Isolation of Cellulolytic Actinomycetes from Marine Sediments

The cellulolytic activity of 36 actinomycetes strains isolated from marine sediments was investigated by the cellulose-azure method. Approximately 50% of the isolates exhibited various degrees of cellulolytic activity.     

Study of the Lytic Activities of Actinomycetes Isolated from Different Soils in Georgia

The lytic activities of 310 cultures from the Collection of Actinomycetes of the Institute of Biochemistry and Biotechnology, National Academy of Sciences of Georgia, were studied; 18% of these strains appeared capable of lysing yeast cell wall. The active producer of the enzyme was selected. This culture was isolated from chestnut soil in Gardabani raion (Central Georgia). Its cultural–morphological, biochemical, and antagonistic properties allowed the culture to be ascribed to the species Geodermatophilus obseurusLuedemann, 1968. The maximal lytic activity under submerged cultivation conditions, exceeding the activity of Actinomyces griseinusby twofold, was observed during the logarithmic growth phase.     

油松菌根根际放线菌的分离与鉴定

为了探讨油松菌根根际放线菌与菌根真菌及油松猝倒病的关系,该实验从油松菌根根际土中分离、纯化得到170株放线菌,采用平板对峙培养法对油松猝倒病病原真菌立枯丝核菌(Rhizoctonia solani)和茄腐皮镰孢霉(Fusarium solani)进行生防放线菌的初筛,得到7株对病原真菌抑制效果较好的放线菌.用初筛选的7株放线菌菌株分别与外生菌根真菌灰鹅膏菌(Amanita vatinata)、灰环粘盖牛肝菌(Suillus laricinus)和血红铆钉菇(Gomphidius viscidus)进行纯培养互作复筛,得到2株放线菌(菌株19#、菌株110#)对外生菌根真菌灰鹅膏菌和血红铆钉菇均具有促生作用.将菌根真菌灰鹅膏菌、放线菌菌株19#、病原真菌立枯丝核菌进行纯培养互作试验表明,菌株19#能协同灰鹅膏菌拮抗立枯丝核菌.经形态特征、培养特征、生理生化特性测定和16SrDNA序列分析,确定菌株19#为苯胺紫链霉菌(Streptomyces mauvecolor).     

Isolation and properties of macrotetrolide synthase from the mycelium of Actinomycetes

The formation of cyclic polyester antibiotics (macrotetrolides) from nactinic acids in a cell-free system in the presence of a mycelium homogenate of Streptomyces chryzomallus var. macrotetrolidi, a producer of a complex of homologous macrotetrolide antibiotics, was demonstrated. An enzyme catalyzing the formation of an ester macrotetrolide ring and possessing a specific activity of 360 mumol/min/mg of protein has been isolated for the first time from the mycelium homogenate and purified 176-fold with a 18% yield. Macrotetrolide synthase represents a macromolecular complex with a molecular mass of 360 kDa formed by several heterogeneous polypeptides. The effects of physico-chemical environmental factors on the stability and activity of the enzyme were demonstrated. The optimal conditions for the manifestation of the synthase activity (10 mM Tris-HCl, pH 8.0, 10% ethanol, 2% glycerol, 200 micrograms/ml of nactinic acids) were selected. The kinetic parameters of the enzyme-catalyzed macrotetrolide synthesis reaction (Km = 2.9.10(-4) M, V = 22.3 microM/min) were determined.     

Oligotrophy is Helpful for the Isolation of Bioactive Actinomycetes

It is necessary to develop new methods for the isolation of unknown actinomycetes from soils. To evaluate the effects of oligotrophic medium on the isolation of soil actinomycetes and develop a new isolation method, the Gause’s synthetic medium was diluted to one tenth the recommended concentration in the present study. Soil dilution plate technique was used to isolate actinomycetes from the soil samples. Oligotrophy decreased actinomycete and streptomycete counts, as well as the number of antagonistic actinomycete species. Oligotrophy also decreased the number of actinomycete species in five samples. Some actinomycete species were cultured only on the oligotrophic medium, whereas other species could not be cultured. Oligotrophy decreased actinomycete counts more significantly for soils with organic matter content >40 g/kg. We used 16S rRNA sequence analysis to identify 22 actinomycete species that were only cultured on the oligotrophic medium. Oligotrophic medium was helpful for the isolation of Streptomyces spp., Micromonospora spp. and Streptosporangium spp. Slightly more than 80 % of the identified actinomycete species were biologically active. Therefore, we could draw a conclusion that oligotrophic medium could be helpful for the discovery of new antibiotic producers and the exploitation and utilization of new, biologically active compounds.     

