Biology subjects

van Wezel, G.

Publications and source records attributed to van Wezel, G..

At least 19 recordsLinked to original sources

bioRxiv2026

Reconstructing plant beneficial bacterial consortia by integrating dilution-to-extinction microbiome perturbation with genome-resolved synthetic ecology

Across the biosphere, microbiomes play essential roles in shaping the health of their host. One notable example of such a microbiome-associated phenotype is disease-suppressive soils, where susceptible plant hosts enrich and activate specific rhizosphere microbial consortia for protection against fungal root pathogens. However, identifying and reconstructing microbial consortia responsible for host protection remains challenging, given the inherent taxonomic and functional complexity of microbiomes. Here, we integrated metagenomic profiling of disease-suppressive microbiomes perturbed by dilution-to-extinction (DTE) with comprehensive culturing and synthetic ecology to identify the key bacterial taxa conferring suppressiveness to the fungal wheat pathogen Fusarium culmorum. Metagenomics of wheat rhizosphere samples along the DTE trajectory revealed bacterial taxa and functions associated with the disease-suppressive phenotype. Crosslinking these DTE metagenome data with a genome-sequenced collection of 336 rhizobacterial isolates from the suppressive soil allowed the reconstruction of synthetic communities (SynComs) of 11 de-replicated strains negatively associated with disease severity. Upon re-introduction in sterilized suppressive soils, this SynCom consistently reproduced the disease-suppressive phenotype. Paired time-series metagenomics and metatranscriptomics of the SynComs pinpointed candidate biosynthetic gene clusters, including a novel non-alpha poly-amino-acid (NAPAA) gene cluster from Arthrobacter, upregulated in presence of F. culmorum. Chemically synthesized NAPAA variants {varepsilon}-poly-L-lysine and {delta}-poly-L-ornithine significantly inhibited F. culmorum hyphal growth. Collectively, our work establishes a transformative strategy for reconstructing microbial consortia that recapitulates beneficial microbiome-associated phenotypes in plant and animal kingdoms.

microbiology↗

bioRxiv2025

Morphological plasticity of endophytic Chitinophaga pinensis

Environmental changes, whether due to climate change or human influences, compromise the resilience of plants to biotic and abiotic stresses, such as pathogens, drought and heat. Plant microbiota are known to promote plant resilience. To be able to harness the power of the plant microbiome we need to identify microbiota with health-promoting properties. Recent studies have demonstrated that the bacterium Chitinophaga pinensis enhances plant health and increases resistance to fungal infections. Here, we show that C. pinensis exhibits an unusually high morphological plasticity, switching between a filamentous and a spherical cell state, each of which is characterized by a distinct transcriptional profile. Despite these transcriptional differences, spherical cells remained metabolically active and replicating, while lacking structural characteristics typically associated with dormant states. Furthermore, the spherical cell morphology of C. pinensis facilitates hitchhiking behaviour and motility via surfactin cheating, potentially influencing its dispersal and interactions within the plant microbiome. To investigate the structural dynamics and transcriptional adaptation of this plant endophyte, we applied a combination of microscopy and culture-based techniques. Taken together, our study provides new insights into the morphological flexibility and transcriptional regulation of the plant-beneficial C. pinensis.

microbiology↗

bioRxiv2025

Defining the minimal structural requirements of DivIVA in filamentous Actinomycetota

The morphogenetic protein DivIVA exhibits diverse functions across bacterial phyla. In Bacillota, DivIVA is primarily involved in cell division, whereas in Actinomycetota, it plays a central role in coordinating polar growth. Due to its essential nature, gaining insight into DivIVA function is challenging. Here we report on the functionality of truncated DivIVA proteins, using a unique divIVA deletion mutant created in cell wall-deficient Kitasatospora viridifaciens L-forms. DivIVA comprises an N-terminal domain, two coiled-coil regions separated by an intercoil linker, and a C-terminal domain. Deleting either the intercoil or the C-terminal region impacted branching dynamics. We also created a minimized variant wherein both were deleted simultaneously, containing the N-terminus and fused coiled-coils, resembling DivIVA from unicellular bacteria. Expression of this minimized variant resulted in severe growth defects. Cells exhibited a strong increase in hyphal width and cell wall thickness, accompanied by frequent tip bursting. Finally, we successfully introduced chimeric DivIVA from the unicellular actinobacterium Mycolicibacterium smegmatis with an N-terminal domain of Kitasatospora viridifaciens, demonstrating functional conservation within the phylum. In contrast, a chimeric DivIVA from Bacillus subtilis could not support growth, underscoring that polar growth is encoded within Actinomycetota-specific amino acid motifs encoded in the first and second coiled-coil. These findings enhance our understanding of the structure-function relationship for DivIVA and present new opportunities to study polar growth. ImpactDivIVA is essential for polar growth in Actinomycetota. In Streptomycetaceae, this membrane-binding protein localizes at growing hyphal tips and along lateral hyphal walls where new branches emerge. Due to its essentiality, the structural relationship of DivIVA between unicellular and multicellular species remains elusive. Using a Kitasatospora viridifaciens L-form divIVA deletion mutant, we expressed truncated DivIVA variants to identify essential regions. Deleting two large unstructured domains produced a minimized variant containing the N-terminus and fused coiled-coils, resembling DivIVA from unicellular bacteria. This strongly impacted morphogenesis, increasing hyphal width and cell wall thickness, and leading to hyphal tip bursting. Finally, we successfully substituted DivIVA of K. viridifaciens with that from Mycolicibacterium smegmatis. Furthermore, Bacillus subtilis DivIVA could not facilitate reversion, showing that polar growth depends on amino acid motifs unique to Actinomycetota. These findings enhance our understanding of the structure-function relationship of DivIVA and offer new opportunities to study polar growth.

