scholarly journals Microbiological and molecular insights on rare Actinobacteria harboring bioactive prospective

2020 ◽  
Vol 44 (1) ◽  
Author(s):  
Dina H. Amin ◽  
Nagwa A. Abdallah ◽  
Assem Abolmaaty ◽  
Sahar Tolba ◽  
Elizabeth M. H. Wellington
Keyword(s):  
Bioinformatics Analysis ◽  
Genetic Manipulation ◽  
Antibiotic Production ◽  
Gene Clusters ◽  
Bioactive Molecules ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Pcr Screening ◽  
Molecular Approaches ◽  
Shed Light

Abstract Background Actinobacteria is as a group of advanced filamentous bacteria. Rare Actinobacteria are of special interest as they are rarely isolated from the environments. They are a major source of important bioactive compounds. Determining the proper strategy for the identification of Actinobacteria harboring biosynthetic gene clusters and producing bioactive molecules is a challenging platform. Methodology In this review, we discuss a consequence of microbiological and molecular methods for the identification of rare Actinobacteria. In addition to that, we shed light on rare Actinobacteria’s significance in antibiotic production. We also clarified molecular approaches for the manipulation of novel biosynthetic gene clusters via PCR screening, fosmid libraries, and Illumina whole-genome sequencing in combination with bioinformatics analysis. Conclusion Perceptions of the conventional and molecular identification of Actinobacteria were conducted. This will open the door for the genetic manipulation of novel antibiotic gene clusters in heterologous hosts. Also, these conclusions will lead to constructing new bioactive molecules via genetically engineering biosynthetic pathways.

scholarly journals Identification of a New Antimicrobial, Desertomycin H, Utilizing a Modified Crowded Plate Technique

Marine Drugs ◽  
2021 ◽  
Vol 19 (8) ◽  
pp. 424
Author(s):  
Osama G. Mohamed ◽  
Sadaf Dorandish ◽  
Rebecca Lindow ◽  
Megan Steltz ◽  
Ifrah Shoukat ◽  
...  
Keyword(s):  
Antibiotic Production ◽  
Gene Clusters ◽  
Multidrug Resistant ◽  
Microbial Interactions ◽  
Mass Spectrometry Data ◽  
Metagenomic Data ◽  
Resistant Bacteria ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Plate Technique

The antibiotic-resistant bacteria-associated infections are a major global healthcare threat. New classes of antimicrobial compounds are urgently needed as the frequency of infections caused by multidrug-resistant microbes continues to rise. Recent metagenomic data have demonstrated that there is still biosynthetic potential encoded in but transcriptionally silent in cultivatable bacterial genomes. However, the culture conditions required to identify and express silent biosynthetic gene clusters that yield natural products with antimicrobial activity are largely unknown. Here, we describe a new antibiotic discovery scheme, dubbed the modified crowded plate technique (mCPT), that utilizes complex microbial interactions to elicit antimicrobial production from otherwise silent biosynthetic gene clusters. Using the mCPT as part of the antibiotic crowdsourcing educational program Tiny Earth®, we isolated over 1400 antibiotic-producing microbes, including 62, showing activity against multidrug-resistant pathogens. The natural product extracts generated from six microbial isolates showed potent activity against vancomycin-intermediate resistant Staphylococcus aureus. We utilized a targeted approach that coupled mass spectrometry data with bioactivity, yielding a new macrolactone class of metabolite, desertomycin H. In this study, we successfully demonstrate a concept that significantly increased our ability to quickly and efficiently identify microbes capable of the silent antibiotic production.

scholarly journals Phytoplankton trigger the production of cryptic metabolites in the marine actinobacteria Salinispora tropica

2020 ◽  
Author(s):  
Audam Chhun ◽  
Despoina Sousoni ◽  
Maria del Mar Aguiló-Ferretjans ◽  
Lijiang Song ◽  
Christophe Corre ◽  
...  
Keyword(s):  
Natural Products ◽  
Secondary Metabolites ◽  
Antibiotic Production ◽  
Antimicrobial Effect ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Marine Actinobacteria ◽  
Salinispora Tropica ◽  
Eukaryotic Phytoplankton

