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Micromonospora strains proved to be a model organism for drug discovery and plant growth promotion (PGP). Strain DSM 115977 T was subjected to polyphasic taxonomic analysis and genome mining for biosynthetic gene clusters and PGP-associated genes in order to determine its taxonomic rank and assess its biosynthetic potential. The strain was found to form a novel species within the evolutionary radiation of the genus Micromonospora . The strain contained glucose, mannose, xylose, and ribose as whole-cell sugars and the isomer DL-diaminopimelic acid in its peptidoglycan. Strain DSM 115977 T had iso -C 15:0 , iso -C 16:0 , C 17:1 cis 9, C 17:0 , iso -C 17:0 , and 10-methyl - C 17:0 as fatty acid profile (>5%) and MK10-H 4 and MK10-H 6 as the predominant menaquinones (>10%). The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, glycophosphatidylinositol, glycophospholipids, phosphoaminolipid, unidentified lipids, and phospholipids. The genome of the strain had a size of 7.0 Mbp with a DNA G + C content of 73.4%. It formed a well-supported sub-clade with its close phylogenomic neighbor, Micromonospora echinofusca DSM 43913 T (98.7%). Digital DNA-DNA hybridization and average nucleotide identity derived from sequence comparisons between the strain and its close phylogenomic neighbors were below the thresholds of 70 and 95-96% for prokaryotic species demarcation, respectively. Based on these findings, strain DSM 115977 T (Asg4 T = KCTC 59188 T ) merits to be considered as the type strain of a new species for which the name Micromonospora reichwaldensis sp. nov. is proposed. Genome mining for biosynthetic gene clusters encoding specialized secondary metabolites highlighted its ability to produce potentially novel therapeutic compounds. The strain is rich in plant growth-promoting genes whose predicted products directly and indirectly affect the development and immune system of the plant., Importance: In view of the significant pharmaceutical, biotechnological, and ecological potentials of micromonosporae, it is particularly interesting to enhance the genetic diversity of this genus by focusing on the isolation of novel strain from underexplored habitats, with the promise that novel bacteria will lead to new chemical entities. In this report, modern polyphasic taxonomic study confirmed the assignment of strain DSM 115977 T to a novel species for which the name Micromonospora reichwaldensis sp. nov. is proposed. The strain harbors in its genomic sequence several biosynthetic gene clusters for secondary metabolites and genes associated with plant growth-promoting features. The results of this study provide a very useful basis for launching more in-depth research into agriculture and/or drug discovery.

Large-scale fermentations (»100 m³) often encounter concentration gradients which may significantly affect microbial activities and production performance. Reliably investigating such scenarios in silico would allow to optimize bioproduction. But related simulations are very rare in particular for large bubble columns. Here, we pioneer the spatially resolved investigation of a 600 m³ bubble column operating for Escherichia coli based l-phenylalanine fed-batch production. Microbial kinetics are derived from experimental data. Advanced Euler-Lagrange (EL) computational fluid dynamics (CFD) simulations are applied to track individual bubble dynamics that result from a recently developed bubble breakage model. Thereon, the complex nonlinear characteristics of hydrodynamics, mass transfer, and microbial activities are simulated for large scale and compared with real data. As a key characteristic, zones for upriser, downcomer, and circulation cells were identified that dominate mixing and mass transfer. This results in complex gradients of glucose, dissolved oxygen, and microbial rates dividing the bioreactor into sections. Consequently, alternate feed designs are evaluated splitting real feed rates in two feeds at different locations. The opposite reversed installation of feed spots and spargers improved the product synthesis by 6.24% while alternate scenarios increased the growth rate by 11.05%. The results demonstrate how sophisticated, spatially resolved simulations of hydrodynamics, mass transfer, and microbial kinetics help to optimize bioreactors in silico., (© 2024 The Author(s). Biotechnology and Bioengineering published by Wiley Periodicals LLC.)

Specialized or secondary metabolites are small molecules of biological origin, often showing potent biological activities with applications in agriculture, engineering and medicine. Usually, the biosynthesis of these natural products is governed by sets of co-regulated and physically clustered genes known as biosynthetic gene clusters (BGCs). To share information about BGCs in a standardized and machine-readable way, the Minimum Information about a Biosynthetic Gene cluster (MIBiG) data standard and repository was initiated in 2015. Since its conception, MIBiG has been regularly updated to expand data coverage and remain up to date with innovations in natural product research. Here, we describe MIBiG version 4.0, an extensive update to the data repository and the underlying data standard. In a massive community annotation effort, 267 contributors performed 8304 edits, creating 557 new entries and modifying 590 existing entries, resulting in a new total of 3059 curated entries in MIBiG. Particular attention was paid to ensuring high data quality, with automated data validation using a newly developed custom submission portal prototype, paired with a novel peer-reviewing model. MIBiG 4.0 also takes steps towards a rolling release model and a broader involvement of the scientific community. MIBiG 4.0 is accessible online at https://mibig.secondarymetabolites.org/., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Actinomycetes are a prolific source of bioactive natural compounds many of which are used as antibiotics or other drugs. In this study we investigated the genomic and biochemical diversity of 32 actinobacterial strains that had been deposited at the DSMZ-German Collection of Microorganisms and Cell Cultures decades ago. Genome-based phylogeny and in silico DNA-DNA hybridization supported the assignment of these strains to 26 novel species and two novel subspecies and a reclassification of a Streptomyces species. These results were consistent with the biochemical, enzymatic, and chemotaxonomic features of the strains. Most of the strains showed antimicrobial activities against a range of Gram-positive and Gram-negative bacteria, and against yeast. Genomic analysis revealed the presence of numerous unique biosynthetic gene clusters (BGCs) encoding for potential novel antibiotic and anti-cancer compounds. Strains DSM 41636 T and DSM 61640 T produced the antibiotic compounds A33853 and SF2768, respectively. Overall, this reflects the significant pharmaceutical and biotechnological potential of the proposed novel type strains and underlines the role of prokaryotic systematics for drug discovery. In order to compensate for the gender gap in naming prokaryotic species, we propose the eponyms for all newly described species to honour female scientists., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Streptomyces sp. DSM 41014, DSM 41527, and DSM 41981 are three strains from the DSMZ strain collection. Here, we present the draft genome sequences of DSM 41014, DSM 41527, and DSM 41981 with a size of 9.09 Mb, 8.45 Mb, and 9.23 Mb, respectively., Competing Interests: The authors declare that there are no conflicts of interest., (Copyright © 2024 The Authors.)

