Rimosus Research Articles

"Microbial Wars" in a Stirred Tank Bioreactor: Investigating the Co-Cultures of Streptomyces rimosus and Aspergillus terreus, Filamentous Microorganisms Equipped With a Rich Arsenal of Secondary Metabolites.

Microbial co-cultivation is an approach frequently used for the induction of secondary metabolic pathways and the discovery of novel molecules. The studies of this kind are typically focused on the chemical and ecological aspects of inter-species interactions rather than on the bioprocess characterization. In the present work, the co-cultivation of two textbook producers of secondary metabolites, namely Aspergillus terreus (a filamentous fungus used for the manufacturing of lovastatin, a cholesterol-lowering drug) and Streptomyces rimosus (an actinobacterial producer of an antibiotic oxytetracycline) in a 5.5-L stirred tank bioreactor was investigated in the context of metabolic production, utilization of carbon substrates and dissolved oxygen levels. The cultivation runs differed in terms of the applied co-culture initiation strategy and the composition of growth medium. All the experiments were performed in three bioreactors running in parallel (corresponding to a co-culture and two respective monoculture controls). The analysis based upon mass spectrometry and liquid chromatography revealed a broad spectrum of more than 40 secondary metabolites, including the molecules identified as the oxidized derivatives of rimocidin and milbemycin that were observed solely under the conditions of co-cultivation. S. rimosus showed a tendency to dominate over A. terreus, except for the runs where S. rimosus was inoculated into the already developed bioreactor cultures of A. terreus. Despite being dominated, the less aggressive strain still had an observable influence on the production of secondary metabolites and the utilization of substrates in co-culture. The monitoring of dissolved oxygen levels was evaluated as a fast approach of identifying the dominant microorganism during the co-cultivation process.

Streptomyces tardus sp. nov.: A Slow-Growing Actinobacterium Producing Candicidin, Isolated From Sediments of the Trondheim Fjord.

Marine environments are home to an extensive number of microorganisms, many of which remain unexplored for taxonomic novelty and functional capabilities. In this study, a slow-growing Streptomyces strain expressing unique genomic and phenotypic characteristics, P38-E01T, was described using a polyphasic taxonomic approach. This strain is part of a collection of over 8,000 marine Actinobacteria isolates collected in the Trondheim fjord of Norway by SINTEF Industry (Trondheim, Norway) and the Norwegian University of Science and Technology (NTNU, Trondheim, Norway). Strain P38-E01T was isolated from the sediments of the Trondheim fjord, and phylogenetic analyses affiliated this strain with the genus Streptomyces, but it was not closely affiliated with other described species. The closest related type strains were Streptomyces daliensis YIM 31724T (98.6%), Streptomyces rimosus subsp. rimosus ATCC 10970T (98.4%), and Streptomyces sclerotialus NRRL ISP-5269T (98.3%). Predominant fatty acids were C16:0 iso, C16:0, and Summed Feature 3, and the predominant respiratory quinones were MK-10(H6), MK-10(H4), and MK9(H4). The main polar lipids were identified as diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, and phosphoglycolipid. The whole-cell sugars were glucose, ribose, and in minor amounts, mannose. The cell wall peptidoglycan contained LL-diaminopimelic acid. The draft genome has a size of 6.16 Mb, with a %G + C content of 71.4% and is predicted to contain at least 19 biosynthetic gene clusters encoding diverse secondary metabolites. Strain P38-E01T was found to inhibit the growth of the pathogenic yeast Candida albicans ATCC 90028 and a number of Gram-positive bacterial human and plant pathogens. Metabolites extracted from cultures of P38-E01T were analyzed by mass spectrometry, and it was found that the isolate produced the antifungal compound candicidin. Phenotypic and chemotaxonomic signatures, along with phylogenetic analyses, distinguished isolate P38-E01T from its closest neighbors; thus, this isolate represents a novel species of the genus Streptomyces for which the name Streptomyces tardus sp. nov. (P38-E01T = CCM 9049T = DSM 111582T) is proposed.

