人皮肤菌群分离株与成纤维细胞培养

为研究微生物群(微种群)与宿主的关系,我们在人成纤维细胞的培养物中,定量的加入从皮肤上分离的常住菌和其它菌株的纯培养物进行混合培养,设立不加菌液的对照组,发现加入疮疤丙酸杆菌(A_(14-1)、表皮葡萄球菌(F_(65))等常住菌,以及从皮肤上分离的干酪乳杆菌(ad_3株)、青春双歧杆菌(Bf_1),有促进人成纤维细胞生长,延缓其老化等作用。而加入从临床标本中分离的脆弱美杆菌(A104株)及金黄色葡萄球菌(189株),则出现培养的人成纤维细胞脱核、裂解和死亡等毒害作用。我们的实验表明:皮肤正常菌群对宿主的皮肤有生理性的滋助作用。     

枯草芽孢杆菌活菌体外拮抗6种肠道致病菌的研究

对枯草芽孢杆菌BS 3株在体外对 6种常见肠道致病菌 (肠产毒性大肠杆菌、肠侵袭性大肠杆菌、肠致病性大肠杆菌、鼠伤寒沙门菌、福氏志贺菌和宋内志贺菌 )的拮抗作用进行了研究。结果表明 ,枯草芽孢杆菌BS 3株对宋内志贺菌、肠致病性大肠杆菌及肠产毒性大肠杆菌拮抗作用较为明显。     

葡萄球菌自身菌苗治疗感染性痤疮疗效观察

痤疮是发生于毛囊皮脂腺的慢性炎症性疾病，当继发细菌感染，出现脓疱红肿、疼痛明显，严重易形成萎缩性或增生性瘢痕，具有损容性。有研究表明在感染性痤疮的皮损中，金黄色葡萄球菌为主要菌种，还可以分离到表皮葡萄球菌、腐生葡萄球菌等，金黄色葡萄球菌为主要病原菌，而痤疮丙酸杆菌、表皮葡萄球菌为皮肤的常住菌，为条件致病菌。     

一株功能性益生菌FQ（15）生物学特性的研究

目的研究一株新分离的功能性益生菌FQ15的生物学特性。方法通过药敏试验研究FQ15对21种抗生素的耐药性;同时,利用耐酸、耐受胆盐等抗逆性实验确定其是否具备功能性的基本要求,最后采用平板抑菌实验研究FQ15对8种常见致病菌是否具有抑菌效果,并通过混合培养方法,确定其对致病性大肠埃希菌及金黄色葡萄球菌的拮抗作用。结果FQ15对7种常见的抗生素有耐药性,在pH不低于3.0的环境下能够正常的生长,能够耐受0.1%-0.7%浓度的胆盐,对8种常见致病菌均具有很好的抑菌效果,并且在与致病菌混合培养中36 h内可以彻底清除金黄色葡萄球菌,48 h内可以完全消灭大肠埃希菌。结论新分离的功能性益生菌FQ15具有良好的生物学特性,而且对临床常见的病原菌具有较强的体外抑制作用。     

双歧杆菌的粘附特性及其对肠道致病菌的体外拮抗作用

本文采用体外细胞培养法, 从实验室现有的20株不同生境来源的双歧杆菌中筛选具有较强粘附能力的菌株, 并通过混合培养和牛津杯方法, 研究了具有粘附特性的双歧杆菌对肠道致病菌的体外拮抗作用。结果显示, 长寿老人源菌株A03和I06的粘附能力最强, 其菌液对金黄色葡萄球菌及大肠杆菌均具有显著的抑制性, 但是菌体及中和后的发酵液均没有抑菌性, 说明受试菌主要通过其代谢产物中的有机酸来发挥其抑菌性能; 此外, 通过分析比较受试菌和肠道致病菌分别与Caco-2细胞粘附后释放的乳酸脱氢酶量, 证实双歧杆菌与致病菌对细胞的作用具有本质上的区别, 双歧杆菌的粘附能减缓致病菌对细胞所造成的损害。     

