具有多种胞外酶的对虾肠道黏附菌的分离和鉴定

用对虾饲料培养基从健康凡纳滨对虾肠道分离出500株黏附细菌,以产淀粉酶、脂肪酶和蛋白酶能力为指标,筛选出产该3种消化酶的细菌90株,占总菌株的18%.对其中生长较快的69株进行16S rDNA 基因测序,确定其分类地位.结果显示,69株菌分别属于不动杆菌属(Acinetobacter)、芽孢杆菌属(Bacillus)、葡萄球菌属(Staphylococcus)、假交替单胞菌属(Pseudoalteromonas)、气单胞菌属(Aeromonas)、嗜盐单胞菌属(Halomonas)、利斯顿氏菌属(Listonella)、莫拉氏菌属(Moraxella)等,其中数量最多是芽胞杆菌属,占鉴定细菌总数的53.62%,数量最少是气单胞菌属和嗜盐单胞菌属,均占鉴定细菌总数的2.90%.表明对虾肠道黏附菌群中具有较多能分泌多种消化酶的细菌,可进一步开发为促进对虾消化功能的益生菌     

Effect of carbon and nitrogen ratio control on Artemia growth,water quality,biofloc microbial diversity under high salinity and zero-water exchange culture condition

Biofloc technology has been applied successfully in the intensive aquaculture of several fish and shrimp species.The growth of heterotrophic microorganisms can be stimulated through adding extra carbon,which reduces the nitrogen level in the water and provides microbial protein to the animals.However,most of the studies and practical applications have been conducted in freshwater and marine environment.This paper focused on brine shrimp Artemia that lives in high salinity environment together with other halophilic or halotolerant microorganisms.The effect of carbon supplementation on Artemia growth,water quality,and microbial diversity of bioflocs was studied in the closed culture condition without any water exchange.The salinity of the culture medium was 100.A 24-d culture trial was conducted through supplementing sucrose at carbon/nitrogen(C/N)ratio of 5,15,and 30(Su5,Sul5,and Su30),respectively.The culture without adding sucrose was used as a control.Artemia was fed formulated feed at a feeding ration of 60% recommended feeding level.The results showed that sucrose supplementation at higher C/N ratio(15 and 30)significantly improved the Artemia survival,growth and water quality(P0.05).Addition of sucrose at C/N ratio of 15 and 30 significantly increased biofloc volume(BFV)(P0.05).The Illumina MiSeq sequencing analysis showed that supplementing carbon at C/N ratio of 15 had a better total bacterial diversity and richness,and shaped the microbial composition at genera level.This study should provide information for studying the mechanism of biofloc technology and its application in high salinity culture conditions.     

Effects of culture conditions on growth and docosahexaenoic acid production from Schizochytrium limacinum

The effects of temperature, initial pH, salinity of culture medium, and carbon and nitrogen sources on growth and docosahexaenoic acid （C22：6 n-3, DHA） production from Schizochytrium limacinum OUC88 were investigated in the present study. The results revealed that the optimal temperature, initial pH and salinity level of the medium for DHA production were 23 ℃, 7.0 and 18, respectively. Glucose was proved the best carbon source for the growth and DHA production from S. limacinum. Among the nitrogen sources tested, soybean cake hydrolysate, a cheap by-product, was found to be effective for the accumulation of DHA in S. limacinum cells. In addition, increasing the concentration of carbon sources in the medium caused a significant increase in cell biomass; however, accumulation of DHA in cells was mainly stimulated by the ratio of C/N in the medium. Under the optimal culture conditions, the maximum DHA yield achieved in flasks was 4.08 g L^-1 after 5 d of cultivation     

Nitrogen Budget in Recirculating Aquaculture and Water Exchange Systems for Culturing <Emphasis Type="Italic">Litopenaeus vannamei</Emphasis>

