Extracts Of Streptomyces Research Articles

Antifungal efficacy of photodynamic therapy on Cryptococcus and Candida species is enhanced by Streptomyces spp. extracts in vitro.

The research on actinobacteria isolated from traditional medicinal plants is limited. Here, four new Streptomyces isolates (Ha1, Pp1, UzK and UzM) were obtained from the rhizospheres of Helianthus annuus, Pongamia pinnata and Ziziphus mauritiana, frequently utilized in Indian traditional medicine. The Streptomyces isolates aqueous extracts were studied alone against the growth of the Cryptococcus neoformans H99 reference strain, the fluconazole-tolerant T1-5796 and 89-610 strains, three histone deacetylase (HDAC) genes mutant strains, C. gattii NIH198, Candida albicans, C. glabrata, C. parapsilosis and C. tropicalis to determine minimum inhibitory concentration (MIC). Next, the extracts were employed in combination with aluminium-phthalocyanine chloride nanoemulsion-mediated photodynamic therapy to evaluate a possible interaction. We demonstrated that the C. neoformans T1-5796 fluconazole-tolerant strain was more severely inhibited by the Pp1 isolate extract (MIC: 6mg mL-1) than H99, which was not inhibited. Growth inhibition of the HDAC null mutants was more prominent for the extract of the UzM isolate, showing inhibition at 2mg mL-1. The UzM extract was also the most effective in hindering the Candida species proliferation, with MIC values ranging from 10 to 40mg mL-1. The four Streptomyces extracts, especially UzK and UzM, significantly enhanced the antifungal effect of the photodynamic therapy. Our results indicate these Streptomyces isolates as sources of novel metabolites which could potentiate the effect of photodynamic therapy in controlling yeasts superficial infections.

Evaluation of the Cytotoxicity of Secondary Bioactive Compounds Produced by Streptomyces in Soil against a Colon Cancer Cell Line.

Colorectal cancer is one of the most serious malignancies affecting humans. In this study, Streptomyces bioactivechemicals extracted from soil were analyzed for their anti-colorectal-cancer and antibacterial properties. A total of 100 soil samples were collected from Kerman-Iran, incubated in SCA media and the antimicrobial properties were tested using the cross-streak method. Three strains were cultured in ISP4 medium to obtain secondary bioactive compounds. After studying the effects of the bioactive compounds on the HT29 and human foreskin fibroblast (HFF) cell lines, the expression of the p53, p21, BAX, BCL2, Casp3 and Casp8 genes was analyzed by real-time PCR and flow cytometry to detect the presence of apoptosis.The isolates show high degree of identification with Streptomyces rochei, Streptomyces fungicidicus and Streptomyces maritimus due to 16SrDNA sequence homology. Compared to HT-29 cells, Streptomyces extracts had lower cytotoxicity against normal cells (SI=5.88), followed by HFF (SI=4.14). The cell lines demonstrated a dose-dependent significant increase in DNA fragmentation, an increase in the proportion of cells in sub-G1 phase and caused G2/M cell cycle arrest in HT-29 and HFF cells.The bacterial extracts obtained displayed strong antibacterial properties and inhibited the proliferation of HT-29 and HFF cell lines. The treated cells exhibited morphological changes caused by the activation of caspase and p53/p21 proteins. This confirms that Streptomyces-induced apoptosis is mediated by the activation of p21/p53. Anti-apoptotic Bcl-2 gene expression was downregulated by treatment with the extracts. Further studies are needed to understand the antimicrobial properties of Streptomyces.