乳酸菌代谢产物对常见致病菌的体外抑菌试验

为了探讨乳杆菌和乳酶生菌的制菌机理,笔者用其代谢产物对致病菌作了体外抑菌试验。一、试验用菌种和培养基:乳杆菌是经鉴定的嗜酸乳杆菌,乳酪生菌是桂林制药厂生产的乳酶生片经纯化获得。致病菌是从鸡葡萄球菌病分离的金     

地衣芽胞杆菌对白色念珠菌等的拮抗作用

目的了解地衣芽胞杆菌在试管内与阴道正常菌群共生关系的情况。方法将地衣芽胞杆菌菌液分别与葡萄球菌、大肠埃希菌、白色念珠菌、德氏乳杆菌混合培养,定量计数各菌在不同时间内单独培养和混合培养时各菌的活菌数。结果地衣芽胞杆菌生长不受金黄色葡萄球菌、白色念珠菌和大肠埃希菌的影响,金黄色葡萄球菌和白色念珠菌在有地衣芽胞杆菌存在的情况下,其生长受到明显的抑制（P〈0.05）;乳杆菌在12-48 h内,有显著的抑制地衣芽胞杆菌生长的作用,而乳杆菌的生长不受地衣芽胞杆菌的存在与否而正常生长。结论地衣芽胞杆菌对金黄色葡萄球菌及白色念珠菌在体外具有明显的拮抗作用,地衣芽胞杆菌对大肠埃希菌、乳杆菌无明显的体外拮抗作用。     

枯草杆菌BS_(224)菌体外拮抗试验

为了探讨枯草杆菌BS_(224)菌对烧伤创面感染的几种常见致病菌的拮抗效应,我们分别将BS_(224)菌与金黄色葡萄球菌、绿脓杆菌及大肠杆菌的对数生长期的菌,按等量比例混合于肉汤管内,经28℃不同时间的培养和菌落计数后进行统计分析。结果显示BS_(224)菌对金黄色葡萄球菌、绿脓杆菌及大肠杆菌在体外均有极显著拮抗作用。     

侧孢短芽孢杆菌AMCC 100017的分离鉴定及其生防潜力

【目的】从土壤中分离、筛选和鉴定具有抑制病原真菌活性等生防效果的菌株,为进一步开发利用具有良好定殖能力的生防菌株奠定基础。【方法】利用对峙试验筛选拮抗菌并评价其拮抗性能,根据其形态特征、生理生化特性、16S r RNA基因序列测定及Gen Bank序列相似性分析进行分离菌株的分类鉴定,并通过福林酚法测定该菌株的蛋白酶活力。【结果】从山东泰安各种类型土壤中分离得到侧孢短芽孢杆菌(Brevibacillus laterosporus),保藏编号为AMCC100017。平板对峙试验结果表明该菌株对多种植物病原真菌均有较强的拮抗作用,尤其是对镰刀菌属致病菌拮抗效果明显。另外,本试验还初步验证该菌能产生较高活性的胞外蛋白酶。【结论】侧孢短芽孢杆菌AMCC 100017在作物真菌病害生物防治方面,有较好的开发和利用潜力,并可望应用于线虫生物防治。     

阴道卷曲乳酸杆菌对阴道致病菌的体外生物拮抗作用

目的研究阴道卷曲乳酸杆菌对阴道致病菌的体外生物拮抗作用。方法将阴道卷曲乳酸杆菌分别与3株阴道致病菌（金黄色葡萄球菌、大肠埃希菌、粪肠球菌）混合接种于GAM液体培养基中进行厌氧培养,通过平板菌落计数法计算阴道致病菌的菌数。结果经混合培养后阴道致病菌的菌数明显下降。结论阴道卷曲乳酸杆菌A7在体外能明显抑制3株肠道致病菌的生长繁殖。     

部分正常菌群分离菌株的超氧化歧化酶活性的测定

为了进一步研究部分正常菌群的生理和病理机制,我们测定了这些菌株的SOD活性和脂质过氧化物(LPO)变化,发现皮肤常住菌的分离株的混合培养时,SOD活性增加,说明它们具有协同作用。此外,对其它菌株SOD活性测定结果可知,类杆菌和双歧杆菌的SOD活性最低,其它菌株也有一定的SOD活性,如乳杆菌的SOD活性为47.47～98.98μg/ml,大肠杆菌的SOD活性为127.12μg/ml肠球菌的SOD活性为172.5μg/ml。     

Microbial colonization of an in vitro model of a tissue engineered human skin equivalent--a novel approach.

