Bacterium Streptomyces Research Articles

The roseoflavin producer Streptomyces davaonensis has a high catalytic capacity and specific genetic adaptations with regard to the biosynthesis of riboflavin.

The bacterium Streptomyces davaonensis synthesizes the antibiotic roseoflavin in the stationary phase of growth. The starting point for roseoflavin biosynthesis is riboflavin (vitamin B2 ) and four enzymes (RibCF, RosB, RosA and RosC) are necessary to convert a vitamin (riboflavin) into a potent, broad-spectrum antibiotic (roseoflavin). In S. davaonensis, seven enzymatic functions are required to synthesize the roseoflavin precursor riboflavin from the central building blocks GTP and ribulose 5-phosphate. When compared with other bacterial and in particular Streptomyces genomes the S. davaonensis genome contains an unusual high number (21) of putative riboflavin biosynthetic genes (rib genes), including a rib gene encoding an additional riboflavin synthase originating from an Archaeon. We show by complementation analyses and enzyme assays that 17 out of these 21 putative rib genes indeed encode for riboflavin biosynthetic enzymes. Biochemical analyses of selected enzymes support this finding. Transcriptome analyses show that all of the rib genes are expressed either in the exponential or in the stationary phase of growth and thus do not represent silent genes. We conclude that the Rib enzymes produced in the stationary phase represent a physiological adaptation to support roseoflavin biosynthesis.

Telomeric and sub-telomeric regions undergo rapid turnover within a Streptomyces population

Genome dynamics was investigated within natural populations of the soil bacterium Streptomyces. The exploration of a set of closely related strains isolated from micro-habitats of a forest soil exhibited a strong diversity of the terminal structures of the linear chromosome, i.e. terminal inverted repeats (TIRs). Large insertions, deletions and translocations could be observed along with evidence of transfer events between strains. In addition, the telomere and its cognate terminal protein complexes required for terminal replication and chromosome maintenance, were shown to be variable within the population probably reflecting telomere exchanges between the chromosome and other linear replicons (i.e., plasmids). Considering the close genetic relatedness of the strains, these data suggest that the terminal regions are prone to a high turnover due to a high recombination associated with extensive horizontal gene transfer.

The substrate selectivity of the two homologous SGNH hydrolases from Streptomyces bacteria: Molecular dynamics and experimental study

Two extracellular enzymes of the SGNH hydrolase superfamily reveal highly homologous 3D structures, but act on different substrates; one is a true phospholipase A1 from Streptomyces albidoflavus (SaPLA1, EC: 3.1.1.32, PDB code: 4HYQ), whereas the promiscuous enzyme from Streptomyces rimosus (SrLip, EC: 3.1.1.3, PDB code: 5MAL) exhibits lipase, phospholipase, esterase, thioesterase, and Tweenase activities. To get insight into binding modes of phospholipid and triglyceride substrates in both enzymes and understand their chain-length preferences, we opted for computational approach based on in silico prepared enzyme-substrate complexes. Docking procedure and molecular dynamics simulations at microsecond time scale were applied. The modelled complexes of SaPLA1 and SrLip enzymes revealed substrate accommodation: a) the acyl-chain attached to sn-1 position fits into the hydrophobic pocket, b) the acyl-chain attached to sn-2 position fits in the hydrophobic cleft, whereas c) the sn-3 bound acyl chain of the triglyceride or polar head of the glycerophospholipid fits into the binding groove. Moreover, our results pinpointed subtle amino acid differences in the hydrophobic pockets of these two enzymes which accommodate the acyl chain attached to sn-1 position of glycerol to be responsible for the chain length preference. Slight differences in the binding grooves of SaPLA1 and SrLip, which accommodate the acyl chain attached to sn-3 position are responsible for exclusive phospholipase and both phospholipase/lipase activities of these two enzymes, respectively. The results of modelling correlate with the experimentally obtained kinetic parameters given in the literature and are important for protein engineering that aims to obtain a variant of enzyme, which would preferably act on the substrate of interest.

Food/Feed and Environmental Risk Assessment of Insect Resistant Genetically Modified Maize 1507 for Cultivation, Import, Processing, Food and Feed Uses under Directive 2001/18/EC and Regulation (EC) No 1829/2003 (C/ES/01/01, C/NL/00/10, EFSA/GMO/NL/2004/02)

Food/Feed and Environmental Risk Assessment of Insect Resistant Genetically Modified Maize 1507 for Cultivation, Import, Processing, Food and Feed Uses under Directive 2001/18/EC and Regulation (EC) No 1829/2003 (C/ES/01/01, C/NL/00/10, EFSA/GMO/NL/2004/02)

Soil smell is bacterial lure

The delightful smell of soil is caused by small molecules like geosmin produced by the bacteria living there. In fact, every species of Streptomyces bacteria makes geosmin—but it’s not clear why th...