Mechanism of action of secondary metabolites from marine-derived Streptoymces on bacterial isolates by membrane permeability

This study was designed to further investigate the potential mechanism of action of our previously characterized and identified marine-derived Streptomyces extracts (ESC003 and ESC012) on selected bacterial isolates from our culture bank. Time-kill kinetics, protein and lipid leakages assay, cell membrane permeability, phosphate and potassium ions efflux assay, extracellular adenosine triphosphate (ATP) concentration and membrane potential (MP) were all carried out using the marine-derived Streptomyces extract to determine and understand the probable mode of action at which the extract inhibit or kill bacterial cell. The MIC of ESC003 and ESC012 ranged from 0.16 to 6.25 mg/mL while the MBC ranged from 1.25 to >10 mg/mL. On the time-kill kinetics, a reduction in mean viable cell amount was detected at respective time studied. For the impermeability of the bacterial isolates, the relative electric conductivity increased with increase in concentration and time interval of exposure. As regards the protein leakage, lipid leakage, 260 nm absorbing materials leakage, phosphate and potassium ions efflux; considerable amount of these products were leaked with increase in concentration and time of exposure to the bacterial isolates. The extracellular ATP concentration from respective bacterial isolates increased appreciably with increased concentration of exposure with a simultaneous decrease in membrane potential. Findings from this study revealed that the Streptomyces extracts revealed a significant breakthrough against susceptible bacterial isolates via the permeability of the bacterial cell membrane, and thus resulted in the outflow of ATP, electrolytes, DNA materials, and proteins. These changes lead to disruption, and eventually cell death, which were proportional to a concurrent decrease in the viability of the bacterial cell.

Antagonistic activity of the Streptomyces species isolated from rhizospheric soil of selected medicinal important plants.

A total eleven streptomyces species were isolated from the plant rhizosphere soils. Extract were prepared by using Mineral medium and tested for their antibacterial activity against the Enterobacteriaceae members. Seven isolates (63%) of the streptomyces extracts were active against five pathogenic bacteria. 5 isolates (45%) of the Streptomyces extracts doesn’t showed any activity against the 5 pathogenic organisms. 80% of the inhibition was showed by strain no 6 & 10 in the organisms E. coli , K. pneumoniae and S. flexineri . 70-75% of inhibition was showed by strain no 3, 4, 5 and 11 on the organisms K. pneumoniae, S. typhi, E. coli and S. flexineri .

Targeted Dereplication of Microbial Natural Products by High-Resolution MS and Predicted LC Retention Time.

A new strategy for the identification of known compounds in Streptomyces extracts that can be applied in the discovery of natural products is presented. The strategy incorporates screening a database of 5555 natural products including 5098 structures from Streptomyces sp., using a high-throughput LCMS data processing algorithm that utilizes HRMS data and predicted LC retention times (tR) as filters for rapid identification of known compounds in the natural product extract. The database, named StrepDB, contains for each compound the structure, molecular formula, molecular mass, and predicted LC retention time. All identified compounds are annotated and color coded for easier visualization. It is an indirect approach to quickly assess masses (which are not annotated) that may potentially lead to the discovery of new or novel structures. In addition, a spectral database named MbcDB was generated using the ACD/Spectrus DB Platform. MbcDB contains 665 natural products, each with structure, experimental HRESIMS, MS/MS, UV, and NMR spectra. StrepDB was used to screen a mutant Streptomyces albus extract, which led to the identification and isolation of two new compounds, legonmaleimides A and B, the structures of which were elucidated with the aid of MbcDB and spectroscopic techniques. The structures were confirmed by computer-assisted structure elucidation (CASE) methods using ACD/Structure Elucidator Suite. The developed methodology suggests a pipeline approach to the dereplication of extracts and discovery of novel natural products.

Fourier transform infrared spectroscopy as a tool for identification of crude microbial extracts with anti-malarial potential.

Fourier transform infrared (FT-IR) spectroscopy is an indispensable tool for identifying biologically active functional groups in uncharacterized crude samples. Here, using FT-IR spectrum analysis, we identified crude extracts of Streptomyces that have anti-malarial activities and conducted a statistical analysis of their spectra. Among the three crude microbial extracts tested herein, an aromatic amine C-N stretching functional group was observed in the spectra of Streptomyces sp. BJSG1 and BJSG4 crude extracts. These extracts showed promising activity against Plasmodium falciparum in vitro cultures with IC50 values of 0.5 for BJSG1 and 0.4μg/mL for BJSG4. The present results showed that FT-IR analysis is necessary for the primary analysis of unknown samples in anti-malarial drug development.

