枯草芽孢杆菌培养基配方优化的研究

枯草芽孢杆菌培养基的组分是由葡萄糖、蛋白胨、酵母膏、磷酸二氢钾、碳酸钙等5种原料构成，采用L1（4^5）正交表，将5种原料作为枯草芽孢杆菌培养有影响的因素，通过试验数理统计的直观和理论分析，对其配方进行优化：试验结果表明，当培养基的配方为无水葡萄糖3．50g／L、蛋白胨0．83g／L、酵母膏0．50g／L、磷酸二氢钾0．35g／L和碳酸钙0．25g／L，温度（34±2）℃时，培养16h即可达到生长峰值。由于枯草芽孢杆菌光学密度（optical density，OD）与活菌数量的关系：Y=0．5182x^2＋1．4462x＋1．9714（r=0．996343），接种的枯草芽孢杆菌由1．9714×10^8cfu／mL提高到3．3124×10^8cfu／mL。优化的枯草芽孢杆菌培养基条件能有效促进枯草芽孢杆菌生长。     

一株水产用益生枯草芽孢杆菌液体发酵初步研究

为解决工业化生产中枯草芽孢杆菌(Bacillus subtilis)活菌数量低、成本高的问题,通过正交试验、单因素试验和100 L发酵罐中试,确定了枯草芽孢杆菌(Bacillus subtilis)HJ02工业生产用的优化液体培养基和发酵工艺条件。最佳培养基配方为:葡萄糖10 g/L、玉米粉8 g/L、豆粕粉25 g/L、硫酸锰30.8 mg/L、硫酸镁5 g/L、磷酸二氢钠0.52 g/L、磷酸氢二钠2.33 g/L;最优发酵工艺条件为:培养温度30℃,初始pH 7.0;最佳接种菌龄是18 h。优化后培养基成本降低了50%以上,细菌浓度达到了7.19×109个/mL。     

一株海洋红酵母的鉴定及其培养基的优化

通过菌落形态、培养特征和生理生化特征的测定,初步确定海洋红酵母HN-3为红酵母属的胶红酵母。比较不同的碳源、氮源、酵母膏质量分数和海水盐度对该酵母生长的影响,优化其培养基。试验结果表明,最佳培养基为2%葡萄糖、1.5%蛋白胨、1%酵母膏,盐度30。在该培养条件下,酵母菌HN-3的产菌量达到4.80×107cfu/mL,比初始发酵条件下的产菌量(4.20×107cfu/mL)提高了14.3%。     

海洋红酵母RH1菌株发酵培养条件的研究

该研究比较了不同碳源、氮源、无机盐对海洋红酵母菌（Rhodotorula sp.）RH1菌株发酵产量的影响,通过葡萄糖、蛋白胨、酵母膏和硫酸镁（MgSO4）4因素3水平的正交试验,确定了RH1菌株的最优培养基为蛋白胨10 g.L-1,酵母膏15 g.L-1,葡萄糖20 g.L-1,MgSO40.25 g.L-1,磷酸二氢钾（KH2PO4）0.25 g.L-1,氯化钠（NaCl）10 g.L-1。结果显示,该菌株最佳摇瓶发酵条件为接种量10%,初始pH 5.0,摇瓶装液量80 mL/500 mL三角瓶,培养温度28℃,经48 h培养,菌量可达10.46×108cfu.mL-1,比优化条件前提高23.9%。还进行了RH1菌株25 L发酵罐扩大培养试验,在接种量为8%、初始pH 5.0、搅拌速率350～400 r.min-1、通气量9 L.min-1、温度28℃的条件下,经28 h的培养,菌量可达33.6×108cfu.mL-1。预计通过连续补料的方式进行培养,有望进一步提高菌量。     

来自鳗鲡的二株益生菌适宜培养基的研究

用正交试验法,通过研究培养基基础成分、主要添加成分和无机盐对养殖鳗鲡(Anguilla japonica)肠道益生菌枯草芽孢杆菌(A40209CDC4)和少动鞘氨醇单胞菌(A31009NA)的生长影响,确定了优化的培养基。菌株A31009NA的优化配方为:蛋白胨1.0%,牛肉膏0.1%,食盐0.5%,酵母膏0.5%,葡萄糖1.0%,硫酸铵0.05%,MgSO4.7H2O 0.1 g/L,CuSO4.5 H2O 0.15 mg/L,MnSO4.H2O 1.5 mg/L,CoCl.H2O 0.15 mg/L;A40209CDC4优化培养基配方为:蛋白胨1.0%,牛肉膏0.1%,食盐1.0%,酵母膏0.5%,硫酸铵0.05%,MgSO4.7 H2O 0.1 g/L,MnSO4.H2O 1.5 mg/L,CoCl.H2O 0.15 mg/L,CaCl20.02 g/L。优化培养基与普通营养肉汤(蛋白胨1.0%,牛肉膏0.3%,食盐0.5%)培养对照试验结果表明,优化培养基有效地提高了细菌产量。     