几种选择性分离稀有放线菌的方法

由于从常见放线菌中发现新化合物的几率越来越小,人们开始将目光集中于稀有放线菌。作者介绍了近年来出现的胞外多糖胶与动孢溶液相结合的分离方法、再水化-离心法、极高频辐射法、噬菌体定向分离法和蔗糖梯度离心法等用于稀有放线菌选择性分离的方法,以及这些稀有放线菌在产生生物活性物质方面的潜力。     

Selective Isolation of Aerobic Actinomycetes

The composition of an arginine-glycerol-salt medium (AGS), suitable for the selective isolation of aerobic actinomycetes, was given. When soil samples were treated with calcium carbonate and plated on the AGS medium, higher total and relative plate counts of actinomycetes were obtained than when other media and methods were used.     

西沙群岛海域海洋放线菌的分离及其抗菌活性

分离西沙群岛海域放线菌并研究其抗菌活性。采用干燥、辐射、冷冻及加热处理等11种样品预处理方式和10种培养基对海洋放线菌进行分离,对代表性菌株进行鉴定,并考察分离放线菌生长的海水依赖性。进一步以金黄色葡萄球菌、大肠埃希菌、啤酒酵母和扩展青霉为指示菌考察分离放线菌的抗菌活性。从西沙群岛海域样品中分离获得放线菌383株,其中专性海洋放线菌23株。选定93株代表菌株进行鉴定,93株菌隶属于9个科,11个属。不同培养基对分离放线菌菌株的数量及种类影响显著。6株放线菌对4种指示菌均有抑菌活性,其中4株为专性海洋放线菌,表明海洋环境具有丰富的放线菌资源,这些放线菌特别是专性海洋放线菌有望为新型抗菌物质的发现与开发提供菌种来源。     

Isolation of Actinomycetes from Soil Using Extremely High Frequency Radiation

A new method employing extremely high frequencies (EHFs) is proposed for the selective isolation of actinomycetes from soil. The pretreatment of soil suspensions with EHF wavelengths of 5.6 and 7.1 mm led to a nonselective isolation of actinomycetes. At the same time, the irradiation of soil suspensions within wavelength bands of 3.8–5.8 and 8–11.5 mm considerably augmented the total number of isolated actinomycetes and increased the fraction of the isolated rare genera by 2 and 7 times, respectively. The rare actinomycete genera were represented by Actinomadura, Microtetraspora, Nonomuraea, Micromonospora, Amycolatopsis, Pseudonocardia, Saccharotrix, and Streptosporangium.     

Some Approaches to the Selective Isolation of Actinomycetes of the Genus Actinomadura from Soil

Some approaches to the selective isolation of actinomycetes of the genus Actinomadura from soil are described. The approach that involves the thermal treatment of soil samples and their plating onto Gauze 1 medium with the antibiotics nystatin, nalidixic acid, and rubomycin provides for an increased amount of actinomaduras isolated from the soil actinomycete complex and for a decreased amount of streptomycetes.     

Isolation of antibiotic-producing Actinomycetes from soil samples exposed to UV light

The use of nonroutine means in isolation of microorganisms from natural substrates extended the possibilities of detecting new cultures which often appear to be producers of previously unknown antibiotics. A new procedure for isolating actinomyces of definite groups was developed. It implies preliminary exposure of soil suspensions to UV light. With the use of the procedure, 2539 strains of actinomycetes belonging to different genera were isolated. There was a marked decrease after the irradiation in isolation of cultures belonging to Streptomyces, a genus most widely distributed in nature and studied in detail while isolation of cultures belonging to other genera, promising as sources of novel antibiotics, increased. Micromonospora, Amycolatopsis and Nocardia proved to be the most stable to the effect of UV light. With the use of the procedure it is possible to increase 2-3-fold isolation of cultures belonging to Micromonospora, a genus known as a producer of many antibiotics including those used clinically.     

