Characterization and comparative studies ofStreptomyces sp. 246 producing a cytostatic antibiotic

The authors submit the results of taxonomic comparative studies of the strainStreptomyces sp. 246, which produces a polypeptide type cytostatic antibiotic. Strain 246 is characterized by tufts of straight sporophores of the “Rectus-Flexibilis” type, smooth spores arranged in chains (over 10 spores in a chain), yellow aerial and substrate mycelium, a negative test for melanin synthesis, utilization of glucose, arabinose, xylose, mannitol, fructose and rhamnose and inability to grow on sucrose, inositol, raffinose and cellulose. The taxonomic characters ofStreptomyces sp. 246 are identical with those of the strainStreptomyces chrysomallus JA 1449-1 and differ manifestly from those ofStreptomyces antibioticus strains (producing actinomycins, antimycin A and oleandomycin), fromStreptomyces cinereoruber ETH 7451 (producing rhodomycin) and from the strainStreptomyces sp. 4127 (producing actinomycin D).     

链霉菌IMS002的分类鉴定及其抗真菌活性物质解析

【目的】对一株分离自植物根际土壤的具有抗真菌活性的链霉菌IMS002进行菌株分类鉴定,通过活性追踪分离纯化并鉴定有机相中的活性物质。【方法】通过16S rDNA和5个不同基因(atpD,gyrB,recA,rpoB,trpB)串联聚类分析以及生理生化实验分析,对链霉菌IMS002进行菌株分类鉴定,用扫描电子显微镜观察该株链霉菌的菌丝及孢子形态,以尖孢镰刀菌(Fusarium oxysporum)为指示菌进行生物活性追踪,通过硅胶柱层析、凝胶柱层析及高压液相色谱(HPLC)对活性物质进行分离和纯化,使用液质联用高分辨质谱仪、500 MHz核磁共振波谱仪以及圆二色光谱仪确定该物质的化学结构。【结果】IMS002经初步鉴定与产二素链霉菌(Streptomycesambofaciens)具有较近的亲缘关系,其发酵液对尖孢镰刀菌具有良好的抑菌效果,经分离和纯化以及现代波谱技术分析,确定有机相中的抑菌活性组分为Borrelidin。【结论】链霉菌IMS002能够产生化合物Borrelidin,该化合物对尖孢镰刀菌具有抑制活性。     

Streptomyces sp. ASBV‐1 reduces aflatoxin accumulation by Aspergillus parasiticus in peanut grains

Aims: To evaluate the ability of Streptomyces sp. (strain ASBV‐1) to restrict aflatoxin accumulation in peanut grains. Methods and Results: In the control of many phytopathogenic fungi the Streptomyces sp. ASBV‐1 strain showed promise. An inhibitory test using this strain and A. parasiticus was conducted in peanut grains to evaluate the effects of this interaction on spore viability and aflatoxin accumulation. In some treatments the Streptomyces sp ASBV‐1 strain reduced the viability of A. parasiticus spores by c. 85%, and inhibited aflatoxin accumulation in peanut grains. The values of these reductions ranged from 63 to 98% and from 67% to 96% for aflatoxins B₁ and G₁, respectively. Conclusions: It was demonstrated that Streptomyces sp. ASBV‐1 is able to colonize peanut grains and thus inhibit the spore viability of A. parasiticus, as well as reducing aflatoxin production. Significance and Impact of the Study: The positive finding for aflatoxin accumulation reduction in peanut grains seems promising and suggests a wider use of this actinobacteria in biological control programmes.     

Biological control of anthracnose (Colletotrichum gloeosporioides) in yam by Streptomyces sp.MJM5763

Aim: To find a suitable biocontrol agent for yam anthracnose caused by Colletotrichum gloeosporioides. Methods and Results: An actinobacterial strain, MJM5763, showing strong antifungal activity, multiple biocontrol and plant growth‐promoting traits was isolated from a yam cultivation field in Yeoju, South Korea. Based on morphological and physiological characteristics and analysis of the 16S rDNA sequence, strain MJM5763 was identified as a novel strain of Streptomyces and was designated as Streptomyces sp. MJM5763. Treatment with MJM5763 and the crude culture filtrate extract (CCFE) was effective in suppressing anthracnose in detached yam leaves in vitro and reduced incidence and severity of anthracnose in yam plants under greenhouse conditions. The CCFE treatment was the most effective of all the treatments and reduced the anthracnose severity by 85–88% and the incidence by 79–81%, 90 days after inoculation with the pathogen. CCFE treatment was also effective under field conditions and showed a reduction of 86 and 75% of anthracnose severity and incidence, respectively. Conclusion: Streptomyces sp. strain MJM5763 was effective in biocontrolling anthracnose in yam caused by C. gloeosporioides. Significance and Impact of the Study: Streptomyces sp. MJM5763 is a potential alternative to chemical fungicides for reducing yield losses to anthracnose in yam.     