In order to investigate the culture characteristics of two indoor intensive Litopenaeus vannamei farming modes, recirculating aquaculture system (RAS) and water exchange system (WES), this study was carried out to analyze the water quality and nitrogen budget including various forms of nitrogen, microorganism and chlorophyll-a. Nitrogen budget was calculated based on feed input, shrimp harvest, water quality and renewal rate, and collection of bottom mud. Input nitrogen retained in shrimp was 23.58% and 19.10% respectively for WES and RAS, and most of nitrogen waste retained in water and bottom mud. In addition, most of nitrogen in the water of WES was TAN (21.32%) and nitrite (15.30%), while in RAS was nitrate (25.97%), which means that more than 76% of ammonia and nitrite were removed. The effect of microalgae in RAS and WES was negligible. However, bacteria played a great role in the culture system considering the highest cultivable cultivable bacterial populations in RAS and WES were 1.03×10¹⁰ cfu mL^?1 and 2.92×10⁹ cfu mL^?1, respectively. Meanwhile the proportion of bacteria in nitrogen budget was 29.61% and 24.61% in RAS and WES, respectively. RAS and WES could realize shrimp high stocking culture with water consuming rate of 1.25 m³ per kg shrimp and 3.89 m³ per kg shrimp, and power consuming rates of 3.60 kw h per kg shrimp and 2.51 kw h per kg shrimp, respectively. This study revealed the aquatic environment and nitrogen budget of intensive shrimp farming in detail, which provided the scientific basis for improving the industrial shrimp farming.     

海洋短小芽孢杆菌ZH1-6菌株的鉴定及生物学特性

从新鲜近江牡蛎中分离到一株拮抗细菌,命名为ZH1-6,该菌对金黄色葡萄球菌、乙型副伤寒沙门氏菌及大肠杆菌等多种病原菌有较强的抑制作用。通过形态学观察、常规生理生化指标测定、16SrDNA序列测定和同源性分析,鉴定该菌株为短小芽孢杆菌。研究表明:ZH1-6能利用多种碳、氮源,其生长温度范围为10-50℃,生长pH范围为pH3-10,最适初始pH为6.0。     

A Study on the contribution of different food sources to shrimp growth in an intensive <Emphasis Type="Italic">Fenneropenaeus chinensis</Emphasis> pond

Stable isotope methods can be used to determine the food sources and prey items of aquatic organisms accurately and reliably. This study examined the relative contribution of artificial foods (the formulated feed and Artemia) and natural foods to shrimp growth in an intensive Fenneropenaeus chinensis pond by using carbon and nitrogen stable isotopes. The results showed that the nutrition utilization efficiency of the harvested shrimp was low, only 33.18% of feed nitrogen and 21.73% of feed carbon being converted to shrimp flesh. Our stable isotope results showed that the shrimp obtained nutrition for maximum growth from artificial foods, whose contribution was 93.5%, with the remaining attributed to the natural foods. However, there was 0.94 t harvested shrimp derived from natural foods (the rest of 13.56 t harvested shrimp derived from artificial foods) in 1ha intensive pond with a shrimp production of 14.50 t ha⁻¹. Therefore, unit area shrimp production can be increased by increasing the contribution proportion of natural foods in intensive shrimp farming.     

Production of Pacific White Shrimp Polycultured with Swimming Crab at Different Densities,and Nutrient Budget in the Enclosure System

This study was to evaluate the effect of stocking densities on the nitrogen and phosphorus budgets for a polyculture of the crab Portunus trituberculatus and the Pacific white shrimp Litopenaeus vannamei.The shrimps(initial weight,0.012 g)were cultured at a density of 45 shrimp m^-2,and juvenile crabs(initial weight,0.024 g)were cultured at five densities of 0,3,6,9,and 12 crabs m^-2.The treatments were grouped as C0S45,C3S45,C6S45,C9S45,and C12S45,respectively.Water quality parameters,growth of shrimp and crabs,and nitrogen and phosphorus budget were measured.The results indicated that the shrimp performances in polyculture treatments C3S45 and C6S45 were superior to those in the crab-free treatment(C0S45).The crab in treatments C3S45 and C6S45 exhibited a significantly higher final mean weight and carapace width/length than those in treatments C9S45 and C12S45.The final size and survival of crabs had a negative correlation with the increasing crab stocking density.The contents of total phos-phorus and total nitrogen and the comprehensive contamination index values were higher in the C9S45 and C12S45 treatments than in the other treatments.The conversion ratios of nitrogen for crab and shrimp growth in treatment C3S45 were significantly higher than those in the crab-free treatment.These findings indicate that polyculturing shrimp with crabs at suitable densities can improve productivity,profitability,nutrient utilization,and the environmental quality.From the 60-day treatments,the optimal culture densi-ties were 3-6 crabs m^-2 and 45 shrimps m^-2.     