一种培养光合细菌的方法

<正>目前市场上有很多的光合细菌产品,尤其是淘宝网上都能便捷地购买到高浓度的光合细菌。但这些光合细菌产品是浮游态生长的,并且这些培养基的配方均是商业机密。而公开的光合细菌的培养液中需要添加多种成分,尤其是营养物质丰富的酵母膏或牛肉膏等,配置过程相对繁琐。本文介绍一种便捷的、原料来源丰富的光合细菌培养方法,其原料是渔用配合饲料,并且可生长出附着态的光合细菌。养殖户可自行在养殖过程中培养光合细菌并应用到养殖水体中。     

富集光合细菌培养基的优化研究

采用正交实验设计,对一种富集光合细菌的培养基进行优化,试验结果表明酵母膏和NaHCO3在培养基中的浓度对富集红螺菌科的光合细菌有显著的影响。若以达到相同生长量(OD660值)做指标,应用优选出来的培养基进行富集比原培养基要提前5～6d。     

一株刺参益生菌的发酵配方及其发酵条件优化

以健康刺参肠道的益生茵——枯草芽孢杆菌(Bacillus subtilis)为研究对象,以豆粕、葡萄糖和氯化钠为基础发酵培养基,通过单因素和正交试验,确定该菌株适用于工业生产的最佳发酵培养基配方;通过单因素试验,确定最佳发酵条件;同时在生物反应器中进行扩大培养试验,得到其最佳种龄和发酵时间.试验确立该茵株的最佳培养基配方为:氯化钠10g/L,葡萄糖2.5g/L,豆粕20g/L;最佳发酵条件为:培养温度28℃,初始pH 7.5,转速170r/min,菌种接种量5％.以最佳发酵条件和培养基,在10L生物反应器中进行扩大培养试验,得到最佳种龄为24h、最佳发酵时间为32h,菌体细胞浓度最高达到8.5×109cfu/mL,芽孢生成率达到91％.研究结果为刺参微生态制剂的工业化生产提供基础数据,为海水养殖专用微生态制剂的开发提供参考.     

海带表面降油细菌的分离鉴定及降油性能研究

用柴油为唯一碳源的培养基选择分离海带表面具有降解柴油性能的细菌;对分离菌株进行了形态学、生理生化特性以及16SrDNA序列分析并测定其生长特性;测定了接菌不同浓度(7×107 cfu/mL、7×108cfu/mL、7×109 cfu/mL)分离菌对柴油的降解率;另外,测定了加入不同浓度葡萄糖(0.5g/L、1g/L、2g/L、4g/L、6g/L、8g/L、10g/L)作用7d后分离菌对柴油的降解率。结果显示:从海带表面分离到2株降油细菌,编号为:HD-4和HD-6。HD-4菌株菌落形态圆形、直径1.5mm、黄色、不透明、边缘整齐,HD-6菌株菌落圆形,直径1.0mm,浅黄色、透明、边缘整齐。两株菌均为革兰氏阴性短杆菌。生理生化特征和16SrDNA序列分析确定HD-4菌株为假交替单胞菌(Pseudoalteromonas sp.),其16SrDNA同源性为99%;HD-6菌株为交替单胞菌(Alteromonas_sp.),其16SrDNA同源性为98%。2株菌最适生长温度均为15℃。HD-4和HD-6最适生长pH分别为9和8。适宜NaCl浓度分别为2%和4%;在25℃振荡培养(150r/min)7d,接菌量为7×107 cfu/mL、7×108 cfu/mL、7×109 cfu/mL时,HD-4对柴油的初始降解率分别为8.0%,22.1%和27.6%;HD-6初始降解率分别为23.7%、38.8%和43.2%。加入葡萄糖后2株菌的降油率均有所增高,加入4g/L葡萄糖时达最高值,3个接种浓度下HD-4菌株分别为85.4%、82.3%和80.4%,HD-6菌株分别为86.8%、93.7%和89.3%。当接菌量为7×107 cfu/mL,HD-4在葡萄糖含量为4g/L时作用7d,降油率可达到最大值86.72%,HD-6菌株在葡萄糖浓度6g/L达到最大值67.64%。葡萄糖浓度超过4g/L和6g/L时HD-4和HD-6菌株的降油性能有所下降。     