Presence of the Weakly Pathogenic Fusarium poae in the Fusarium Head Blight Disease Complex Hampers Biocontrol and Chemical Control of the Virulent Fusarium graminearum Pathogen

Fusarium head blight (FHB) in wheat (Triticum aestivum L.) is caused by a consortium of mutually interacting Fusarium species. In the field, the weakly pathogenic F. poae often thrives on the infection sites of the virulent F. graminearum. In this ecological context, we investigated the efficacy of chemical and biocontrol agents against F. graminearum in wheat ears. For this purpose, one fungicide comprising prothioconazole + spiroxamine and two bacterial biocontrol strains, Streptomyces rimosus LMG 19352 and Rhodococcus sp. R-43120 were tested for their efficacy to reduce FHB symptoms and mycotoxin (deoxynivalenol, DON) production by F. graminearum in presence or absence of F. poae. Results showed that the fungicide and both actinobacterial strains reduced FHB symptoms and concomitant DON levels in wheat ears inoculated with F. graminearum. Where Streptomyces rimosus appeared to have direct antagonistic effects, Rhodococcus and the fungicide mediated suppression of F. graminearum was linked to the archetypal salicylic acid and jasmonic acid defense pathways that involve the activation of LOX1, LOX2 and ICS. Remarkably, this chemical- and biocontrol efficacy was significantly reduced when F. poae was co-inoculated with F. graminearum. This reduced efficacy was linked to a suppression of the plant’s intrinsic defense system and increased levels of DON. In conclusion, our study shows that control strategies against the virulent F. graminearum in the disease complex causing FHB are hampered by the presence of the weakly pathogenic F. poae. This study provides generic insights in the complexity of control strategies against plant diseases caused by multiple pathogens.

Multiple copies of the oxytetracycline gene cluster in selected Streptomyces rimosus strains can provide significantly increased titers

BackgroundNatural products are a valuable source of biologically active compounds that have applications in medicine and agriculture. One disadvantage with natural products is the slow, time-consuming strain improvement regimes that are necessary to ensure sufficient quantities of target compounds for commercial production. Although great efforts have been invested in strain selection methods, many of these technologies have not been improved in decades, which might pose a serious threat to the economic and industrial viability of such important bioprocesses.ResultsIn recent years, introduction of extra copies of an entire biosynthetic pathway that encodes a target product in a single microbial host has become a technically feasible approach. However, this often results in minor to moderate increases in target titers. Strain stability and process reproducibility are the other critical factors in the industrial setting. Industrial Streptomyces rimosus strains for production of oxytetracycline are one of the most economically efficient strains ever developed, and thus these represent a very good industrial case. To evaluate the applicability of amplification of an entire gene cluster in a single host strain, we developed and evaluated various gene tools to introduce multiple copies of the entire oxytetracycline gene cluster into three different Streptomyces rimosus strains: wild-type, and medium and high oxytetracycline-producing strains. We evaluated the production levels of these engineered S. rimosus strains with extra copies of the oxytetracycline gene cluster and their stability, and the oxytetracycline gene cluster expression profiles; we also identified the chromosomal integration sites.ConclusionsThis study shows that stable and reproducible increases in target secondary metabolite titers can be achieved in wild-type and in high oxytetracycline-producing strains, which always reflects the metabolic background of each independent S. rimosus strain. Although this approach is technically very demanding and requires systematic effort, when combined with modern strain selection methods, it might constitute a very valuable approach in industrial process development.