microbiology↗

bioRxiv2023

Finding needles in haystacks: identification of novel conserved PETase enzymes in Streptomyces

The rising use of plastic results in an appalling amount of waste which scatters into the environment affecting environmental, animal, and human health. One of these plastics is PET which is mainly used for bottles and textiles. In this research, we investigate the PET degrading ability of the IsPETase homolog ScLipA from Streptomyces coelicolor. Of 96 different Streptomyces strains screened, 18 % were able to degrade the model substrate BHET. Three different variants of lipase A, named ScLipA, S2LipA and S92LipA were identified and analyzed in detail. The lipA gene was deleted from S. coelicolor M145 using CRISPR/Cas9, resulting in reduced BHET degradation. LipA overexpression in the knock-out background significantly enhanced BHET degradation. All three enzymes were expressed in E. coli BL21 for protein purification and biochemical analysis, showing that enzymatic activity most likely resides in a dimeric form of the enzyme. The optimum pH and temperature were determined to be pH 7 and 25 {degrees}C for all three variants. Using these conditions, the activity on BHET and amorphous PET film was investigated. S2LipA efficiently degraded BHET and caused roughening and small indents on the surface of PET films, consistent with PET-degrading activity. The frequent occurrence of the S2LipA variant in Streptomyces suggests an environmental advantage towards the degradation of more hydrophobic substrates such as these polluting plastics in the environment.

microbiology↗

bioRxiv2023

Probiotics supplementation to adult human small intestinal stoma microbiota causes dynamic increase in the community resistance to perturbations and nutrient utilization

The gut microbiota plays a pivotal role in health and disease. The use of probiotics as microbiota-targeted therapies is a promising strategy to improve host health. However, dynamic molecular mechanisms are often not elucidated, especially when targeting the small intestinal microbiota. Here, we show that supplementation of a probiotic formula (Ecologic(R)825) to the adult human small intestinal ileostoma microbiota counteracts the growth of Enterococcaceae and Enterobacteriaceae and reduces ethanol production, leading to major changes in nutrient utilization and resistance to perturbations. The observed alterations coincided with an initial increase in lactate production and decrease in pH by the probiotics, followed by a sharp increase in the levels of butyrate and propionate. Additionally, increased production of multiple N-acyl amino acids was detected in the stoma samples supplemented with the probiotic formula. Overall, this study shows how network theory can be used to improve the current or identify novel microbiota-targeted therapies. The outcome may help further understand the reported effects of these probiotic formula on the host.

microbiology↗

bioRxiv2022

The plant stress hormone jasmonic acid evokes defensive responses in streptomycetes

Actinobacteria are prevalent in the rhizosphere and phyllosphere of diverse plant species where they help to enhance tolerance of plants against biotic and abiotic stresses. Here, we show that the plant hormones jasmonic acid (JA) and methyljasmonate (MeJA) alter growth, development and specialized metabolism of Streptomyces. Challenge of Streptomyces coelicolor with JA or MeJA led to strongly enhanced production of the polyketide antibiotic actinorhodin. JA is toxic to Streptomycetaceae, whereby members of the genus Streptacidiphilus are generally more sensitive than streptomycetes. As a defensive response, extensive amino acid conjugation of JA was observed; the most prevalent conjugation was with glutamine (Gln), while conjugates with Val, Tyr, Phe and Leu/Ile were identified after longer exposure to JA. Synthetic JA conjugates failed to activate antibiotic production and had strongly reduced toxicity, strongly suggesting that conjugation inactivates JA and serves to detoxify the hormone. Thus, for the first time we provide evidence that plant hormones modulate growth, development and secondary metabolism of streptomycetes, whereby amino acid conjugation serves as a defense strategy by the bacteria to circumvent plant hormone toxicity. IMPORTANCEMicroorganisms that live on or inside plants greatly influence plant health. Streptomycetes are considered to have an important role in defense against plant diseases, but the mechanisms through which they protect plants are currently not fully understood. It has been suggested that streptomycetes respond to changes in the plants physiology, among others by producing protective molecules; however, little is known of the signal transduction from plant to bacterium. We here demonstrate that the plant hormones jasmonic acid (JA) and methyljasmonate (MeJA) directly influence the life cycle of streptomycetes by modulating antibiotic synthesis and promoting faster development. Moreover, the plant hormones specifically stimulate the synthesis of the polyketide antibiotic actinorhodin in Streptomyces coelicolor. Jasmonic acid is then modified in the cell by amino acid conjugation, which reduces the bioactivity of the hormone and thus quenches the signal. To the best of our knowledge, this has not been reported previously. Collectively, these results suggest a relationship between plant physiological changes and the response of streptomycetes in multiple ways.

