Cycloheximide-Producing Streptomyces Associated with Xyleborinus saxesenii and Xyleborus affinis Fungus-Farming Ambrosia Beetles

AbstractSymbiotic microbes help a myriad of insects acquire nutrients. Recent work suggests that insects also frequently associate with actinobacterial symbionts that produce molecules to help defend against parasites and predators. Here we explore a potential association between Actinobacteria and two species of fungus-farming ambrosia beetles, Xyleborinus saxesenii and Xyleborus affinis. We isolated and identified actinobacterial and fungal symbionts from laboratory reared nests, and characterized small molecules produced by the putative actinobacterial symbionts. One 16S rRNA phylotype of Streptomyces (XylebKG-1) was abundantly and consistently isolated from the nests and adults of X. saxesenii and X. affinis nests. In addition to Raffaelea sulphurea, the symbiont that X. saxesenii cultivates, we also repeatedly isolated a strain of Nectria sp. that is an antagonist of this mutualism. Inhibition bioassays between S. griseus XylebKG-1 and the fungal symbionts from X. saxesenii revealed strong inhibitory activity of the actinobacterium towards the fungal antagonist Nectria sp. but not the fungal mutualist R. sulphurea. Bioassay guided HPLC fractionation of S. griseus XylebKG-1 culture extracts, followed by NMR and mass spectrometry identified cycloheximide as the compound responsible for the observed growth inhibition. A biosynthetic gene cluster putatively encoding cycloheximide was also identified in S. griseus XylebKG-1. The consistent isolation of a single 16S phylotype of Streptomyces from two species of ambrosia beetles, and our finding that a representative isolate of this phylotype produces cycloheximide, which inhibits a parasite of the system but not the cultivated fungus, suggests that these actinobacteria may play defensive roles within these systems.

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Pentaminomycins F and G, First Non-Ribosomal Peptides Containing 2-Pyridylalanine

10.26434/chemrxiv.13142786 ◽

2020 ◽

Cited By ~ 1

Author(s):

Daniel Carretero Molina ◽

Francisco Javier Ortiz-Lopez ◽

Jesús Martín ◽

Ignacio González ◽

Marina Sánchez-Hidalgo ◽

...

Keyword(s):

Mass Spectrometry ◽

Amino Acid ◽

Gene Cluster ◽

Biosynthetic Gene Cluster ◽

Biosynthetic Gene ◽

Peptide Synthetase ◽

Non Ribosomal Peptides

Pentaminomycins F-H, a group of three new hydroxyarginine-containing cyclic pentapeptides, were isolated from cultures of a Streptomyces cacaoi subsp. cacaoi strain along with the known pentaminomycins A-E. The structures of the new peptides were determined by a combination of mass spectrometry and NMR and Marfey's analyses. Among them, pentaminomycins F and G were shown to contain in their structures the rare amino acid 3-(2-pyridyl)-alanine. This finding represents the first reported example of non-ribosomal peptides containing this residue. The LDLLD chiral sequence found for the three compounds was in agreement with that reported for previously isolated pentaminomycins and consistent with the epimerization domains present in the putative non-robosomal peptide synthetase (NRPS) biosynthetic gene cluster.

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Unexpected distribution of the 4-formylaminooxyvinylglycine (FVG) biosynthetic pathway in Pseudomonas and beyond

PLoS ONE ◽

10.1371/journal.pone.0247348 ◽

2021 ◽

Vol 16 (4) ◽

pp. e0247348

Author(s):

Edward W. Davis ◽

Rachel A. Okrent ◽

Viola A. Manning ◽

Kristin M. Trippe

Keyword(s):

Mass Spectrometry ◽

Gene Cluster ◽

Biosynthetic Pathway ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Species Group ◽

