产胞外多糖植物内生菌的分离和鉴定

利用平板涂布法从胡萝卜、芦荟、土豆、地瓜和紫薯样品中分离得到24株细菌,利用苯酚硫酸法测定菌株胞外多糖的产量,最终得到3株高产胞外多糖的细菌菌株25-Z-1、25-Z-3和12-L-6。经16S rDNA序列测定及系统发育分析,从紫薯样品中分离得到的25-Z-1与Bacillus pumilus strain ST277处于同一分支,相似性达到100%;25-Z-3与Bacillus cereus strain Se05处于同一分支,相似性达到99%;从芦荟中分离得到的12-L-6与Bacillus sp.GZT处于同一分支,相似性为99%。在LB液体培养基中,发酵温度30℃,摇床转数170 r/min条件下,发酵7 d后胞外多糖产量可分别达30.3 mg/L、29.1 mg/L及28.2 mg/L。     

竹黄产胞外多糖液态培养条件优化

目的:优化竹黄Shiraia sp. SUPER-H168产胞外多糖的液态培 养基组成和发酵条件.方法:通过单因素和正交试验优化培养基组成和发酵条件,在65L气升式发酵罐中进行验证试验.结果:经分析确定最佳培 养基组成为(g/L)葡萄糖40,玉米浆粉3,KH2PO4 1,MgSO4*7H2O 0.05;最佳发酵条件为起始pH 6.0,温度30℃,装液量75mL/250mL,接种量10%(v/v).在此条件下,摇瓶中 生物量及多糖分别达到11.7g/L和1.12g/L,验证试验中分别可达到12.1g/L和1.35g/L.结论:放大试验结果优于摇瓶水平,实验数据对生产有一定指导作用.     

产灵芝胞外多糖培养基的优化及发酵指标变化

经单因素和正交试验优化,灵芝胞外多糖最佳发酵培养基各成分质量分数为:麦芽糖2%,黄豆粉1%,FeSO4·7H2O0.02%,KH2PO40.1%,土豆汁体积分数30%,pH自然,产量可达到86.36g·L-1(湿重)。灵芝胞外多糖产量受发酵过程各因素的影响,发酵过程中pH、总糖、还原糖和氨基氮有一定的相关性。灵芝多糖整个发酵过程需要144h左右,第6d达到发酵终点。     

灵芝胞外多糖的发酵条件及其组份的初步研究

本文报道了液体培养灵芝Gl8801菌林产生胞外多糖的最佳发酵条件和胞外多搪的化学组成。适宜的液体培养基(g/L):饴糖40,花生饼粉30,KH_2PO_41.5,(NH_4)_2SO_41.5,MgSO_4·7H_2O_(0.75),CaCO_32,最适起始pH5.5。300ml三角瓶装培养基60ml,发酵温度28℃。摇瓶(165rpm)培养7～8天,发酵液纯多糖含量可达880mg/L。灵芝胞外多糖分为水溶性多糖和水不溶性多糖两种。水溶性多糖的单糖组份为半乳糖、葡萄糖、阿拉伯糖和木糖,所含单糖重量比为6:4:4:1。不溶性多糖为葡聚糖。灵芝胞外多糖的糖环构型为吡喃糖,末端糖基构型,半乳糖为α型,葡萄糖为β型。     

啤酒酵母胞外多糖发酵条件的研究

以啤酒酵母S－12为出发菌株,用紫外线＋氯化锂作为复合诱变剂,获得一株产胞外多糖量较高的变株S－12－4,比出发菌株提高33．3％,同时对变株进行了最佳培养条件的研究,结果表明：最适碳源和氮源分别为大米糖3％、酵母粉0．37％及NH4Cl0．32％,最适发酵条件为起始PH6．0、培养温度26℃,发酵周期为30h,在此基础上进行培养,变株S－12－4产胞外多糖最高可达38.2mg/100mL,比初始条件提高了54％。     

西藏灵菇胞外多糖组分对小鼠免疫调节作用及机制的研究

摘要：【目的】研究数均分子量为0.1×105~3.0×105（组分1）及1.8×103（组分2）的西藏灵菇胞外多糖组分对正常小鼠免疫功能的影响,并探讨其影响机制。【方法】依据卫生部保健食品功能学评价程序和检验方法,灌胃给药,剂量分别120 mg/kg体重、80 mg/kg体重、40 mg/kg体重,检测脏器／体重比值、半数溶血值（HC50）、自然杀伤细胞（NK）活性、迟发型变态反应（DTH）、腹腔巨噬细胞吞噬功能。采用免疫印迹法,测定小鼠腹腔巨噬细胞中Erk蛋白及COX-2酶的表达量。【结果】组分1能够明     

