world as fungicide, ectoparasiticide and antiseptic in the aquaculture. Current datas suggest that both MG and its metabolite leucomalachite green (LMG) may be carcinogenic and mutagenic. For those reasons MG has been banned in aquaculture industry in European Commission and U.S.A, and is also not approved in China. However, owing to low cost and marked effect, MG may still be used illegally. Additionally, the environment has been polluted because of MG’s wide use before. Much of the pollutants have been transferred to sediment. In order to guarantee the safety of aquaculture, a sensitive and selective analytical method is needed for the determination of MG in aquaculture sediment. The current researches mainly focus on the determination of MG in aquatic products, rarely in sediment. Due to the matrix complex of sediment, sample pretreatment technology is the key technology. As a new type of functional material, muti-walled nanotubes (MWCNTs) have extensive prospect in the field of analytical chemistry. Now several articles have illustrated that MWCNTs have been used in the pretreatment of heavy metals and organic pollutants. Few literatures available were involved in determination of MG in sediment. During the past research, much more expensive materials such as cation exchange columns or alumina columns were mostly applied as solid-phase extraction adsorbents to determine MG in aquatic products and water. However, with advantages of lower price and good chemical properties, MWCNTs still have not been well used in analysis of MG. Now, a method for the determination of MG in sediment was established by using MWCNTs as solid-phase extraction adsorbents combined with ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The sediment sample was extracted by acetonitrile and cleaned up on MWCNTs solid phase extraction (SPE) cartridges. The effects of experimental conditions, including SPE cartridge type, diameter and loading amounts of MWCNTs, as well as type and volume of the eluent on the extraction efficiency were investigated. Chromatographic separation was carried out on UPLC C18 column (2.1 mm×50 mm, 1.7 μm). The mass spectrometer was performed using multiple reaction monitoring (MRM) with electrospray ionization in positive mode. Under the optimized conditions, the analytes were in good linearity in the range of 0.0510.0 μg/L with the correlation coefficients (r) all above 0.995. The limits of detection were 0.03, 0.07 μg/kg for MG and LMG, and limits of quantity were 0.10, 0.20 μg/kg, respectively. Spiked at three concentration levels of 0.50, 2.00 and 5.00 μg/kg, the recovery rates of MG, LMG were 97.1%108.3% and 94.7%110.5% and the relative standard deviations (RSD) were all within 8.3%. In conclusion, MWCNTs have good performance in the enrichment and purification. The proposed method has advantages of good reproducibility, high recovery, simplicity, and low cost, and can be successfully applied for the determination of malachite green in aquaculture sediment.

Recently, intensive culture of Scophthalmus maximus has been seriously influenced by various diseases. To solve the problem, different kinds of antibiotics were incorporated in fish diets, which may produce potential drug-resistant pathogens and become a growing menace to aquatic animals as well as mankind. Therefore, it is urgent to develop new and safe additives in aquaculture. Saccharicterpenin is a bioactive compound derived from the plants of camellia seed, and its effective components are triterpenoid saponins and polysaccharides. Previous studies showed that saccharicterpenin could improve growth performance and immune function in freshwater fish. Thus, the aim of the study was examine the effects of dietary administration of saccharicterpen on growth performance, immunity and heat shock protein 70 (HSP-70) content of juvenile turbot, S. maximus (with initial body mass (55.0±0.2) g). Six isocaloric and isonitrogenous diets were formulated containing increasing levels (0, 25, 50, 75, 100, 200 mg/kg) of saccharicterpenin and named as D0, D25, D50, D75, D100, D200, respectively. Each diet was randomly assigned to triplicate groups of 30 juveniles. The experiment was conducted in plastic cylinder tanks in an indoor recirculated feeding system for 64 days. At the termination of the feeding trial, growth performance, serum and liver immunity and metabolic enzymes were determined. The remaining fishes were challenged intraperitoneally with Vibrio anguillarum. The results showed that fish fed with diets of D25, D50, and D75 showed significantly higher mass gain (MG), specific growth rate (SGR), protein efficiency ratio (PER), condition factor (CF), hepato-somatic index (HSI) and spleen-somatic index (SSI) (P<0.05), and obviously decreased feed conversion ratio (FCR) (P<0.05). However, viscero-somatic