Biosynthetic Gene ◽

Additional Gene ◽

Genetic Potential ◽

Lines Of Evidence

The biological herbicide and antibiotic 4-formylaminooxyvinylglycine (FVG) was originally isolated from several rhizosphere-associated strains of Pseudomonas fluorescens. Biosynthesis of FVG is dependent on the gvg biosynthetic gene cluster in P. fluorescens. In this investigation, we used comparative genomics to identify strains with the genetic potential to produce FVG due to presence of a gvg gene cluster. These strains primarily belong to two groups of Pseudomonas, P. fluorescens and P. syringae, however, a few strains with the gvg cluster were found outside of Pseudomonas. Mass spectrometry confirmed that all tested strains of the P. fluorescens species group produced FVG. However, P. syringae strains did not produce FVG under standard conditions. Several lines of evidence regarding the transmission of the gvg cluster including a robust phylogenetic analysis suggest that it was introduced multiple times through horizontal gene transfer within the Pseudomonas lineage as well as in select lineages of Thiomonas, Burkholderia and Pantoea. Together, these data broaden our understanding of the evolution and diversity of FVG biosynthesis. In the course of this investigation, additional gene clusters containing only a subset of the genes required to produce FVG were identified in a broad range of bacteria, including many non-pseudomonads.

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Antimicrobial Activity and Identification of the Biosynthetic Gene Cluster of X-14952B From Streptomyces sp. 135

Frontiers in Microbiology ◽

10.3389/fmicb.2021.703093 ◽

2021 ◽

Vol 12 ◽

Author(s):

Na Li ◽

Simin Chen ◽

Zhiqiang Yan ◽

Jinhua Han ◽

Yongquan Ta ◽

...

Keyword(s):

Antimicrobial Activity ◽

Antifungal Activity ◽

16S Rrna ◽

16S Rrna Gene ◽

Gene Cluster ◽

Genome Mining ◽

Biosynthetic Gene Cluster ◽

Rrna Gene ◽

Biosynthetic Gene ◽

Type Strains

The bacterial genus Streptomyces is an important source of antibiotics, and genome mining is a valuable tool to explore the potential of microbial biosynthesis in members of this genus. This study reports an actinomycete strain 135, which was isolated from Qinghai-Tibet Plateau in China and displayed broad antimicrobial activity. The fermentation broth of strain 135 displayed strong antifungal activity (>70%) against Sclerotinia sclerotiorum, Botrytis cinerea, Valsa mali, Phytophthora capsici, Glomerella cingulata, Magnaporthe grisea, Bipolaris maydis, Exserohilum turcicum in vitro, meanwhile possessed significant preventive and curative efficacy against S. sclerotiorum, Gaeumannomyces graminis, and P. capsici on rape leaves (54.04 and 74.18%), wheat (90.66 and 67.99%), and pepper plants (79.33 and 66.67%). X-14952B showed the greatest antifungal activity against S. sclerotiorum and Fusarium graminearum which the 50% inhibition concentration (EC50) were up to 0.049 and 0.04 μg/mL, respectively. Characterization of strain 135 using a polyphasic approach revealed that the strain displayed typical features of the genus Streptomyces. 16S rRNA gene sequencing and phylogenetic analysis demonstrated that the isolate was most closely related to and formed a clade with Streptomyces huasconensis HST28T (98.96% 16S rRNA gene sequence similarity). Average nucleotide identity (ANI) and DNA-DNA hybridization (DDH) values in strain 135 and related type strains were both below the threshold of species determination (91.39 and 56.5%, respectively). OrthoANI values between strain 135 and related type strains are under the cutoff of determining species (<95%). The biosynthetic gene cluster (BGC) designated to X-14952B biosynthesis was identified through genome mining and the possible biosynthesis process was deduced.