假单胞菌胞外多糖发酵条件的研究

研究了假单胞菌（Ｐｓｅｕｄｏｍｏｎａｓｓｐ．６）利用木糖产胞外多糖的发酵条件。实验表明ＫＨ２ＰＯ４,Ｏ２和高碳氮比对多糖合成有促进作用,在发酵后期补加木糖有助于多糖产量的提高     

刚果红平板法筛选血链球菌胞外多糖缺失突变株

细菌胞外多糖不但能够保护细菌,防止外界环境变化和侵蚀者对菌体细胞的侵害,而且在医学微生物学中还具有特有的生物学活性, 包括抗辐射、抗氧化等特性。尽管细菌胞外多糖受到了学者的广泛关注,但胞外多糖的生物合成机制还不十分清楚。以血链球菌为模式生物,以随机插入突变机制构建的血链球菌胞外多糖形成基因文库为受试对象,建立了一种简单易行的刚果红平板法用于筛选血链球菌细胞外粘多糖形成缺失突变株的方法,为日后研究细菌胞外多糖形成机制以及临床上筛选胞外多糖缺失突变株奠定了基础。     

The identification of vahlkampfiid amoebae by ITS sequencing

We have determined the internal transcribed spacer (ITS) sequences (including the 5.8S ribosomal DNA) of 30 strains of 14 species belonging to eight vahlkampfiid genera. Each previously described species has a specific ITS sequence, except for Tetramitus aberdonicus, Tetramitus thorntoni, and Tetramitus jugosus, which have identical ITS sequences. The latter three may therefore constitute a single species despite their apparent phenotypic differences. The ITS sequence appears to be conserved within a species. The species Willaertia magna appears to be ubiquitous. The 5.8S rDNA sequences of Singhamoeba horticola and Learamoeba waccamwensis indicate that they do not represent different genera, but both belong to the genus Tetramitus. The ITS sequences of 16 undescribed vahlkampfiid isolates were determined. Based on these sequences, seven isolates were identified as belonging to described species, while nine probably represent seven new species. Five of these presumed new species belong to the genus Tetramitus, and one each to the genera Vahlkampfia and Paravahlkampfia.     

云南程海湖酵母菌多样性及应用

【目的】针对云南丽江永胜县境内程海湖环境的特殊性,研究高原湖泊环境中酵母菌的多样性,初步探索程海湖环境中酵母菌的利用价值。【方法】对程海湖的湖水和其周边土壤样品中的酵母菌进行分离;应用26S rDNA的D1/D2区域序列分析,并结合形态及生理生化指标对分离获得的酵母菌进行鉴定;采用筛选培养基对已鉴定酵母菌进行产酶定性实验,分析高原湖泊中酵母菌的多样性及可应用性。【结果】分离得到酵母菌64株,对其中63株进行鉴定,归属于9个属22个种(包括4个疑似新种或新变种);地霉属Geotrichum和隐球酵母属Cryptococcus是2种环境中的共有属;在产酶活性筛选中发现有9株产胞外酶活性的菌株,其中YM24373既产蛋白酶又可产淀粉酶。【结论】研究结果显示程海湖中酵母菌组成具有较为丰富的多样性,其应用价值值得进一步研究。     

Distribution and identification of red yeasts in deep-sea environments around the northwest Pacific Ocean

We isolated 99 yeast strains, including 40 red yeasts, from benthic animals and sediments collected from the deep-sea floor in various areas in the northwest Pacific Ocean. Comparing the yeast isolates from animals and sediments collected from shallow locations, the proportion of red yeasts differed considerably, comprising 81.5% and 10.6% of the isolates from animals and sediments, respectively. All of the red yeast isolates belonged to the genera Rhodotorula and Sporobolomyces. On the basis of morphological and physiological characteristics, the isolates were identified as R. aurantiaca, R. glutinis, R. minuta and R. mucilaginosa of the genus Rhodotorula, and S. salmonicolor and S. shibatanus of the genus Sporobolomyces. Only R. glutinis and R. mucilaginosa were isolated from sediments. All of the others were isolated from animal sources. Phylogenetic analyses based on internal transcribed spacer (ITS) regions and 5.8S rRNA gene sequences allowed us to establish the precise taxonomic placement of each of the isolates and thereby investigate the intraspecific relationships among the isolates. Twenty-two strains identified as members of R. glutinis, which showed a wide distribution in the deep-sea, and five isolates identified as R. minuta, which were isolated only from benthic animals, showed substantial heterogeneity within the species. The isolates phenotypically identified as Sporobolomyces species and R. mucilaginosa phylogenetically occupied the placements corresponding to these species. Some strains assigned to known species on the basis of phenotypic features should be regarded as new species as suggested by the results of molecular analysis.     