Screening the Antibacterial Activities of Streptomyces Extracts against Phytopathogens Xanthomonas oryzae pathovar oryzae, Xanthomonas campestris pathovar vesicatoria, and Pectobacterium carotovorum pathovar carotovorum

Xanthomonas oryzae pv. oryzae (Xoo), X. campestris pv. vesicatoria (Xcv), and Pectobacterium carotovorum pv. carotovorum (Pcc) are the causative agents of bacterial blight in rice, bacterial spot in pepper, and bacterial soft rot in carrot and cabbage, respectively. To isolate novel microbial extracts with antimicrobial activities against these bacteria, approximately 5,300 different Streptomyces extracts were prepared and tested. Microbial cultures from various Streptomyces strains isolated from the Jeju Island, Baekam, Mankyoung river, Jiri mountain etc. in Korea were extracted into three different factions -secreted hydrophobic, secreted hydrophilic, and mycelia- using ethyl acetate, water, and methanol. Initially, 34, 29, and 10 extracts were selected as having antibacterial activities against Xoo, Xcv, and Pcc, respectively. Extracts 1169G4, 1172E9, and 1172E10 had the highest growth inhibition activities against both Xoo and Xcv, and extracts 1151H7 and 1152H7 showed the highest growth inhibition activities against Pcc.

Antiproliferative and antiplasmodial compounds from selected Streptomyces species.

In continuation of our ongoing search for bioactive compounds from microbial extracts, we performed antiproliferative and/or antimalarial assays on extracts of 806 microbial species isolated from Madagascan marine organisms, on 1317 species isolated from Madagascan soil samples and on a Streptomyces species (S.4) from a marine sponge collected from the Florida Keys. This work identified active extracts from four Streptomyces isolates (S.1, S.2, S.3 and S.4). The extracts of Streptomyces S.1 and S.2 showed antiproliferative activity against the A2780 ovarian cancer cell line, while those of S.3 and S.4 displayed both antiproliferative and antimalarial activity. Bioassay-guided fractionation coupled with dereplication of the active extracts led to the identification and isolation of nonactin (1), monactin (2), dinactin (3), ±-nonactic acid (4), toyocamycin (5), piperafizine A (6) and a new dipeptide named xestostreptin (7). The structures of all isolated compounds 1-7 were elucidated by analyses of their NMR spectroscopic and mass spectrometric data, and were confirmed by comparison with the data reported in the literature. Compound 6 was crystallized and subjected to X-ray diffraction analysis to confirm its structure as piperafizine A (6). Compounds 1-3 displayed strong antiproliferative activity against A2780 ovarian cancer cells (IC50 values of 0.1, 0.13 and 0.2 μM, respectively), A2058 melanoma cells (IC50 values of 0.2, 0.02 and 0.02 μM, respectively), and H522-T1 non small-cell cancer lung cells (IC50 values of 0.1, 0.01 and 0.01 μM, respectively), while compounds 4 and 7 exhibited weak antiplasmodial activity against the Dd2 strain of Plasmodium falciparum, with IC50 values of 6.5 and 50 μM, respectively.

Antitumor activity of two Streptomyces extracts (Ag18 & Ag20) on Ehrlich ascites tumor in mice in vitro and in vivo studies

Antitumor activity of two Streptomyces extracts (Ag18 & Ag20) on Ehrlich ascites tumor in mice in vitro and in vivo studies