Developmental Regulation of Susceptibility and Tolerance to Fusarium Root Rot in Beans

The soil-borne fungus, Fusarium solani f. sp. phaseoli, attacks roots and hypocotyls of bean (Phaseolus vulgaris) plants causing a devastating disease called root and foot rot. In a study of the host-pathogen relationship it was found that young bean roots, with the radicle just emerging, were highly tolerant to the pathogen, whereas older bean seedlings, with a fully developed root system, were completely susceptible. Investigations by low-temperature scanning electron microscopy demonstrated that significantly fewer spores and hyphae were present on the root surface of young bean seedlings as compared to older ones. A similar pattern of attachment was found when bean roots were inoculated with spores of F. solani f. sp. pisi, a related pathogen causing disease on peas but not on beans. Light microscopic studies showed that F. solani f. sp. pisi did not penetrate the root but rapidly formed thick-walled resting spores on the root surface. F. solani f. sp. phaseoli on the other hand quickly penetrated the root and formed an extensive network of fungal hyphae. These results demonstrate that the ability of fungal propagules to adhere to and to penetrate host tissues are two distinct processes. Furthermore, the data indicate that young bean roots lack a surface component necessary for attachment of fungal spores which may help explain their tolerance to Fusarium root rot.     

Septins enforce morphogenetic events during sexual reproduction and contribute to virulence of Cryptococcus neoformans

Septins are conserved, cytoskeletal GTPases that contribute to cytokinesis, exocytosis, cell surface organization and vesicle fusion by mechanisms that are poorly understood. Roles of septins in morphogenesis and virulence of a human pathogen and basidiomycetous yeast Cryptococcus neoformans were investigated. In contrast to a well‐established paradigm in S. cerevisiae, Cdc3 and Cdc12 septin homologues are dispensable for growth in C. neoformans yeast cells at 24°C but are essential at 37°C. In a bilateral cross between septin mutants, cells fuse but the resulting hyphae exhibit morphological abnormalities, including lack of properly fused specialized clamp cells and failure to produce spores. Interestingly, post‐mating hyphae of the septin mutants have a defect in nuclear distribution. Thus, septins are essential for the development of spores, clamp cell fusion and also play a specific role in nuclear dynamics in hyphae. In the post‐mating hyphae the septins localize to discrete sites in clamp connections, to the septa and the bases of the initial emerging spores. Strains lacking CDC3 or CDC12 exhibit significantly reduced virulence in a Galleria mellonella model of infection. Thus, C. neoformans septins are vital to morphology of the hyphae and contribute to virulence.     

Characterization of an extracellular glycoprotein inhibitor of trypsin (TI-23) produced by Streptomyces sp. 23

Summary The production of an extracellular trypsin inhibitor, TI-23, was found to parallel the growth of Streptomyces sp. 23 at different cultivation temperatures, reaching a maximum level at late exponential phase. Although the different temperatures (18°, 28° and 37°C) did not greatly affect the growth of the microorganism, they proved to be an important factor for extracellular inhibitory activity. Maximum specific rates of both cell growth and production of the inhibitor were recorded during the cultivation of Streptomyces sp. 23 at 37°C. TI-23 proved to be a monomeric glycoprotein containing 17% carbohydrate and differing in amino acid composition from the known extracellular proteinase inhibitors of streptomycetes. The molecular mass of the inhibitor was estimated to be about 13 kDa and the isoelectric point 4.3. The inhibition spectrum of TI-23 included trypsin as well as some microbial alkaline proteinases.     

Isolation and Characterization of a Spore Germination Inhibitor from Streptomyces sp. CB-1-1, a Phytopathogen Causing Root Tumor of Melon

The germination rate and activation conditions of spores were examined for four strains of Streptomyces sp., a phytopathogen causing root tumor of melon. An inhibitor was isolated from the agar-cultured material of strain CB-1-1 and then characterized. The inhibitor selectively acted on spore germination and did not affect hyphal growth, and inhibition was abolished by washing the spores in water. The inhibitor was produced by an agar culture, and most of the inhibitor existed in the spores. The IC₅₀ value for the inhibitor was approximately 0.25 μg/ml.     