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Linking Genes to Molecules in Eukaryotic Sources: An Endeavor to Expand Our Biosynthetic Repertoire

Molecules ◽

10.3390/molecules25030625 ◽

2020 ◽

Vol 25 (3) ◽

pp. 625

Author(s):

Jack G. Ganley ◽

Emily R. Derbyshire

Keyword(s):

Small Molecules ◽

Natural Product ◽

Biosynthetic Gene Cluster ◽

Compelling Evidence ◽

Biosynthetic Gene ◽

Guided Discovery ◽

Natural Product Discovery ◽

Chemical Structures ◽

Synthetic Utility ◽

Fungal Genomes

The discovery of natural products continues to interest chemists and biologists for their utility in medicine as well as facilitating our understanding of signaling, pathogenesis, and evolution. Despite an attenuation in the discovery rate of new molecules, the current genomics and transcriptomics revolution has illuminated the untapped biosynthetic potential of many diverse organisms. Today, natural product discovery can be driven by biosynthetic gene cluster (BGC) analysis, which is capable of predicting enzymes that catalyze novel reactions and organisms that synthesize new chemical structures. This approach has been particularly effective in mining bacterial and fungal genomes where it has facilitated the discovery of new molecules, increased the understanding of metabolite assembly, and in some instances uncovered enzymes with intriguing synthetic utility. While relatively less is known about the biosynthetic potential of non-fungal eukaryotes, there is compelling evidence to suggest many encode biosynthetic enzymes that produce molecules with unique bioactivities. In this review, we highlight how the advances in genomics and transcriptomics have aided natural product discovery in sources from eukaryotic lineages. We summarize work that has successfully connected genes to previously identified molecules and how advancing these techniques can lead to genetics-guided discovery of novel chemical structures and reactions distributed throughout the tree of life. Ultimately, we discuss the advantage of increasing the known biosynthetic space to ease access to complex natural and non-natural small molecules.

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Unexpected distribution of the 4-formylaminooxyvinylglycine (FVG) biosynthetic pathway in Pseudomonas and beyond

10.1101/2021.02.08.430221 ◽

2021 ◽

Author(s):

Edward W. Davis ◽

Rachel A. Okrent ◽

Viola A. Manning ◽

Kristin M. Trippe

Keyword(s):

Mass Spectrometry ◽

Gene Cluster ◽

Biosynthetic Pathway ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Species Group ◽

Biosynthetic Gene ◽

Additional Gene ◽

Genetic Potential ◽

Lines Of Evidence

ABSTRACTThe biological herbicide and antibiotic 4-formylaminooxyvinylglycine (FVG) was originally isolated from several rhizosphere-associated strains of Pseudomonas fluorescens. Biosynthesis of FVG is dependent on the gvg biosynthetic gene cluster in P. fluorescens. In this investigation, we used comparative genomics to identify strains with the genetic potential to produce FVG due to presence of a gvg gene cluster. These strains primarily belong to two groups of Pseudomonas, P. fluorescens and P. syringae, however, a few strains with the gvg cluster were found outside of Pseudomonas. Mass spectrometry confirmed that all tested strains of the P. fluorescens species group produced FVG. However, P. syringae strains did not produce FVG under standard conditions. Several lines of evidence regarding the transmission of the gvg cluster including a robust phylogenetic analysis suggest that it was introduced multiple times through horizontal gene transfer within the Pseudomonas lineage as well as in select lineages of Thiomonas, Burkholderia and Pantoea. Together, these data broaden our understanding of the evolution and diversity of FVG biosynthesis. In the course of this investigation, additional gene clusters containing only a subset of the genes required to produce FVG were identified in a broad range of bacteria, including many non-pseudomonads.

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Signalling and Bioactive Metabolites from Streptomyces sp. RK44

Molecules ◽

10.3390/molecules25030460 ◽

2020 ◽

Vol 25 (3) ◽

pp. 460 ◽

Cited By ~ 4

Author(s):

Qing Fang ◽

Fleurdeliz Maglangit ◽

Linrui Wu ◽

Rainer Ebel ◽

Kwaku Kyeremeh ◽

...