Automated Genome Mining of Ribosomal Peptide Natural Products

Ribosomally synthesized and posttranslationally modified peptides (RiPPs), especially from microbial sources, are a large group of bioactive natural products that are a promising source of new (bio)chemistry and bioactivity.1 In light of exponentially increasing microbial genome databases and improved mass spectrometry (MS)-based metabolomic platforms, there is a need for computational tools that connect natural product genotypes predicted from microbial genome sequences with their corresponding chemotypes from metabolomic data sets. Here, we introduce RiPPquest, a tandem mass spectrometry database search tool for identification of microbial RiPPs, and apply it to lanthipeptide discovery. RiPPquest uses genomics to limit search space to the vicinity of RiPP biosynthetic genes and proteomics to analyze extensive peptide modifications and compute p-values of peptide-spectrum matches (PSMs). We highlight RiPPquest by connecting multiple RiPPs from extracts of Streptomyces to their gene clusters and by the discovery of a new class III lanthipeptide, informatipeptin, from Streptomyces viridochromogenes DSM 40736 to reflect that it is a natural product that was discovered by mass spectrometry based genome mining using algorithmic tools rather than manual inspection of mass spectrometry data and genetic information. The presented tool is available at cyclo.ucsd.edu.

Dietary microbial toxins and type 1 diabetes.

Toxins may promote type 1 diabetes by modifying or damaging the beta cell causing release of autoantigens. Streptomyces is a common soil bacterium that produces many toxic compounds. Some Streptomyces can infect vegetables, raising the possibility of dietary exposure to toxins. We aimed to identify toxins that erode cellular proton gradients in extracts of Streptomyces and infested vegetables and to establish the effect of low doses of these toxins on pancreatic islets in mice. The vacuolar ATPase inhibitors, bafilomycin and concanamycin, and the ionophore, nigericin, were identified in extracts from 4 of 13 Streptomyces isolated from infested potatoes and in potatoes themselves. Injection of bafilomycin A1 into mice impaired glucose tolerance, reduced islet size, and decreased relative beta cell mass. Thus, exposure to small quantities of bafilomycin in the diet may contribute to the cause of type 1 diabetes.

New 1-O-acyl alpha-L-rhamnopyranosides and rhamnosylated lactones from Streptomyces sp., inhibitors of 3 alpha-hydroxysteroid-dehydrogenase (3alpha-HSD).

Chemical screening with extracts of Streptomyces sp. (strain GT 61150) resulted in the detection, isolation, and structure elucidation of two new acyl alpha-L-rhamnopyranosides (1 and 2) and three new rhamnosyllactones A, B1 and B2 (3 approximately 5). Rhamnosyllactones B1 and B2 were obtained as a 5:1 mixture. The structures were confirmed by spectroscopic analysis, especially 2D-NMR techniques. The rhamnosyltransferase of our strain is able to connect the sugar moiety to heteroaromatic carboxylic acids and enols. The metabolites 1 and 4/5 as well as previously reported acylrhamnosides 6 approximately 11 inhibit the enzyme 3alpha-hydroxysteroid-dehydrogenase (3alpha-HSD).

Why Metronidazole Is Active against both Bacteria and Parasites

Metronidazole is one of the rare examples of a drug developed against a parasite which has since gained broad use as an antibacterial agent ([24][1]). Briefly, at Rhone-Poulenc labs in France, extracts of Streptomyces spp. were screened for activity against Trichomonas vaginalis , a cause of vaginal

Identification and characterization of a heat-labile type I glutamine synthetase fromStreptomyces cinnamonensis