适冷溶菌酶产生菌的分子鉴定及其酶学性质的研究

采用比浊法对本试验室保存的126株南极微生物产适冷溶菌酶进行测定,发现7株菌产溶菌酶.并对其中的菌株NJ147进行ITS-5.8 S rDNA基因序列的同源性和系统发育分析,结果表明:菌株NJ147属于Debaryomyces hansenii属.该菌株所分泌溶菌酶的最适作用温度为30 ℃,在0 ℃时酶活性是最高活性的...     

西藏曲拉和云南乳饼中酵母菌的鉴定及其生物多样性

【目的】探讨西藏曲拉和云南乳饼中酵母菌的生物多样性及其分布特征,为我国传统乳制品中酵母菌资源的利用提供基础数据。【方法】从西藏和云南分别采集的5份曲拉样品和8份乳饼样品中分离出41株酵母菌,利用26SrDNAD1/D2区域序列分析对这些菌株进行了分类鉴定。【结果】曲拉和乳饼样品中酵母菌的总数分别在106-107cfu/g和102-106cfu/g之间,曲拉样品的酵母菌平均数比乳饼样品中的高34倍。共鉴定出10属12种,其中西藏曲拉的优势菌株为发酵毕赤氏酵母(Pichia fermentans)和酿酒酵母(Saccharomyces cerevisiae);云南乳饼的优势菌株为类筒假丝酵母(Candida zeylanoides)和喜仙人掌毕赤氏酵母(Pichia cactophila)。毕赤氏酵母属(Pichia)是曲拉和乳饼的共同优势属。【结论】西藏曲拉和云南乳饼中的酵母菌都具有丰富的生物多样性,但其差异性很大。     

一株产冠菌素新菌种的分离与鉴定

【目的】从不同样本中分离筛选性能稳定的产冠菌素菌株。【方法】根据冠菌素引起植物叶片产生弥散性黄萎病、块茎膨大的特性,采集各种植物病叶、病枝及感病植物的土壤,采用穿刺法与系列稀释法分离筛选菌株;液相色谱测定菌株产生的冠菌素;在电子和光学显微镜下观察菌体形态;根据生理生化试验、(G+C)mol%含量、16S rDNA序列分析等对菌株进行鉴定;对分离提纯的发酵产物进行紫外、质谱和红外分析。【结果】菌株BBC933为革兰氏阴性菌,端生鞭毛,短杆状,无芽孢。在41℃下不生长,细胞内有聚β-羟基丁酸盐颗粒积累,没有精氨酸双水解酶和氧化酶,不能水解淀粉、明胶,不进行硝酸还原及反硝化作用,过氧化氢酶反应呈阳性。菌株的(G+C)mol%含量为67.2%,根据该菌株16S rDNA序列的同源性分析,构建系统发育树。【结论】菌株BCC933鉴定为洋葱伯克霍尔德氏菌(Burkholder cepacia),具有产冠菌素性能。国内外未曾见报道洋葱伯克霍尔德氏菌产冠菌素。     

鮸鱼弧菌病病原菌(哈维氏弧菌)的分离与鉴定

【目的】2009年春季,浙江省舟山地区养殖鮸鱼暴发弧菌病。症状主要表现为体表病灶部位出血、肌肉溃烂、内脏器官有白斑等。【方法】从病鱼体表溃疡部位及内脏分离出优势菌株090212,经人工感染证实该菌即为致病菌。通过API系统和菌体常规形态特征、培养特性和生理生化反应指标测定以及16S rRNA测序分析等综合鉴定,【结果】确认090212为哈维氏弧菌(Vibrio harveyi),该菌为革兰氏阴性,菌体呈短杆状,极生单鞭毛。该菌对氟苯尼考、四环素等5种抗生素敏感。【结论】哈维氏弧菌是海水养殖鱼类的常见致病菌,但作为养殖鮸鱼的病原菌尚属首次报道,将对鮸鱼的病害防治和健康养殖具有重要的指导意义。     