Production and statistical optimization of Paromomycin by Streptomyces rimosus NRRL 2455 in solid state fermentation

BackgroundParomomycin is a 2-deoxystreptamine aminocyclitol aminoglycoside antibiotic with broad spectrum activity against Gram-negative, Gram-positive bacteria and many protozoa. This study introduces a strategy for paromomycin production through solid-state fermentation using Streptomyces rimosus subsp. paromomycinus NRRL 2455. Solid state fermentation has gained enormous attention in the development of several products because of their numerous advantages over submerged liquid fermentation. After selecting the best solid substrate, a time course study of paromomycin production was carried out followed by optimization of environmental conditions using response surface methodology. Paromomycin yields obtained using this technique were also compared to those obtained using submerged liquid fermentation.ResultsUpon screening of 6 different substrates, maximum paromomycin concentration (0.51 mg/g initial dry solids) was obtained with the cost-effective agro-industrial byproduct, corn bran, impregnated with aminoglycoside production media. Optimization of environmental conditions using D-optimal design yielded a 4.3-fold enhancement in paromomycin concentration reaching 2.21 mg/g initial dry solids at a pH of 8.5, inoculum size of 5% v/w and a temperature of 30 °C.ConclusionCompared to submerged liquid fermentation, solid state fermentation resulted in comparable paromomycin concentrations, cost reduction of raw materials, less energy consumption and waste water discharge, which have major implications in industrial fermentation. Therefore, solid state fermentation is a promising alternative to submerged liquid fermentation for paromomycin production. To the best of our knowledge, this is the first report on the optimized paromomycin production through solid state fermentation process.

Molecular Biology Methods in Streptomyces rimosus, a Producer of Oxytetracycline.

Streptomyces rimosus is used for production of the broad-spectrum antibiotic oxytetracycline (OTC). S. rimosus belongs to Actinomyces species, a large group of microorganisms that produce diverse set of natural metabolites of high importance in many aspects of our life. In this chapter, we describe specific molecular biology methods and a classical homologous recombination approach for targeted in-frame deletion of a target gene or entire operon in S. rimosus genome. The presented protocols will guide you through the design of experiment and construction of homology arms and their cloning into appropriate vectors, which are suitable for gene-engineering work with S. rimosus. Furthermore, two different protocols for S. rimosus transformation are described including detailed procedure for targeted gene replacement via double crossover recombination event. Gene deletion is confirmed by colony PCR, and colonies are further characterized by cultivation and metabolite analysis. As the final step, we present in trans complementation of the deleted gene, to confirm functionality of the engineering approach achieved by gene disruption. A number of methodological steps and protocols are optimized for S. rimosus strains including the use of the selected reporter genes. Protocols described in this chapter can be applied for studying function of any individual gene product in diverse OTC-producing Streptomyces rimosus strains.

A newly isolated Streptomyces rimosus strain capable of degrading deltamethrin as a pesticide in agricultural soil

Chemical pesticides or insecticides with complex structures are highly abundant in the biosphere and have inevitable side effects on farmland, natural resources, and human health. Deltamethrin is the most popular and widely used pesticide that disrupts the cellular calcium channels. In the present study, isolated strains of bacteria were examined to determine the ones that were capable of degrading deltamethrin. Different species of bacteria were evaluated in terms of the capability to degrade deltamethrin. It is important to note that Streptomyces rimosus was able to degrade up to 200 mg/L deltamethrin concentration and could be grown in mineral salt medium agar containing deltamethrin to be used as a source of carbon and energy. The results demonstrated that there is a diversity of deltamethrin-degrading bacteria in agricultural soil ecosystems. The application of these bacteria, especially S. rimosus, might be used as a bioremediation technique to decrease pesticide contamination of the ecosystem.

Effect of microbial non-volatile metabolites on growth of Fusarium oxysporum f. sp. ciceri and lini

Rhizospheric fungal antagonists activity against phytopathogen by production of volatile and non-volatile metabolites. In vitro assessments revealed their antibiosis and mycoparasitic ability to affect growth of the pathogen. Soil borne fungi, bacteria and actinomycetes were identified and used as antagonistic microbes against some soil borne pathogens. The efficacy of volatile and non-volatile metabolites produced by 14 selected dominant microbes against the common fungal test pathogens (F. oxysporum f. sp. ciceri & lini) were suppressed with colony interaction, volatile, and non-volatile metabolites emanated from different leaf surface microbes like Aspergillus niger, A. terreus, A. luchuensis, Trichoderma harzianum, Penicillium citrinum, Streptomyces rimosus, and S. aureofaciens and prior application of these antagonists effectively reduced the development of Fusarium wilt in chick pea and linseed.