The taxonomic position of three actinobacterial strains, BCCO 10&#95;0061 T , BCCO 10&#95;0798 T , and BCCO 10&#95;0856 T , recovered from bare soil in the Sokolov Coal Basin, Czech Republic, was established using a polyphasic approach. The multilocus sequence analysis based on 100 single-copy genes positioned BCCO 10&#95;0061 T in the same cluster as Lentzea waywayandensis , strain BCCO 10&#95;0798 T in the same cluster as Lentzea flaviverrucosa , Lentzea californiensis , Lentzea violacea , and Lentzea albidocapillata , and strain BCCO 10&#95;0856 T clustered together with Lentzea kentuckyensis and Lentzea alba . Morphological and chemotaxonomic characteristics of these strains support their assignment to the genus Lentzea . In all three strains, MK-9(H 4 ) accounted for more than 80 % of the isoprenoid quinone. The diagnostic diamino acid in the cell-wall peptidoglycan was meso -diaminopimelic acid. The whole-cell sugars were rhamnose, ribose, mannose, glucose, and galactose. The major fatty acids (>10 %) were iso-C 15 : 0 , anteiso-C 15 : 0 , iso-C 16 : 0 , and C 16 : 0 . The polar lipids were diphosphatidylglycerol, methyl-phosphatidylethanolamine, phosphatidylethanolamine, hydroxy-phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylinositol. The genomic DNA G+C content of strains (mol%) was 68.8 for BCCO 10&#95;0061 T , 69.2 for BCCO 10&#95;0798 T , and 68.5 for BCCO 10&#95;0856 T . The combination of digital DNA-DNA hybridization results, average nucleotide identity values and phenotypic characteristics of BCCO 10&#95;0061 T , BCCO 10&#95;0798 T , and BCCO 10&#95;0856 T distinguishes them from their closely related strains. Bioinformatic analysis of the genome sequences of the strains revealed several biosynthetic gene clusters (BGCs) with identities >50 % to already known clusters, including BGCs for geosmin, coelichelin, ε-poly-l-lysine, and erythromycin-like BGCs. Most of the identified BGCs showed low similarity to known BGCs (<50 %) suggesting their genetic potential for the biosynthesis of novel secondary metabolites. Based on the above results, each strain represents a novel species of the genus Lentzea , for which we propose the name Lentzea sokolovensis sp. nov. for BCCO 10&#95;0061 T (=DSM 116175 T ), Lentzea kristufekii sp. nov. for BCCO 10&#95;0798 T (=DSM 116176 T ), and Lentzea miocenica sp. nov. for BCCO 10&#95;0856 T (=DSM 116177 T ).

Strain collections are a treasure chest of numerous valuable and taxonomically validated bioresources. The Leibniz Institute DSMZ is one of the largest and most diverse microbial strain collections worldwide, with a long tradition of actinomycetes research. Actinomycetes, especially the genus Streptomyces , are renowned as prolific producers of antibiotics and many other bioactive natural products. In light of this, five Streptomyces strains, DSM 40971 T , DSM 40484 T , DSM 40713 T , DSM 40976 T , and DSM 40907 T , which had been deposited a long time ago without comprehensive characterization, were the subject of polyphasic taxonomic studies and genome mining for natural compounds based on in vitro and in silico analyses. Phenotypic, genetic, and phylogenomic studies distinguished the strains from their closely related neighbors. The digital DNA-DNA hybridization and average nucleotide identity values between the five strains and their close, validly named species were below the threshold of 70% and 95%-96%, respectively, determined for prokaryotic species demarcation. Therefore, the five strains merit being considered as novel Streptomyces species, for which the names Streptomyces kutzneri sp. nov., Streptomyces stackebrandtii sp. nov. , Streptomyces zähneri sp. nov. , Streptomyces winkii sp. nov., and Streptomyces kroppenstedtii sp. nov. are proposed. Bioinformatics analysis of the genome sequences of the five strains revealed their genetic potential for the production of secondary metabolites, which helped identify the natural compounds cinerubin B from strain DSM 40484 T and the phosphonate antibiotic phosphonoalamide from strain DSM 40907 T and highlighted strain DSM 40976 T as a candidate for regulator-guided gene cluster activation due to the abundance of numerous " Streptomyces antibiotic regulatory protein" (SARP) genes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Nouioui, Zimmermann, Hennrich, Xia, Rössler, Makitrynskyy, Pablo Gomez-Escribano, Pötter, Jando, Döppner, Wolf, Neumann-Schaal, Hughes and Mast.)

Strain TÜ4103 T was originally sampled from Java, Indonesia and deposited in the Tübingen strain collection under the name ' Streptomyces sp.'. The strain was found to be an antibiotic producer as strain TÜ4103 T showed bioactivity against Gram-positive bacteria, such as Bacillus subtilis and Kocuria rhizophila in bioassays. Strain TÜ4103 T showed 16S rRNA gene sequence similarity of 99.65 % to Kitasatospora cheerisanensis DSM 101999 T and 98.82 % to Kitasatospora niigatensis DSM 44781 T and Kitasatospora cineracea DSM 44780 T . Genome-based phylogenetic analysis revealed that strain TÜ4103 T is closely related to K. cineracea DSM 44780 T and K. niigatensis DSM 44781 T . The digital DNA-DNA hybridization values between the genome sequences of strain TÜ4103 T and its closest phylogenomic relatives, strains DSM 44780 T and DSM 44781 T , were 43.0 and 42.9 %, respectively. Average nucleotide identity (ANI) values support this claim, with the highest ANI score of 91.14 % between TÜ4103 T and K. niigatensis being closely followed by an ANI value of 91.10 % between K. cineracea and TÜ4103 T . The genome of TÜ4103 T has a size of 7.91 Mb with a G+C content of 74.05 mol%. Whole-cell hydrolysates of strain TÜ4103 T are rich in meso -diaminopimelic acid, and rhamnose, galactose and mannose are characteristic as whole-cell sugars. The phospholipid profile contains phosphatidylethanolamine, diphosphatidylglycerol and glycophospholipid. The predominant menaquinones (>93.5 %) are MK-9(H 8 ) and MK-9(H 6 ). Based on the phenotypic, genotypic and genomic characteristics, strain TÜ4103 T (=DSM 114396 T =CECT 30712 T ) merits recognition as the type strain of a novel species of the genus Kitasatospora , for which the name Kitasatospora fiedleri sp. nov. is proposed.