Developmentally regulated volatiles geosmin and 2-methylisoborneol attract a soil arthropod to Streptomyces bacteria promoting spore dispersal

Volatile compounds emitted by bacteria are often sensed by other organisms as odours, but their ecological roles are poorly understood1,2. Well-known examples are the soil-smelling terpenoids geosmin and 2-methylisoborneol (2-MIB)3,4, which humans and various animals sense at extremely low concentrations5,6. The conservation of geosmin biosynthesis genes among virtually all species of Streptomyces bacteria (and genes for the biosynthesis of 2-MIB in about 50%)7,8, suggests that the volatiles provide a selective advantage for these soil microbes. We show, in the present study, that these volatiles mediate interactions of apparent mutual benefit between streptomycetes and springtails (Collembola). In field experiments, springtails were attracted to odours emitted by Streptomyces colonies. Geosmin and 2-MIB in these odours induce electrophysiological responses in the antennae of the model springtail Folsomia candida, which is also attracted to both compounds. Moreover, the genes for geosmin and 2-MIB synthases are under the direct control of sporulation-specific transcription factors, constraining emission of the odorants to sporulating colonies. F. candida feeds on the Streptomyces colonies and disseminates spores both via faecal pellets and through adherence to its hydrophobic cuticle. The results indicate that geosmin and 2-MIB production is an integral part of the sporulation process, completing the Streptomyces life cycle by facilitating dispersal of spores by soil arthropods.

Assessing anti‐cancer potential of novel staurosporine analogs

Staurosporine, a compound isolated from Streptomyces bacteria, is known to inhibit cell growth and induce apoptosis in cells, though it is a pan‐kinase inhibitor and therefore too toxic to be an anti‐cancer agent. However, staurosporine analogs with increased specificity have been tested in clinical trials. While most groups focus on modifying the sugar portion of staurosporine, we have explored the biological activity of novel analogs consisting of a central five‐membered ring structure with different substituents located at the 3 and 4 position. These molecules are structurally similar to the five‐membered pyrrolinone group and bis‐indole complex contained within staurosporine. Our most potent first generation analogs had a low micromolar IC50 value (concentration that killed 50% of histiocytic lymphoma U‐937 cells) and provided initial structural rules for effective cell targeting. We now report on a second generation of compounds with nanomolar potency. Unlike staurosporine, preliminary direct kinase binding data showed that our most potent analogs weakly bind to protein kinase C, indicating a unique mode of action. Compound analysis by the National Cancer Institute against a panel of 60 different cancer cell lines should allow us to narrow the cellular target. Using fluorescent microscopy analysis, we are confirming that the compounds induce cell death via apoptosis. Thus, our new findings aid the cytotoxicity studies in explaining the impact of our compounds on cancer cells.

An Engineered E. coli Strain for Direct in Vivo Fluorination.

Selectively fluorinated compounds are found frequently in pharmaceutical and agrochemical products where currently 25-30 % of optimised compounds emerge from development containing at least one fluorine atom. There are many methods for the site-specific introduction of fluorine, but all are chemical and they often use environmentally challenging reagents. Biochemical processes for C-F bond formation are attractive, but they are extremely rare. In this work, the fluorinase enzyme, originally identified from the actinomycete bacterium Streptomyces cattleya, is engineered into Escherichia coli in such a manner that the organism is able to produce 5'-fluorodeoxyadenosine (5'-FDA) from S-adenosyl-l-methionine (SAM) and fluoride in live E. coli cells. Success required the introduction of a SAM transporter and deletion of the endogenous fluoride efflux capacity in order to generate an E. coli host that has the potential for future engineering of more elaborate fluorometabolites.

Targeted induction of a silent fungal gene cluster encoding the bacteria-specific germination inhibitor fumigermin.

Microorganisms produce numerous secondary metabolites (SMs) with various biological activities. Many of their encoding gene clusters are silent under standard laboratory conditions because for their activation they need the ecological context, such as the presence of other microorganisms. The true ecological function of most SMs remains obscure, but understanding of both the activation of silent gene clusters and the ecological function of the produced compounds is of importance to reveal functional interactions in microbiomes. Here, we report the identification of an as-yet uncharacterized silent gene cluster of the fungus Aspergillus fumigatus, which is activated by the bacterium Streptomyces rapamycinicus during the bacterial-fungal interaction. The resulting natural product is the novel fungal metabolite fumigermin, the biosynthesis of which requires the polyketide synthase FgnA. Fumigermin inhibits germination of spores of the inducing S. rapamycinicus, and thus helps the fungus to defend resources in the shared habitat against a bacterial competitor.