A Study on the Contribution of Different Food Sources to Shrimp Growth in an Intensive Fenneropenaeus chinensis Pond

Stable isotope methods can be used to determine the food sources and prey items of aquatic organisms accurately andreliably.This study examined the relative contribution of artificial foods (the formulated feed and Artemia) and natural foods toshrimp growth in an intensive Fenneropenaeus chinensis pond by using carbon and nitrogen stable isotopes.The results showed that the nutrition utilization efficiency of the harvested shrimp was low,only 33.18% of feed nitrogen and 21.73% of feed carbon beingconverted to shrimp flesh.Our stable isotope results showed that the shrimp obtained nutrition for maximum growth from artificialfoods,whose contribution was 93.5%,with the remaining attributed to the natural foods.However,there was 0.94 t harvested shrimpderived from natural foods (the rest of 13.56t harvested shrimp derived from artificial foods) in lha intensive pond with a shrimpproduction of 14.50 tha-1.Therefore,unit area shrimp production can be increased by increasing the contribution proportion of natu-ral foods in intensive shrimp farming.     

Optimization of culture conditions and medium composition for the marine algicidal bacterium Alteromonas sp. DH46 by uniform design

Harmful algal blooms (HABs) have led to extensive ecological and environmental issues and huge economic losses. Various HAB control techniques have been developed, and biological methods have been paid more attention. Algicidal bacteria is a general designation for bacteria which inhibit algal growth in a direct or indirect manner, and kill or damage the algal cells. A metabolite which is strongly toxic to the dinoflagellate Alexandrium tamarense was produced by strain DH46 of the alga-lysing bacterium Alteromonas sp. The culture conditions were optimized using a single-factor test method. Factors including carbon source, nitrogen source, temperature, initial pH value, rotational speed and salinity were studied. The results showed that the cultivation of the bacteria at 28°C and 180 r min^?1 with initial pH 7 and 30 salt contcentration favored both the cell growth and the lysing effect of strain DH46. The optimal medium composition for strain DH46 was determined by means of uniform design experimentation, and the most important components influencing the cell density were tryptone, yeast extract, soluble starch, NaNO₃ and MgSO₄. When the following culture medium was used (tryptone 14.0g, yeast extract 1.63g, soluble starch 5.0 g, NaNO₃ 1.6 g, MgSO₄ 2.3 g in 1L), the largest bacterial dry weight (7.36 g L^?1) was obtained, which was an enhancement of 107% compared to the initial medium; and the algal lysis rate was as high as 98.4% which increased nearly 10% after optimization.     

Carbon cycle in shrimp polyculture mesocosm

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Ecological effects of co-culturing the sea cucumber Apostichopus japonicus with the Chinese white shrimp Fenneropenaeus chinensis in an earthen pond