Classification of novel soil streptomycetes as Streptomyces aureus sp. nov., Streptomyces laceyi sp. nov. and Streptomyces sanglieri sp. nov

The taxonomic positions of soil isolates known as Streptomyces groups A, B and C were clarified. Comparative 16S rDNA sequence studies indicated that representatives of all three taxa formed distinct phyletic lines within the Streptomyces tree though the group A strains were shown to be related to Streptomyces griseus and associated validly described species. The taxonomic integrity of all three groups was highlighted by DNA:DNA relatedness and ribotype data though the group A strains encompassed a higher degree of genetic variation than the group B and C strains. In light of these and earlier phenotypic data it is proposed that Streptomyces groups A, B and C be given species status as Streptomyces sanglieri sp. nov., Streptomyces aureus sp. nov. and Streptomyces laceyi sp. nov., respectively. This revised version was published online in June 2006 with corrections to the Cover Date.     

Zur Kenntnis thermophiler Actinomyceten

Zusammenfassung Für Thermomonospora curvata und Streptomyces rectus=Actinomyces thermophilus sensu Gilbert werden Neotypen vorgeschlagen. Für Streptomyces thermoviolaceus ssp. thermoviolaceus und ssp. apingens, S. thermovulgaris, Pseudonocardia thermophila und Microbispora bispora werden Typusstämme ausgewählt. Thermomonospora curvata wird als Typusart für die Gattung Thermomonospora gewählt. Die Gattung Thermopolyspora wird Synonym von Microbispora. Von den Thermopolyspora-Arten kommt nur Th. bispora zur Gattung Microbispora. Für Th. polyspora, Th. flexuosa und Th. rectivirgula ist später eine neue Gattung aufzustellen.Folgende Sporenfeinstruktur wurde für die thermophilen Arten festgestellt: Bei Streptomyces thermovulgaris, Pseudonocardia thermophila und Microbispora bispora sind die Spore glatt, sie sind stachelig bei Streptomyces rectus und Thermomonospora curvata und warzig bei Streptomyces thermoviolaceus. Die Warzen sind in der Mitte vertieft.Ultradünnschnitte wurden von den Sporen von Thermomonospora curvata hergestellt. Die Stacheln werden durch die Falten einer Hülle gebildet, welche locker die dickwandige, glatte Spore umgibt. Die Ähnlichkeit der Bilder mit Microellobosporia ist vielleicht ein Hinweis auf eine mögliche Verwandtschaft der beiden Gattungen.Bei Pseudonocardia thermophila wurden das Mycelwachstum und die Sporenbildung im einzelnen verfolgt. Sowohl die Substrat- als auch die Lufthyphen sin septiert. Viererlei Sporen werden gebildet: Zerfallssporen im Substratmycel, Zerfallssporen im Luftmycel, Segmentationssporen in den Lufthyphen=Entstehung nach dem Pseudonocardia-Typ und Sporenbildung in den Lufthyphen nach dem Streptomyces-Typ.Fünf Formen der Luftsporenbildung werden diskutiert: der Streptomyces-Typ, der Microbispora-Typ, der Thermomonospora-Typ, der Pseudonocardia-Typ und die Sporenbildung, wie sie Krassilnikov u. Agre für Thermopolyspora-Arten beschrieben.

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Studies on thermophilic actinomycetes

Summary Typecultures are designated for the species Streptomyces thermoviolaceus ssp. thernoviolaceus and ssp. apingens, S. thermovulgaris, Pseudonocardia thermophila and Microbispora bispora; neotypes are proposed for Thermonospora curvata and Streptomyces rectus=Actinomyces thermophilus sensu Gilbert. Thermonospora curvata is chosen as type species of the genus Thermonospora. The genus Thermopolyspora becomes a synonym of Microbispora. A new genus will have to be erected for Thermopolyspora polyspora, Th. flexuosa and Th. rectivirgula, which do not fit into the genus Microbispora.The fine structure of the spores of some thermophilic species is described. Streptomyces thermovulgaris, Pseudonocardia thermophila and Microbispora bispora have smooth spore walls; spines are developed in Streptomyces rectus and Thermomonospora curvata, and warts in Streptomyces thermoviolaceus. The warts have been found to possess a median depression.Ultrathin sections were cut of the spores and hyphae of Thermomonospora curvata. The spine bearing layer forms a loose sheath around the smooth, thick wall of the spore as is reported in Microellobosporia. This may indicate relationship between these two genera.In Pseudonocardia thermophila the development of the substrate and aerial mycelia and the spore formation has been studied in detail. Both the substrate and aerial hyphae are septate. Four kinds of spores were observed: fragmentation spores in the substrate hyphae, fragmentation spores in the aerial hyphae, segmentation spores in the aerial hyphae produced according to the Pseudonocardia-type, and spores formed in the aerial hyphae according to the Streptomyces-type.Among the non-sporangiate actinomycetes five types of aerial spore formation are discussed, the Streptomyces-type, Microbispora-type, Thermomonospora-type, Pseudonocardia-type, and the formation of spores described by Krassilnikov and Agre for two species of Thermopolyspora.