Chelex-100快速提取放线菌DNA作为PCR扩增模板

旨在建立有效扩增16S rRNA基因序列的放线菌DNA快速提取的方法。采用Chelex-100法提取放线菌DNA,使用PCR扩增16S rRNA基因序列评价提取核酸的质量。结果显示,Chelex-100法能够在10 min之内从放线菌中快速提取DNA,所提取的DNA可以直接用于PCR扩增反应,PCR扩增产物电泳条带清晰,符合理论预期结果。因此,Chelex-100法提取放线菌DNA可以作为16S rRNA基因序列PCR扩增的模板,该方法具有经济、简便、快速的特点,适合于放线菌菌株大规模地筛选和分类鉴定。     

Isolation and Diversity of Actinomycetes in the Chesapeake Bay

Chesapeake Bay was investigated as a source of actinomycetes to screen for production of novel bioactive compounds. The presence of relatively large populations of actinoplanetes (chemotype II/D actinomycetes) in Chesapeake Bay sediment samples indicates that it is an eminently suitable ecosystem from which to isolate actinomycetes for screening programs. Actinomycetes were isolated from sediment samples collected in Chesapeake Bay with an isolation medium containing nalidixic acid, which proved to be more effective than heat pretreatment of samples. Actinomycete counts ranged from a high of 1.4 × 10⁵ to a low of 1.8 × 10² CFU/ml of sediment. Actinomycetes constituted 0.15 to 8.63% of the culturable microbial community. The majority of isolates from the eight stations studied were actinoplanetes (i.e., chemotype II/D), and 249 of these isolates were obtained in a total of 298 actinomycete isolates. Antimicrobial activity profiles indicated that diverse populations of actinoplanetes were present at each station. DNA hybridization studies showed considerable diversity among isolates between stations, but indicated that actinoplanete strains making up populations at nearby stations were more similar to each other than to populations sampled at distant stations. The diversity of actinoplanetes and the ease with which these organisms were isolated from Chesapeake Bay sediments make this a useful source of these actinomycetes.     

Isolation of Hydrocarbon-Oxidizing Psychroactive Bacteria from Oil-Polluted Soils

Microorganisms growing on a mineral medium with crude oil and its light fractions as the only source of carbon and energy have been isolated from samples of oil-polluted soils collected in the Usa district (Komi Republic, Russia). Hydrocarbon-oxidizing psychroactive bacteria of the genus Cytophaga, which are clearly capable of consuming crude oil hydrocarbons, have been identified for the first time. A method for cultivating microorganisms on porous plastic is proposed.     

红树林放线菌分离鉴定及活性代谢产物研究

以采集自四个红树林地点的16份混合土壤为研究材料,选用7种选择性培养基,共分离获得330株放线菌。其中217株菌经16SrRNA基因序列分析,发现近75%菌株属于小单孢菌属(Micromonospora),其他还包括多形态孢菌属(Polymorphospora),疣孢菌属(Verrucosispora)等小单孢菌科的2个属和非小单孢菌科的9个属。采用美蓝酶标仪法对所分离到的放线菌进行抗菌活性检测,共50株菌表现出对金黄色葡萄球菌(Staphylococcus aureus ATCC 51650)、大肠杆菌(Escherichia coli ATCC 25922)和白色念珠菌(Candida albicans ATCC 10231)有不同程度抗性。然后利用高效液相色谱(HPLC)和液质联用技术(LC-MS)对有生物活性的菌株进行化学筛选,最后确定了5株可能产新颖化合物的小单孢菌。     

Selective Isolation and Enumeration of Actinomycetes using Rifampicin

Addition of rifampicin (5µg/ml) to a number of media allowed the selective isolation of Actinomadura strains from soil and enabled large numbers of ' Thermomonospora (Thm.) chromogena ' and Streptomyces (St.) albus to be isolated from hay and straw. Saccharomonospora (S.) viridis was also tolerant to the antibiotic but was often inhibited by other isolates, while other bacteria including Micropolyspora faeni, 'Thm. fusca', Thermoactinomyces (Tha.) vulgaris and some other Streptomyces spp. were mostly sensitive, even to 2·5 µg/ml. Heating soil samples at 100°C for 15 min further decreased the number of unwanted bacteria growing on glucose-yeast extract agar supplemented with 5 µg/ml of rifampicin.