Harnessing the intracellular triacylglycerols for titer improvement of polyketides in Streptomyces.

Pharmaceutically important polyketides such as avermectin are mainly produced as secondary metabolites during the stationary phase of growth of Streptomyces species in fermenters. The source of intracellular metabolites that are funneled into polyketide biosynthesis has proven elusive. We applied multi-omics to reveal that intracellular triacylglycerols (TAGs), which accumulates in primary metabolism, are degraded during stationary phase. This process could channel carbon flux from both intracellular TAGs and extracellular substrates into polyketide biosynthesis. We devised a strategy named 'dynamic degradation of TAG' (ddTAG) to mobilize the TAG pool and increase polyketide biosynthesis. Using ddTAG we increased the titers of actinorhodin, jadomycin B, oxytetracycline and avermectin B1a in Streptomyces coelicolor, Streptomyces venezuelae, Streptomyces rimosus and Streptomyces avermitilis. Application of ddTAG increased the titer of avermectin B1a by 50% to 9.31 g l-1 in a 180-m3 industrial-scale fermentation, which is the highest titer ever reported. Our strategy could improve polyketide titers for pharmaceutical production.

Extracellular production of the engineered thermostable protease pernisine from Aeropyrum pernix K1 in Streptomyces rimosus

BackgroundThe thermostable serine protease pernisine originates from the hyperthermophilic Archaeaon Aeropyrum pernix and has valuable industrial applications. Due to its properties, A. pernix cannot be cultivated in standard industrial fermentation facilities. Furthermore, pernisine is a demanding target for heterologous expression in mesophilic heterologous hosts due to the relatively complex processing step involved in its activation.ResultsWe achieved production of active extracellular pernisine in a Streptomyces rimosus host through heterologous expression of the codon-optimised gene by applying step-by-step protein engineering approaches. To ensure secretion of fully active enzyme, the srT signal sequence from the S. rimosus protease was fused to pernisine. To promote correct processing and folding of pernisine, the srT functional cleavage site motif was fused directly to the core pernisine sequence, this way omitting the proregion. Comparative biochemical analysis of the wild-type and recombinant pernisine confirmed that the enzyme produced by S. rimosus retained all of the desired properties of native pernisine. Importantly, the recombinant pernisine also degraded cellular and infectious bovine prion proteins, which is one of the particular applications of this protease.ConclusionFunctional pernisine that retains all of the advantageous properties of the native enzyme from the thermophilic host was successfully produced in a S. rimosus heterologous host. Importantly, we achieved extracellular production of active pernisine, which significantly simplifies further downstream procedures and also omits the need for any pre-processing step for its activation. We demonstrate that S. rimosus can be used as an attractive host for industrial production of recombinant proteins that originate from thermophilic organisms.

Production of antibiotic of Microlides group from a local isolate of the bacterium Streptomyces rimosus

Three local isolates of the genus Streptomyces were obtained from ten soil samples collected from different places of Niniva governorate. The isolates showed bioactivity against test Gram positive and negative bacteria and two Genera of fungi. Streptomyces isolates were identified according to microscopic and morphological tests. The Streptomyces isolates were subjected to ultra violet ray for different time periods. The treated isolates showed little increase in inhibition zone and treated period of (5) minutes showed the best inhibition of the test bacteria. The Streptomyces isolates treated with N – methyl – N – nitro – N – nitroso guaniction at the concentration of (50) showed clear increase of antibiotic production. The isolate Streptomyces III showed clear inhibition against Escherichia coli, Pseudomonas aeruginosa and Bacillus subtilis and the diameter inhibition zone was (20,29,20)mm respectively.

Reclassification of Streptomyces rimosus subsp. paromomycinus as Streptomyces paromomycinus sp. nov.

