肺炎克雷伯杆菌疫苗中试生产中细菌培养工艺的改进及优化

目的中试生产中对肺炎克雷伯杆菌培养工艺进行改进及优化。方法采用液体综合培养基代替半综合培养基在10L和100L中国丽生物反应器中对肺炎克雷伯杆菌进行培养,在10L中国丽生物反应器探讨不同的培养基配方、pH值、培养温度、搅拌转速、溶氧,工艺参数稳定后,扩大培养到100L中国丽生物反应器,并探讨培养过程中补加葡萄糖的浓度及补加方式等对细菌浓度及荚膜多糖含量的影响。结果肺炎克雷伯杆菌液体综合培养基可代替半综合培养基用于该菌的培养,培养过程中维持pH值7．2、温度37℃、通气60L／h、搅拌转速250r／min、培养到2h时开始以恒速补加30mL／L40％葡萄糖溶液、培养时间为5h,细菌长势最好,收获的荚膜多糖含量最高。结论肺炎克雷伯杆菌的培养工艺放大到100L中国丽生物反应器中,经过多次试验初步建立了稳定的肺炎克雷伯杆菌中试培养工艺。     

采用计算流体力学技术研究搅拌对兽疫链球菌发酵生产透明质酸的影响

搅拌是影响透明质酸(HA)发酵的一个重要因素,然而有关搅拌对HA发酵影响的认识存在较大争议。本研究采用计算流体力学(CFD)技术深入研究了搅拌对菌体生长和HA合成的影响。结果表明,菌体量和HA产量受搅拌转速的影响很小,而HA分子量随着转速的增加呈现出先增加后降低的趋势。分阶段控制转速研究表明转速对HA分子量的影响主要体现在HA合成阶段。CFD计算结果表明随着搅拌转速的增加,混合时间降低的同时反应器内部的剪切速率明显增加。最终通过改变搅拌桨组合方式的手段有效地解决了上述矛盾,并使得HA分子量提高23.9%。     

折流元件对环流反应器性能影响及用于苏云金茅杆菌发酵的效果

本文讨论了折流元件对环流反应器的流体力学性能的影响及用作生物反应器的研究结果。实验是在外筒规格Φ0 .2 19( 0 .2 10 )× 1.6m ,内筒规格Φ0 .153( 0 .150 )× 1m反应器内进行的 ,材质为不锈钢 ,括号内表示的是导流筒的内径。内筒装有具有强化传递过程作用的折流元件。BT发酵实验表明折流元件可改善环流反应器的发酵性能。将研究结果放大到 2 0m3的气升式环流反应器中 ,与工厂同等规模的搅拌式发酵罐相比 ,在完全省去搅拌功耗又不增加无菌空气用量的条件下 ,发酵液毒力效价高于在机械搅拌式发酵罐内的发酵结果约 10 %并且节电 30 %以上。     

发酵法生产谷氨酰胺转胺酶(MTG)的中试研究

比较了摇瓶和 2 .5L种子罐所培养的种子液 ,并在 5L罐上考察了以种子罐培养种子时不同接种量对发酵过程的影响。在将 5L小罐实验结果放大到 30 0L和 3m3 罐的中试规模时 ,分别采用以vvm相等和KLa相等的原则 ,对通风量进行放大 ,以P V相等为依据对搅拌转速进行放大。结果表明 ,当采用种子罐培养种子液时 ,适宜的种龄为 18～ 2 0h ,接种量为 2 % ,30 0L罐中酶活和生产强度分别达到 3.12u mL和 78.0u L·h ,均高于 5L罐的水平 ,3m3 罐酶活为 2 .70u mL ,接近于 5L罐的酶活水平 ,表明本研究所采用的通气量、搅拌转速等放大原则是合理的     

氧对膜生物反应器短程硝化的影响

为了研究膜生物反应器的短程硝化性能以及氧对短程硝化的影响,通过对比耗氧率和供氧率,提出了膜生物反应器短程硝化的控制优化建议。在膜生物反应器硝化过程中,DO小于1 mg/L开始出现亚硝氮积累;DO降到0.5 mg/L,出水氨氮浓度与亚硝氮浓度之比接近1∶1;DO调控在0.5-1 mg/L范围内,有利于前置硝化反应器与后续厌氧氨氧化反应器衔接。膜生物反应器中污泥浓度可达20 g/L,耗氧能力可达19.86 mg O2/(L·s),但最大供氧能力仅为0.369 mg O2/(L·s),供氧成为反应器运行的制约瓶颈,"低DO高流量"曝气是继续提高短程硝化效能的控制策略。     

出芽短梗霉发酵过程溶氧控制的研究

在搅拌罐式生物反应器中,通过控制DO（溶氧浓度）的变化,对出芽短梗霉（Aureobasidium pullulans）发酵过程的控制进行了研究。以100g/L玉米粉水解液做碳源,比较了不同溶氧控制条件下发酵参数的变化及其对出芽短梗霉发酵结果的影响。结果表明,过低的DO对菌体生长和多糖生产都不利,过高的DO使培养液中糖大部分消耗在菌体的生长上,也不利于多糖的生产,通过控制搅拌速度和通气量能将DO维持在较合适的水平。     

甘草细胞放大培养中搅拌式反应器操作策略优化

为获得甘草细胞在反应器中放大培养的最佳条件,在建立稳定的甘草细胞搅拌式生物反应器放大培养体系的基础上,分别以单因素和正交实验获得的数据为样本,以细胞净增长生物量为考察指标,运用BP神经网络耦合遗传算法对反应器操作策略进行优化。结果表明,接种量6.4%、摇床转速89r/min、通气速率0.1vvm是甘草细胞进行反应器培养的最优条件;与传统的正交实验方法相比,这种基于神经网络耦合遗传算法的优化方法使反应器中细胞生物量的积累提高了6.9%。     