Streptogramins are the last line of defense antimicrobials with pristinamycin as a representative substance used as therapeutics against highly resistant pathogenic bacteria. However, the emergence of (multi)drug-resistant pathogens renders these valuable antibiotics useless; making it necessary to derivatize compounds for new compound characteristics, which is often difficult by chemical de novo synthesis due to the complex nature of the molecules. An alternative to substance derivatization is mutasynthesis. Herein, we report about a mutasynthesis approach, targeting the phenylglycine (Phg) residue for substance derivatization, a pivotal component of streptogramin antibiotics. Mutasynthesis with halogenated Phg(-like) derivatives altogether led to the production of two new derivatized natural compounds, as there are 6-chloropristinamycin I and 6-fluoropristinamycin I based on LC-MS/MS analysis. 6-Chloropristinamycin I and 6-fluoropristinamycin I were isolated by preparative HPLC, structurally confirmed using NMR spectroscopy and tested for antimicrobial bioactivity. In a whole-cell biotransformation approach using an engineered E. coli BL21(DE3) pET28- hmo /pACYC- bcd-gdh strain, Phg derivatives were generated fermentatively. Supplementation with the E. coli biotransformation fermentation broth containing 4-fluorophenylglycine to the pristinamycin mutasynthesis strain resulted in the production of 6-fluoropristinamycin I, demonstrating an advanced level of mutasynthesis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (This journal is © The Royal Society of Chemistry.)

Strains USC-21046 T and USC-21048 T were isolated from foaming coastal marine waters on the Sunshine Coast, Queensland, Australia. Both strains displayed growth and morphological characteristics typical for members belonging to the genus Nocardia . The major polar lipids were diphosphatidylglycerol and phosphatidylethanolamine, and the major fatty acids were C 16 : 0 , C 18 : 1  ω9 c , C 18 : 0 and C 18 : 0 10-methyl. The mycolic acids of strains USC-21046 T and USC-21048 T consisted of chain lengths between 50-64 and 56-68, respectively. Moreover, both of those strains contained meso -diaminopimelic acid and ribose, arabinose, glucose and galactose as whole cell sugars. Based on the phylogenomic results, both strains belonged to the genus Nocardia with strain USC-21046 T showing an 80.4 % genome similarity to N. vinacea NBRC 16497 T and N. pseudovaccinii NBRC 100343 T , whereas USC-21048 T strain showed an 83.6 % genome similarity to N. aobensis NBRC 100429 T . Both strains were delineated from their closely related relatives based on physiological (e.g. growth on sole carbon source) and chemotaxonomic (e.g. cellular fatty composition) differences. The digital DNA-DNA hybridization (dDDH) values between USC-21046 T and USC-21048 T and their closely related relatives were below the dDDH threshold value of ≤70 % used for the taxonomic classification of novel species status. The genome length of strains USC-21046 T and USC-21048 T were 6 878 863 and 7 066 978 bp, with G+C contents of 65.2 and 67.8 mol%, respectively. For the novel isolates, we propose the names Nocardia australiensis sp. nov. with the type strain USC-21046 T (=DSM 111727 T =NCCB 100867 T ) and Nocardia spumae sp. nov. with the type strain USC-21048 T (=DSM 111726 T =NCCB 100868 T ).

Today, one of the biggest challenges in antibiotic research is a targeted prioritization of natural compound producer strains and an efficient dereplication process to avoid undesired rediscovery of already known substances. Thereby, genome sequence-driven mining strategies are often superior to wet-lab experiments because they are generally faster and less resource-intensive. In the current study, we report on the development of a novel in silico screening approach to evaluate the genetic potential of bacterial strains to produce protein synthesis inhibitors (PSI), which was termed the protein synthesis inhibitor ('psi') target gene footprinting approach = Ψ-footprinting. The strategy is based on the occurrence of protein synthesis associated self-resistance genes in genome sequences of natural compound producers. The screening approach was applied to 406 genome sequences of actinomycetes strains from the DSMZ strain collection, resulting in the prioritization of 15 potential PSI producer strains. For twelve of them, extract samples showed protein synthesis inhibitory properties in in vitro transcription/translation assays. For four strains, namely Saccharopolyspora flava DSM 44771, Micromonospora aurantiaca DSM 43813, Nocardioides albertanoniae DSM 25218, and Geodermatophilus nigrescens DSM 45408, the protein synthesis inhibitory substance amicoumacin was identified by HPLC-MS analysis, which proved the functionality of the in silico screening approach., (© The Author(s) 2022. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Actinobacteria is an ancient phylum of Gram-positive bacteria with a characteristic high GC content to their DNA. The ActinoBase Wiki is focused on the filamentous actinobacteria, such as Streptomyces species, and the techniques and growth conditions used to study them. These organisms are studied because of their complex developmental life cycles and diverse specialised metabolism which produces many of the antibiotics currently used in the clinic. ActinoBase is a community effort that provides valuable and freely accessible resources, including protocols and practical information about filamentous actinobacteria. It is aimed at enabling knowledge exchange between members of the international research community working with these fascinating bacteria. ActinoBase is an anchor platform that underpins worldwide efforts to understand the ecology, biology and metabolic potential of these organisms. There are two key differences that set ActinoBase apart from other Wiki-based platforms: [1] ActinoBase is specifically aimed at researchers working on filamentous actinobacteria and is tailored to help users overcome challenges working with these bacteria and [2] it provides a freely accessible resource with global networking opportunities for researchers with a broad range of experience in this field.