Streptomycetes have two distinct glutamine synthetases (GS): a heat-stable dodecameric GSI and a heat-labile octameric GSII. A heat-inactivated GS activity was detected in crude extracts ofStreptomyces cinnamonensis cells grown with nitrate or glutamate as the nitrogen source. The purified enzyme obtained from crude extracts of the nitrate-grown cells after affinity and anion-exchange chromatography was also heat-labile; it was inactivated by 80 % when incubated at 50 °C for 1 h. However, the enzyme has properties typical of GSI and similar with those of the heat-stable GSI purified fromS. aureofaciens: It is composed of twelve subunits, each ofM 55 kDa, and has a native molar mass of 625 kDa and an isoelectric point at pH 4.2. In addition, its activity is regulated by reversible adenylylation. Mg2+ and NaCl but not Mn2+ protected the purified enzyme from thermal inactivation, and both NaCl and Mn2+ or Mg2+ stabilized its activity at 4–8 °C. As compared with GSI fromS. aureofaciens, theS. cinnamonensis enzyme was cleaved more extensively during SDS-PAGE, was less sensitive to feedback inhibitors, and similarly affected by divalent cations. TheKm values were 12.5 mmol/L forl-glutamate, 0.1 for NH4+, 1.25 for ATP, 18.5 forl-glutamine, 3.3 for hydroxylamine and 0.087 for ADP. To our best knowledge, this is the first report of a heatlabile GSI from any source.

DoesStreptomyces aureofaciens contain two alanine dehydrogenase isozymes ?

Two NAD-dependent dehydrogenases exhibiting activity with L-alanine were detected in crude extracts ofStreptomyces aureofaciens. The dominant enzyme was found to be an alanine dehydrogenase with 6 or 4 subunits, each ofM, 45,000. The minor enzyme exhibited high activities with branched-chain amino acids, suggesting it is a valine dehydrogenase. The results demonstrate thatS. aureofaciens has only one alanine dehydrogenase, which is catalytically active in a six- or four-subunit structure.

Purification of macrolide 2′-phosphotransferase fromStreptomyces coelicolor Müller

An enzyme that catalyzes 2′-O-phosphorylation of oleandomycin and several other macrolide antibiotics has been purified approximately 47-fold from cell-free extracts ofStreptomyces coelicolor Muller, NRRL 3532 (UC™ 5240). The reaction product was verified as being oleandomycin-2′-O-phosphate by mass spectrometry. As a result of purification, the enzyme was separated from two lincosaminide inactivating enzyme activities also present in the cell-free extract.

Enzymic degradation of bromoxynil by cell-free extracts of Streptomyces felleus.

A method is described for spectrophotometric monitoring the degradation of the herbicide bromoxynil by cell-free extracts of Streptomyces felleus. The method involves a decrease in absorbance at 286 nm (absorption maximum of bromoxynil) that can be ascribed most probably to the cleavage of the aromatic ring of the bromoxynil molecule. Conditions necessary for measuring this degradation together with physico-chemical features of the degradation indicate that the reaction(s) is seemingly catalyzed by an Fe2+-dependent dioxygenase whose activity was not, however, detected in cell-free extracts of a bromoxynil-sensitive mutant of S. felleus as well as other bromoxynil-sensitive streptomycete strains.

Streptomycin Resistance in a Streptomycin-Producing Microorganism

Cell-free extracts of Streptomyces bikiniensis contain an adenosine 5'-triphosphate-dependent kinase which inactivates streptomycin (Sm) and dihydrostreptomycin by phosphorylation. The products have been identified as streptomycin 6-phosphate and dihydrostreptomycin 6-phosphate. Activity was not present in logarithmic-phase cells, which were susceptible to 25 mug of Sm per ml. In stationary-phase cells, activity appeared 12 h before detectable Sm in the medium. These cells were resistant to more than 200 mug of Sm per ml. Certain S. bikiniensis isolates selected from cultures treated with acriflavine or ethidium bromide lost the ability to produce Sm and became susceptible to 10 mug of Sm per ml throughout their growth. Cell-free extracts of the dye-treated isolates did not inactivate Sm and lacked streptomycin kinase activity at all stages in development. Ribosomes from resistant cells bound the same amount of [(3)H]dihydrostreptomycin as ribosomes from susceptible cells, and there was no correlation between the uptake of [(3)H]dihydrostreptomycin and resistance. The Sm-inactivating enzyme was identified as streptomycin-6-kinase. These results suggest that phosphorylation by streptomycin-6-kinase is a major factor in resistance in S. bikiniensis.

