木棉花对乳酸菌生长及保存活力的影响

通过实验研究木棉花对与人类关系密切的植物乳杆菌、两歧双歧杆菌、嗜热链球菌的生长及保存活力的影响。结果表明:木棉花对植物乳杆菌和两歧双歧杆菌的生长及保存活力有较大促进作用,而且此促进作用随着木棉花浓度的增大而增大。木棉花对嗜热链球菌的生长和保存活力的促进作用很小。因此,可考虑将木棉花适当添加到含有植物乳杆菌、两歧双歧杆菌的食品中,用以促进菌体的生长及保存活力,还可考虑利用木棉花制微生态制剂。     

桃花粉低温和超低温保存方法比较研究

桃(Prunus persica(L.)Batsch)是我国重要的无性繁殖作物种质资源,目前主要保存于3个国家无性繁殖作物种质圃。随着以茎尖、花粉、休眠芽为保存载体的超低温保存技术的发展,超低温保存已成为无性繁殖作物重要备份保存方式。本研究以15份桃种质花粉为研究对象,开展含水量、回湿处理和保存温度(4℃低温保存和液氮超低温保存)对保存后花粉离体萌发率的影响研究。研究结果:明确了桃种质花粉超低温保存的含水量;揭示了回湿处理对部分桃种质花粉超低温保存产生显著影响;超低温保存后花粉离体萌发率最高可达83%;4℃低温保存和超低温保存比较研究结果表明,超低温保存4年后14份桃种质花粉离体萌发率仍可保持30%以上,11份桃种质花粉离体萌发率与保存前花粉离体萌发率相比无显著变化甚至显著提高,而4℃低温保存的花粉离体萌发率降至0。该研究为国家种质库建立花粉规模化超低温保存提供技术支撑。     

温度、pH、溶氧对添加植物乳杆菌构建生物絮凝系统的影响

生物絮凝技术和微生物制剂在水产养殖业中广泛应用,但向生物絮凝系统中添加益生菌最优条件研究甚少。本试验采取L_9(3~3)安排3因素3水平关于溶氧(厌氧0.5 mg/L,有氧5 mg/L,厌氧-有氧交替)、pH (5.5, 7, 8.5)、温度(25℃, 30℃, 35℃)正交试验,开展35 d以植物乳杆菌为添加剂培养生物絮团试验,通过比较三态氮的变化情况、脱氢酶活性(dehydrogenase activity, DHA)及微生物种群结构,得出以植物乳杆菌为添加剂培养生物絮凝的最佳工艺。结果表明:在有氧、pH值8.5、30℃时,生物絮凝培养用时最短,为13 d,生物脱氮效率高;温度和溶氧对脱氢酶活性有显著性影响,25℃,厌氧-好氧交替,pH值7条件下培养的生物絮团脱氢酶活性最高;微生物群落结构分析表明生物絮团中主要为变形菌门(70%),其次为拟杆菌门;植物乳杆菌在有氧、35℃、pH值5.5条件下丰度最高,为1.8%。综合实际生产,添加植物乳杆菌构建生物絮凝系统最佳条件为30℃、p H值7、有氧条件(DO5 mg/L)。     

可抑制致病菌的益生菌筛选及抑菌物质的初步确定

为了得到可抑制致病菌的益生菌并初步确定其抑菌物质。从土壤、饲料添加剂和肥料添加剂3种样品中初筛能产生抑菌圈的菌株。用牛津杯法,对初筛得到的菌株和实验室保存的菌株进行复筛。对复筛的未知菌株进行形态学、16S r RNA基因序列和生理生化特征鉴定。对复筛菌株的发酵上清液进行酸碱作用验证、高温处理、蛋白酶处理、盐析处理和透析处理,再进行抑菌活力测定。结果显示复筛中有8株对大肠杆菌、金黄色葡萄球菌和藤黄微球菌都有较强抑制作用的菌株。8株菌中有4株已知菌,4株未知菌。已知菌株分别为保加利亚乳杆菌、干酪乳杆菌、鼠李糖乳杆菌和植物乳杆菌。4株未知菌中3株被鉴定为地衣芽孢杆菌,1株被鉴定为乳酸片球菌。地衣芽孢杆菌能产生多种耐高温的抑菌物质,其中包含能透过8-14 k D透析袋的小分子和不能透过8-14 k D透析袋的大分子,初步判定为多肽类物质。5株产酸菌株产生的抑菌物质主要是酸性物质。     