A chemical reinvestigation of the Indonesian strain Streptomyces sp. SHP 22-7 led to the isolation of three new pyrimidine nucleosides, along with six known analogues and zincphyrin. The structures of the new compounds ( 6 , 7 , 10 ) were elucidated by employing spectroscopic techniques (NMR, MS, CD, and IR) as well as enantioselective analyses of methyl branched side chain configurations. Application of the precursor-directed feeding approach led to the production and partial isolation of nine further pyrimidine analogues. The new compounds 6 , 7 , and 11 and three of the known compounds ( 2 - 4 ) were found to possess antimycobacterial and cytotoxic properties.

Biopharmaceutical production processes often use mammalian cells in bioreactors larger than 10,000 L, where gradients of shear stress, substrate, dissolved oxygen and carbon dioxide, and pH are likely to occur. As former tissue cells, producer cell lines such as Chinese hamster ovary (CHO) cells sensitively respond to these mixing heterogeneities, resulting in related scenarios being mimicked in scale-down reactors. However, commonly applied multi-compartment approaches comprising multiple reactors impose a biasing shear stress caused by pumping. The latter can be prevented using the single multi-compartment bioreactor (SMCB) presented here. The exchange area provided by a disc mounted between the upper and lower compartments in a stirred bioreactor was found to be an essential design parameter. Mimicking the mixing power input at a large scale on a small scale allowed the installation of similar mixing times in the SMCB. The particularities of the disc geometry may also be considered, finally leading to a converged decision tree. The work flow identifies a sharply contoured operational field comprising disc designs and power input to install the same mixing times on a large scale in the SMCB without the additional shear stress caused by pumping. The design principle holds true for both nongassed and gassed systems., Competing Interests: The authors have declared no conflicts of interest., (© 2022 The Authors. Engineering in Life Sciences published by Wiley‐VCH GmbH.)

Amycolatopsis sp. strain DSM 110486 and Pseudonocardia sp. strain DSM 110487 are two novel actinomycete species that were isolated from Hengam Island beach sand from the Persian Gulf. Here, we present the complete genome sequences of DSM 110486 and DSM 110487, with sizes of 10.98 Mbp and 10.33 Mbp, respectively.

Strain M2 T was isolated from the beach of Cuxhaven, Wadden Sea, Germany, in course of a program to attain new producers of bioactive natural products. Strain M2 T produces litoralimycin and sulfomycin-type thiopeptides. Bioinformatic analysis revealed a potential biosynthetic gene cluster encoding for the M2 T thiopeptides. The strain is Gram-stain-positive, rod shaped, non-motile, spore forming, showing a yellow colony color and forms extensively branched substrate mycelium and aerial hyphae. Inferred from the 16S rRNA gene phylogeny strain M2 T affiliates with the genus Streptomonospora. It shows 96.6% 16S rRNA gene sequence similarity to the type species Streptomonospora salina DSM 44593  T and forms a distinct branch with Streptomonospora sediminis DSM 45723  T with 97.0% 16S rRNA gene sequence similarity. Genome-based phylogenetic analysis revealed that M2 T is closely related to Streptomonospora alba YIM 90003  T with a digital DNA-DNA hybridisation (dDDH) value of 26.6%. The predominant menaquinones of M2 T are MK-10(H 6 ), MK-10(H 8 ), and MK-11(H 6 ) (> 10%). Major cellular fatty acids are iso-C 16:0, anteiso C 17:0 and C 18:0 10-methyl. The polar lipid profile consisted of diphosphatidylglycerol phosphatidyl glycerol, phosphatidylinositol, phosphatidylcholine, phosphatidylethanolamine, three glycolipids, two unknown phospholipids, and two unknown lipids. The genome size of type strain M2 T is 5,878,427 bp with 72.1 mol % G + C content. Based on the results obtained from phylogenetic and chemotaxonomic studies, strain M2 T (= DSM 106425  T  = NCCB 100650  T ) is considered to represent a novel species within the genus Streptomonospora for which the name Streptomonospora litoralis sp. nov. is proposed., (© 2021. The Author(s).)

A big challenge in natural product research of today is rapid dereplication of already known substances, to free capacities for the exploration of new agents. Prompt information on bioactivities and mode of action (MOA) speeds up the lead discovery process and is required for rational compound optimization. Here, we present a bioreporter approach as a versatile strategy for combined bioactivity- and MOA-informed primary screening for antimicrobials. The approach is suitable for directly probing producer strains grown on agar, without need for initial compound enrichment or purification, and works along the entire purification pipeline with culture supernatants, extracts, fractions, and pure substances. The technology allows for MOA-informed purification to selectively prioritize activities of interest. In combination with high-resolution mass spectrometry, the biosensor panel is an efficient and sensitive tool for compound deconvolution. Concomitant information on the affected metabolic pathway enables the selection of appropriate follow-up assays to elucidate the molecular target., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Indonesia is one of the most biodiverse countries in the world and a promising resource for novel natural compound producers. Actinomycetes produce about two thirds of all clinically used antibiotics. Thus, exploiting Indonesia's microbial diversity for actinomycetes may lead to the discovery of novel antibiotics. A total of 422 actinomycete strains were isolated from three different unique areas in Indonesia and tested for their antimicrobial activity. Nine potent bioactive strains were prioritized for further drug screening approaches. The nine strains were cultivated in different solid and liquid media, and a combination of genome mining analysis and mass spectrometry (MS)-based molecular networking was employed to identify potential novel compounds. By correlating secondary metabolite gene cluster data with MS-based molecular networking results, we identified several gene cluster-encoded biosynthetic products from the nine strains, including naphthyridinomycin, amicetin, echinomycin, tirandamycin, antimycin, and desferrioxamine B. Moreover, 16 putative ion clusters and numerous gene clusters were detected that could not be associated with any known compound, indicating that the strains can produce novel secondary metabolites. Our results demonstrate that sampling of actinomycetes from unique and biodiversity-rich habitats, such as Indonesia, along with a combination of gene cluster networking and molecular networking approaches, accelerates natural product identification.