AbstractBacteria from the Actinomycete family are a remarkable source of natural products with pharmaceutical potential. The discovery of novel molecules from these organisms is, however, hindered because most of the biosynthetic gene clusters (BGCs) encoding these secondary metabolites are cryptic or silent and are referred to as orphan BGCs. While co-culture has proven to be a promising approach to unlock the biosynthetic potential of many microorganisms by activating the expression of these orphan BGCs, it still remains an underexplored technique. The marine actinobacteria Salinispora tropica, for instance, produces valuable compounds such as the anti-cancer molecule salinosporamide A but half of its putative BGCs are still orphan. Although previous studies have looked into using marine heterotrophs to induce orphan BGCs in Salinispora, the potential impact of co-culturing marine phototrophs with Salinispora has yet to be investigated. Following the observation of clear antimicrobial phenotype of the actinobacterium on a range of phytoplanktonic organisms, we here report the discovery of novel cryptic secondary metabolites produced by S. tropica in response to its co-culture with photosynthetic primary producers. An approach combining metabolomics and proteomics revealed that the photosynthate released by phytoplankton influences the biosynthetic capacities of S. tropica with both production of new molecules and the activation of orphan BGCs. Our work pioneers the use of phototrophs as a promising strategy to accelerate the discovery of novel natural products from actinobacteria.ImportanceThe alarming increase of antimicrobial resistance has generated an enormous interest in the discovery of novel active compounds. The isolation of new microbes to untap novel natural products is currently hampered because most biosynthetic gene clusters (BGC) encoded by these microorganisms are not expressed under standard laboratory conditions, i.e. mono-cultures. Here we show that co-culturing can be an easy way for triggering silent BGC. By combining state-of-the-art metabolomics and high-throughput proteomics, we characterized the activation of cryptic metabolites and silent biosynthetic gene clusters in the marine actinobacteria Salinispora tropica by the presence of phytoplankton photosynthate. We further suggest a mechanistic understanding of the antimicrobial effect this actinobacterium has on a broad range of prokaryotic and eukaryotic phytoplankton species and reveal a promising candidate for antibiotic production.

scholarly journals Challenges and Advances in Genome Editing Technologies in Streptomyces

Biomolecules ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 734 ◽  
Author(s):  
Yawei Zhao ◽  
Guoquan Li ◽  
Yunliang Chen ◽  
Yinhua Lu
Keyword(s):  
Natural Product ◽  
Genome Editing ◽  
Genetic Manipulation ◽  
Chemical Compounds ◽  
Gene Clusters ◽  
Future Research ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Research Focus ◽  
Experimental Conditions

The genome of Streptomyces encodes a high number of natural product (NP) biosynthetic gene clusters (BGCs). Most of these BGCs are not expressed or are poorly expressed (commonly called silent BGCs) under traditional laboratory experimental conditions. These NP BGCs represent an unexplored rich reservoir of natural compounds, which can be used to discover novel chemical compounds. To activate silent BGCs for NP discovery, two main strategies, including the induction of BGCs expression in native hosts and heterologous expression of BGCs in surrogate Streptomyces hosts, have been adopted, which normally requires genetic manipulation. So far, various genome editing technologies have been developed, which has markedly facilitated the activation of BGCs and NP overproduction in their native hosts, as well as in heterologous Streptomyces hosts. In this review, we summarize the challenges and recent advances in genome editing tools for Streptomyces genetic manipulation with a focus on editing tools based on clustered regularly interspaced short palindrome repeat (CRISPR)/CRISPR-associated protein (Cas) systems. Additionally, we discuss the future research focus, especially the development of endogenous CRISPR/Cas-based genome editing technologies in Streptomyces.

scholarly journals Modular and Integrative Vectors for Synthetic Biology Applications in Streptomyces spp.

2019 ◽  
Vol 85 (16) ◽  
Author(s):  
Céline Aubry ◽  
Jean-Luc Pernodet ◽  
Sylvie Lautru
Keyword(s):  
Synthetic Biology ◽  
Anticancer Agents ◽  
Gene Clusters ◽  
Bioactive Molecules ◽  
Dna Assembly ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Dna Cloning ◽  
Specialized Metabolism ◽  
Assembly Method