     

Isolation and Physiological Activities of Piericidin A,A Natural Insecticide Produced by Streptomyces

In the course of screening research on new insecticides among metabolites of microorganisms, two active components were isolated from mycellia of Streptomyces sp. 16–22. The names, Piericidin A and B, were proposed to these active metabolites. Isolation procedure, physical and chemical properties and physiological activities of Piericidin A were presented. From the results of taxonomic study on Streptomyces sp. 16–22, it was found that this is a new species, to which the name, Streptomyces mobaraensis Nagatsu et Suzuki, was proposed.     

链霉菌的功能及其在农业上的应用

链霉菌是自然界中大量存在的一类微生物,具有多种多样的功能,其基内菌丝多核有间隔,气生菌丝上附着孢子链,从孢子萌发到孢子释放完成整个生命周期。链霉菌以菌丝体的形式定殖于多种植物体的根、茎、叶等部位而发挥功能,能分泌多种具有促进植物生长与生物防治功能的代谢物。对近几年链霉菌在提升植物营养吸收、促进植物生长、增强植物应对逆境能力、改善土壤结构、恢复污染水体等方面的研究进行了综述,并对今后链霉菌的研究方向和应用前景进行了展望。     

An Inhibitor of S-Hemolysin,SHI, Produced by Streptomyces sp. Strain No. 6288

Streptomyces sp. strain No. 6288 produces S-Hemolysin, which is a unique phospholipase C with a high substrate specificity for sphingomyelin. Moreover, the strain also produced two kinds of phospholipase inhibitors, designated as SHI and S-PLI, in different phases of cultivation. The purified SHI was shown to be a basic substance containing an amino group and glycoside moiety, and it was a more effective inhibitor of S-Hemolysin and sphingomyelinase from Streptomyces sp. with a higher substrate specificity for sphingomyelin than α-toxin from Clostridium perfringens.     

Biologically Active Endophytic Streptomycetes from <Emphasis Type="Italic">Nothofagus</Emphasis> spp. and Other Plants in Patagonia

Endophytic streptomycetes have been isolated and characterized from several species of Nothofagus and other plants growing in the southern reaches of Patagonia. No endophytic streptomycete was obtained from any plant species studied in Northern Patagonia. However, from Southern Patagonia, biologically active Streptomyces spp. from several plant species were isolated. Each isolate, as studied by scanning electron microscopy (SEM), has small hyphae, some produce typical barrel-shaped spores in culture and each has some unique hyphal surface structures. Interestingly, although none has any detectable antibacterial killing properties, each has demonstrable killing activity against one or more pathogenic fungi including representative plant pathogenic organisms such as Phytophthora erythroseptica, Pythium ultimum, Sclerotinia sclerotiorum, Mycosphaerella fijiensis, and Rhizoctonia solani. The 16S rDNA sequences of the isolates were distinct from all other genetic accessions of Streptomyces in GenBank. However, isolate C-2 from Chiliotrichum diffusum (Compositae) is identical, in all respects, to isolate C-4 obtained from Misodendrum punctulatum (Loranthaceae). These results confirm that endophytic streptomycetes represent a novel source of biologically active microorganisms.     

Pyrindicin,a New Alkaloid from a Streptomyces Strain

In the course of our examination for the alkaloid productivities of Streptomyces strains, Streptomyces sp. NA–15 was found to produce a new alkaloid, pyrindicin, in the culture medium. The strain NA–15 was found to be a variant of Streptomyces griseoflavus and was designated as S. griseoflavus var. pyr indie us nov. var.