Isolate 4NS15 T was isolated from a neglected arid habitat in Kerman, Iran. The strain showed 16S rRNA gene sequence similarity values of 98.9 % to the type strains of Kibdelosporangium aridum subsp. aridum , Kibdelosporangium phytohabitans and Kibdelosporangium philippinense and 98.6 % to the type strain K. aridum subsp. largum , respectively. Genome-based phylogenetic analysis revealed that isolate 4NS15 T is closely related to Kibdelosporangium aridum subsp. aridum DSM 43828 T . The digital DNA-DNA hybridization value between the genome sequences of 4NS15 T and strain DSM 43828 T is 29.8 %. Strain 4NS15 T produces long chains of spores without a sporangium-like structure which can be distinguished from other Kibdelosporangium species. Isolate 4NS15 T has a genome size of 10.35 Mbp with a G+C content of 68.1 mol%. Whole-cell hydrolysates of isolate 4NS15 T are rich in meso -diaminopimelic acid and cell-wall sugars such as arabinose, galactose, glucose and ribose. Major fatty acids (>10 %) are C 16 : 0 , iso-C 16 : 0 and iso-C 15 : 0 . The phospholipid profile contains diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylhydroxyethanolamine, aminolipid and glycoaminolipid. The predominant menaquinone is MK-9(H 4 ). Based on its phenotypic and genotypic characteristics, isolate 4NS15 T (NCCB 100701=CIP 111705=DSM 110728) merits recognition as representing a novel species of the genus Kibdelosporangium , for which the name Kibdelosporangium persicum sp. nov. is proposed.

Antibiotic producers have mainly been isolated from soil, which often has led to the rediscovery of known compounds. In this study, we identified the freshwater snail Physa acuta as an unexplored source for new antibiotic producers. The bacterial diversity associated with the snail was characterized by a metagenomic approach using cultivation-independent high-throughput sequencing. Although Actinobacteria represented only 2% of the bacterial community, the focus was laid on the isolation of the genus Streptomyces due to its potential to produce antibiotics. Three Streptomyces strains (7NS1, 7NS2 and 7NS3) were isolated from P. acuta , and the antimicrobial activity of the crude extracts were tested against a selection of Gram-positive and Gram-negative bacteria and fungi. 7NS3 showed the strongest activity against Gram-positive bacteria and, thus, was selected for genome sequencing and a phylogenomic analysis. 7NS3 represents a novel Streptomyces species, which was deposited as Streptomyces sp. DSM 110735 at the Leibniz Institute-German Collection of Microorganisms and Cell Cultures (DSMZ). Bioassay-guided high-performance liquid chromatography (HPLC) and high-resolution electrospray ionization-mass spectrometry (HR-ESI-MS) analyses of crude extract fractions resulted in the detection of four compounds, one of which matched the compound characteristics of emycin A, an angucycline-like aromatic polyketide. Genome mining studies based on the whole-genome sequence of 7NS3 resulted in the identification of a gene cluster potentially coding for emycin A biosynthesis. Our study demonstrates that freshwater snails like P. acuta can represent promising reservoirs for the isolation of new antibiotic-producing actinobacterial species.

Pristinamycin biosynthesis in Streptomyces pristinaespiralis is governed by a complex hierarchical signaling cascade involving seven different transcriptional regulators (SpbR, PapR1, PapR2, PapR3, PapR4, PapR5, and PapR6). The signaling cascade is triggered by γ-butyrolactone (GBL)-like effector molecules, whereby the chemical structure of the effector, as well as its biosynthetic origin is unknown so far. Three of the pristinamycin transcriptional regulators (SpbR, PapR3, and PapR5) belong to the type of γ-butyrolactone receptor (GBLR). GBLRs are known to either act as "real" GBLRs, which bind GBLs as ligands or as "pseudo" GBLRs binding antibiotics or intermediates thereof as effector molecules. In this study, we performed electromobility shift assays (EMSAs) with SpbR, PapR3, and PapR5, respectively, in the presence of potential ligand samples. Thereby we could show that all three GBLRs bind synthetic 1,4-butyrolactone but not pristinamycin as ligand, suggesting that SpbR, PapR3, and PapR5 act as "real" GBLRs in S. pristinaespiralis . Furthermore, we identified a cytochrome P450 monooxygenase encoding gene snbU as potential biosynthesis gene for the GBLR-interacting ligand. Inactivation of snbU resulted in an increased pristinamycin production, which indicated that SnbU has a regulatory influence on pristinamycin production. EMSAs with culture extract samples from the snbU mutant did not influence the target binding ability of SpbR, PapR3, and PapR5 anymore, in contrast to culture supernatant samples from the S. pristinaespiralis wild-type or the pristinamycin deficient mutant papR2::apra , which demonstrates that SnbU is involved in the synthesis of the GBLR-interacting ligand., (Copyright © 2020 Handel, Kulik and Mast.)

Streptomyces sp. TÜ 2975 and TÜ 3180 are two strains from the Tübingen Actinomycetes strain collection. Here, we present the draft genome sequences of TÜ 2975 and TÜ 3180, with sizes of 7.62 Mb and 8.63 Mb, respectively., (Copyright © 2020 Hennrich et al.)

L-phenylglycine (L-Phg) is a rare non-proteinogenic amino acid, which only occurs in some natural compounds, such as the streptogramin antibiotics pristinamycin I and virginiamycin S or the bicyclic peptide antibiotic dityromycin. Industrially, more interesting than L-Phg is the enantiomeric D-Phg as it plays an important role in the fine chemical industry, where it is used as a precursor for the production of semisynthetic β-lactam antibiotics. Based on the natural L-Phg operon from Streptomyces pristinaespiralis and the stereo-inverting aminotransferase gene hpgAT from Pseudomonas putida, an artificial D-Phg operon was constructed. The natural L-Phg operon, as well as the artificial D-Phg operon, was heterologously expressed in different actinomycetal host strains, which led to the successful production of Phg. By rational genetic engineering of the optimal producer strains S. pristinaespiralis and Streptomyces lividans, Phg production could be improved significantly. Here, we report on the development of a synthetic biology-derived D-Phg pathway and the optimization of fermentative Phg production in actinomycetes by genetic engineering approaches. Our data illustrate a promising alternative for the production of Phgs.