Using net enclosures in an earthen pond, we established three culture treatments with the sea cucumber Apostichopus japonicus and the Chinese white shrimp Fenneropenaeus chinensis : monoculture of sea cucumbers(C), monoculture of shrimp(S), and co-culture of the two species(CS). We measured levels of suspended particulate matter in the water column; total organic matter, total organic carbon, total nitrogen, and carbon/nitrogen ratios in both settling particles and the sediment; and chlorophyll a levels in the sediment. We then compared these variables between the three treatments. We also examined growth, survival, and yield of the two species in the different treatments. From June to September, the mean monthly suspended particulate matter sedimentation rates in the CS and S treatments were significantly( P 0.05) greater than those in the C treatment. From August to November, the mean monthly total organic matter, total organic carbon, total nitrogen, and chlorophyll a contents in the sediment in the CS and S treatments were significantly( P 0.05) greater than those in the C treatment. Final wet weight, specific growth rate, survival rate, and total yield of sea cucumbers in co-culture were all significantly greater than those of sea cucumbers in monoculture. There were no significant differences among any of these variables for shrimp reared in the two systems. The bioturbation of the sediment and fecal production of the shrimp likely supplied natural food for the sea cucumbers. Co-culture of the two species is a viable option for increasing yield per unit area, maximizing use of the water body, and diversifying crop production.     

一株促藻生长细菌培养条件的优化

研究分离自对虾养殖池的一株促藻生长细菌的最佳培养条件,通过单因子实验得到该菌株的最佳碳源为葡萄糖,最佳氮源为酵母膏,初始pH值为7.5,最佳氯化钠质量浓度5 g/L;在其他培养条件不变的情况下,通过对酵母膏、葡萄糖、初始pH值做三因子三水平的正交实验,得到培养基最佳组合为:酵母膏10 g/L,葡萄糖1 g/L,初始pH值7.5。     

Isolation and characterization of a marine bacterium producing protease from Chukchi Sea, Arctic

A Gram negative bacterium Ar/W/b/75°25＇N/1 producing extracellular alkaline protease was isolated from surface water of latitude 75°25＇N, and longitude 162°25＇W in Chukchi sea, Arctic. The strain can grow at the temperature range from 7℃ to 30℃, and grow better at 30(℃. It can not grow at 40℃. Keeping certain salinity concentration in medium is necessary for cell growth. It grows well in medium containing salinity concentration from 0. 5 % to 10 % sodium chloride. Glucose, sucrose and soluble starch can be utilized by the strain, among which glucose is the optimal carbon source. Peptone is the optimal organic nitrogen source for cell growth and protease producing, and ammonium nitrate is the optimal inorganic nitrogen source.About 75.7% of total protease of the strain are extracellular enzyme. Optimal temperature for proteolytic activity is at 40℃. Protease of the strain keeps stable below 40℃, and shows high proteolytic activity within the pH range from 7 to 11.     

Effect of Zinc Oxide Nanoparticles on Denitrification and Denitrifying Bacteria Communities in Typical Estuarine Sediments

For revealing the effects of increasing of zinc oxide nanoparticles(ZnO NPs) on denitrification and denitrifying bacteria communities in estuarine sediments, the surface sediments of two typical estuaries(the Yangtze River Estuary and the Yellow River Estuary) were added with medium concentration(170 mg L~(-1)) and high concentration(1700 mg L~(-1)) of ZnO NPs for anaerobic culture in laboratory. The concentration of NO_3~-and NO_2~-, the reductase activity and denitrification rate were measured by physicochemical analysis, nir S gene abundance and denitrifying bacteria communities by molecular biological methods. The results showed that ZnO NPs inhibited NO_3~-, NO_2~-reduction process and NO_3~-, NO_2~-reductase activity, and a stronger inhibition effect resulting from the higher ZnO NPs concentration. ZnO NPs decreased nirS gene abundance and community diversity of denitrifying bacteria. In addition, the inhibition degree of ZnO NPs on the denitrification process of sediments in different estuaries was different. These results were of great significance for evaluating the potential ecological toxicity and risks of nanomaterials in estuaries.     

