超声波诱变选育乳链菌肽(Nisin)高产菌株

目的：从乳酸乳球菌中选育Nisin高产菌株。方法：利用超声波对乳酸乳球菌进行诱变,并用琼脂扩散法检测其效价。结果：获得一突变菌株S-1,该菌株的生物效价为343.53IU/mL,比原始菌株提高31.52%。结论：突变株S-1遗传性能稳定,为进一步菌种选育奠定基础。     

梅岭霉素产生菌抗药性突变标志诱变筛选模型的初步研究

本文通过梅岭霉素 (Meilingmycin)产生菌南昌链霉菌NS 4 1 80菌株孢子对 6种抗生素敏感性测定 ,采用诱变剂EMS四种不同诱变剂量对菌株孢子进行诱变处理 ,诱变处理的孢子涂布在含致死浓度链霉素的高氏平板上。然后从抗药性突变标志菌株中进一步筛选梅岭霉素高产菌株。在 150 0多个抗药性突变株中通过初筛获得了比诱变出发菌株的产素能力提高 50 %以上菌株。通过诱变剂量分别与抗药性突变率和突变株产素产量的变势统计分析表明 ,菌株抗药性突变与产素突变密切相关 ,产素突变的EMS诱变剂量高于抗药性突变诱变剂量 ,在 0 .0 3mol/LEMS剂量作用下 ,菌株致死率为 99.4 3% ,抗药性突变率为 0 .0 4 4 0 % ,建立了梅岭霉素产生菌抗药性突变标志诱变推理性筛选模型。为南昌链霉菌高产菌种选育研究作了有益的尝试 ,并有助于其它链霉菌属的抗生素产生菌育种研究。     

捷安肽素高产菌的紫外诱变模型的建立及选育

从新疆棉株上分离得到一株细菌ZK,发酵生产抗病原真菌肽类物质-捷安肽素。以此株菌作为出发菌株,通过紫外线诱变处理获得了高产突变株UV146,在摇瓶试验中,该变异株产捷安肽素的量高于亲株。     

乳链菌肽高产菌株的选育及其基因定位

以乳酸乳酸球菌7962为原始菌株,用紫外线、LiCl、(60)~Co及8-MOP+NUV等多种理化诱变剂对其进行诱变处理,获得一株乳链菌肽高产突变株AL2。其效价稳定在2300～2500Iu/ml。经DNA杂交证实,编码乳链菌肽的前体基因位于染色体上,其遗传性状是稳定的。毒理试验表明AL2及其产物属于实际无毒类物质。     

一株抗G- 菌和酵母菌的乳酸乳球菌的分离鉴定与抗菌活性

以G+ 菌金黄色葡萄球菌(Staphylococcus aureus)作为指示菌, 通过抑菌筛选法从生牛奶中初筛得到具有抑菌活性的14株细菌菌株, 然后通过个体形态与培养特征观测、部分生理生化反应、G + C mol%测定、16S rDNA序列比对分析、PCR扩增特异性N-乙酰胞壁酸水解酶基因和序列对比分析等鉴定, 确定其中的一株具有较高抑菌活性的分离株为乳酸乳球菌乳酸亚种(Lactococcus lactis subsp. lactis)菌株, 命名为MB191。对多种G+ 细菌、G- 细菌、酵母菌和丝状真菌的对峙培养抗性测定结果表明, MB191除对供试G+ 细菌具有较高的抑菌活性以外, 还对丁香假单胞菌(Pseudomonas syringae)、荧光假单胞菌(P. fluorescens)等G- 细菌和汉逊德巴利酵母(Debaryomyces hansenii)等具有明显的抑菌活性。乳酸乳球菌的这一特性目前尚未见文献报道。     

瑞拉菌素产生菌RL-2的诱变育种

本研究以委内瑞拉链霉菌秦岭变种RL-2(Streptomyces venezuelaevar. qinlingensis RL-2)为出发菌株, 分别采用紫外线、氯化锂及紫外线加氯化锂的复合诱变方式对其孢子悬液进行诱变处理。在复合诱变的紫外线照射时间为45 s和氯化锂浓度为0.4%的情况下, 获得一株瑞拉菌素的高产突变菌株UVL-108且连续传接6代其遗传特性较稳定。采用双向培养及生长速率法对其进行初筛和复筛, 结果表明: UVL-108的拮抗性能较出发菌株提高了77%, 其发酵液对稻瘟病菌的抑菌毒力是出     