This was a preliminary investigation to define the conditions of colonization of a human skin equivalent (SE) model with cutaneous microorganisms. SEs of 24 mm diameter were constructed with a dermal matrix of fibrin containing fibroblasts and a stratified epidermis. Microbial colonization of the SEs was carried out in a dry environment, comparable to 'in vivo' skin, using a blotting technique to remove inoculation fluid. The microbial communities were sampled by scrub washing and viable cells enumerated on selective growth medium. Staphylococcus epidermidis, Propionibacterium acnes and Malassezia furfur (human skin commensals) and Staphylococcus aureus (transient pathogen) were colonized at inoculum densities of 10(2)-10(6) CFU SE(-1) on the surface of replicate SEs. Growth of all species was supported for upto 72-120 h, with recovery densities of between 10(4)-10(9) CFU SE(-1). A novel, real-time growth monitoring method was also developed, using S. aureus containing a lux cassette. Light output increased from 20 to 95 h, and colonization increased from 10(2) to 10(8) CFU SE(-1), as confirmed by conventional recovery. Thus, the SE model has potential to investigate interactions between resident and transient microbial communities with themselves and their habitat, and for testing treatments to control pathogen colonization of human skin.     

Microbial colonization of an in vitro model of a tissue engineered human skin equivalent – a novel approach

This was a preliminary investigation to define the conditions of colonization of a human skin equivalent (SE) model with cutaneous microorganisms. SEs of 24 mm diameter were constructed with a dermal matrix of fibrin containing fibroblasts and a stratified epidermis. Microbial colonization of the SEs was carried out in a dry environment, comparable to ' in vivo ' skin, using a blotting technique to remove inoculation fluid. The microbial communities were sampled by scrub washing and viable cells enumerated on selective growth medium. Staphylococcus epidermidis , Propionibacterium acnes and Malassezia furfur (human skin commensals) and Staphylococcus aureus (transient pathogen) were colonized at inoculum densities of 10²–10⁶ CFU SE⁻¹ on the surface of replicate SEs. Growth of all species was supported for upto 72–120 h, with recovery densities of between 10⁴–10⁹ CFU SE⁻¹. A novel, real-time growth monitoring method was also developed, using S. aureus containing a lux cassette. Light output increased from 20 to 95 h, and colonization increased from 10² to 10⁸ CFU SE⁻¹, as confirmed by conventional recovery. Thus, the SE model has potential to investigate interactions between resident and transient microbial communities with themselves and their habitat, and for testing treatments to control pathogen colonization of human skin.     

Maintenance of the normal flora of human skin grafts transplanted to mice

Full-thickness human cadaver skin was maintained on the dorso-lateral thoracic region of hairless mice whose immune rejection mechanism was suppressed using anti-mouse-thymocyte globulin. The bacterial profile of the pregrafted skin did not differ significantly from the normal human microflora. In contrast, the murine skin exhibited quantitative and qualitative differences from the human flora, in particular by the complete absence of Propionibacterium acnes, the dominant bacterium on sebum-rich areas of human skin. The normal microbial profile of the human grafts was maintained throughout the experimental period despite the novel environmental milieu. There was little contamination of the grafts from the normal murine flora. It was concluded that the grafted human skin would provide a realistic model for studying the ecology of human cutaneous micro-organisms.     