渔源解淀粉芽孢杆菌CQN-2菌株培养基及发酵条件优化

CQN-2是一株具有抗病、促生长等多种功能的肠道内源性解淀粉芽孢杆菌。为研究其最佳培养条件,从而为规模化生产发酵提供依据,本文以菌体生物量为指标,采用单因素试验和正交试验对CQN-2菌株的最适培养基及发酵条件进行优化。结果表明,CQN-2菌株的最适碳源为可溶性淀粉,最适氮源为细菌学蛋白胨,经优化后的最佳培养基配方为可溶性淀粉20 g/L、细菌学蛋白胨30 g/L、糖蜜50 g/L、KH_2PO_4 0.5 g/L、MgSO_4 0.5 g/L、NaCl 0.3 g/L、Mn~(2+) 5 mmol/L、Al~(3+) 1 mmol/L;最佳发酵培养条件为初始pH值7.0,接种量为3%、装液接种量25 mL/250 mL;使用优化后的培养基将CQN-2接种于5 L发酵罐进行高密度发酵培养36 h后的活菌数为3.03×10~(12) CFU/mL。     

Association between the interannual variation in the oceanic environment and catch rates of bigeye tuna (Thunnus obesus) in the Atlantic Ocean

The environmental processes associated with variability in the catch rates of bigeye tuna in the Atlantic Ocean are largely unexplored. This study used generalized additive models (GAMs) fitted to Taiwanese longline fishery data from 1990 to 2009 and investigated the association between environmental variables and catch rates to identify the processes influencing bigeye tuna distribution in the Atlantic Ocean. The present findings reveal that the year (temporal factor), latitude and longitude (spatial factors), and major regular longline target species of albacore catches are significant for the standardization of bigeye tuna catch rates in the Atlantic Ocean. The standardized catch rates and distribution of bigeye tuna were found to be related to environmental and climatic variation. The model selection processes showed that the selected GAMs explained 70% of the cumulative deviance in the entire Atlantic Ocean. Regarding environmental factors, the depth of the 20 degree isotherm (D20) substantially contributed to the explained deviance; other important factors were sea surface temperature (SST) and sea surface height deviation (SSHD). The potential fishing grounds were observed with SSTs of 22–28°C, a D20 shallower than 150 m and negative SSHDs in the Atlantic Ocean. The higher predicted catch rates were increased in the positive northern tropical Atlantic and negative North Atlantic Oscillation events with a higher SST and shallow D20, suggesting that climatic oscillations affect the population abundance and distribution of bigeye tuna.     

Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China

Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae(GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the ?rst time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in ?ve provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding36.9%(62/168), 37.5%(63/168) and 19.6%(33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia(95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010,but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during2007–2011, III onlyoccurred in 2012. 14 different PFGE types, including 4 new types(N, O,P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9%(141/168) of the total GBS strains. Ia corresponded to 11 PFGE types(A–H, N–P), in which type D predominated(51%). Ib represented 3 genotypes(I, J and Q) and III harbored only 2genotypes(N and F). Type N and Fsynchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years.Although Iastillpredominated, new rareserotypeIII alreadyoccurred in China.     

Growth performance,digestive enzyme activity and immune response of Japanese sea bass,Lateolabrax japonicus fed with fructooligosaccharide