Streptomyces antibiotic regulatory protein (SARP) family regulators are well-known activators of antibiotic biosynthesis in streptomycetes. The respective genes occur in various types of antibiotic gene clusters encoding, e.g., for polyketides, ribosomally and non-ribosomally synthesized peptides, or β-lactam antibiotics. We found that overexpression of the SARP-type regulator gene papR2 from Streptomyces pristinaespiralis in Streptomyces lividans leads to the activation of the silent undecylprodigiosin (Red) gene cluster. The activation happens upon the inducing function of PapR2, which takes over the regulatory role of RedD, the latter of which is the intrinsic SARP regulator of Red biosynthesis in S. lividans . Due to the broad abundance of SARP genes in different antibiotic gene clusters of various actinomycetes and the uniform activating principle of the encoded regulators, we suggest that this type of regulator is especially well suited to be used as an initiator of antibiotic biosynthesis in actinomycetes. Here, we report on a SARP-guided strategy to activate antibiotic gene clusters. As a proof of principle, we present the PapR2-driven activation of the amicetin/plicacetin gene cluster in the novel Indonesian strain isolate Streptomyces sp. SHP22-7., (Copyright © 2020 Krause, Handayani, Blin, Kulik and Mast.)

Streptomyces sp. strain I6 is a novel strain isolated from an Indonesian mangrove sediment sample. Bioinformatic analysis of the genome sequence of Streptomyces sp. I6 revealed 23 biosynthetic gene clusters. One of them encodes the synthesis of a putative phosphonate secondary metabolite, a class of underexplored natural compounds with great pharmaceutical potential.

Streptomyces sp. SHP22-7 is a novel strain isolated from a mangrove sample on Enggano Island, Indonesia. Here, we present the 7.9-Mbp genome sequence of SHP22-7, which will provide insight into its natural compound biosynthetic potential.

Streptomyces sp. strain SW4 exhibited broad-spectrum antibacterial activity toward Gram-positive and Gram-negative pathogens. The 7.5-Mb draft genome sequence gives insight into the complete secondary metabolite production capacity and reveals genes putatively responsible for its antibacterial activity.

l-phenylglycine is a rare non-proteinogenic amino acid, which only occurs in a few natural compounds, such as the streptogramin antibiotics pristinamycin I and virginiamycin S or the bicyclic peptide antibiotic dityromycin. Here we report on the biochemical characterization of the aminotransferase PglE that catalyzes the transamination from phenylglyoxylate to l-phenylglycine, which represents the final reaction step during phenylglycine biosynthesis. Enzyme assays with the purified PglE enzyme revealed that l-phenylalanine is used as an amino group donor for the transamination reaction, leading to the formation of phenylpyruvate, which may re-enter phenylglycine biosynthesis as a precursor. Based on these results, we postulate a novel l-phenylglycine biosynthetic pathway., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The strain Streptomyces sp. BSE7F, a novel Streptomyces strain isolated from Indonesian mangrove sediment, displays antimicrobial activities against Gram-positive bacteria, Gram-negative bacteria, and yeast. Bioinformatic analysis of the genome sequence revealed the occurrence of 22 biosynthetic gene clusters disclosing the secondary metabolite capacity of strain BSE7F., (Copyright © 2018 Handayani et al.)

Potatoes are a promising system for industrial production of the biopolymer cyanophycin as a second compound in addition to starch. To assess the efficiency in the field, we analysed the stability of the system, specifically its sensitivity to environmental factors. Field and greenhouse trials with transgenic potatoes (two independent events) were carried out for three years. The influence of environmental factors was measured and target compounds in the transgenic plants (cyanophycin, amino acids) were analysed for differences to control plants. Furthermore, non-target parameters (starch content, number, weight and size of tubers) were analysed for equivalence with control plants. The huge amount of data received was handled using modern statistical approaches to model the correlation between influencing environmental factors (year of cultivation, nitrogen fertilization, origin of plants, greenhouse or field cultivation) and key components (starch, amino acids, cyanophycin) and agronomic characteristics. General linear models were used for modelling, and standard effect sizes were applied to compare conventional and genetically modified plants. Altogether, the field trials prove that significant cyanophycin production is possible without reduction of starch content. Non-target compound composition seems to be equivalent under varying environmental conditions. Additionally, a quick test to measure cyanophycin content gives similar results compared to the extensive enzymatic test. This work facilitates the commercial cultivation of cyanophycin potatoes.

Pristinamycin, produced by Streptomyces pristinaespiralis , which is a streptogramin-like antibiotic consisting of two chemically unrelated components: pristinamycin I (PI) and pristinamycin II (PII), shows potent activity against many antibiotic-resistant pathogens. However, so far pristinamycin production titers are still quite low, particularly those of PI. In this study, we constructed a PI single component producing strain by deleting the PII biosynthetic genes ( snaE1 and snaE2 ). Then, two metabolic engineering approaches, including deletion of the repressor gene papR3 and chromosomal integration of an extra copy of the PI biosynthetic gene cluster (BGC), were employed to improve PI production. The final engineered strain ΔPIIΔ papR3 /PI produced a maximum PI level of 132 mg/L, with an approximately 2.4-fold higher than that of the parental strain S. pristinaespiralis HCCB10218. Considering that the PI biosynthetic genes are clustered in two main regions in the 210 kb "supercluster" containing the PI and PII biosynthetic genes as well as a cryptic polyketide BGC, these two regions were cloned separately and then were successfully assembled into the PI BGC by the transformation-associated recombination (TAR) system. Collectively, the metabolic engineering approaches employed is very efficient for strain improvement in order to enhance PI titer.

Due to the threat posed by the increase of highly resistant pathogenic bacteria, there is an urgent need for new antibiotics; all the more so since in the last 20 years, the approval for new antibacterial agents had decreased. The field of natural product discovery has undergone a tremendous development over the past few years. This has been the consequence of several new and revolutionizing drug discovery and development techniques, which is initiating a 'New Age of Antibiotic Discovery'. In this review, we concentrate on the most significant discovery approaches during the last and present years and comment on the challenges facing the community in the coming years., (© 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.)