Diversity and geographic distribution of soil streptomycetes with antagonistic potential against actinomycetoma-causing Streptomyces sudanensis in Sudan and South Sudan

BackgroundProduction of antibiotics to inhibit competitors affects soil microbial community composition and contributes to disease suppression. In this work, we characterized whether Streptomyces bacteria, prolific antibiotics producers, inhibit a soil borne human pathogenic microorganism, Streptomyces sudanensis. S. sudanensis represents the major causal agent of actinomycetoma – a largely under-studied and dreadful subcutaneous disease of humans in the tropics and subtropics. The objective of this study was to evaluate the in vitro S. sudanensis inhibitory potential of soil streptomycetes isolated from different sites in Sudan, including areas with frequent (mycetoma belt) and rare actinomycetoma cases of illness.ResultsUsing selective media, 173 Streptomyces isolates were recovered from 17 sites representing three ecoregions and different vegetation and ecological subdivisions in Sudan. In total, 115 strains of the 173 (66.5%) displayed antagonism against S. sudanensis with different levels of inhibition. Strains isolated from the South Saharan steppe and woodlands ecoregion (Northern Sudan) exhibited higher inhibitory potential than those strains isolated from the East Sudanian savanna ecoregion located in the south and southeastern Sudan, or the strains isolated from the Sahelian Acacia savanna ecoregion located in central and western Sudan. According to 16S rRNA gene sequence analysis, isolates were predominantly related to Streptomyces werraensis, S. enissocaesilis, S. griseostramineus and S. prasinosporus. Three clusters of isolates were related to strains that have previously been isolated from human and animal actinomycetoma cases: SD524 (Streptomyces sp. subclade 6), SD528 (Streptomyces griseostramineus) and SD552 (Streptomyces werraensis).ConclusionThe in vitro inhibitory potential against S. sudanensis was proven for more than half of the soil streptomycetes isolates in this study and this potential may contribute to suppressing the abundance and virulence of S. sudanensis. The streptomycetes isolated from the mycetoma free South Saharan steppe ecoregion show the highest average inhibitory potential. Further analyses suggest that mainly soil properties and rainfall modulate the structure and function of Streptomyces species, including their antagonistic activity against S. sudanensis.

Genome mining of biosynthetic and chemotherapeutic gene clusters in Streptomyces bacteria

Streptomyces bacteria are known for their prolific production of secondary metabolites, many of which have been widely used in human medicine, agriculture and animal health. To guide the effective prioritization of specific biosynthetic gene clusters (BGCs) for drug development and targeting the most prolific producer strains, knowledge about phylogenetic relationships of Streptomyces species, genome-wide diversity and distribution patterns of BGCs is critical. We used genomic and phylogenetic methods to elucidate the diversity of major classes of BGCs in 1,110 publicly available Streptomyces genomes. Genome mining of Streptomyces reveals high diversity of BGCs and variable distribution patterns in the Streptomyces phylogeny, even among very closely related strains. The most common BGCs are non-ribosomal peptide synthetases, type 1 polyketide synthases, terpenes, and lantipeptides. We also found that numerous Streptomyces species harbor BGCs known to encode antitumor compounds. We observed that strains that are considered the same species can vary tremendously in the BGCs they carry, suggesting that strain-level genome sequencing can uncover high levels of BGC diversity and potentially useful derivatives of any one compound. These findings suggest that a strain-level strategy for exploring secondary metabolites for clinical use provides an alternative or complementary approach to discovering novel pharmaceutical compounds from microbes.

Cyclic di-GMP in Streptomycetes: A New Conformation, New Binding Mode, New Receptor, and a New Mechanism to Control Cell Development.

A recent paper by Gallagher etal. (2020) demonstrates that c-di-GMP controls spore formation in Streptomyces venezuelae through sequestering the sporulation sigma factor σWhiG and presents the crystal structure of a ternary complex between c-di-GMP, σWhiG, and its anti-sigma factor, RsiG.

Final Health and Environmental Risk Assessment of Genetically Modified Maize 59122

Final Health and Environmental Risk Assessment of Genetically Modified Maize 59122

Food, Feed and Environmental Risk Assessment of Glufosinatetolerant Genetically Modified Oilseed Rape T45 for Food and Feed Uses, Import and Processing Under Regulation (EC) No 1829/2003 (Application EFSA/GMO/UK/2005/25)

Food, Feed and Environmental Risk Assessment of Glufosinatetolerant Genetically Modified Oilseed Rape T45 for Food and Feed Uses, Import and Processing Under Regulation (EC) No 1829/2003 (Application EFSA/GMO/UK/2005/25)

Comprehensive Analyses of Cytochrome P450 Monooxygenases and Secondary Metabolite Biosynthetic Gene Clusters in Cyanobacteria.