青少年痤疮面部皮肤微生物群落结构变化

【背景】青少年痤疮是一种最常见的慢性炎症性损容性皮肤病,与痤疮丙酸杆菌的异常增殖有关。【目的】探究痤疮皮损区与附近无明显皮损区微生物组成与健康对照的差异,为从微生态角度防治痤疮提供理论基础。【方法】利用细菌16S rRNA基因V1-V2区和真菌TIS1高通量测序技术分析北京地区16岁青少年面部痤疮皮肤细菌和真菌群落结构,将痤疮皮损区与附近无明显皮损区微生物组成与健康组进行比较,寻找差异菌群。【结果】痤疮患者面部皮损区与附近无明显皮损区细菌多样性(Shannon指数)较健康对照组显著性降低(P0.001),主要与丙酸杆菌(痤疮丙酸杆菌)和葡萄球菌(表皮葡萄球菌PM221)显著性上升相关,而痤疮皮损区与附近未明显皮损区细菌组成无显著性差异。痤疮患者皮损区与附近无明显皮损区较健康对照组真菌丰富度(Chao1指数)显著性上升(P0.05),与限制性马拉色菌的显著上升相关。【结论】面部皮肤微生物变化与青少年痤疮的发生相关。本研究为从微生物角度防治痤疮提供理论依据。     

The probiotic potential against vibriosis of the indigenous microflora of rainbow trout

The antibacterial properties of the indigenous microflora of rainbow trout ( Oncorhynchus mykiss Walbaum) and the potential use of inhibitory bacteria as fish probiotics were investigated. A total of 1018 bacteria and yeasts were isolated on tryptone soy agar (TSA) from skin, gills and intestine. Forty-five of these inhibited growth of the fish pathogenic bacterium Vibrio anguillarum in a well diffusion assay. The antagonism was most prominent among Pseudomonas spp., as 28 (66%) of the antagonistic bacteria belonged to this genus, despite constituting only 15% of the total tested flora. As pseudomonads are typically siderophore producers, chrome azurol S (CAS) agar was used as a semi-selective medium for isolation of antagonistic bacteria. On this medium, 75% of the iron-chelating strains were inhibitory to V. anguillarum . Eight strains out of a subset of 11 antagonists caused a 3–6 log unit reduction in the density of V. anguillarum [measured by polymerase chain reaction (PCR) detection in a most probable number (MPN) regimen] in a broth co-culture assay. Survival of rainbow trout infected with vibriosis was improved 13–43% by six out of nine antagonistic strains tested in vivo. All disease-protecting strains were pseudomonads, isolated from CAS plates, whereas two Carnobacterium spp. that were antagonistic in in vitro well diffusion assays did not alter the accumulated mortality of rainbow trout. The addition of live bacterial cultures to fish-rearing water may thus improve survival of the fish; however, in vitro antagonism could not completely predict an in vivo effect. Further studies on the underlying mechanism of activity are required to design appropriate selection criteria for fish probiotic bacteria.     

Friends and foes: streptomycetes as modulators of plant disease and symbiosis

The ecological role of soil streptomycetes within the plant root environment is currently gaining increased attention. This review describes our recent advances in elucidating the complex interactions between streptomycetes, plants, pathogenic and symbiotic microorganisms. Streptomycetes play diverse roles in plant-associated microbial communities. Some act as biocontrol agents, inhibiting plant interactions with pathogenic organisms. Owing to the antagonistic properties of streptomycetes, they exert a selective pressure on soil microbes, which may not always be for plant benefit. Others promote the formation of symbioses between plant roots and microbes, and this is in part due to their direct positive influence on the symbiotic partner, expressed as, e.g., promotion of hyphal elongation of symbiotic fungi. Recently, streptomycetes have been identified as modulators of plant defence. By repressing plant responses to pathogens they facilitate root colonisation with pathogenic fungi. In contrast, other strains induce local and systemic resistance against pathogens or enhance plant growth. In conclusion, while streptomycetes have a clear potential of acting as biocontrol agents, care has to be taken to avoid strains that select for virulent pathogens or enhance disease development. We argue towards the use of an integrated screening approach in the search for efficient biocontrol agents, including assays on in vitro antagonism, plant growth, and disease suppression.     