Pyrosequencing of nirS gene revealed spatial variation of denitrifying bacterial assemblages in response to wetland desertification at Tibet plateau

Amplicon sequencing of functional genes is a powerful technique to explore the diversity and abundance of microbes involved in biogeochemical processes. One such key process, denitrification, is of particular importance because it can transform nitrate(NO3-) to N2 gas that is released to the atmosphere. In nitrogen limited alpine wetlands, assessing bacterial denitrification under the stress of wetland desertification is fundamental to understand nutrients, especially nitrogen cycling in alpine wetlands, and thus imperative for the maintenance of healthy alpine wetland ecosystems. We applied amplicon sequencing of the nirS gene to analyze the response of denitrifying bacterial community to alpine wetland desertification in Zoige, China. Raw reads were processed for quality, translated with frameshift correction, and a total of 95,316 nirS gene sequences were used for rarefaction analysis, and 1011 OTUs were detected and used in downstream analysis. Compared to the pristine swamp soil, edaphic parameters including water content, organic carbon, total nitrogen, total phosphorous, available nitrogen, available phosphorous and potential denitrification rate were significantly decreased in the moderately degraded meadow soil and in severely degraded sandy soil. Diversity of the soil nirS-type denitrifying bacteria communities increased along the Zoige wetland desertification, and Proteobacteria and Chloroflexi were the dominant denitrifying bacterial species. Genus Cupriavidus(formerly Wautersia), Azoarcus, Azospira, Thiothrix, and Rhizobiales were significantly(P0.05) depleted along the wetland desertification succession. Soil available phosphorous was the key determinant of the composition of the nirS gene containing denitrifying bacterial communities. The proportion of depleted taxa increased along the desertification of the Zoige wetland, suggesting that wetland desertification created specific physicochemical conditions that decreased the microhabitats for bacterial denitrifiers and the denitrification related genetic diversity.     

一种户外开放式微藻培养体系优化

【目的】建立可用于大规模培养的户外开放式微藻培养体系。【方法】设计一种微藻平面开放浅层培养(Flat,open and shallow,FOS,简称浮法)体系,以小球藻(Chlorella sorokiniana)为模式藻种,探讨温度、光照、pH值和培养基营养成分等因素在该培养体系中对小球藻生长的影响,开展900 L体系的户外培养试验。【结果】浮法体系主要由塑料袋和垫板组成,在该培养体系中的小球藻在20~40℃范围内均可生长,最适生长温度为30~35℃。小球藻生长有明显的照度依赖性,无光时几不生长,随着照度升高,生长速度加快,在较高的照度下表现有光饱和现象。以TAP为基础培养基,在碳源或氮源缺乏时小球藻几不生长。pH 7.5左右有利于小球藻生长。900 L体系的户外培养试验的生物量(干物质得率)为0.15 g/(L·d)。【结论】在此新型户外平面开放浅层微藻培养体系中,温度、照度、pH值及营养成分等对小球藻的生长均有不同程度的影响。该体系有成本低、操作简便、容易控制、自然资源利用率高等特点,有大面积推广潜力。     

Alkaloids from an algicolous strain of <Emphasis Type="Italic">Talaromyces</Emphasis> sp.

Compounds isolated and identified in a culture of the alga-endophytic fungus Talaromyces sp. cf-16 included two naturally occurring alkaloids, 2-[(S)-hydroxy(phenyl)methyl]-3-methylquinazolin-4(3H)-one (1a) and 2-[(R)-hydroxy(phenyl)methyl]-3-methylquinazolin-4(3H)-one (1b), that were identified for the first time. In addition, seven known compounds (2–8) were obtained from the culture. Following chiral column chromatography, compounds 1a and 1b were identified as enantiomers by spectroscopic analyses and quantum chemical calculations. Bioassay results showed that 5 was more toxic to brine shrimp than the other compounds, and that 3–6 could inhibit Staphylococcus aureus.     

Ecological effects of co-culturing the sea cucumber <Emphasis Type="Italic">Apostichopus japonicus</Emphasis> with the Chinese white shrimp <Emphasis Type="Italic">Fenneropenaeus chinensis</Emphasis> in an earthen pond