The prokaryotic phylum Cyanobacteria are some of the oldest known photosynthetic organisms responsible for the oxygenation of the earth. Cyanobacterial species have been recognised as a prosperous source of bioactive secondary metabolites with antibacterial, antiviral, antifungal and/or anticancer activities. Cytochrome P450 monooxygenases (CYPs/P450s) contribute to the production and diversity of various secondary metabolites. To better understand the metabolic potential of cyanobacterial species, we have carried out comprehensive analyses of P450s, predicted secondary metabolite biosynthetic gene clusters (BGCs), and P450s located in secondary metabolite BGCs. Analysis of the genomes of 114 cyanobacterial species identified 341 P450s in 88 species, belonging to 36 families and 79 subfamilies. In total, 770 secondary metabolite BGCs were found in 103 cyanobacterial species. Only 8% of P450s were found to be part of BGCs. Comparative analyses with other bacteria Bacillus, Streptomyces and mycobacterial species have revealed a lower number of P450s and BGCs and a percentage of P450s forming part of BGCs in cyanobacterial species. A mathematical formula presented in this study revealed that cyanobacterial species have the highest gene-cluster diversity percentage compared to Bacillus and mycobacterial species, indicating that these diverse gene clusters are destined to produce different types of secondary metabolites. The study provides fundamental knowledge of P450s and those associated with secondary metabolism in cyanobacterial species, which may illuminate their value for the pharmaceutical and cosmetics industries.

Lethal and sublethal effects of spirotetramat and abamectin on predatory beetles (Menochilus sexmaculatus) via prey (Agonoscena pistaciae) exposure, important for integrated pest management in pistachio orchards

Menochilus sexmaculatus Fabricius (Coleoptera: Coccinellidae) is an important biological control agent in pistachio orchards, especially against Agonoscena pistaciae Burckhardt and Lauterer (Hemiptera: Psyllidae), which is the most damaging pest of pistachio. In this project we exposed M. sexmaculatus adults to two important commonly-used insecticides through feeding on treated prey (A. pistaciae) to evaluate the side-effects on this predator. We tested spirotetramat, which belongs to the keto-enol group inhibiting lipid biosynthesis in insects, at 2/1, 1/1 and 1/2 of the maximum field recommended concentration (MFRC), and abamectin, which is a mixture of avermectins and a natural fermentation product of the bacterium Streptomyces avermitilis, at 1/1, 1/2, 1/4, 1/8 and 1/16 of its MFRC. Spirotetramat did not affect adult survival of M. sexmaculatus at all three concentrations when ingested via treated prey, while in marked contrast abamectin caused 100% adult mortality of M. sexmaculatus when ingested via treated prey at 1/1, 1/2, 1/4 and 1/8 of the MFRC. At sublethal levels, spirotetramat reduced total and daily fecundity of M. sexmaculatus at all three concentrations tested, but did not affect egg hatching at 1/1 and 1/2 of the MFRC. Moreover, prey consumption was decreased when beetles were exposed to the prey treated with spirotetramat at 1/1 and 2/1 of the MFRC concentrations. With abamectin, even at 1/16 of the MFRC, total fecundity, daily fecundity and prey consumption of M. sexmaculatus adults were significantly affected. In conclusion, no acute toxicity was observed on M. sexmaculatus by ingestion of prey treated with spirotetramat, although reproduction parameters and prey consumption were affected at MFRC and lower concentrations. In marked contrast, abamectin was notably very harmful at its MFRC and also at lower concentrations. This research highlighted the importance of toxicity risk assessments, including lethal and sublethal effects, to obtain a more accurate estimation of the compatibility of insecticides in current integrated pest management (IPM) programs.

A new serine protease family with elastase activity is produced by Streptomyces bacteria.