Cp-S316菌株抗细菌活性物质对大肠杆菌的抑制作用及高产突变株选育

通过测定大肠杆菌K12 (Escherichia coli K12)菌悬液的OD260的变化, 研究了多粘类芽孢杆菌(Paenibacillus polymyxa)Cp-S316抗细菌活性物质对其细胞膜完整性的影响, 结果表明Cp-S316抗细菌活性物质可损伤大肠杆菌K12的细胞膜, 从而引起胞内RNA、DNA等大分子物质的泄漏。为获得抗细菌活性物质高产菌株, 以Cp-S316为出发菌株, 通过紫外诱变以及对自身产生的抗细菌活性物质的抗性筛选法进行预筛、摇瓶初筛和复筛, 获得突变株多粘类芽孢杆菌A17, 其发酵效价比出发菌株Cp-S316提高91%, 该突变株的高产遗传性状稳定。     

南昌霉素高产菌株的链霉素抗性基因突变诱变筛选研究

通过对链霉素对南昌霉素（Nanchangmycin)产生菌NS－41－80菌株孢子的致死浓度测定基础上,采用诱变剂甲基磺酸乙酯（EMS）的不同诱发剂量对菌株孢子进行诱变处理,诱变处理的孢子涂布在含链霉素（10ug/mL)致死浓度的高氏平板上,获得了大量的链霉素抗性基因（str)突变株。然后从3,000株链霉素抗性基因（str)突变株中通过初筛获得比诱变出发菌株产素能力提高20％以上的菌株202株,再进一步通过摇瓶复筛,获得比出发菌株产素能力分别提高100％,200％,300％高产菌株为48株,7株和1株,分别为复筛菌和初筛菌株的23．76％和1．60％,3．46％和0．23％,0．5％和0．03％,将产素能力提高240％以上5个菌株连同出发菌株连续3批次进行摇瓶发酵结果,5个突变株的产素能力均比出发菌株的产素能力提高57％－96．4％,其中突变株80－5．3－165菌株摇瓶发酵单位达6,000ug/mL以上,3批次摇瓶平均发酵单位达5,855ug/mL,建立了南昌霉素高产菌株的链霉素抗性基因突变诱变快速高效的筛选方法。     

利用特比萘芬抗性筛选赤霉素高产菌株及相关发酵特性的研究

【目的】诱变、筛选赤霉素高产菌株,并明确该菌遗传稳定性及部分发酵特性。【方法】利用亚硝基胍、60Co-γ射线以及复合诱变的方法,结合抑真菌剂-特比萘芬抗性筛选,定向选育赤霉素高产突变株;通过琼脂斜面传代实验确定遗传稳定性;发酵罐培养了解其部分发酵性状。【结果】藤仓赤霉菌单细胞悬液先后经过终浓度为300 mg/L亚硝基胍30 min和60Co-γ射线300戈瑞下复合诱变,得到抗120μg/L特比萘芬的赤霉菌突变株。摇瓶复筛,确定赤霉素突变菌株的赤霉素合成能力,其中ZNL13-3菌产赤霉素效价为2 215±35 mg/L,较之诱变前发酵效价平均提高了11.87%。ZNL13-3菌株经连续15次试管斜面传代考察,菌株赤霉素合成的稳定性较好,能维持原发菌株93.2%,具有良好的遗传稳定性和生产应用前景。5 L发酵罐培养,比较菌丝浓度和产物浓度随发酵时间变化趋势,ZNL13-3菌比生产速率优于原发菌株。【结论】赤霉菌抗特比萘芬的变异特性与赤霉素高产之间存在某种对应关系,为进一步定向优化筛选优良菌株提供参考。     

提高展青霉(Penicillium patulum)产灰黄霉素效价的诱变育种研究

为进一步筛选高产灰黄霉素的工业生产菌株,分别对前期采用紫外线-氯化锂（UV-LiCl）、半导体激光（LD laser）及CO2激光（CO2laser）对展青霉FS80-1复合诱变获得三株高产菌株进行液体发酵和固体培养比较。结果表明,通过UV-LiCl复合诱变获得突变菌株GM120-43的液体发酵产灰黄霉素效价11 982μg/mL,比出发菌株提高37.52%,固体培养效价为89 496μg/g（干重）,比出发菌株提高80.04%。;半导体激光诱变获得突变株LD100-1的液体发酵效价9 440μg/mL,固体培养效价119 766μg/g干重,比出发菌株FS80-1提高了140%;两个突变株的生物学特性均发生不同程度的变化,突菌株GM120-43适合于液体发酵生产,突变株LD100-1适合于固体发酵培养。     

Characterization of a bacteriocin-like substance produced by Bacillus amyloliquefaciens isolated from the Brazilian Atlantic forest.