ABSTRACT With the development of synthetic biology in the field of (actinobacterial) specialized metabolism, new tools are needed for the design or refactoring of biosynthetic gene clusters. If libraries of synthetic parts (such as promoters or ribosome binding sites) and DNA cloning methods have been developed, to our knowledge, not many vectors designed for the flexible cloning of biosynthetic gene clusters have been constructed. We report here the construction of a set of 12 standardized and modular vectors designed to afford the construction or the refactoring of biosynthetic gene clusters in Streptomyces species, using a large panel of cloning methods. Three different resistance cassettes and four orthogonal integration systems are proposed. In addition, FLP recombination target sites were incorporated to allow the recycling of antibiotic markers and to limit the risks of unwanted homologous recombination in Streptomyces strains when several vectors are used. The functionality and proper integration of the vectors in three commonly used Streptomyces strains, as well as the functionality of the Flp-catalyzed excision, were all confirmed. To illustrate some possible uses of our vectors, we refactored the albonoursin gene cluster from Streptomyces noursei using the BioBrick assembly method. We also used the seamless ligase chain reaction cloning method to assemble a transcription unit in one of the vectors and genetically complement a mutant strain. IMPORTANCE One of the strategies employed today to obtain new bioactive molecules with potential applications for human health (for example, antimicrobial or anticancer agents) is synthetic biology. Synthetic biology is used to biosynthesize new unnatural specialized metabolites or to force the expression of otherwise silent natural biosynthetic gene clusters. To assist the development of synthetic biology in the field of specialized metabolism, we constructed and are offering to the community a set of vectors that were intended to facilitate DNA assembly and integration in actinobacterial chromosomes. These vectors are compatible with various DNA cloning and assembling methods. They are standardized and modular, allowing the easy exchange of a module by another one of the same nature. Although designed for the assembly or the refactoring of specialized metabolite gene clusters, they have a broader potential utility, for example, for protein production or genetic complementation.

scholarly journals Competition sensing alters antibiotic production in Streptomyces

2020 ◽  
Author(s):  
Sanne Westhoff ◽  
Alexander Kloosterman ◽  
Stephan F. A. van Hoesel ◽  
Gilles P. van Wezel ◽  
Daniel E. Rozen
Keyword(s):  
Genetic Relatedness ◽  
Cell Damage ◽  
Antibiotic Production ◽  
Interference Competition ◽  
Resource Limitation ◽  
Gene Clusters ◽  
Regulatory Control ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
The Social

ABSTRACTOne of the most important ways that bacteria compete for resources and space is by producing antibiotics that inhibit competitors. Because antibiotic production is costly, the biosynthetic gene clusters coordinating their synthesis are under strict regulatory control and often require “elicitors” to induce expression, including cues from competing strains. Although these cues are common, they are not produced by all competitors and so the phenotypes causing induction remain unknown. By studying interactions between 24 antibiotic-producing streptomycetes we show that inhibition between competitors is common and occurs more frequently if strains are closely related. Next, we show that antibiotic production is more likely to be induced by cues from strains that are closely related or that share secondary metabolite biosynthetic gene clusters (BGCs). Unexpectedly, antibiotic production is less likely to be induced by competitors that inhibit the growth of a focal strain, indicating that cell damage is not a general cue for induction. In addition to induction, antibiotic production often decreased in the presence of a competitor, although this response was not associated with genetic relatedness or overlap in BGCs. Finally, we show that resource limitation increases the chance that antibiotic production declines during competion. Our results reveal the importance of social cues and resource availability in the dynamics of interference competition in streptomycetes.SIGNIFICANCE STATEMENTBacteria secrete antibiotics to inhibit their competitors, but the presence of competitors can determine whether these toxins are produced. Here, we study the role of the competitive and resource environment on antibiotic production in Streptomyces, bacteria renowned for their production of antibiotics. We show that Streptomyces are more likely to produce antibiotics when grown with closely related competitors or that share biosynthetic pathways for secondary metabolites, but not when they are threatened by competitor’s toxins, in contrast to predictions of the competition sensing hypothesis. Streptomyces also often reduce their output of antibiotics when grown with competitors, especially under nutrient limitation. Our findings highlight that interactions between the social and resource environments strongly regulate antibiotic production in these medicinally important bacteria.

scholarly journals Whole-genome sequence of bioactive streptomycete derived from mangrove forest in Malaysia, Streptomyces sp. MUSC 14

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Hooi-Leng Ser ◽  
Loh Teng-Hern Tan ◽  
Wen-Si Tan ◽  
Wai-Fong Yin ◽  
Kok-Gan Chan
Keyword(s):  
East Coast ◽  
Genetic Manipulation ◽  
Genome Mining ◽  
Gene Clusters ◽  
Peninsular Malaysia ◽  
Whole Genome Sequence ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Streptomyces Sp ◽  
A Genome

The contribution of streptomycetes to human health is undeniably important and significant, given that these filamentous microbes can produce interesting compounds that can be used to cure deadly infections and even cancer. Isolated from the east coast of Peninsular Malaysia, Streptomyces sp. MUSC 14 has shown significant antioxidant capacity. The current study explores the genomic potential of MUSC 14 via a genome mining approach. The genome size of MUSC 14 is 10,274,825 bp with G + C content of 71.3 %. AntiSMASH analysis revealed a total of nine biosynthetic gene clusters (with more than 80 % similarities to known gene clusters). This information serves as an important foundation for subsequent studies, particularly the purification and isolation of bioactive compounds by genetic manipulation techniques.