index (VSI) and survival rate (SR) revealed no noticeable difference among all treatments　(P>0.05). Fish fed with diet of D50 showed the highest serum lysozyme activity. Superoxide dismutase (SOD) activity were significant enhanced in fish fed diets D75, D100 and D200 compared with other treatments　(P<0.05). Nitric oxide content in serum was significant reduced with the increasing level of saccharicterpenin (P<0.05). HSP-70 contents in serum showed no significantly difference among all treatments　(P>0.05). Aspartate transaminase (AST) of serum in D200 was significantly higher than the other groups　(P<0.05), but alanine aminotransferase (ALT) showed no significantly difference　(P>0.05）. ALT and AST of liver significantly decreased with the increasing level of saccharicterpenin (P<0.05). There was no difference in total antioxidant capacity
(T-AOC) in liver among all groups (P>0.05). Liver malondialdehyde (MDA) content in group D25 was significantly lower than that of D100　(P<0.05), with no change among other groups　(P>0.05). The challenge study showed that fish fed with diet of D50 presented lower cumulative mortality rate than other groups　(P<0.05), and higher immune protective rate of 42.31% compared with those fed with diets of D25 (19.23%) and D75 (15.39%) after 48 hours, respectively. In conclusion, the addition of saccharicterpenin can improve the growth performance, immunity and disease resistance in S. maximus. Quadratic regression of SGR indicates that the optimal level of saccharicterpenin in diet for juvenile turbot is 74.58 mg/kg.

Squid liver meal is an attractant used in aquafeed widely. However, the fatty acid rancidification and high content of cadmium in squid liver meal may affect the animal health. Soluble fish protein concentrates are produced from the waste water of fish meal production, and have many attractive materials. Chinese soft-shelled turtle, Pelodiscus sinensis, is an important economic animal in China, and fishmeal is one main protein resource in turtle feed. Squid liver meal and soluble fish protein concentrates maybe the alternatives to fishmeal in turtle feed because of their high protein content and low price. No study has reported the effects of squid liver meal on health of turtle and the utilization of soluble fish protein concentrates in turtle feed. Therefore, this experiment has the aim of exploring the effects of the supplementation of squid liver meal and soluble fish protein concentrates on the growth and liver function of P. sinensis.
Three experimental groups were designed, control group (white fishmeal group,WFM), squid liver meal group (SLM) and soluble fish protein concentrates group (SFP). Nutritional compositions of three diets were similar. For the diets of SLM and SFP, 5% white fishmeal in the control diet was replaced with squid liver meal and soluble fish protein concentrates respectively. The feeding experiment lasted for eight weeks. Chinese turtles with initial average body mass of 6.8 g were fed twice a day at (30±1) ℃. Feed intake, growth and feed utilization were determined, and the activities of glutamic-oxalacetic transminase (GOT) and glutamic-pyruvic transminase (GPT) for liver and serum and the contents of serum triglyceride and cholesterol were analyzed. No significant difference was found in feeding rate, specific growth rate, viscera-somatic index, hepatosomatic index among three experimental groups. Feed coefficient of SFP was significantly higher than WFM and SLM, and there was no significant difference in feed coefficient between WFM and SLM. Digestibilities of dry mass and lipid of SLM were both significantly higher than those of WFM. No difference in protein digestibility was found between SLM and WFM. Digestibilities of dry mass, protein and lipid of SFP were all significantly higher than those of WFM. Hepatic GOT activity of SLM was significantly lower than that of WFM, and serum GPT activity of SLM was significantly higher than that of WFM. There were no significant differences in GPT and GOT activities between SFP and WFM for both liver and serum tissues. No significant difference was found in the contents of serum triglyceride and cholesterol among three experimental groups. These results indicated that 5% dietary supplementation of squid liver meal may cause slight hepatic damage to turtle, but had no evident effect on growth; and 5% dietary supplementation of soluble fish protein concentrates could reduce feed utilization, but had no adverse effects on liver function of turtle. Since the price of squid liver meal is lower than that of soluble fish protein concentrates, squid liver meal may be more suitable than soluble fish protein concentrates to replace fishmeal in turtle feed, and the recommended supplementation level should be less than 5%.