Keyword(s):

Mass Spectrometry ◽

Proliferative Activity ◽

Structural Diversity ◽

Biosynthetic Gene Cluster ◽

Bioactive Metabolites ◽

Ionization Mass ◽

Biosynthetic Gene ◽

Preliminary Screening ◽

Streptomyces Sp

Streptomyces remains one of the prolific sources of structural diversity, and a reservoir to mine for novel natural products. Continued screening for new Streptomyces strains in our laboratory led to the isolation of Streptomyces sp. RK44 from the underexplored areas of Kintampo waterfalls, Ghana, Africa. Preliminary screening of the metabolites from this strain resulted in the characterization of a new 2-alkyl-4-hydroxymethylfuran carboxamide (AHFA) 1 together with five known compounds, cyclo-(L-Pro-Gly) 2, cyclo-(L-Pro-L-Phe) 3, cyclo-(L-Pro-L-Val) 4, cyclo-(L-Leu-Hyp) 5, and deferoxamine E 6. AHFA 1, a methylenomycin (MMF) homolog, exhibited anti-proliferative activity (EC50 = 89.6 µM) against melanoma A2058 cell lines. This activity, albeit weak is the first report amongst MMFs. Furthermore, the putative biosynthetic gene cluster (ahfa) was identified for the biosynthesis of AHFA 1. DFO-E 6 displayed potent anti-plasmodial activity (IC50 = 1.08 µM) against P. falciparum 3D7. High-resolution electrospray ionization mass spectrometry (HR ESIMS) and molecular network assisted the targeted-isolation process, and tentatively identified six AHFA analogues, 7–12 and six siderophores 13–18.

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Pentaminomycins F and G, First Non-Ribosomal Peptides Containing 2-Pyridylalanine

10.26434/chemrxiv.13142786.v1 ◽

2020 ◽

Author(s):

Daniel Carretero Molina ◽

Francisco Javier Ortiz-Lopez ◽

Jesús Martín ◽

Ignacio González ◽

Marina Sánchez-Hidalgo ◽

...

Keyword(s):

Mass Spectrometry ◽

Amino Acid ◽

Gene Cluster ◽

Biosynthetic Gene Cluster ◽

Biosynthetic Gene ◽

Peptide Synthetase ◽

Non Ribosomal Peptides

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Anacyclamide D8P, a prenylated cyanobactin from a Sphaerospermopsis sp. cyanobacterium

10.26434/chemrxiv.5346739.v1 ◽

2017 ◽

Cited By ~ 1

Author(s):

Joana Martins ◽

Niina Leikoski ◽

Matti Wahlsten ◽

Joana Azevedo ◽

Jorge Antunes ◽

...

Keyword(s):

Gene Cluster ◽

Cyclic Peptides ◽

Biosynthetic Gene Cluster ◽

The Novel ◽

Tyrosine Residue ◽

Biosynthetic Gene ◽

Post Translational Modification ◽

Biological Assays ◽

Mass Spectrometric Data ◽

Spectrometric Data

Cyanobactins are a family of linear and cyclic peptides produced through the post-translational modification of short precursor peptides. Anacyclamides are macrocyclic cyanobactins with a highly diverse sequence that are common in the genus Anabaena. A mass spectrometry-based screening of potential cyanobactin producers led to the discovery of a new prenylated member of this family of compounds, anacyclamide D8P (1), from Sphaerospermopsis sp. LEGE 00249. The anacyclamide biosynthetic gene cluster (acy) encoding the novel macrocyclic prenylated cyanobactin, was sequenced. Heterologous expression of the acy gene cluster in Escherichia coli established the connection between genomic and mass spectrometric data. Unambiguous establishment of the type and site of prenylation required the full structural elucidation of 1 using Nuclear Magnetic Resonance (NMR), which demonstrated that a forward prenylation occurred on the tyrosine residue. Compound 1 was tested in pharmacologically or ecologically relevant biological assays and revealed moderate antimicrobial activity towards the fouling bacterium Halomonas aquamarina CECT 5000.

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