拉格啤酒酵母菌物种和株系的快速分子鉴定

近年来的基因组学研究结果已证实拉格啤酒酵母Saccharomyces pastorianus是一个由艾尔啤酒酵母S. cerevisiae和真贝氏酿酒酵母S. eubayanus杂交而成的杂交种,并可根据地域传承和染色体倍性分为两个株系,即I型/Saaz系和II型/Frohberg系。前者主要为异源3倍体,后者则主要为异源4倍体。为了探讨中国啤酒酿造酵母菌的物种和菌系归属,我们根据拉格啤酒酵母及其两个菌系的基因组特性,制定了一套基于IntFR片段种特异性扩增和ITS-RFLP分析的精确但简便易行的拉格啤酒酵母菌物种和株系鉴定新方法,并以酿酒酵母属内相关种的模式或权威菌株和部分酒精及面包酵母为参照,对保藏于中国普通微生物菌种保藏中心（CGMCC）的41株啤酒酿造酵母菌进行了重新鉴定和分型。这些菌株除1株原定名为贝氏酿酒酵母S. bayanus外,其余菌株的原定名均为S. cerevisiae。研究结果确认了S. bayanus菌株鉴定的正确性,但在其余的40株啤酒酵母菌株中,21株属于S. cerevisiae,1株属于葡萄汁酿酒酵母S. uvarum,18株属于S. pastorianus。菌系鉴定和流式细胞测定结果显示在确认的S. pastorianus菌株中,1株为I型/Saaz系,3倍体;17株为II型/Frohberg系,其中9株为4倍体,两株为3倍体,5株介于3倍至4倍体之间。啤酒酵母物种和株系的确认对优化发酵工艺和菌种选育及遗传改造等具有重要意义。     

微囊藻毒素降解菌的筛选、鉴定及其降解活性研究

采用从巢湖水华蓝藻细胞中提取、提纯的藻毒素（Microcystins,MCs）为微生物生长的唯一碳源和氮源,通过平板分离纯化,从巢湖底泥中分离出5株能够降解藻毒素的菌株,并对其中降解活性较高的一株进行分子鉴定。应用PCR技术克隆到16S rDNA片段,核苷酸序列分析结果表明,该菌的16S rDNA的全序列与吉氏库特菌kurthia gib-soniistrain HC050630C-1的相似性达99%。微囊藻毒素降解实验结果表明,用15mg/L乙醇作为外加碳源时可显著提高菌株M9降解MCs的能力,在48h内对初始浓度分别为17.1mg/L的MC-RR和11.3mg/L的MC-LR的降解率分别达到70.0%和81.6%。而葡萄糖对菌株M9的生长有明显抑制作用。     

Phylogenetic analysis of isolated biofuel yeasts based on 5.8S-ITS rDNA and D1/D2 26S rDNA sequences

The utilization of agro-industrial wastes such as whey as raw materials for the production of bio-ethanol is gaining importance as a result of the attractiveness of renewable fuel alternatives due to exhaustion of fossil fuel sources coupled with the positive impact to the environment. Here, we report the isolation of two Kluyveromyces spp. designated as BM4 and P41, able to produce ethanol as main fermentation product from fermenting whey. Three different molecular biological approaches including, the RFLP analysis of the 5.8S-ITS rDNA, the sequence of the 5.8S-ITS rDNA region and the sequence of the D1/D2 domain of the 26S rRNA gene were applied for accurate identification. While RFLP analysis of 5.8S-ITS region failed to accurate the differentiation between the two species, sequencing of this region and D1/D2 region of the 26S rRNA gene verified the identification. PCR amplification and sequence analysis of 5.8S-ITS rRNA and D1/D2 domain of the 26S rRNA genes revealed that the isolates BM4 and P41 were highly related to Kluyveromyces marxianus and Kluyveromyces lactis with homology of 99% for both. In addition, phylogenetic analysis indicated that both BM4 and P41 shared a cluster with K. marxianus and K. lactis, respectively. The fermentative performance of both strains on cheese whey to produce ethanol was evaluated at different parameters such as incubation temperature, initial pH, whey sugar concentrations, and yeast concentrations. Results show that the maximum ethanol productions achieved at pH 4.5 and 35 °C were 5.52% and 5.05% for K. marxianus and K. lactis, respectively. Our results demonstrated that K. marxianus and K. Lactis could be recommended for cheese whey bioremediation in the environment and produce renewable biofuel.