猪乳铁蛋白在4种重组乳杆菌中的表达及抑菌活性比较

为了获得优化的猪乳铁蛋白乳杆菌表达系统,并比较重组猪乳铁蛋白的抑菌活性,根据乳杆菌使用密码子的偏嗜性优化合成猪乳铁蛋白成熟肽编码序列,将其克隆到乳杆菌表达载体pPG612.1的XhoⅠ/BamHⅠ位点,获得了plf乳杆菌表达载体质粒pPG612.1-plf。将获得的重组质粒分别电转化入干酪乳杆菌ATCC393、戊糖乳杆菌KLDS1.0413、植物乳杆菌KLDS1.0344和副干酪乳杆菌KLDS1.0652细胞内,获得4种表达猪乳铁蛋白的重组乳杆菌。经木糖诱导,通过Western blotting和激光共聚焦检测重组猪乳铁蛋白的表达,用ELISA方法检测和比较4种重组菌上清中表达猪乳铁蛋白的量,并用琼脂孔穴扩散抑菌法检测4种重组乳杆菌表达乳铁蛋白的抑菌活性。结果表明,乳铁蛋白在4种重组乳杆菌中均得到正确表达,其产物分子量约73 kDa,重组干酪乳杆菌、重组戊糖乳杆菌、重组植物乳杆菌和重组副干酪乳杆菌的重组猪乳铁蛋白表达量分别为9.6μg/mL、10.8μg/mL、12.5μg/mL、9.9μg/mL。重组猪乳铁蛋白对大肠杆菌、金黄色葡萄球菌、鼠伤寒沙门氏菌、巴氏杆菌和李氏杆菌均有一定的抑菌作用,对金黄色葡萄球菌的抑菌作用最强,且4种重组乳杆菌中重组植物乳杆菌表达产物的抑菌效果优于其他重组菌的表达产物。结果表明在4种乳杆菌中重组猪乳铁蛋白的最佳表达系统为植物乳杆菌,该结果为猪乳铁蛋白的乳杆菌表达系统进一步开发与应用奠定了基础。     

改良皮片4℃低温保存方法在创面二次植皮治疗中的应用研究

目的:探讨改良皮片4℃低温保存方法在创面二次植皮治疗中的应用效果。方法:①动物实验:健康成年豚鼠3只,处死后,取背部皮肤做成32个1 cm×1 cm小皮样,随机分为新鲜皮组、庆大盐水保存组、RPMI保存组,改良RPMI保存组进行4℃保存;1周后测定皮肤活力;②临床实验:观察自2018年10月至2018年12月,二期植皮患者33例,应用改良RPMI 4℃低温保存皮肤二期回植的患者16例,与重新取皮植皮患者17例比较其皮片成活率。结果:动物实验证实:改良RPMI保存组4℃保存组在皮肤储存1周时皮肤平均活力较庆大盐水保存组、RPMI保存组高(P0.05);临床实验证实:改良RPMI保存组与重新植皮的皮片平均成活率没有显著性差别(P0.05)。结论:改良4℃断层皮片低温保存方法可短期内保存皮肤较高的活力,是皮片再利用的一种有效方法。     