A Bacillus strain producing a bacteriocin-like substance was characterized by biochemical profiling and 16S rDNA sequencing. Phylogenetic analysis indicated that the strain has high sequence similarity with Bacillus amyloliquefaciens. The antimicrobial substance was inhibitory to pathogenic and food-spoilage bacteria, such as Listeria monocytogenes, Bacillus cereus, Serratia marcescens, and Pasteurella haemolytica. It was stable over a wide temperature range, but lost activity when the temperature reached 121 degrees C/15 min. Maximum activity was observed at acidic and neutral pH values, but not at alkaline pH. The antimicrobial substance was sensitive to the proteolytic action of trypsin, papain, proteinase K, and pronase E. Except for iturins, other antimicrobial peptides have not been described for B. amyloliquefaciens. The identification of a bacteriocin-like inhibitory substance active against L. monocytogenes addresses an important aspect of food protection.     

In vitro study on bacteriocin production of Enterococci associated with chickens

In recent years, the approach of using innovative strategies such as probiotics or bacteriocins for the prevention or treatment of bacterial infections has come into focus. The present study was undertaken to check in vitro ability of Enterococci-isolated from the gastrointestinal tract of chickens-to produce a bacteriocin-like substance and to describe some further probiotic properties in five selected Enterococcus faecium strains. All strains (n=17) were found to produce bacteriocin-like substances against 14 out of 20 indicator bacteria of animal, food or environmental origin. Selected E. faecium strains expressed sufficient survival by pH 3.0 after 3h, in the presence of 1% bile after 24h and they were sensitive to most of antimicrobials tested. All tested strains adhere to the human, canine and porcine intestinal mucus (between 1.5% and 9.2%). However, better adhesion ability was observed for the canine mucus. PCR detection of enterocin structural genes determined presence of enterocins A and P genes in all selected strains. Characterization of bacteriocin substance in detail was performed in E. faecium EF55. The EF55 strain produced a bacteriocin-like substance (during the late logarithmic and early stationary growth phase) with inhibitory activity mostly against Gram-positive bacteria (100-51,200 AU/mL) including Listeria monocytogenes. Proteinaceous character of the bacteriocin substance was confirmed (its inhibitory activity was lost after its treatment with proteases), it was found to be stable after heating (100 degrees C 10 min) and during 12 months storage at -20 degrees C. The highest inhibitory activity of bacteriocin produced by EF55 strain (growing in MRS) broth was achieved between pH 7.0 and 9.0.     

Characterization of a broad range antimicrobial substance from Bacillus cereus

AIMS: The aim of this research was to isolate and characterize an antimicrobial substance from the Bacillus cereus type strain ATCC 14579. METHODS AND RESULTS: A substance with antimicrobial activity was isolated from B. cereus ATCC 14579. The substance was produced during late exponential growth and well into the stationary phase with a maximum 9 h after inoculation. The inhibitory substance was purified by reverse-phase HPLC and shown to be highly active against closely related Bacillus spp. Clinically relevant species such as Staphylococcus aureus and Micrococcus luteus were also inhibited. The substance was characterized as a bacteriocin-like inhibitory substance (BLIS) with a molecular mass of ca 3.4 kDa. The BLIS was very heat stable, and sensitive only to pronase E and proteinase K. Antimicrobial activity was stable and high in the pH range of 2.0-9.0, and relatively unaffected by organic chemicals. CONCLUSIONS: An antimicrobial substance produced by the B. cereus type strain ATCC 14579 was characterized, with a wide spectrum of activity and the potential to be applied as a control agent against pathogenic bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study is the first report of a substance with antimicrobial activity from the B. cereus type strain.     

Antibacterial activity among Pseudomonas strains of meat origin

Almost 750 Pseudomonas strains from meat, fish, milk or plant were screened for inhibitory potential towards 10 selected Pseudomonas fragi isolated from meat. Only strains of fish origin and a reference Ps. putida strain produced an inhibitory substance which was either a siderophore or a bacteriocin-like substance. The two systems were efficient in meat extract medium suggesting potential use for controlling meat quality.     