Antagonistic action of Pseudomonas aeruginosa against Staphylococci, coliform bacilli and various types of Proteus

Antagonistic activity of 2 fresh isolates and 3 collection strains of Pseudomonas aeruginosa against 177 microbial strains was determined with the method of late antagonism. Among the microbial strains there were 56 staphylococcal strains isolated from patents and carriers. 38 nontypable colon bacilli isolated from healthy persons, 59 enteropathogenic colon bacilli of various serogroups, 12 strains of Proteus and 12 colon bacilli, carriers of multiple drug resistance factors (R factors). All the cultures were sensitive to the antagonistic action of 5 or at least 3 strains of Pseudomonas used in the study. The most active antagonists were the fresh isolates of Pseudomonas as compared to the collection strains. Among the staphylococci S. aureus proved to be the most resistant to the antagonistic action of Pseudomonas as compared to S. epidermidis, the same as the strains isolated from carriers as compared to the strains isolated from patients. As for the enteric bacilli the most resistant were the strains of Proteus. Acquiring of transmissive R factors by the colon bacilli markedly increased their sensitivity to the antagonistic action of Pseudomonas.     

Antagonistic Effect of Nonpathogenic Fusarium oxysporum Fo47 and Pseudobactin 358 upon Pathogenic Fusarium oxysporum f. sp. dianthi

Pseudobactin production by Pseudomonas putida WCS358 significantly improves biological control of fusarium wilt caused by nonpathogenic Fusarium oxysporum Fo47b10 (P. Lemanceau, P. A. H. M. Bakker, W. J. de Kogel, C. Alabouvette, and B. Schippers, Appl. Environ. Microbiol. 58:2978-2982, 1992). The antagonistic effect of Fo47b10 and purified pseudobactin 358 was studied by using an in vitro bioassay. This bioassay allows studies on interactions among nonpathogenic F. oxysporum Fo47b10, pathogenic F. oxysporum f. sp. dianthi WCS816, and purified pseudobactin 358, the fluorescent siderophore produced by P. putida WCS358. Both nonpathogenic and pathogenic F. oxysporum reduced each other's growth when grown together. However, in these coinoculation experiments, pathogenic F. oxysporum WCS816 was relatively more inhibited in its growth than nonpathogenic F. oxysporum Fo47b10. The antagonism of nonpathogenic F. oxysporum against pathogenic F. oxysporum strongly depends on the ratio of nonpathogenic to pathogenic F. oxysporum densities: the higher this ratio, the stronger the antagonism. This fungal antagonism appears to be mainly associated with the competition for glucose. Pseudobactin 358 reduced the growth of both F. oxysporum strains, whereas ferric pseudobactin 358 did not; antagonism by pseudobactin 358 was then related to competition for iron. However, the pathogenic F. oxysporum strain was more sensitive to this antagonism than the nonpathogenic strain. Pseudobactin 358 reduced the efficiency of glucose metabolism by the fungi. These results suggest that pseudobactin 358 increases the intensity of the antagonism of nonpathogenic F. oxysporum Fo47b10 against pathogenic F. oxysporum WCS816 by making WCS816 more sensitive to the glucose competition by Fo47b10.     

Characteristics of the Propionibacterium strains isolated from acne patients

Propionibacterium acnes is a component of physiological flora of human skin. It colonizes the outlets of sebaceous glands and participates in the pathogenesis of inflammatory acne. Acne vulgaris is a common skin disease. It is found in more or less exacerbated form in approximately 85% of adolescent population. The main purpose of the research was to confirm the hypothesis of Propionibacterium bacteria participation in the aetiopathogenesis of acne vulgaris. The researches have proved the presence of Propionibacterium acnes on the surface of the skin both of people with acne-related changes and these with whom such changes were not found. Statistically significant differences were found in the number of P. acnes bacteria per 1 square centimeter of healthy and disease-affected skin as well as in the diversity of biochemical types. The highest number of P. acnes bacteria have been found in fresh changes with visible symptoms of inflammation. In order to confirm the hypothesis of the participation of Propionibacterium bacteria in the aetiopathogenesis of acne, a detailed phenotypical analysis of isolated P. acnes strains have been conducted. Type, biotype, resistance pattern, proteolytic and lipolytic properties have been determined.