scholarly journals Genomics-Driven Activation of Silent Biosynthetic Gene Clusters in Burkholderia gladioli by Screening Recombineering System

Molecules ◽  
2021 ◽  
Vol 26 (3) ◽  
pp. 700
Author(s):  
Hanna Chen ◽  
Tao Sun ◽  
Xianping Bai ◽  
Jie Yang ◽  
Fu Yan ◽  
...  
Keyword(s):  
Genetic Manipulation ◽  
Structural Diversity ◽  
Gene Clusters ◽  
Unnatural Amino Acid ◽  
Biosynthetic Gene ◽  
Assembly Lines ◽  
Biosynthetic Gene Clusters ◽  
Genome Modification ◽  
Burkholderia Gladioli ◽  
Peptide Synthetase

The Burkholderia genus possesses ecological and metabolic diversities. A large number of silent biosynthetic gene clusters (BGCs) in the Burkholderia genome remain uncharacterized and represent a promising resource for new natural product discovery. However, exploitation of the metabolomic potential of Burkholderia is limited by the absence of efficient genetic manipulation tools. Here, we screened a bacteriophage recombinase system Redγ-BAS, which was functional for genome modification in the plant pathogen Burkholderia gladioli ATCC 10248. By using this recombineering tool, the constitutive promoters were precisely inserted in the genome, leading to activation of two silent nonribosomal peptide synthetase gene clusters (bgdd and hgdd) and production of corresponding new classes of lipopeptides, burriogladiodins A–H (1–8) and haereogladiodins A–B (9–10). Structure elucidation revealed an unnatural amino acid Z- dehydrobutyrine (Dhb) in 1–8 and an E-Dhb in 9–10. Notably, compounds 2–4 and 9 feature an unusual threonine tag that is longer than the predicted collinearity assembly lines. The structural diversity of burriogladiodins was derived from the relaxed substrate specificity of the fifth adenylation domain as well as chain termination conducted by water or threonine. The recombinase-mediating genome editing system is not only applicable in B. gladioli, but also possesses great potential for mining meaningful silent gene clusters from other Burkholderia species.

scholarly journals Endophytic microbes from Nigerian ethnomedicinal plants: a potential source for bioactive secondary metabolites—a review

2021 ◽  
Vol 45 (1) ◽  
Author(s):  
Chijioke E. Ezeobiora ◽  
Nwamaka H. Igbokwe ◽  
Dina H. Amin ◽  
Udoma E. Mendie
Keyword(s):  
Infectious Diseases ◽  
Anticancer Agents ◽  
Gene Clusters ◽  
Emerging Diseases ◽  
Bioactive Molecules ◽  
Main Body ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Ethnomedicinal Plants ◽  
Bacteria And Fungi

Abstract Background Endophytes are highly beneficial species of microbes that live in symbiosis with plant tissues in the setting. Endophytes are difficult to isolate in their natural environment, and they are understudied despite being a rich source of bioactive molecules. There are varieties of new infectious diseases emerging across the world, necessitating a constant and expanded search for newer and more efficient bioactive molecules. Nigeria is known for its biodiversity in ethnomedicinal plants, yet these plants are understudied for endophytic microbes harbouring novel bioactive molecules. Main body Endophytes are a source of novel organic natural molecules and are thought to be drug discovery frontiers. Endophyte research has contributed to the discovery of possible anticancer agents following the discovery of taxol. Endophyte research has contributed to the discovery of possible drug compounds with antimicrobial, antioxidant, antiviral, antidiabetic, anti-Alzheimers disease and immunosuppressive properties among others. These breakthroughs provide hope for combating incurable diseases, drug resistance, the emergence of new infectious diseases, and other human health issues. Finding new medicines that may be effective candidates for treating newly emerging diseases in humans has a lot of promise. Most studies have been on fungi endophytes, with just a few reports on bacterial endophytes. The biology of endophytic bacteria and fungi, as well as endophytic microbes isolated from Nigerian medicinal plants, their isolation methods, identification by morphological and molecular methods, fermentation, purification, identification of bioactive compounds and biosynthetic gene clusters are all covered in this study. Conclusion In Nigeria, the sourcing and isolation of endophytes harboring biosynthetic gene clusters are still understudied, necessitating a rigorous quest for bioactive molecules in endophytes inhabiting various ethnomedicinal plants.

scholarly journals The Antibiotic Andrimid Produced by Vibrio coralliilyticus Increases Expression of Biosynthetic Gene Clusters and Antibiotic Production in Photobacterium galatheae