表达量高、血凝活性好的H3N2流感病毒血凝素（HA）蛋白疫苗的筛选研究

摘要 目的：筛选表达量高、血凝活性好的H3N2流感病毒血凝素（HA）重组蛋白。方法：以A/MINNESOTA/41/2019（H3N2）毒株的HA为基础,将HA胞外区N端融合GP67信号肽,C端融合三聚化基序,分别构建HA胞外区全长及近膜区截短2个氨基酸的HA重组杆状病毒,并构建相应的不含三聚化基序的HA重组杆状病毒;经昆虫细胞表达,Western blot鉴定,亲和层析纯化后分析重组蛋白的血凝效价及稳定性。结果：四种HA重组蛋白均获得有效表达,其中HA胞外区全长融合三聚化基序的重组蛋白（HA-T）表达水平最高,经Strep tag亲和层析获得高度纯化,产量高达30 mg/L,Ethylene glycolbis交联分析显示为稳定的HA三聚体形式,血凝活性分析显示HA-T的活性最高,血凝效价为29,动态光散射显示HA-T在4℃放置3个月性质稳定。结论：HA-T表达量高、稳定性好、血凝活性高且易于纯化,研究结果为流感重组蛋白疫苗的研发策略提供了参考。     

Vero 细胞流感病毒的大规模培养和纯化

目的：采用反应器技术以细胞为基质培养流感病毒,并探索其纯化工艺。 方法：采用5L反应器无血清条件下以Vero细胞为基质培养流感病毒,病毒收获后,经灭活、澄清,采用阴离子柱去除宿主DNA,亲和柱浓缩流感病毒,最后采用分子筛三步层析法进行纯化。结果：整个纯化工艺HA回收率达102%,蛋白降低率为95.3%,宿主蛋白降低率为99.77%,DNA的降低率为99.99%。结论 本研究成功建立了一种细胞流感病毒的纯化工艺。     

搅拌式生物反应器悬浮培养水母雪莲细胞的研究

应用 2L通气搅拌式生物反应器一步批式培养水母雪莲细胞。采用倾斜式搅拌桨代替透平桨 ,研究了搅拌转速、通气量和接种量对细胞生长和黄酮合成的影响 ,发现在 75r min、70 0～1000L min和 4.0～ 5.0gDCW L接种量下细胞生长和黄酮合成比较好。经过 12d培养细胞干重达 13.8gDCW L ,黄酮产量 416mg L ,黄酮含量占细胞干重的 30%。水母雪莲细胞生长及黄酮合成的进程表明 ,黄酮积累与细胞生长呈正相关。对细胞聚集体分布的研究发现 ,流变压力使细胞聚集体分裂 ,使反应器中细胞生长受到影响 ,黄酮产量较摇瓶中降低     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

Entwicklungsgeschichte und Ultrastruktur von Pollenkitt und Exine bei nahe verwandten entomophilen und anemophilen Angiospermensippen derAlismataceae,Liliaceae, Juncaceae,Cyperaceae, Poaceae undAraceae

Some closely related members of the monocotyledonous familiesAlismataceae, Liliaceae, Juncaceae, Cyperaceae, Poaceae andAraceae with variable modes of pollination (insect- and wind-pollination) were studied in relation to the ultrastructure of pollenkitt and exine (amount, consistency and distribution of pollenkitt on the surface of pollen grains). The character syndromes of pollen cementing in entomophilous, anemophilous and intermediate (ambophilous or amphiphilous) monocotyledons are the same in principal as in dicotyledons. Comparing present with former results one can summarize: 1) The pollenkitt is always produced in the same manner by the anther tapetum in all angiosperm sub-classes. 2) The variable stickiness of entomophilous and anemophilous pollen always depends on the particular distribution and consistency of the pollenkitt, but not its amount on the pollen surface. 3) The mostly dry and powdery pollen of anemophilous plants always contains a variable amount of inactive pollenkitt in its exine cavities. 4) A step-by step change of the pollen cementing syndrome can be observed from entomophily towards anemophily. 5) From the omnipresence of pollenkitt in all wind-pollinated angiosperms studied one can conclude that the ancestors of anemophilous angiosperms probably have been zoophilous (i.e. entomophilous) throughout.

     

Identification and Characterization of the First Equine Parainfluenza Virus 5

正Dear Editor,Parainfluenza virus 5 (PIV5), known as canine parainfluenza virus in the veterinary field, is a negative-sense,nonsegmented, single-stranded RNA virus belonging to the Paramyxoviridae family (Chen 2018). The virus was first reported in primary monkey kidney cells in 1954 (Hsiung1972), then it has been frequently discovered in various     

Pathogenic Characterization and Full Length Genome Sequence of a Reassortant Infectious Bursal Disease Virus Newly Isolated in Pakistan

<正>Dear Editor,Infectious bursal disease (IBD) is one of the most important diseases of the poultry. The IBD virus (IBDV), a nonenveloped virus belonging to the Birnaviridae family with a genome consisting of two segments of double-stranded RNA (segments A and B), targets B lymphocytes of bursa of Fabricious leading to immunosuppression. In Pakistan,poultry farming is the second biggest industry and IBD is the second biggest disease threating the poultry sector.However, there is limited genome information of IBDV