Streptomyces rimosus is currently composed of two subspecies: Streptomyces rimosus subsp. rimosus and Streptomyces rimosus subsp. paromomycinus. The 16S rRNA gene similarity between type strains of these two subspecies is 99.03 %, whereas that between S. rimosus subsp. paromomycinus and Streptomyces chrestomyceticusis 100 %. To assess the taxonomic status of S. rimosus subsp. paromomycinus, genome sequencing was performed on the type strains of S. rimosus subsp. paromomycinus and S. chrestomyceticus. Digital DNA-DNA hybridization values between S. rimosus subsp. paromomycinus NBRC 15454T and S. rimosus subsp. rimosus ATCC 10970T and between S. rimosus subsp. paromomycinus NBRC 15454T and S. chrestomyceticus NBRC 13444T were 35.4 and 59.9 %, respectively, which are less than the thresholds for bacterial species delineation and indicate that S. rimosus subsp. paromomycinus is not S. rimosus, but an independent species different from S. rimosus and S. chrestomyceticus. In addition, phenotypic data also support that S. rimosus subsp. paromomycinus is distinct from S. chrestomyceticus. Therefore, S. rimosus subsp. paromomycinus should be reclassified as a novel species, for which we propose the name Streptomyces paromomycinus sp. nov. The type strain is NBRC 15454T (=ATCC 14827T=DSM 41429T=JCM 4541T=JCM 4871T=NRRL 2455T=VKM Ac-605T).

Sequential improvement of rimocidin production in Streptomyces rimosus M527 by introduction of cumulative drug-resistance mutations.

Rimocidin is a polyene macrolide that exhibits a strong inhibitory activity against a broad range of plant-pathogenic fungi. In this study, fermentation optimization and ribosome engineering technology were employed to enhance rimocidin production in Streptomyces rimosus M527. After the optimization of fermentation, rimocidin production in S. rimosus M527 increased from 0.11 ± 0.01 to 0.23 ± 0.02g/L during shake-flask experiments and reached 0.41 ± 0.05g/L using 5-L fermentor. Fermentation optimization was followed by the generation of mutants of S. rimosus M527 through treatment of the strain with different concentrations of gentamycin (Gen) or rifamycin. One Genr mutant named S. rimosus M527-G37 and one Rifr mutant named S. rimosus M527-R5 showed increased rimocidin production. Double-resistant (Genr and Rifr) mutants were selected using S. rimosus M527-G37 and S. rimosus M527-R5, and subsequently tested. One mutant, S. rimosus M527-GR7, which was derived from M527-G37, achieved the greatest cumulative improvement in rimocidin production. In the 5-L fermentor, the maximum rimocidin production achieved by S. rimosus M527-GR7 was 25.36% and 62.89% greater than those achieved by S. rimosus M527-G37 and the wild-type strain S. rimosus M527, respectively. Moreover, in the mutants S. rimosus M527-G37 and S. rimosus M527-GR7 the transcriptional levels of ten genes (rimAsr to rimKsr) located in the gene cluster involved in rimocidin biosynthesis were all higher than those in the parental strain M527 to varying degrees. In addition, after expression of the single rimocidin biosynthetic genes in S. rimosus M527 a few recombinants showed an increase in rimocidin production. Expression of rimE led to the highest production.

Biosorption of Lead(II) from Aqueous Solution by Raw and Treated <i>Streptomyces rimosus</i>: Equilibrium, Kinetic and Thermodynamic Studies

This study examines the elimination potential of bacterial biomass (Streptomyces rimosus) against lead (II). The biomass was used in a raw state and treated with NaOH. The adsorption was influenced by pH, temperature and contact time. Treated biomass eliminates the largest amount of lead (II) compared to raw biomass. The Langmuir isotherm is the most appropriate to describe our results with maximum capacity of 47.63 mg/g for lead at 45°C. These results suggest that microbial species as bacterial biomass can be used successfully in the treatment of wastewater contaminated by pollutants such as heavy metals.