Inhibition of Vibrio parahaemolyticus by a bacteriocin-like inhibitory substance (BLIS) produced by Vibrio mediterranei 1

AIMS: The aim of this research was to identify and partially purify new bacteriocin-like substances from strains of halophilic 'non-cholera' vibrios isolated from food sources. METHODS AND RESULTS: Forty-five halophilic Vibrio spp. strains were screened for antimicrobial production. Vibrio mediterranei 1, a nonpathogenic strain, showed antimicrobial activity towards Vibrio parahaemolyticus spp. and related species. The bacteriocin-like inhibitory substance (BLIS), released by the bacteria into growth media, was concentrated by ultrafiltration and characterized. BLIS was sensitive to proteinase K, was stable in the pH range 5-9, was resistant to organic solvents and was heat stable up to 75 degrees C. Initial purification of BLIS by size exclusion chromatography showed an apparent molecular mass of 63-65 kDa. CONCLUSIONS: This study reports the ability of V. mediterranei 1 to produce a bacteriocin-like substance inhibiting growth of V. parahaemolyticus spp. and other closely related bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: The strong activity of BLIS towards the human and fish pathogen V. parahaemolyticus and the persistence of antimicrobial properties under a variety of different conditions suggest its potential application in food microbiology.     

A Novel Bacteriocin-Like Substance Produced by <Emphasis Type="Italic">Enterococcus faecium</Emphasis> 3587

Secretion of a novel bacteriocin-like inhibitory substance from Enterococcus faecium 3587 is described herein for the first time. Whereas some bacteriocins receive their denomination based on the species or genus name of the producer microorganism, the newly discovered bacteriocin-like substance was named “enterocin 3587.” The growth characteristics of the producer strain, as well as the type of production and the primary characteristic of the peptide, were investigated. It was found by sodium dodecyl sulfate–polyacrylamide gel electrophoresis that the molecule possesses a molecular weight <6.5 kDa; its secretion is growth-phase dependent; and it shows activities only against other closely related enterococci but not against other Gram-positive bacteria, such as L. innocua, S. aureus 209, and Lactobacillus delbrueckii B2, nor against some Gram-negative species, such as Escherichia coli HB101.     

Plasmid DNA in Strains of Pediococcus cerevisiae and Pediococcus pentosaceus

Five parental strains of Pediococcus were examined for plasmid content. Each strain contained three to six resident plasmids, ranging in size from 4.5 to 39.5 megadaltons. A bacteriocin-like substance produced by Pediococcus cerevisiae FBB63 was tentatively linked to a 10.5-megadalton plasmid after being cured with novobiocin.     

Coagulin, a bacteriocin-like inhibitory substance produced by Bacillus coagulans I4

A protease-sensitive antibacterial substance produced by Bacillus coagulans I₄ strain, isolated from cattle faeces, was classified as a bacteriocin-like inhibitory substance and named coagulin. The inhibitory spectrum included B. coagulans and unrelated bacteria such as Enterococcus , Leuconostoc , Oenococcus , Listeria and Pediococcus . Coagulin was stable at 60 °C for 90 min, at a pH ranging from 4 to 8 and appeared to be unaffected by α-amylase, lipase or organic solvents (10% v/v). Coagulin exhibited a bactericidal and a bacteriolytic mode of action against indicator cells. The apparent molecular mass was estimated to be about 3–4 kDa by SDS-PAGE. The B. coagulans I₄ strain harbours a plasmid, pI₄, approximately 14 kb in size. Novobiocin curing experiments yielded two derivatives that no longer produced the bacteriocin-like inhibitory substance. Plasmid content of these two derivatives showed that one had lost pI₄,whereas the second harboured a deleted form of this plasmid, thus suggesting a plasmid location for the genes for coagulin production.     

Partial purification and characterization of the bacteriocin produced by Lactobacillus acidophilus YIT 0154

One strain of Lactobacillus acidophilus was found to produce a bacteriocin-like substance in the culture filtrate. The substance was produced in a growth-associated manner, showed heat stability at neutral and acidic pH and exhibited antibacterial activity against various species of Lactobacillus including L. acidophilus itself. The molecular weight of the substance was in the range of 6.2-95 kDa. N-terminal amino acid sequence analysis suggests that the substance may belong to class IIb bacteriocin.