植物乳杆菌CMCC-P0002代谢产物中抑菌物质的研究

目的通过对一株植物乳杆菌代谢产物的抑菌作用以及该产物的某些物理特性的研究,为进一步发现可替代现有抗生素的抗菌物质奠定基础。方法首先利用低温离心和超滤法对植物乳杆菌培养后的上清液进行初步提取,用打孔法行抑菌试验,明确其对铜绿假单胞菌、大肠埃希菌、肺炎克雷伯菌和金黄色葡萄球菌等临床分离菌株的抑菌活性;并进一步经加热、调整pH及过氧化氢酶等处理上清液后,再验证其活性变化。结果植物乳杆菌的培养上清液对铜绿假单胞菌等多株临床分离菌株均具有一定的抑菌活性,且以pH在6以内时该物质抑菌活性较好,过氧化氢酶处理后或加热至100℃、30min,上清液的抑菌活性依然存在。结论在植物乳杆菌的培养上清液中存在着具有抑菌活性的物质,对从临床标本所分离的部分革兰阴性菌及革兰阳性菌有抑菌作用,该物质对热耐受,其活性受pH变化的影响。     

青藏高原乳酸菌对垂穗披碱草青贮饲料发酵品质的影响

摘要:【目的】为了探究青藏高原乳酸菌对垂穗披碱草发酵品质的影响。【方法】本试验将3株从青藏高原垂穗披碱草(Elymus nutans Griseb.)中筛选的耐低温乳酸菌(戊糖片球菌PP-6,植物乳杆菌LP-2和清酒乳杆菌LS-5)作为添加剂加入垂穗披碱草进行青贮,同时对其生理生化特性进行研究,并以低海拔条件下分离得到的相同菌种的商品化乳酸菌制剂为对照,研究所筛选的3 株优良乳酸菌在低温(15 ℃)和常温条件(25 ℃)下对垂穗披碱草青贮饲料发酵品质的影响。【结果】试验结果表明,在碳源利用方面,分离得到的戊糖片球菌PP-6(Pediococcus pentosaceus)发酵棉子糖、乳糖、山梨醇、蜜二糖和蔗糖,清酒乳杆菌LS-5(lactobacillus sakei)可发酵棉子糖、苦杏仁苷、鼠李糖、乳糖、山梨醇、木糖、阿拉伯糖、蜜二糖和蔗糖,而相同菌种的商品菌均不能利用,表明高原乳酸菌具有更广泛的糖源利用特性。将筛选得到的3株低温生长优良菌株进行青藏高原垂穗披碱草青贮发酵试验,青贮50 d后,与对照的商品菌处理相比,在15 ℃时,清酒乳杆菌LS-5显著降低了青贮饲料pH值、丙酸含量和氨态氮/全氮值(P＜0.05),且能够保存更多的水溶性碳水化合物和粗蛋白;植物乳杆菌LP-2(lactobacillus plantarum)和戊糖片球菌PP-6作为复合添加剂显著提高了青贮饲料乳酸含量(P＜0.05),显著降低了氨态氮/全氮值和中性洗涤纤维含量(P＜0.05),能够保存更多的粗蛋白。但在25 ℃时,此3株菌与对照菌种相比均无明显优势。【结论】上述结果表明,从青藏高原分离筛选的3株菌种在低温条件下可有效提高青藏高原垂穗披碱草青贮饲料的发酵品质。     

微贮用植物乳杆菌固定化菌剂的制备

乳酸菌与纤维素降解菌因其可防止微贮饲料酸败、增加秸秆饲料的营养价值等优点,在秸秆微贮过程中起重要作用。但由于乳酸菌的繁殖会抑制纤维素降解菌的活性,如何实现微贮过程中两种微生物分时发挥功能是解决上述问题的关键。文中利用固定化技术将乳酸菌制备成含有玉米秸秆粉的固定化菌剂以达到缓释的目的。首先制作固定化空白小球得出复合固定化载体成球的最佳浓度,利用玉米芯吸附植物乳杆菌S1得到复合固定化载体,以对S1的包埋率、成球效果等为指标,通过对比两种固定化方法 (包埋法与包埋-交联法),得到固定化植物乳杆菌S1的最佳条件。研究表明,使用6%PVA+0.4%SA+0.3%CMC-Na进行包埋-交联时成球效果最好,使用1.2%SA+0.5%CMC-Na进行直接包埋时成球效果最好。通过对比5种固定化工艺,将1.2%SA+0.5%CMC-Na和吸附玉米粉组成的固定化载体混合物逐滴滴入4%氯化钙中直接包埋24 h得到的固定化小球其机械强度以及包埋率均优于其他工艺。因此,利用玉米芯吸附-海藻酸钠包埋的方法可以有效提高植物乳杆菌包埋效率,为使用固定化技术制备微贮饲料菌剂奠定基础。     