Pristinamycin production in Streptomyces pristinaespiralis Pr11 is tightly regulated by an interplay between different repressors and activators. A γ-butyrolactone receptor gene (spbR), two TetR repressor genes (papR3 and papR5), three SARP (Streptomyces antibiotic regulatory protein) genes (papR1, papR2, and papR4), and a response regulator gene (papR6) are carried on the large 210-kb pristinamycin biosynthetic gene region of Streptomyces pristinaespiralis Pr11. A detailed investigation of all pristinamycin regulators revealed insight into a complex signaling cascade, which is responsible for the fine-tuned regulation of pristinamycin production in S. pristinaespiralis., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

The aconitase AcnA from the phosphinothricin tripeptide producing strain Streptomyces viridochromogenes Tü494 is a bifunctional protein: under iron-sufficiency conditions AcnA functions as an enzyme of the tricarboxylic acid cycle, whereas under iron depletion it is a regulator of iron metabolism and oxidative stress response. As a member of the family of iron regulatory proteins (IRP), AcnA binds to characteristic iron responsive element (IRE) binding motifs and post-transcriptionally controls the expression of respective target genes. A S. viridochromogenes aconitase mutant (MacnA) has previously been shown to be highly sensitive to oxidative stress. In the present paper, we performed a comparative proteomic approach with the S. viridochromogenes wild-type and the MacnA mutant strain under oxidative stress conditions to identify proteins that are under control of the AcnA-mediated regulation. We identified up to 90 differentially expressed proteins in both strains. In silico analysis of the corresponding gene sequences revealed the presence of IRE motifs on some of the respective target mRNAs. From this proteome study we have in vivo evidences for a direct AcnA-mediated regulation upon oxidative stress.

Streptogramins are potent drugs against numerous highly resistant pathogens and therefore are used as antibiotics of last-resort human therapy. They consist of a mixture of two different types of chemical substances - the group A streptogramins, which are polyunsaturated macrolactones, and the group B streptogramins, representing cyclic hexadepsipeptides. Streptogramins are unique in their mode of action: each component alone exhibits a moderate bacteriostatic activity by binding to the bacterial 50S ribosomal subunit and thereby blocking translation, whereas the synergic combination of both substances is up to hundred fold more effective than the single compounds, resulting in a bactericidal activity. The streptogramin biosynthetic genes are organized as large antibiotic superclusters. These clusters harbour numerous regulatory genes, which encode different types of regulators that together form a complex hierarchical signalling system, which governs the regulation of streptogramin biosynthesis. Resistance is also regulated by this cascade. However, whereas resistance against streptogramins is quite well understood in diverse pathogenic organisms, only little is known about how the natural producer strains protect themselves against these toxic compounds. Here, we give an overview about the recent advances in streptogramin investigations with a main focus on the best-studied representatives, pristinamycin and virginiamycin. We concentrate on the biosynthesis of these compounds, their regulation and resistance determinants as well as their application in medicine and food industry., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

The two-component system AfsQ1/Q2 of Streptomyces coelicolor was identified in our previous work as a pleiotropic regulator for antibiotic biosynthesis and morphological differentiation under the condition of a minimal medium supplemented with 75 mM glutamate. In this work, we report the dissection of the mechanism underlying the function of AfsQ1/Q2 on antibiotic production and also the identification of the AfsQ1/Q2 regulon. The results showed that AfsQ1/Q2 stimulated antibiotic ACT, RED and CDA production directly through the pathway-specific activator genes actII-ORF4, redZ and cdaR respectively. In addition, expression of sigQ that encodes a sigma factor and is divergently transcribed from afsQ1 was also subject to direct regulation by AfsQ1/Q2. The precise AfsQ1 binding sites in the upstream regions of these target genes were determined by DNase I footprinting assays coupled with site-directed DNA mutagenesis. By computational prediction and functional analysis, at least 17 new AfsQ1 targets were identified, including pstS gene encoding a high-affinity phosphate-binding protein and two developmental genes whiD, bldM. For the AfsQ1/Q2 regulon, an AfsQ1 binding motif comprising the sequence GTnAC-n(6) -GTnAC has been defined. Interestingly, we found from electrophoretic mobility shift assays and transcriptional analysis that AfsQ1/Q2 can also function as a repressor for nitrogen assimilation, and AfsQ1 can compete with GlnR for the promoter regions of glnA and nirB, suggesting the cross-regulation between AfsQ1/Q2 and GlnR in nitrogen metabolism. These findings suggested that AfsQ1/Q2 is important not only for antibiotic biosynthesis but also in maintaining the metabolic homeostasis of nutrient utilization under the stress of high concentration of glutamate in S. coelicolor., (© 2012 Blackwell Publishing Ltd.)

In many organisms, aconitases have dual functions; they serve as enzymes in the tricarboxylic acid cycle and as regulators of iron metabolism. In this study we defined the role of the aconitase AcnA in Streptomyces viridochromogenes Tü494, the producer of the herbicide phosphinothricyl-alanyl-alanine, also known as phosphinothricin tripeptide or bialaphos. A mutant in which the aconitase gene acnA was disrupted showed severe defects in morphology and physiology, as it was unable to form any aerial mycelium, spores nor phosphinothricin tripeptide. AcnA belongs to the iron regulatory proteins (IRPs). In addition to its catalytic function, AcnA plays a regulatory role by binding to iron responsive elements (IREs) located on the untranslated region of certain mRNAs. A mutation preventing the formation of the [4Fe-4S] cluster of AcnA eliminated its catalytic activity, but did not inhibit RNA-binding ability. In silico analysis of the S. viridochromogenes genome revealed several IRE-like structures. One structure is located upstream of recA, which is involved in the bacterial SOS response, and another one was identified upstream of ftsZ, which is required for the onset of sporulation in streptomycetes. The functionality of different IRE structures was proven with gel shift assays and specific IRE consensus sequences were defined. Furthermore, RecA was shown to be upregulated on post-transcriptional level under oxidative stress conditions in the wild-type strain but not in the acnA mutant, suggesting a regulatory role of AcnA in oxidative stress response., (© 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.)