Using net enclosures in an earthen pond, we established three culture treatments with the sea cucumber Apostichopus japonicus and the Chinese white shrimp Fenneropenaeus chinensis: monoculture of sea cucumbers (C), monoculture of shrimp (S), and co-culture of the two species (CS). We measured levels of suspended particulate matter in the water column; total organic matter, total organic carbon, total nitrogen, and carbon/nitrogen ratios in both settling particles and the sediment; and chlorophyll a levels in the sediment. We then compared these variables between the three treatments. We also examined growth, survival, and yield of the two species in the different treatments. From June to September, the mean monthly suspended particulate matter sedimentation rates in the CS and S treatments were significantly (P<0.05) greater than those in the C treatment. From August to November, the mean monthly total organic matter, total organic carbon, total nitrogen, and chlorophyll a contents in the sediment in the CS and S treatments were significantly ( P <0.05) greater than those in the C treatment. Final wet weight, specific growth rate, survival rate, and total yield of sea cucumbers in co-culture were all significantly greater than those of sea cucumbers in monoculture. There were no significant differences among any of these variables for shrimp reared in the two systems. The bioturbation of the sediment and fecal production of the shrimp likely supplied natural food for the sea cucumbers. Co-culture of the two species is a viable option for increasing yield per unit area, maximizing use of the water body, and diversifying crop production.     

Production of polyhydroxybutyrate by the marine photosynthetic bacterium Rhodovulum sulfidophilum P5

The effects of different NaCl concentrations, nitrogen sources, carbon sources, and carbon to nitrogen molar ratios on biomass accumulation and polyhydroxybutyrate (PHB) production were studied in batch cultures of the marine photosynthetic bacterium Rhodovulum sulfidophilum P5 under aerobic-dark conditions. The results show that the accumulation of PHB in strain P5 is a growth-associated process. Strain P5 had maximum biomass and PHB accumulation at 2%-3% NaCl, suggesting that the bacterium can maintain growth and potentially produce PHB at natural seawater salinity. In the nitrogen source test, the maximum biomass accumulation (8.10±0.09 g/L) and PHB production (1.11±0.13 g/L and 14.62%±2.25% of the cell dry weight) were observed when peptone and ammonium chloride were used as the sole nitrogen source. NH 4 + -N was better for PHB production than other nitrogen sources. In the carbon source test, the maximum biomass concentration (7.65±0.05 g/L) was obtained with malic acid as the sole carbon source, whereas the maximum yield of PHB (5.03±0.18 g/L and 66.93%±1.69% of the cell dry weight) was obtained with sodium pyruvate as the sole carbon source. In the carbon to nitrogen ratios test, sodium pyruvate and ammonium chloride were selected as the carbon and nitrogen sources, respectively. The best carbon to nitrogen molar ratio for biomass accumulation (8.77±0.58 g/L) and PHB production (6.07±0.25 g/L and 69.25%±2.05% of the cell dry weight) was 25. The results provide valuable data on the production of PHB by R. sulfidophilum P5 and further studies are on-going for best cell growth and PHB yield.     

Summary of studies on closed-polyculture of penaeid shrimp with fishes and moluscans

ＩＮＴＲＯＤＵＣＴＩＯＮＴｈｅｗｏｒｌｄ’ｓｓｈｒｉｍｐｃｕｌｔｕｒｅｉｎｄｕｓｔｒｙｈａｓｂｅｅｎｓｕｆｆｅｒｉｎｇｆｒｏｍｖｉｒａｌｅｐｉｄｅｍｉｃｄｉｓｅａｓｅｓａｎｄｆｅｌｌｉｎｔｏａｄｉｆ ｆｉｃｕｌｔｐｏｓｉｔｉｏｎｆｏｒｙｅａｒｓ.Ｉｔｉｓｇｅｎｅｒａｌｌｙａｃｃｅｐｔｅｄｔｏｄａｙｔｈａｔｔｈｅｄｉｓｅａｓｅｓｗｅｒｅｃａｕｓｅｄｍａｉｎｌｙｂｙｗａｔｅｒｐｏｌｌｕ ｔｉｏｎｃｒｅａｔｅｄｂｙｓｈｒｉｍｐｃｕｌｔｕｒｅｉｔｓｅｌｆ,ｂｏｔｈｉｎｓｈｒｉｍｐｐｏｎｄｓａｎｄｉｎｃｏａｓ…