2020 ◽  
Vol 11 ◽  
Author(s):  
Yannick Buijs ◽  
Thomas Isbrandt ◽  
Sheng-Da Zhang ◽  
Thomas Ostenfeld Larsen ◽  
Lone Gram
Keyword(s):  
Stress Response ◽  
Antibiotic Production ◽  
Gene Clusters ◽  
Multidrug Resistant ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Gene Encoding ◽  
Chemical Measurements ◽  
Low Concentrations ◽  
Vibrio Coralliilyticus

The development and spread of multidrug resistant pathogens have reinforced the urgency to find novel natural products with antibiotic activity. In bacteria, orphan biosynthetic gene clusters (BGCs) far outnumber the BGCs for which chemistry is known, possibly because they are transcriptionally silent under laboratory conditions. A strategy to trigger the production of this biosynthetic potential is to challenge the microorganism with low concentrations of antibiotics, and by using a Burkholderia genetic reporter strain (Seyedsayamdost, Proc Natl Acad Sci 111:7266–7271), we found BGC unsilencing activity for the antimicrobial andrimid, produced by the marine bacterium Vibrio coralliilyticus. Next, we challenged another marine Vibrionaceae, Photobacterium galatheae, carrier of seven orphan BGCs with sub-inhibitory concentrations of andrimid. A combined approach of transcriptional and chemical measurements of andrimid-treated P. galatheae cultures revealed a 10-fold upregulation of an orphan BGC and, amongst others, a 1.6–2.2-fold upregulation of the gene encoding the core enzyme for biosynthesis of holomycin. Also, addition of andrimid caused an increase, based on UV-Vis peak area, of 4-fold in production of the antibiotic holomycin. Transcriptional measurements of stress response related genes in P. galatheae showed a co-occurrence of increased transcript levels of rpoS (general stress response) and andrimid induced holomycin overproduction, while in trimethoprim treated cultures attenuation of holomycin production coincided with a transcriptional increase of recA (SOS stress response). This study shows that using antimicrobial compounds as activators of secondary metabolism can be a useful strategy in eliciting biosynthetic gene clusters and facilitate natural product discovery. Potentially, such interactions could also have ecological relevant implications.

scholarly journals Competition Sensing Changes Antibiotic Production in Streptomyces

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sanne Westhoff ◽  
Alexander M. Kloosterman ◽  
Stephan F. A. van Hoesel ◽  
Gilles P. van Wezel ◽  
Daniel E. Rozen
Keyword(s):  
Genetic Relatedness ◽  
Cell Damage ◽  
Antibiotic Production ◽  
Interference Competition ◽  
Resource Limitation ◽  
Gene Clusters ◽  
Regulatory Control ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
The Social

ABSTRACT One of the most important ways that bacteria compete for resources and space is by producing antibiotics that inhibit competitors. Because antibiotic production is costly, the biosynthetic gene clusters coordinating their synthesis are under strict regulatory control and often require “elicitors” to induce expression, including cues from competing strains. Although these cues are common, they are not produced by all competitors, and so the phenotypes causing induction remain unknown. By studying interactions between 24 antibiotic-producing strains of streptomycetes, we show that strains commonly inhibit each other’s growth and that this occurs more frequently if strains are closely related. Next, we show that antibiotic production is more likely to be induced by cues from strains that are closely related or that share secondary metabolite biosynthetic gene clusters (BGCs). Unexpectedly, antibiotic production is less likely to be induced by competitors that inhibit the growth of a focal strain, indicating that cell damage is not a general cue for induction. In addition to induction, antibiotic production often decreases in the presence of a competitor, although this response was not associated with genetic relatedness or overlap in BGCs. Finally, we show that resource limitation increases the chance that antibiotic production declines during competition. Our results reveal the importance of social cues and resource availability in the dynamics of interference competition in streptomycetes. IMPORTANCE Bacteria secrete antibiotics to inhibit their competitors, but the presence of competitors can determine whether these toxins are produced. Here, we study the role of the competitive and resource environment on antibiotic production in Streptomyces, bacteria renowned for their production of antibiotics. We show that Streptomyces cells are more likely to produce antibiotics when grown with competitors that are closely related or that share biosynthetic pathways for secondary metabolites, but not when they are threatened by competitor’s toxins, in contrast to predictions of the competition sensing hypothesis. Streptomyces cells also often reduce their output of antibiotics when grown with competitors, especially under nutrient limitation. Our findings highlight that interactions between the social and resource environments strongly regulate antibiotic production in these medicinally important bacteria.

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