Actinomycetes are a rich source for the synthesis of medically and technically useful natural products. The genes encoding the enzymes for their biosynthesis are normally organized in gene clusters, which include also the information for resistance (in the case of antibacterial compounds), regulation, and transport. This facilitates the manipulation of such pathways by molecular genetic techniques. Recent advances in DNA sequencing and analytical chemistry revealed that not only new strains isolated from yet unexplored habitats, but also already known strains possess a large potential for the synthesis of novel compounds. Synthetic Biology now offers a new perspective to exploit this potential further by generating novel pathways, and thereby novel products, by combining different biosynthetic steps originating from different bacteria. The supply of precursors, which are subsequently incorporated into the final product, is often already organized in a modular manner in nature and may directly be exploited for Synthetic Biology. Here we report examples for the synthesis of building blocks and possibilities to modify and optimize antibiotic biosynthesis, exemplary for the synthesis of the manipulation of the synthesis of the glycopeptide antibiotic balhimycin., (Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Determining transcriptional regulator activities is a major focus of systems biology, providing key insight into regulatory mechanisms and co-regulators. For organisms such as Escherichia coli, transcriptional regulator binding site data can be integrated with expression data to infer transcriptional regulator activities. However, for most organisms there is only sparse data on their transcriptional regulators, while their associated binding motifs are largely unknown. Here, we address the challenge of inferring activities of unknown regulators by generating de novo (binding) motifs and integrating with expression data. We identify a number of key regulators active in the metabolic switch, including PhoP with its associated directed repeat PHO box, candidate motifs for two SARPs, a CRP family regulator, an iron response regulator and that for LexA. Experimental validation for some of our predictions was obtained using gel-shift assays. Our analysis is applicable to any organism for which there is a reasonable amount of complementary expression data and for which motifs (either over represented or evolutionary conserved) can be identified in the genome.

GlnR is the global transcriptional regulator of nitrogen assimilation in Streptomyces coelicolor. Under nitrogen starvation, GlnR controls the transcription of at least nine genes associated with nitrogen metabolism. In this study, we identified a new GlnR target gene, SCO2958, named nnaR (nitrate/nitrite assimilation regulator). In silico analysis of NnaR revealed the presence of two distinct domains: an N-terminal uroporphyrinogen-III synthase (HemD)-like enzymatic domain and a C-terminal DNA binding domain. Complementation experiments with a haemin auxotroph Escherichia coli ΔhemD mutant strain revealed that NnaR has no HemD activity. Physiological studies of an S. coelicolor nnaR : : Tn5062 mutant showed that NnaR is involved in regulating nitrite reduction. By electrophoretic mobility shift assays the functionality of the NnaR DNA binding domain was confirmed, and it was found that NnaR binds in front of the genes narK (putative nitrate extrusion protein), nirB (nitrite reductase), nirA (putative nitrite/sulphite reductase) and nasA (putative nitrate reductase), which are associated with nitrate/nitrite assimilation. Furthermore, a cooperative binding of NnaR together with GlnR to the nirB promoter was observed, suggesting that NnaR may act as a GlnR co-activator.

GlnK is an important nitrogen sensor protein in Streptomyces coelicolor. Deletion of glnK results in a medium-dependent failure of aerial mycelium and spore formation and loss of antibiotic production. Thus, GlnK is not only a regulator of nitrogen metabolism but also of morphological differentiation and secondary metabolite production. Through a comparative transcriptomic approach between the S. coelicolor wild-type and a S. coelicolor glnK mutant strain, 142 genes were identified that are differentially regulated in both strains. Among these are genes of the ram and rag operon, which are involved in S. coelicolor morphogenesis, as well as genes involved in gas vesicle biosynthesis and ectoine biosynthesis. Surprisingly, no relevant nitrogen genes were found to be differentially regulated, revealing that GlnK is not an important nitrogen sensor under the tested conditions.

Pristinamycin, produced by Streptomyces pristinaespiralis Pr11, is a streptogramin antibiotic consisting of two chemically unrelated compounds, pristinamycin I and pristinamycin II. The semi-synthetic derivatives of these compounds are used in human medicine as therapeutic agents against methicillin-resistant Staphylococcus aureus strains. Only the partial sequence of the pristinamycin biosynthetic gene cluster has been previously reported. To complete the sequence, overlapping cosmids were isolated from a S. pristinaespiralis Pr11 gene library and sequenced. The boundaries of the cluster were deduced, limiting the cluster size to approximately 210 kb. In the central region of the cluster, previously unknown pristinamycin biosynthetic genes were identified. Combining the current and previously identified sequence information, we propose that all essential pristinamycin biosynthetic genes are included in the 210 kb region. A pristinamycin biosynthetic pathway was established. Furthermore, the pristinamycin gene cluster was found to be interspersed by a cryptic secondary metabolite cluster, which probably codes for a glycosylated aromatic polyketide. Gene inactivation experiments revealed that this cluster has no influence on pristinamycin production. Overall, this work provides new insights into pristinamycin biosynthesis and the unique genetic organization of the pristinamycin gene region, which is the largest antibiotic 'supercluster' known so far., (© 2010 The Authors. Journal compilation © 2010 Society for Applied Microbiology and Blackwell Publishing Ltd.)

GlnR is the global regulator of nitrogen assimilation in Streptomyces coelicolor M145 and other actinobacteria. Two-dimensional polyacrylamide gel electrophoresis analyses were performed to identify new GlnR target genes by proteomic comparison of wild-type S. coelicolor M145 and a ΔglnR mutant. Fifty proteins were found to be differentially regulated between S. coelicolor M145 and the ΔglnR mutant. These spots were identified by nanoHPLC-ESI-MS/MS and classified according to their cellular role. Most of the identified proteins are involved in amino acid biosynthesis and in carbon metabolism, demonstrating that the role of GlnR is not restricted to nitrogen metabolism. Thus, GlnR is supposed to play an important role in the global metabolic control of S. coelicolor M145.