饲料乳酸菌的分离鉴定及优良菌株筛选

对从饲料玉米、高粱、麦秆及棉花中筛选出的乳酸菌进行分类鉴定和综合性分析。用MRS+CaCO3固体培养基从棉花中分离出乳酸菌18株、高粱中30株、饲料玉米中18株、麦秆中18株。经形态学、生理生化试验进行初步鉴定并按产酸试验,耐盐及耐酸试验挑选出32株产酸率强的乳酸菌对其进行16S rDNA分子鉴定。结果显示,32株菌都具有良好的耐盐、耐酸能力;经生理生化和16S rDNA基因序列鉴定可知32株乳酸菌分属于两个属,即乳杆菌属、肠球菌属,4个种,即干酪乳杆菌(Lactobacilluscasei)、肠道球菌(Entercoccus faecium)、植物乳杆菌(Lactobacillus plantarum)、海氏肠球菌(Entercoccus hirae)。4种饲料原料中肠道球菌普遍存在。除了这种乳酸菌以外,棉花有干酪乳杆菌、植物乳杆菌、海氏肠球菌,玉米和麦秆内有植物乳杆菌。从饲料中筛选出4株具有较强产酸能力的乳酸菌,可进一步研发成青贮饲料添加剂。     

A Simple Method for Long-term Preservation of Stock Cultures of Lactic Acid Bacteria

Strains of Lactobacillus sp., Leuconostoc sp. and Streptococcus sp. were preserved for 6–24 months at ambient temperature, partially dehydrated on granular pumice stone. To date, all the cultures stored by this technique have remained viable and no changes in the cultural or biochemical characteristics were observed.     

A群C群脑膜炎球菌结合疫苗稳定性

目的评价A群C群脑膜炎球菌结合疫苗原液和成品的稳定性。方法分别将A群、C群脑膜炎球菌结合疫苗原液及A群C群脑膜炎球菌结合疫苗各选取连续3批,分别放置于37℃、20~25℃和2~8℃3种温度下,在一定的时间取样进行主要项目测定,在关键时间点进行全面检测。结果 A群结合疫苗原液于2~8℃保存9个月,20~25℃保存4周,37℃保存4 d;C群结合疫苗原液于2~8℃保存9个月,20~25℃保存6个月,37℃保存4周;A群C群脑膜炎球菌结合疫苗于2~8℃保存2年3个月,20~25℃保存6个月,37℃可以保存9周;各项检测指标均符合质量标准的要求。结论在2~8℃条件下,A群、C群脑膜炎球菌结合疫苗原液存放6个月,A群C群脑膜炎球菌结合疫苗存放2年,其质量稳定。     

益生乳酸菌Lactobacillus casei Zhang和Lactobacillus plantarum P8对全价饲料pH及微生物类群变化的研究

研究了益生乳酸菌干酪乳杆菌Zhang(Lactobacillus casei Zhang)和植物乳杆菌P8(Lactobacillus planta-rum P8)对全价饲料pH及微生物类群变化的影响。分别将L.casei Zhang、L.plantarum P8单一菌种及复合菌种(11)以6.30 lg cfu/g的接种总量发酵全价饲料,测定25℃10 d发酵期间全价饲料pH和微生物类群的变化,应用选择培养基测定发酵饲料中的乳酸菌及杂菌(酵母菌、霉菌、大肠菌群、芽胞杆菌和梭状芽胞杆菌)的动态变化,应用RT-PCR技术测定试验组中的L.casei Zhang和L.plantarum P8的动态变化。结果显示,试验组pH下降显著,发酵10 d时,L.casei Zhang、L.plantarum P8单一菌种和复合菌种发酵饲料的pH分别为4.23、4.24和4.22,显著低于对照组(P0.05);L.casei Zhang、L.plantarum P8单一菌种和复合菌种发酵饲料中的L.casei Zhang、L.plantarum P8活菌数分别为8.91、8.89、6.58和8.69 lg cfu/g。发酵期间,试验组中酵母菌、霉菌、大肠菌群、芽胞杆菌及梭状芽胞杆菌活菌数显著低于对照组(P0.05),其中L.plantarum P8单一菌种发酵和复合菌种发酵对杂菌抑制效果显著优于L.casei Zhang单一菌种发酵(P0.05)。结果表明,全价饲料经L.casei Zhang、L.plantarum P8发酵可以显著降低其pH,抑制其中杂菌的生长,同时L.casei Zhang、L.plantarum P8在饲料中具有良好的稳定性。     

Preservation of Phakopsora pachyrhizi Uredospores

This study compared different temperatures and dormancy‐reversion procedures for preservation of Phakopsora pachyrhizi uredospores. The storage temperatures tested were room temperature, 5°C, ?20°C and ?80°C. Dehydrated and non‐dehydrated uredospores were used, and evaluations for germination (%) and infectivity (no. of lesions/cm²) were made with fresh harvested spores and after 15, 29, 76, 154 and 231 days of storage. The dormancy‐reversion procedures evaluated were thermal shock (40°C/5 min) followed or not by hydration (moist chamber/24 h). Uredospores stored at room temperature were viable only up to a month of storage, regardless of their hydration condition. Survival of uredospores increased with storage at lower temperatures. Dehydration of uredospores prior to storage increased their viability, mainly for uredospores stored at 5°C, ?20°C and ?80°C. At 5°C and ?20°C, dehydrated uredospores showed increases in viability of at least 47 and 127 days, respectively, compared to non‐dehydrated spores. Uredospore germination and infectivity after storage for 231 days (7.7 months), could only be observed at ?80°C, for both hydration conditions. At this storage temperature, dehydrated and non‐dehydrated uredospores exhibited 56 and 28% of germination at the end of the experiment, respectively. Storage at ?80°C also maintained uredospore infectivity, based upon levels of infection frequency, for both hydration conditions. Among the dormancy‐reversion treatments applied to spores stored at ?80°C, those involving hydration allowed recoveries of 85 to 92% of the initial germination.     

Bioremediation of olive oil mill wastewater and production of microbial biomass

Olive oil mill wastewater (OMWW) was used as a substrate for the culture of a mixture of edible fungi in order to obtain a potentially useful microbial biomass and to induce a partial bioremediation of this fastidious waste. Before fermentation, the OMWW underwent an alkaline-oxidative treatment with the aim of decreasing the polyphenolic content which is the main cause of its toxicity. The fungal mixture grew fairly well in the treated OMWW and reached a maximum of biomass production within about 14 days of fermentation at room temperature. Up to 150–160 g of wet biomass was obtained per liter of OMWW. Analysis of the partially dehydrated biomass revealed a protein content of about 13 g% and 6 g% of row fiber. A relevant presence of unsaturated fatty acids was found, as well as the presence of significant amounts of vitamins A and E, nicotinic acid, calcium, potassium and iron. The possibility of using the microbial biomass produced from OMWW as an additive to animal feed is discussed.     

Preserving young‐of‐the‐year Perca fluviatilis in ethanol,formalin, or in a frozen state and the consequences for measuring morphometrics

As it is often not possible to immediately analyse individuals sampled in the field, captured fish are preserved and stored for later investigation. The objective of this study was to assess the effects of various preservation methods on subsequent changes in morphometric parameters while also providing correction factors to re‐calculate the original body dimensions when sampled fish are measured at a later date. In this study, juvenile perch (Perca fluviatilis, 66.5 ± 10.2 mm total length) were measured directly after capture, then either frozen at ?20°C, preserved in 70% ethanol, or in 4% formalin. They were again measured after 24 h, 3 days, 7 days, and thereafter on a weekly basis for 8 weeks. Ethanol‐preserved perch greatly decreased length and weight; formalin preservation also led to a comparable length reduction, but increased the weight of the perch. In contrast, frozen perch showed less shrinkage and only moderate weight loss. Of the three preservation methods, freezing clearly caused the fewest distortions. Hence, freezing is recommend as the most preferable preservation method, especially in multi‐disciplinary studies on fish ecology; for all other preservation methods the correction factor over time will have to be pre‐determined for each species and size class.     

大肠杆菌DH5α菌株感受态制备及转化率变化研究

通过氯化钙法制备大肠杆菌DH5α菌株感受态,讨论了不同保存温度和保存时间对感受态转化率的影响。结果表明,在4℃下保存,8h达到最高转化率;在-20℃和-70℃下保存,均为48h达到最高转化率。通过氯化钙法制备的DH5α菌株感受态细胞,在-20℃条件下简单保存,20d内完全可以满足一般转化研究的要求,不需要复杂的甘油、液氮处理及超低温要求。     

A Polymethyl Methacrylate Method for Large Specimens of Mineralized Bone with Implants

A simple modified polymethyl methacrylate method is described for large mineralized bone specimens with implants and bioactive materials which produces consistently good histological preservation of the interface between bone and implant. Human femoral heads, whole rabbit condyles and canine tibias and femurs containing implants consisting of hydroxyapatite, smooth polyethylene, porous polyethylene and carbon were dehydrated in ascending grades of ethanol and cleared with xylene on an automated tissue processor which alternated vacuum and pressure for 22 hr. Infiltration was done with washed polymethyl methacrylate at 4 C under vacuum for 13 days. Polymerization was carried out in wide-mouth glass jars at 38 C for 36 hr so that the total processing time was less than 20 days. The only important modification was in the polymethyl methacrylate, which had less plasticizer than usual in order to give a harder block. This enabled production of 4 μm sections with good preservation of mineralized and cellular areas for the study of metabolic bone diseases, morphometry, fluorochrome labelling and interface analysis with the implant in situ.     

阿里山潜蝇茧蜂的羽化率及死亡率与储存温度的拟合关系

本文通过室内模拟低温,研究了阿里山潜蝇茧蜂雌成蜂、9日龄蛹的致死中温度和线性拟合。结果表明,在不同温度下雌蜂的砜分别为4℃下6．6149天,7℃下8．1235天,100℃下9．6161天,13℃下5．0257天,且在所有温度下都有死亡率陡升过程。9日龄蛹在不同温度下的LT50分别为4℃下8．4172天,7℃下8．1235天,10℃下13．9949天,13℃下18．4107天;同一温度下的蛹羽化率与保存天数的关系适宜用二次方程拟合。研究认为蛹的耐寒性强于雌成蜂,可以作为低温保存的虫态。     

Liposome dehydration on nitrocellulose and its application in a biotin immunoassay.

The feasibility of utilizing dehydrated liposomes in the development of a simple immunoassay device for point-of-care diagnostics or field assays was demonstrated. The recovery of liposomes after a cycle of dehydration and rehydration was studied using biotin-tagged, dye-loaded liposomes with antibiotin antibodies immobilized in a defined zone on nitrocellulose strips. Liposomes were vacuum-dehydrated on the strip at a location below the antibiotin zone. The strip was placed in a tube containing a carrier solution and capillary action brought the solution to the dehydrated liposomes, rehydrated them, and caused them to migrate to the antibody zone where intact liposomes were captured and measured optically. High concentrations of either trehalose or sucrose external to the liposomes and both polyvinylpyrrolidone and gelatin in the membrane blocking reagent were essential for preservation of the dehydrated/rehydrated liposomes on nitrocellulose. Between 70 and 80% of the liposomes were recovered on the nitrocellulose strips after a cycle of dehydration and rehydration. The dehydrated liposomes on the strips were stable for at least 1 year when stored in vacuum-sealed plastic bags at 4 degrees C. The technique was successfully applied to the development of a rapid one-step strip immunoassay for biotin.