Within-Species Genomic Variation and Variable Patterns of Recombination in the Tetracycline Producer Streptomyces rimosus.

Streptomyces rimosus is best known as the primary source of the tetracycline class of antibiotics, most notably oxytetracycline, which have been widely used against many gram-positive and gram-negative pathogens and protozoan parasites. However, despite the medical and agricultural importance of S. rimosus, little is known of its evolutionary history and genome dynamics. In this study, we aim to elucidate the pan-genome characteristics and phylogenetic relationships of 32 S. rimosus genomes. The S. rimosus pan-genome contains more than 22,000 orthologous gene clusters, and approximately 8.8% of these genes constitutes the core genome. A large part of the accessory genome is composed of 9,646 strain-specific genes. S. rimosus exhibits an open pan-genome (decay parameter α = 0.83) and high gene diversity between strains (genomic fluidity φ = 0.12). We also observed strain-level variation in the distribution and abundance of biosynthetic gene clusters (BGCs) and that each individual S. rimosus genome has a unique repertoire of BGCs. Lastly, we observed variation in recombination, with some strains donating or receiving DNA more often than others, strains that tend to frequently recombine with specific partners, genes that often experience recombination more than others, and variable sizes of recombined DNA sequences. We conclude that the high levels of inter-strain genomic variation in S. rimosus is partly explained by differences in recombination among strains. These results have important implications on current efforts for natural drug discovery, the ecological role of strain-level variation in microbial populations, and addressing the fundamental question of why microbes have pan-genomes.

Paromomycin production from Streptomyces rimosus NRRL 2455: statistical optimization and new synergistic antibiotic combinations against multidrug resistant pathogens

BackgroundResponse surface methodology (RSM) employing Box-Behnken design was used to optimize the environmental factors for the production of paromomycin, a 2 deoxystreptamine aminocyclitol aminoglycoside antibiotic, (2DOS-ACAGA) from Streptomyces (S.) rimosus NRRL 2455. Emergence of bacterial resistance caught our attention to consider the combination of antimicrobial agents. The effect of paromomycin combination with other antimicrobial agents was tested on some multiple drug resistant isolates. To the best of our knowledge, this is the first report on optimization of paromomycin production from S. rimosus NRRL 2455. A Quadratic model and response surface method were used by choosing three model factors; pH, incubation time and inoculum size. A total of 17 experiments were done and the response of each experiment was recorded. Concerning the effect of combining paromomycin with different antimicrobial agents, it was tested using the checkerboard assay against six multidrug resistant (MDR) pathogens including; Pseudomonas (P.) aeruginosa (2 isolates), Klebsiella (K.) pneumoniae, Escherichia (E.) coli, methicillin sensitive Staphylococcus aureus (MSSA) and methicillin resistant Staphylococcus aureus (MRSA). Paromomycin was tested in combination with ceftriaxone, ciprofloxacin, ampicillin/sulbactam, azithromycin, clindamycin and doxycycline.ResultsThe optimum conditions for paromomycin production were a pH of 6, an incubation time of 8.5 days and an inoculum size of 5.5% v/v using the optimized media (soybean meal 30 g/L, NH4CL 4 g/L, CaCO3 5 g/L and glycerol 40 ml/L), 28 °C incubation temperature, and 200 rpm agitation rate that resulted in 14 fold increase in paromomycin production as compared to preliminary fermentation level using the basal medium. The tested antibiotic combinations showed either synergistic effect on paromomycin activity on most of the tested MDR pathogens (45.83%), additive effect in 41.67% or indifferent effect in 12.5%.ConclusionRSM using multifactorial design was a helpful and a reliable method for paromomycin production. Paromomycin combination with ceftriaxone, ciprofloxacin, ampicillin/sulbactam, azithromycin, clindamycin or doxycycline showed mostly synergistic effect on certain selected clinically important MDR pathogens.