We found an elastolytic activity in the culture supernatant of Streptomyces sp. P-3, and the corresponding enzyme (streptomycetes elastase, SEL) was purified to apparent homogeneity from the culture supernatant. The molecular mass of purified SEL was approximately 18 kDa as judged by SDS-PAGE analysis and gel-filtration chromatography. Utilizing information from N-terminal amino acid sequencing of SEL and mass spectrometry of SEL tryptic fragments, we succeeded in cloning the gene-encoding SEL. The cloned SEL gene contains a 726 bp ORF, which encodes a 241 amino acid polypeptide containing a putative signal peptide for secretion (28 amino acid) and pro-sequence (14 amino acid). Although the deduced primary structure of SEL has sequence similarity to proteins in the S1 protease family, the amino acid sequence shares low identity (< 31.5 %) with any known elastase. SEL efficiently hydrolyses synthetic peptides having Ala or Val in the P1 position such as N-succinyl-Ala-Ala-(Pro or Val)-Ala-p-nitroanilide (pNA), whereas reported proteases by streptomycetes having elastolytic activity prefer large residues, such as Phe and Leu. Compared of kcat/Km ratios for Suc-Ala-Ala-Val-Ala-pNA and Suc-Ala-Ala-Pro-Ala-pNA with subtilisin YaB, which has high elastolytic activity, Streptomyces sp. P-3 SEL exhibits 12- and 121-fold higher, respectively. Phylogenetic analyses indicate that the predicted SEL protein, together with predicted proteins in streptomycetes, constitutes a novel group within the S1 serine protease family. These characteristics suggest that SEL-like proteins are new members of the S1 serine protease family, which display elastolytic activity.

Methodology Development and Validation of Amphotericin B Stability by HPLC‑DAD

The high worldwide consumption of antibiotics and their complex impurity profiles has drawn the attention of the scientific community to the development and validation of stability methods for these drugs. Amphotericin B is an antibiotic that is a natural fermentation product of bacterium Streptomyces nodosus, used as a broad-spectrum antifungal agent, and is highly unstable. For this reason, the objective of this work is the development and validation of an indicative method of stability by high performance liquid chromatography (HPLC) associated to diode array detector (DAD) for amphotericin B (AMB). To achieve this, the chromatographic profiles of acid, basic, oxidative and thermal degradation caused by AMB exposure to water and light were verified by HPLC-DAD, using an isocratic method under the following conditions: C18 chromatographic column (200 × 4.6 mm-5 µm), mobile phase composed of 65:35 of organic phase (methanol/acetonitrile in 41:18)/aqueous phase (2.5 mmol L-1 of disodium edetate, pH 5.0), flow rate of 1.0 mL min-1, injection volume of 20 µL, column temperature 30 ± 2 °C and wavelength of 383 nm. After identification of these profiles, the method was validated according to recommendations of the International Conference on Harmonization guidelines, and then, the active pharmaceutical ingredient (API) peak purity grade, percentage of drug degradation, occurrence of impurities peak and its identification by mass spectrometry (MS) with electrospray ionization (ESI), under positive ionization mode, were evaluated. It is suggested that the main degradation products were amphotericin B (2), degradation product 1 (DP1) and degradation product 2 (DP2) in acid and oxidative medium. Amphotericin B was stable in the presence of water and at 70 ± 2 °C for seven days.

Ivermectin Docks to the SARS-CoV-2 Spike Receptor-binding Domain Attached to ACE2.

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). One drug that has attracted interest is the antiparasitic compound ivermectin, a macrocyclic lactone derived from the bacterium Streptomyces avermitilis. We carried out a docking study to determine if ivermectin might be able to attach to the SARS-CoV-2 spike receptor-binding domain bound with ACE2. We used the program AutoDock Vina Extended to perform the docking study. Ivermectin docked in the region of leucine 91 of the spike and histidine 378 of the ACE2 receptor. The binding energy of ivermectin to the spike-ACE2 complex was -18 kcal/mol and binding constant was 5.8 e-08. The ivermectin docking we identified may interfere with the attachment of the spike to the human cell membrane. Clinical trials now underway should determine whether ivermectin is an effective treatment for SARS-Cov2 infection.

NEW HOPE FOR DEALING WITH “SUPERBUGS”

Introduction: Antimicrobial resistance has become one of the biggest threats to global health. Because of the variety of resistance mechanisms that “superbugs” acquire under the wide antibiotic pressure in medicine, they become difficult to treat with commonly used antibiotics. The spread of so-called ''therapeutic problematic microorganisms'' results in extremely limited treatment options of infections, prolonged hospital stay, increased treatment costs and in many cases, the inability to adequately treat and increased mortality. The new strain of bacteria Streptomyces sp. myrophorea was found to be effective at killing both Gram-positive and Gram-negative multidrug resistant pathogens on the World Health Organization’s priority list. The purpose of this review is to provide information about Streptomyces sp. myrophorea. Materials and methods: a review of scientific literature, studies, and international experience that provide information on Streptomyces sp. myrophorea. Conclusion: The “superbug” problem may be alarming, with news that even our last resort antibiotics are beginning to fail, but there is hope - Streptomyces sp. myrophorea.