microbiology↗

bioRxiv2022

Taxonomic and metabolic diversity of Actinobacteria isolated from faeces of a 28,000-year-old mammoth

Ancient microbial communities of permafrost soils and frozen animal remains represent an archive that has barely been explored. This yet unexplored microbial world is a vast resource that can provide us with new evolutionary insights, metabolic pathways and novel chemistry. Here, we reveal that Actinobacteria isolated from a faecal sample from the intestinal tract of a 28,000-year-old Siberian mammoth are phylogenetically and metabolically distinct from currently known modern siblings. Ancient Micromonospora, Oerskovia, Saccharopolyspora, Sanguibacter and Streptomyces species were successfully revived and their genome sequences resolved. Surprisingly, the genomes of the ancestors show a large phylogenetic distance to strains isolated today and harbour many novel biosynthetic gene clusters that may well represent uncharacterised biosynthetic potential. Metabolic profiles of the strains display production of known molecules like antimycin, conglobatin and macrotetrolides, but the majority of the mass features could not be dereplicated. Our work provides a snapshot into Actinobacteria of the past, yielding unexplored genomic information that is not yet present in current databases.

microbiology↗

bioRxiv2022

Reproducible switching between a walled and cell wall-deficient lifestyle of actinomycetes using gradient agar plates

The cell wall is a shape-defining structure that envelopes almost all bacteria, protecting them from biotic and abiotic stresses. Paradoxically, some filamentous actinomycetes have a natural ability to shed their cell wall under influence of hyperosmotic stress. These wall-deficient cells can revert to their walled state when transferred to a medium without osmoprotection but often lyse due to their fragile nature. Here, we designed plates with an osmolyte gradient to reduce cell lysis and thereby facilitating the transition between a walled and wall-deficient state. These gradient plates allow determining of the osmolyte concentration where switching takes place, thereby enabling careful and reproducible comparison between mutants affected by switching. Exploring these transitions could give valuable insights into the ecology of actinomycetes and their biotechnological applications. HIGHLIGHTS- Using agar plates with a gradient of osmoprotectants, revertant Streptomycetes can gradually revert to a walled state, thereby dramatically decreasing lysis. - Our method allows precise determination of the osmolyte concentration where reversion takes place, allowing careful and reproducible comparison between mutants. - Gradient agar plates can also be used to study chemical differentiation in Streptomycetes as a response to osmotic stress.

microbiology↗

bioRxiv2022

DNA uptake by cell wall-deficient bacteria reveals a putative ancient macromolecule uptake mechanism

Horizontal gene transfer in bacteria is widely believed to occur via three main mechanisms: conjugation, transduction and transformation. These mechanisms facilitate the passage of DNA across the protective cell wall using sophisticated machinery. We present here a new mechanism of DNA uptake that is independent of canonical DNA uptake machineries and is used by bacteria that live without a cell wall. We show that the cell wall-deficient bacteria engulf extracellular material, whereby intracellular vesicles are formed, and DNA is internalized. This mechanism is not specific to DNA, and allows uptake of other macromolecules and even 125 nm lipid nanoparticles (LNPs). Uptake was prevented by molecules known to inhibit eukaryotic endocytosis, suggesting this to be an energy-dependent process. Given that cell wall-deficient bacteria are considered a model for early life forms, our work provides a possible mechanism for primordial cells to acquire new genetic material or food before invention of the bacterial cell wall.

microbiology↗

bioRxiv2021

Role for a lytic polysaccharide monooxygenase in cell wall remodelling

Peptidoglycan is a major constituent of the bacterial cell wall and an important determinant for providing protection to cells. Besides peptidoglycan (PG), many bacteria synthesize other glycans that become part of the cell wall. