产不同细菌素的格氏乳杆菌对小鼠肠道微生态的影响

目前已发现格氏乳杆菌（Lactobacillus gasseri,L. gasseri）产生的多种细菌素对多种致病菌具有抑制作用,但有关细菌素对格氏乳杆菌和宿主肠道微生态之间关系的研究较少。为探究产不同细菌素的格氏乳杆菌及其对小鼠肠道微生态的影响,作者通过细菌素操纵子预测、抑菌试验以及液相色谱－质谱联用法证明了格氏乳杆菌FNMGHLBE6L1、FNMGHLBE20L5、FHNFQ16L5可产不同细菌素,之后通过动物试验探究不同格氏乳杆菌对小鼠肠道微生态的影响。结果表明,3株格氏乳杆菌均对小鼠肠道菌群的多样性及菌群结构产生了显著影响;且格氏乳杆菌显著降低了小鼠结肠中白介素-1β的水平。综上,产不同细菌素的格氏乳杆菌可对宿主肠道菌群产生显著影响,且菌株间的差异与所产细菌素存在相关性。该研究为探究产不同细菌素的格氏乳杆菌与肠道菌群之间的关联提供了依据。     

健康婴儿粪便样本中加氏乳杆菌的分离鉴定及安全性评价

目的:本研究从125例健康婴儿粪便样本中分离鉴定出加氏乳杆菌(Lactobacillus gasseri)并对其安全性和抑菌性能进行评价。方法:本研究以1月龄健康婴儿粪便样本为对象,经过MRS培养基分离纯化、革兰氏染色、生化鉴定和16S rRNA测序技术进行菌种鉴定;随后,对分离得到的加氏乳杆菌利用血琼脂平板进行溶血性评价;利用微量肉汤稀释法对常见抗生素的药物敏性进行测定,并对加氏乳杆菌的安全性进行评价;最后,采用牛津杯法检测对大肠杆菌ATCC 25922和金黄色葡萄球菌ATCC 25923的抑制作用。结果:从125例健康婴儿粪便样本中成功分离并鉴定出15株加氏乳杆菌。溶血性检测结果显示,14株均无溶血性;待测菌株对四环素、万古霉素、利奈唑胺、利福平敏感性较高,对亚胺培南完全敏感,对甲氧苄啶完全耐药;抑菌实验结果显示,有6株菌同时对大肠杆菌和金黄色葡萄球菌具有抑菌效果。结论:本研究从健康婴儿粪便样本中成功分离筛选出3株安全性较高且对大肠杆菌和金黄色葡萄球菌同时具有一定抑制作用的加氏乳杆菌,编号分别为:LGI 6-1、LGI 6-2、LGI 6-3,作为潜在的益生菌资源,为后续深入探究加...     

植物乳杆菌M1-NTG300产细菌素培养条件优化

以植物乳杆菌素的效价作为特征性考察指标,对植物乳杆菌M1-NTG300产细菌素培养条件进行优化。通过响应面分析结合实际值确定培养基最佳组分为:葡萄糖浓度3.5%、蛋白胨浓度3.5%、吐温80浓度0.35%、氯化镁浓度0.20%,此培养条件下植物乳杆菌M1-NTG300所产细菌素的效价为1 269.4IU/mL,细菌素效价提高了19.8%。     

Effects of gassericins A and T, bacteriocins produced by Lactobacillus gasseri, with glycine on custard cream preservation

Lactobacillus gasseri LA39 and LA158 isolated from human-infant feces produce bacteriocins named gassericins A and T, respectively. Both gassericins have high heat stability (121°C, 10 min), good pH tolerance (pH 2-11), and strong bactericidality against many gram-positive bacteria, especially lactic acid bacteria, and thus are expected to be effective food preservatives. A microwell plate assay against 12 strains of custard cream spoilage bacteria showed that the gassericins had broader antibacterial spectra than nisin A. Although the gassericins allowed gram-negative isolates to grow, they successfully inhibited the growth of all tested bacterial strains in microwells with the addition of glycine. Glycine was bacteriostatic against many strains except lactic acid bacteria. For practical use, gassericin A was efficiently produced by cultivation in a food-grade medium improved using cheese whey, nourishing proteose peptone, and surfactant yolk lecithin. The practical preservative effect of gassericin A and glycine was verified from the viability of 4 isolated strains, Bacillus cereus, Lactococcus lactis ssp. lactis, Achromobacter denitrificans, and Pseudomonas fluorescens, in custard creams. Custard cream containing 123 arbitrary units of gassericin A per milliliter entirely growth-inhibited the 2 gram-positive strains. In custard cream containing an insufficient amount of gassericin A (49 arbitrary units/mL), the gram-positive strains gradually grew but were completely inhibited by the addition of 0.5% (wt/wt) glycine. The 2 gram-negative strains did not multiply even in the additive-free custard cream, probably because of the unsuitable growth environment. This is the first report showing the combined effect of bacteriocin and glycine and their application for food preservation, which may be helpful for future use in the food industry.     

罗伊氏乳杆菌产罗伊氏细菌素的工艺优化

罗伊氏乳杆菌(Lactobacillus reuteri)是一种安全、高效生产罗伊氏细菌素的菌株。作者优化了罗伊氏乳杆菌ATCC 53608转化甘油生产罗伊氏细菌素的工艺参数。结果表明:处于初期稳定期(26 h)的L. reuteri,其细胞内甘油脱水酶能达到最大酶活2.2 U/mg,有利于转化甘油生产罗伊氏细菌素。正交实验优化后的最佳生产工艺:甘油浓度400 mmol/L,接种量110 mg/mL,酶转化时间2 h,转化温度30℃,pH 6.2。优化后罗伊氏细菌素的平均产量达到(241.2±3.4) mmol/L。     

冻干保护剂对植物乳杆菌的代谢特性的影响

通过分析植物乳杆菌Lactobacillus plantarum CGMCC 6016(L. P. dy-1)冻干菌粉发酵后胞内游离氨基酸、有机酸、菌体蛋白的谱图变化,对比研究了不同冻干保护剂对L. P. dy-1冻干菌粉代谢特征的影响。结果显示,直接活化组L. P. dy-1的胞内氨基酸总量为625.23 μg/100 μL,10%菊粉保护剂组有所增加,为753.72 μg/100 μL;22.4%菊粉保护剂组、菊粉复合保护剂组和脱脂乳复合保护剂组则相对减少;直接活化组L. P. dy-1的胞内有机酸总量为62.84 mg/L,冻干保护剂组均显著降低,其中菊粉复合保护剂组与直接活化组最为接近,为52.99 mg/L;不同菊粉保护剂组冻干菌粉发酵后,菌体蛋白的谱图较为相近,且与直接活化组差异不大,脱脂乳复合保护剂组较未冻干菌体蛋白谱差异明显,表现为有较多条带的缺失,且谱带较浅。结果表明,与常用的脱脂乳复合保护剂相比,菊粉保护剂,尤其是菊粉复合保护剂的添加,能够使植物乳杆菌冻干菌粉的代谢特性更接近正常组。     

干酪乳杆菌16发酵豆乳工艺优化及抗氧化性研究

采用干酪乳杆菌(Lactobacillus casei)16对豆乳进行发酵,探究发酵时间、发酵温度、冷藏时间对L.casei 16发酵豆乳的pH、总酚含量、游离氨基酸含量、物性指标的影响,并研究其抗氧化能力。结果表明,最佳发酵条件为L.casei 16以2%接种量接种于豆乳,37℃发酵24h,发酵结束后4℃冷藏6h,该条件下发酵豆乳的总酚含量为13.94mg/100mL,游离氨基酸含量为1.33mg/mL,稠度达2 192.32g·s,总还原力和羟自由基清除能力分别为0.31和79.43%,显著高于未发酵豆乳和未优化发酵条件的豆乳(P<0.05)。     

Bacteriocin production by Lactobacillus pentosus B96 can be expressed as a function of temperature and NaCl concentration

Lactobacillus pentosus B96 is a bacteriocin-producing strain that was isolated from fermenting olive brines. The aim of the present work was the optimization of bacteriocin production, using response surface methodology (RS). A two-level screening Plackett–Burman design was used to select influencing factors. Then, a central composite design, with three repetitions in the centre, for pH, NaCl concentration, and temperature was carried out. Finally, an RS, which included the region of maximum accumulated bioactivity, was built as a function of NaCl concentration and temperature. Bioactivity accumulation was always observed during the exponential growth-phase, although no apparent correlation between maximum accumulated bioactivity and biomass formation was found. L. pentosus B96 is known to grow better at about 30 °C, neutral pH, and by the absence of NaCl; however, a suboptimal temperature (22 °C) and a moderate NaCl stress (0.65 mol l⁻¹) stimulated bacteriocin production. The research led to environmental conditions that maximized bacteriocin activity, which can be expressed as a polynomial function of temperature and NaCl concentration. The suboptimal growth conditions, which were found to produce the highest bacteriocin titres, resembled those prevailing during green table olive fermentation. This model can be used to improve “in situ” bacteriocin production thus contributing to the microbiological control of the process.     

Purification, partial characterisation and mode of action of enterococcin EFS2, an antilisterial bacteriocin produced by a strain of Enterococcus faecalis isolated from a cheese

Enterococcus faecalis strain EFS2, isolated from the surface of a traditional cheese, produced a bacteriocin active against Gram-positive bacteria including Listeria spp. and some Staphylococcus aureus strains. The bacteriocin, named enterococcin EFS2, has been purified to homogeneity by ammonium sulphate precipitation and reversed-phase high performance liquid chromatography (RP-HPLC). The molecular weight was determined by mass spectrometry to be 7149.6. The amino acid composition of enterococcin EFS2 revealed that it contained 67 amino acid residues and had a blocked amino-terminal end. Enterococcin EFS2 induced viability loss, efflux of K⁺ ions and ATP, and cell lysis. Kinetic study of bactericidal activity of enterococcin EFS2 on Listeria innocua strain LIN 11 indicated slower cell destruction than by nisin. At pH 7.0, the activity of enterococcin EFS2 was the highest at 35 °C and was lost at 15 °C. The bacteriocin was more active against L. innocua strain LIN11 in broth adjusted to pH 6.0, 7.0 and 8.0 than to pH 4.5 at 30 °C.     

一株抑制轮枝镰孢菌的鼠李糖乳杆菌的筛选及其在玉米防霉中应用

本研究以轮枝镰孢菌JF-5-1(Fusarium verticillium)为指示菌,对实验室保存的132株乳酸菌进行抑菌筛查,并对该菌株产生的抑霉菌物质进行了初步探究。结果表明,17株乳酸菌展现了良好的抑制轮枝镰孢菌特性,对轮枝镰孢菌具有较强抑制作用的为91号菌株,经过16S rDNA鉴定其为鼠李糖乳杆菌(Lactobacillus rhamnosus)。菌株上清液对木瓜蛋白酶和胰蛋白酶不敏感,且热稳定性较好,热处理并未改变抑菌性。当pH=5时,抑菌率急速下降为25.0%,当pH大于等于5.5时,91号菌株抑菌活性全部丧失。故初步推测该抑菌物质可能为有机酸,其最小抑菌浓度为8.0 mg/mL。本研究筛选出来的乳酸菌应用在玉米等粮食防霉中,其可抑制轮枝镰孢菌对粮食的侵染且绿色安全,具有非常广阔的应用前景。     

Biochemical,antimicrobial and molecular characterization of a noncytotoxic bacteriocin produced by Lactobacillus plantarum ST71KS

Lactobacillus (Lb.) plantarum ST71KS was isolated from homemade goat feta cheese and identified using biochemical and molecular biology techniques. As shown by Tricine-SDS-PAGE, this lactic acid bacterium produces a bacteriocin (ST71KS) with an estimated molecular weight of 5.0 kDa. Bacteriocin ST71KS was not affected by the presence of α-amylase, catalase and remained stable in a wide range of pH and after treatment with Triton X-100, Triton X-114, Tween 20, Tween 80, NaCl, SDS, urea and EDTA. This bacteriocin also remained active after being heated at 100 °C for 2 h and even after 20 min at 121 °C; however, it was inactivated by proteolitic enzymes. Production of bacteriocin ST71KS reached 6400 AU/mL during stationary growth phase of Lb. plantarum cultivated in MRS at 30 °C and 37 °C. Bacteriocin ST71KS displayed a bactericidal effect against Listeria monocytogenes strains 603 and 607 and did not adsorb to the producer cells. Lb. plantarum ST71KS harbors two bacteriocin genes with homology to plantaricin S and pediocin PA-1. These characteristics indicate that bacteriocin ST71KS is a class IIa bacteriocin. The peptide presented no toxic effect when tested in vitro with kidney Vero cells, indicating safe technological application to control L. monocytogenes in foods.     

低聚半乳糖对植物乳杆菌发酵乳特性及抗蜡样芽孢杆菌活性的改善

为探究低聚半乳糖对植物乳杆菌发酵乳特性及抗菌活性的影响,本文采用单因素法考察影响发酵乳特性的主要因素,并以响应面法优化发酵乳最佳发酵条件;以产肠毒素蜡样芽孢杆菌HN001为指示菌,探究低聚半乳糖的添加对植物乳杆菌ZDY2013发酵乳抑菌活性的改善作用。结果表明:植物乳杆菌能有效利用低聚半乳糖进行体外代谢,并抑制蜡样芽孢杆菌生长;牛奶中添加适量低聚半乳糖能够增加植物乳杆菌发酵乳中的活菌数、降低发酵乳的pH,并提高其持水力;响应面分析发现低聚半乳糖发酵乳的最佳制备条件为:2.0%的植物乳杆菌接种量、1.0%的低聚半乳糖添加量、发酵时间为24 h及发酵温度为42 ℃;添加低聚半乳糖的发酵乳能有效控制产肠毒素蜡样芽孢杆菌浓度在106 CFU/mL以下。该研究结果为低聚半乳糖及植物乳杆菌ZDY2013在发酵乳中的应用奠定了理论基础。     

添加红枣汁对植物乳杆菌发酵胡萝卜品质的影响

为改善胡萝卜发酵制品的品质,本研究以植物乳杆菌为发酵菌株,探讨添加红枣汁对植物乳杆菌发酵胡萝卜(37 ℃发酵7 d)过程中理化指标和营养成分的影响。结果表明,与对照和未添加红枣汁发酵的胡萝卜相比,红枣汁发酵组的抗坏血酸和胡萝卜素含量显著升高,在发酵7 d后分别升高了8.48 mg/100 g(P<0.01)和1.29 mg/100 g(P<0.05)。红枣汁发酵组的总酚、总酸含量和活菌数显著高于对照和未添加红枣汁组(P<0.01)。红枣汁发酵组的多糖和亚硝酸盐含量降低,但与对照和未添加红枣汁组没有显著差异(P>0.05),同时,3组胡萝卜的质构特性、色差值和pH没有显著性差异(P>0.05)。上述结果显示,添加红枣汁对植物乳杆菌发酵胡萝卜的品质有一定程度的改善。     

Inhibition of Listeria monocytogenes by bovicin HC5, a bacteriocin produced by Streptococcus bovis HC5

Cattle can be infected with Listeria monocytogenes by consuming contaminated plant materials, soil or silage, and farmers have sought ways of preventing this contamination. Recent work indicated that Streptococcus bovis HC5 produced a bacteriocin (bovicin HC5) that could inhibit a variety of gram-positive bacteria, and we examined the ability of bovicin HC5 to inhibit 10 strains of L. monocytogenes that had been isolated from plant materials, soil, silage and infected cattle. Growth experiments indicated that all of the L. monocytogenes strains were inhibited by 100 activity units (AU) of bovicin HC5 ml(-1). L. monocytogenes cultures that were transferred with sublethal doses (12.5 AU ml(-1)) could be adapted in stepwise fashion to higher doses of bovicin HC5. However, even 'adapted' cultures did not grow if 400 AU ml(-1) was added. The effect of bovicin HC5 on L. monocytogenes was bactericidal, and viability decreased 5-7 logs after only 2 h of exposure. Bovicin HC5 caused a nearly complete efflux of intracellular potassium in 15 min but only if the pH was less than 6.0. When the pH was greater than 6.0, the cells maintained their potassium pool. L. monocytogenes cells that were acid-adapted (final pH of 4.6) were as sensitive to bovicin HC5 as those that were not acid-adapted (final pH of 6.3). These results support the idea that bovicin HC5 could be effective in controlling listeria in contaminated silages.     

Flavour compound production by Yarrowia lipolytica, Saccharomyces cerevisiae and Debaryomyces hansenii in a cheese-surface model

A simple cheese model mimicking a cheese surface was developed for the detection of cheese flavour formation of yeasts. A total of 56 flavour compounds were detected by dynamic headspace sampling followed by gas chromatography–mass spectrometry analysis. Yarrowia lipolytica CBS 2075 primarily produced sulphides, furans and short-chain ketones; Saccharomyces cerevisiae D7 primarily produced esters and Debaryomyces hansenii D18335 primarily produced branched-chain aldehydes and alcohols. For several of the detected flavour compounds, an increase in production was observed upon exposure to dairy-relevant environmental stress conditions including high NaCl concentration and low temperature. The predominant yeasts on the cheese surface may be important for development of flavour, and thus the use of yeasts as ripening cultures has the potential to affect the flavour of cheese.     

Characterization and purification of a bacteriocin produced by a potential probiotic culture, Lactobacillus acidophilus 30SC

Lactobacillus acidophilus 30SC was tested for its potential as a probiotic culture. The strain exhibited good acid tolerance in an artificial gastric solution as well as bile resistance in media containing 0.3% bile acids. The strain produced a heat-stable antimicrobial compound that was shown to be proteinaceous in nature and, therefore, referred to as a bacteriocin. The bacteriocin was active over a wide pH range and inhibited a number of gram-positive bacteria including Listeria ivanovii and pathogenic strains. The bacteriocin was purified by 50% ammonium sulfate precipitation followed by hydrophobic interaction column chromatography. The SDS-PAGE of the active fractions resulted in a single band with estimated molecular mass of 3.5 kDa. These results demonstrate the potential of L. acidophilus 30SC as a probiotic culture that can be utilized in the manufacturing of dairy foods and dietary supplements.     

小叶苦丁枝条提取物的植物乳杆菌发酵代谢组分析

小叶苦丁枝条是小叶苦丁茶生产的废弃物,数量巨大,一般以焚烧的方式处理,造成了不可忽视的环境污染和资源浪费。为实现小叶苦丁枝条资源化利用,本课题引入益生菌植物乳杆菌对小叶苦丁枝条水提物进行发酵,采取LC-MS非靶向代谢组研究手段探讨了小叶苦丁枝条提取物发酵前后物质的构成、分布与变化。通过PCA、PLS-DA等多元统计方法分析发现,小叶苦丁枝条提取物经植物乳杆菌发酵前后存在显著差异的代谢物共175种,其中发酵后上调的100种,下调的75种。单变量统计排名前25的差异代谢物中包含肽类 3种,碳水化合物1种,核酸类4种,脂类1种,有机酸1种,黄酮1种,其他14种,这些差异代谢物涉及11条差异显著的代谢途径,其中6条为氨基酸代谢途径。差异代谢物中,发酵后相对百分含量分别提高915倍的川皮苷、11倍的鹅肌肽等具有抗氧化、抗炎、抗癌、降血糖等生理活性。同时抗生素庆大霉素 C1a、林卡霉素发酵后分别提升22 672倍和62倍。由此表明,采用乳酸菌发酵技术资源化利用小叶苦丁枝条具有一定的可行性,但还需进一步的安全评估。     

重离子束辐照选育高产细菌素植物乳杆菌

采用重离子束12C6+对产细菌素植物乳杆菌Lp1进行辐照选育,分析比较了不同辐照剂量下致死率和突变率,并通过双层琼脂扩散法筛选高产细菌素突变菌株.在辐照剂量300 Gy时,出发菌株植物乳杆菌Lp1的致死率和正突变率分别为86.3％和28.28％.在300 Gy辐照菌悬液中,分离筛选出高产细菌素突变菌株Lp092、Lp0...     

Protection of Lactobacillus acidophilus from the low pH of a model gastric juice by incorporation in a W/O/W emulsion

The protective effect of a W/O/W emulsion for Lactobacillus acidophilus from a model gastric juice was investigated in order to develop a method for utilizing the advantages of the probiotics. The bacteria were included in the inner-phase solution of the W/O/W emulsion, and a method for counting the viable-bacteria included in the W/O/W emulsion was developed. The relative viability of the bacteria included in the W/O/W emulsion was 49% at 2 h in the model gastric juice, whereas the viability of the bacteria directly dispersed in the juice declined to 1.3% even at 0.67 h. The relative viabilities of the encapsulated bacteria in the model gastric juice at 2 h were 0.12 and 1.10 for the emulsions having the median diameters of 11.9 and 25.4 μm, prepared with inner-phase volume ratios at 0.03 and 0.45, respectively. The relative viabilities of the bacteria in the W/O/W emulsions with the median diameters of 11 and 27 μm, prepared at the homogenization speed of 2.2×10⁴ and 9.8×10³ rpm, were 0.4 and 0.8, respectively. These results suggested that both the inner-phase volume ratio and the median diameter of oil droplet affected the relative viability of the included bacteria.