After the culture conditions for pyrindicin production were studied, pyrindicin was obtained as its hydrochloride (mp 145°C, decomp.) from the cultured broth. The compound was shown to possess weak antimicrobial and several pharmacological activities. The LD₅₀ of the hydrochloride (ip, in mice) was 87 mg/kg.     

Microbial degradation and physico-chemical alteration of polyhydroxyalkanoates by a thermophilic <Emphasis Type="Italic">Streptomyces</Emphasis> sp.

A thermophilic Streptomyces sp. capable of degrading various aliphatic polyesters was isolated from a landfill site. The isolate, Streptomyces sp. BCC23167, demonstrated rapid aerobic degradation of several polyesters, including polyhydroxyalkanoate copolymers, poly(ɛ-caprolactone) and polybutylene succinate at 50°C and neutral pH. The degrading activity was repressed by glucose and cellobiose, but tolerant to repression by other carbon substrates. Degradation of a commercial poly[(R)-3-hydroxybutyrate-co-3-hydroxyhexanoate] (PHBHx) by Streptomyces sp. BCC23167 progressed from surface to bulk as suggested by the slight decrease in polymer molecular weight. Differential scanning calorimetry analysis of PHBHx film degradation by Streptomyces sp. BCC23167 showed that relative crystallinity of the film increased slightly in the early stage of degradation, followed by a marked decrease later on. The surface morphology of degraded films was analyzed by scanning electron microscopy, which showed altered surface structure consistent with the changes in crystallinity. The isolate is thus of potential for application in composting technology for bio-plastic degradation.     

The effect of different conditions of temperature and acidity on antimicrobial metabolites of Streptomyces sp. isolates C-11 and C-26 against F. subglutinans

In this study, the effects of pH and temperature on active two Streptomyces sp. against F. subglutinans were investigated. Streptomyces sp. isolates C-11 and C-26 were grown in submerged cultures for determination of growth curve and preparation of crude extract for further biological characterisations. Activity reached maximum at 9 and 4?days in rotary cultures for isolates C-26 and C-11, respectively. The results effects of different pH and temperature showed that these isolates were tolerant to high temperatures and the most antagonistic effects on fungi are investigated in alkaline conditions.     

Direct Introduction of Fluorescein Isothiocyanate-conjugated Albumin into Intact Macroconidia of Fusarium oxysporum f. sp. lycopersici by Electroporation

An electroporation procedure for the introduction of fluorescein isothiocyanate-conjugated albumin into intact macroconidia of Fusarium oxysporum f. sp. lycopersici was performed in this paper. FITC-albumin was used to establish an efficient electroporation procedure because its presence in spores could be easily detected by fluorescence microscopy. The uptake of fluorescein isothiocyanate-conjugated albumin into spores was successfully mediated by high voltage electrical pulses at field strength of 10 KV/cm. In electroporation of 10⁶ macroconidia per ml with the present system, 1.2 × 10⁵ spores incorporated the protein, and then 3.1 × 10⁴ normally germinated, elongated vegetative hyphae, and produced spores.     

Endophytic actinomycetes from Pinus thunbergii and their antifungal activity against Cylindrocladium sp.

The inside of Pinus thunbergii could be a reliable screening source for a useful agent in controlling plant disease. Isolation of endophytic actinomycetes from P. thunbergii and their potential as biocontrol agents against the plant pathogen Cylindrocladium sp. were investigated. Two endophytic actinomycetes, Streptomyces sp. and Microbispora sp., were isolated from surface-sterilised root tissues of P. thunbergii seedlings. The recovery test of these two endophytic actinomycetes from pine seedling showed that Streptomyces sp. was isolated from only roots, but Microbispora sp. was isolated from both roots and leaves. Thus, Microbispora sp. is able to move to leaves from roots. Moreover, we evaluated the potential of both strains as biocontrol agents against Cylindrocladium sp. Two weeks after inoculation of Cylindrocladium sp. alone, pine seedlings showed a 50% mortality rate. Co-inoculation of Cylindrocladium sp. and Microbispora sp. did not affect seedling mortality rate. However, inoculation with both Cylindrocladium sp. and Streptomyces sp. reduced seedling mortality to 12%. Streptomyces sp. could be a useful agent in controlling pine disease caused by Cylindrocladium sp. Thus, it seems that Streptomyces sp. may induce a local host defence reaction and Microbispora sp. systemically spreads to aerial parts through the transpiration stream.