Propriétés antimicrobiennes de deux souches de Streptomyces albus isolées de sols d’Afrique. Applications possibles

The antibiotic activity of two strains of S. albus is facilited by culture on slices of potatoes. Streptomyces extracts show an antibiotic activity against Gram positive and Gram negative bacteria and an antifongic activity against nearly all Aspergillus. These antimicrobial effects bring hope, between other utilizations, the control of some mycotoxins pollution, or mold attacks.

Biosynthesis of the Monoguanidinated Inositol Moiety of Bluensomycin, a Possible Evolutionary Precursor of Streptomycin

Abstract The general pattern of biosynthesis of the bluensidine (1D-1-O-carbamoyl-3-guanidino-3-deoxy-scyllo-inositol) moiety of bluensomycin, a monoguanidinated analogue of dihydrostreptomycin, has been studied in extracts of Streptomyces hygroscopicus forma glebosus ATCC 14607 (S. glebosus). Our results are consistent with the following biosynthetic pathway: myo-inositol (C)/→ keto-scyllo-inositol (D)/→ aminodeoxy-scyllo-inositol (E)/→ 1-amino-1-deoxy-scyllo-inositol-4-P (U)/→ 1D-1-O-carbamoyl-3-amino-3-deoxy-scyllo-inositol-6-P (F)/→ 1D-1-O-carbamoyl-3-guanidino-3-deoxy-scyllo-inositol-6-P →→ bluensomycin. A major uncertainty concerns the step (U) at which the carbamoyl group is introduced. Carbamoylation might occur, for example, prior to phosphorylation or following transamidination. myo-Inositol:NAD oxidoreductase activity (C) is reported for the first time in extracts of Streptomyces; activity was assayed by two radiochemical methods, both involving coupling with aminotransferase (D). l-Glutamine:keto-scyllo-inositol aminotransferase (D) was also found to catalyze aminodeoxy-scyllo-inositol:keto-scyllo-inositol, aminodeoxy-scyllo-inositol:pyruvate, and 1,3-diamino-1,3-dideoxy-scyllo-inositol:keto-scyllo-inositol transaminations. l-Arginine:inosamine-P amidinotransferase (F) catalyzed transamidinations with the following compounds as amidino acceptors: NH2OH, 1-amino-1-deoxy-scyllo-inositol-4-P, 1D-1-guanidino-3-amino-1,3-dideoxy-scyllo-inositol-6-P, 1D-1,3-diamino-1,3-dideoxy-scyllo-inositol-6-P, 1D-1,3-diamino-1,2,3-trideoxy-scyllo-inositol-6-P, and a compound present in S. glebosus extracts believed to be 1D-1-O-carbamoyl-3-amino-3-deoxy-scyllo-inositol-6-P. The latter transamidination is markedly enhanced in crude extracts by carbamoyl-P. S. glebosus extracts also have 1-guanidino-1-deoxy-scyllo-inositol-4-P phosphohydrolase activity; neither this enzyme nor the corresponding enzyme from streptomycin producers can dephosphorylate the transamidination product presumed to be bluensidine-6-P. Acid hydrolysis of the latter compound gave a compound which, unlike the unhydrolyzed compound, was converted to 1D-1-guanidino-3-amino-1,3-dideoxy-scyllo-inositol by enzymes from a streptomycin producing strain. S. glebosus cannot carry out the above conversion since it apparently lacks at least two enzymes which occur in streptomycin producers: guanidinodeoxy-(scyllo)-inositol dehydrogenase and l-alanine:1D-1-guanidino-3-keto-1-deoxy-scyllo-inositol aminotransferase. It is suggested that streptomycin producing strains might be descendents of an ancestral strain which, like S. glebosus, produced the monoguanidinated inositol derivative, bluensomycin. It is further suggested that gene duplication and subsequent evolutionary divergence resulted in biosynthesis of the diguanidinated inositol derivative, streptomycin, which is 10 times more effective than bluensomycin as an antibiotic and inhibitor of protein biosynthesis.