Isolation and Screening of some Actinomycetes from Soil from Damietta and Mansoura and its Antimicrobial Activities

Sixty two isolates of actinomycetes were isolated from different eight sandy and clay soils from various locations of Egypt (Damietta and Mansoura) during the year 2016. All isolates of actinomycetes were screened against known different types of bacteria and fungi like: three Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa and Klebsiella pneumoniae), two Gram-positive bacteria (Staphylococcus aureus and Bacillus cereus) and one fungus (Candida albicans). About nineteen strains of actinomycetes were isolated which had different antibacterial and antifungal activity. Two potent actinomycetes isolates were selected to be identified according to their morphological, physiological and biochemical characterization and identified as Streptomyces rimosus.

New Gene of Aminoglycoside Phosphotransferase aph(3'')-Id from Streptomyces rimosus ATCC10970, Encoding Streptomycin Resistance

Previously, in the strain Streptomyces rimosus ATCC10970 (producer of oxytetracycline), the aminoglycoside phosphotransferase AphVIII, determining kanamycin, neomycin, and paromomycin resistance, was identified and characterized. Recently, the authors obtained the 3D structure of AphVIII. The 14 aph genes, including gene aphVIII, were annotated when the genome of S. rimosus ATCC10970 was sequenced. In the present study, a new aph(3'')-Id (aphSR3) gene encoding streptomycin phosphotansferase was first identified in the strain of S. rimosus ATCC10970 using bioinformatic and comparative phylogenetic analysis of the aphSR1-aphSR14 genes with the previously known aph genes from clinical isolates and producer strains of aminoglycoside antibiotics belonging to seven subfamilies. When cloning, it was found that the gene aphSR3 (aph(3'')-Id) in Escherichia coli causes resistance to streptomycin at a concentration of 150 μg/mL. The obtained data can be used in practical terms to study the distribution and features of the functions of genes that determine the natural resistance to aminoglycoside antibiotics in actinobacteria of the genus Streptomyces.

Competitive biosorption of heavy metals from aqueous solutions onto Streptomyces rimosus

The aim of this study is the simultaneous biosorption of Cd+2 and Ni+2 on a dead biomass, Streptomyces rimosus pretreated with NaOH (0,1 M). Kinetic tests were carried out for the binary mixture (cadmium-nickel) during 6 hours contact time to ensure that balance was reached. The amounts adsorbed at equilibrium were 22.8 mg Ni2+/g and 9.86 mg Cd2+/g biomass, respectively. The biosorption depends mainly by some parameters, such as the pH, the initial concentration of metal and the initial concentration of biomass. The isotherm of adsorption according to two models, Langmuir and Freundlich, was carried out in our study. The results of the kinetics of adsorption show that the experimental values are well represented by the kinetic model of pseudo-second order. This enables us to determine the behavior of these adsorbents with respect to a real industrial effluent.

Production of extracellular heterologous proteins in Streptomyces rimosus, producer of the antibiotic oxytetracycline.

Among the Streptomyces species, Streptomyces lividans has often been used for the production of heterologous proteins as it can secrete target proteins directly into the culture medium. Streptomyces rimosus, on the other hand, has for long been used at an industrial scale for oxytetracycline production, and it holds 'Generally Recognised As Safe' status. There are a number of properties of S. rimosus that make this industrial strain an attractive candidate as a host for heterologous protein production, including (1) rapid growth rate; (2) growth as short fragments, as for Escherichia coli; (3) high efficiency of transformation by electroporation; and (4) secretion of proteins into the culture medium. In this study, we specifically focused our efforts on an exploration of the use of the Sec secretory pathway to export heterologous proteins in a S. rimosus host. We aimed to develop a genetic tool kit for S. rimosus and to evaluate the extracellular production of target heterologous proteins of this industrial host. This study demonstrates that S. rimosus can produce the industrially important enzyme phytase AppA extracellularly, and analogous to E. coli as a host, application of His-Tag/Ni-affinity chromatography provides a simple and rapid approach to purify active phytase AppA in S. rimosus. We thus demonstrate that S. rimosus can be used as a potential alternative protein expression system.