In this experiment, a feeding trial was performed to determine the effects of fructooligosaccharide (FOS) on growth performance, digestive enzyme activity and immune response of Japanese sea bass, Lateolabrax japonicus juveniles (initial weight 38.3 ± 0.5 g), and the fish were examined following feeding with six levels of FOS (0, 0.5, 1, 2, 4 and 6 g/kg) for 28 days. Significant enhancement of weight gain (WG) and specific growth rate (SGR) was found in fish fed 1 g/kg FOS incorporated diets (p < .05), while the feed conversion ratio (FCR) in the 1, 2 g/kg FOS groups reduced significantly compared with the control (p < .05). Besides, the crude lipid in the 4, 6 g/kg FOS groups increased significantly compared with the control (p < .05). On the other hand, the erepsin and lipase activities significantly elevated in intestine of fish fed 2 g/kg FOS (p < .05) and the lysozyme activity in serum of fish fed 2 g/kg FOS were significantly higher than that in the control (p < .05). Moreover, the alkaline phosphatase activities in serum of fish fed 0.5, 1, 2 g/kg FOS were significantly higher than in control (p < .05). Regression analysis showed that the relationships between dietary FOS levels and either SGR, FCR, erepsin or lysozyme activities were best expressed by regression equations, and the optimal inclusion levels are 1.37, 1.80, 3.06, 3.11, 1.93 and 1.80 g/kg for SGR, FCR, erepsin, lipase, lysozyme and total superoxide dismutase activities, respectively. Overall, this study revealed that FOS incorporated diets could beneficial for L. japonicus culture in terms of increasing the growth, digestion and immune activities. Under the present experimental condition, the optimal supplementary level of FOS in the diet of L. japonicus is 1–3 g/kg.     

Growth hormone (GH) and reproduction: a review

Interaction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.     

Purification and partial characterization of GtHs (cfLH and cfFSH) from Indian walking catfish (Clarias batrachus) (L.) and development of a homologous ELISA for cfLH

Two gonadotropins (GtH; Qa and Qb) were purified by gel filtration and ion exchange chromatography from the pituitaries of Indian walking catfish (Clarias batrachus). The presence of GtH during purification was assessed by in vitro oocyte maturation and in vivo steroidogenic activity, and their identities were determined by elution profiles, molecular weight, biological activities and yield. The molecular weights of Qa and Qb were 37 and 42 kDa, respectively, and composed of distinct subunits (Qa: 20 and 14 kDa and Qb: 26 and 18 kDa). Polyclonal antibodies raised against Qa immunostained Qa, Qb and pituitary GtH cells. A competitive Qa‐ELISA was developed whose sensitivity was 6.25 ng mL^?1 (1.25 ng well^?1) with intra‐ (3.5%) and inter‐ (12.4%) assay coefficients of variation. Displacement curves parallel to the standard were obtained with plasma and pituitary extracts of catfish, Qb and carp GtHII. The assay was validated by measuring the plasma Qa levels after LHRH treatment and in relation to ovarian growth in the female catfish during different reproductive phases. Based on the results, Qa and Qb corresponded to fish LH and FSH respectively. The findings will increase the knowledge of the mechanisms controlling fish reproduction and identification of sensitive phases in fish in captivity for hormonal manipulation.     

Controlled infection of Poecilia reticulata Peters (guppy) with Tetrahymena by immersion and intraperitoneal injection

Tetrahymena is a protozoan parasite, which infects guppy, Poecilia reticulata Peters, and causes substantial economical losses in commercial farms worldwide. Studies of guppy infected by Tetrahymena require standardized infection protocols. The LD50 for Tetrahymena infection of guppies by intraperitoneal (IP) injection was calibrated, and the level obtained was 946 parasites per fish. Guppy infection with Tetrahymena by immersion, imitating the natural route of infection via the integument, was studied under normal or stress conditions. Exposure to cold and netting (CNI) and to cold only (CI) followed by immersion exposure to 10 000 Tetrahymena per mL resulted in 22.5% and 19.2% mortality, respectively, as compared to 14.2% and 10% in groups that were netted only (NI) or non‐stressed (I). Histopathology revealed that immersion infection resulted in a systemic infection. Lysozyme levels, measured 3 weeks after infection, were significantly higher in the CNI group (288 μg per mg protein) compared with CI‐, NI‐ and I‐treated groups (94.5, 64 and 62.3 μg mg^?1, respectively). There was no evident parasite immobilization activity in body homogenates, suggesting no development of acquired immunity. Re‐infection by IP injection revealed no increase in protection in any of the treatment groups, mortality range of 56.3–75%, higher than in the non‐exposed control (40.6% mortality).     

Involvement of gonadal steroids in final oocyte maturation of white perch (Morone americana) and white bass (M. chrysops): in vivo and in vitro studies

Plasma estradiol-17 (E₂), testosterone (T), 17,20-dihydroxy-4-pregnen-3-one (DHP) and 17,20,21-tri-hydroxy-4-pregnen-3-one (20-S) levels were measured by radioimmunoassay (RIA) in white perch (Morone americana) and white bass (M. chrysops) that were induced to undergo final oocyte maturation (FOM) with human chorionic gonadotropin (hCG). Plasma DHP levels increased in females of both species in association with oocyte germinal vesicle migration (GVM) and germinal vesicle breakdown (GVBD) and decreased thereafter. Plasma 20-S levels also increased with oocyte GVM in white bass, but were several-fold lower than DHP levels. Circulating E₂ and T levels were greatest during GVM and GVBD in both species and decreased to low levels during oocyte hydration and ovulation. Follicles from white perch and white bass which received a priming injection of hCG in vivo, produced both DHP and 20-S in vitro after exposure to hCG and their oocytes underwent GVBD. Ovarian incubates from unprimed fish of either species produced only E₂ and T and their oocytes did not complete GVBD. Oocytes from unprimed bass, but not perch, matured when follicles were exposed to hCG in vitro. Both trilostane and cycloheximide blocked in vitro production of DHP and 20-S and oocyte GVBD by white perch follices. DHP and 20-S were equipotent inducers of FOM in the GVBD bioassay. None of several other structurally-related steroids tested were effective within a physiological range of concentrations. These results indicate a role for DHP and 20-S in the control of FOM in white perch and white bass.     

Cardiac,ventilatory and metabolic responses of two ecologically dissimilar species of fish to waterborne cyanide

Changes in heart rate, ventilatory activity and oxygen consumption were determined in trout (Salmo gairdneri) and brown bullhead catfish (Ictalurus nebulosus) during exposure to a steadily increasing concentration of waterborne cyanide selected to produce death in 8–9 hours for each species. The lethal cyanide concentration for the bullheads was an order of magnitude higher than for trout. Trout developed an immediate and gradually increasing bradycardia throughout the exposure period. Cyanide produced tachycardia in the bullhead followed by a gradual onset of bradycardia as the concentration of cyanide was raised. Pericardial injection of atropine (a muscarinic cholinergic antagonist) indicated that bradycardia in the trout was due initially to increased vagal tone but later due to the direct effect of cyanide on the heart. Hyperventilation in the trout persisted throughout the exposure period, although the rate and amplitude fluctuated and was variable between individual fish. During the last hour of exposure (highest cyanide concentration), ventilation was characterized by rapid, shallow breaths followed by a sudden respiratory arrest. The bullheads exhibited hyperventilation during the first 3 hours of exposure followed by a gradual, linear drop in ventilation rate and amplitude until death occurred. Cardiac and ventilatory responses in both species were attributed to stimulation of central and peripheral chemoreceptors by cyanide. Evidence is presented which suggests the initial response in the bullheads was due, at least in part, to gustatory stimulation by the cyanide. Oxygen consumption of the trout remained above pre-exposure levels for the majority of the test period. Oxygen consumption in the bullhead paralleled the changes in heart and ventilatory rates. Whole-body lactate levels of fingerlings of both species during cyanide exposure were measured to estimate the extent of anaerobiosis. Whole-body lactate levels were much greater in the bullheads than the trout, indicating a higher capacity for anaerobiosis, possibly due to a greater fuel supply. Overall, the trout responded to cyanide in a manner similar to that produced by environmental hypoxia whereas the bullheads experienced a gustatory stimulus which masked the hypoxia-like response.     

Protein and amino acid composition from the mantle of juvenile O ctopus vulgaris exposed to prolonged starvation

Protein and amino acid composition of the mantle of juvenile O ctopus vulgaris (Cuvier, 1797) during fasting for 27 days were determined. Average protein content of octopus mantle was of 711.19 ± 46.80 g kg^?1 DW, and it decreased with increasing fasting days. The non‐essential amino acids content was higher (486.18 ± 11.08 g kg^?1 protein) than essential amino acids (425.82 ± 9.15 g kg^?1 protein) at the start of the experiment (unstarved animals). The results suggest that the amino acid profile of the mantle where the most abundant amino acids are Arg, His, Lys, Gly, Leu and Pro could indicate a prolonged fasting condition (>20 days) or poor nutrition of O . vulgaris. This study supports the idea of using mantle for metabolic needs of starved O . vulgaris suggesting that the degradation pathway of amino acids to pyruvate and tricarboxylic acid cycle intermediates was favoured contrary to the degradation pathway of ketogenic amino acids. Special considerations should be taken concerning Thr, Ile, Ser, Ala, Asx (Asp, Asn), Glx (Glu, Gln) (because of their fast intake) and Lys and His (due to their stable contents) during a prolonged period of fasting.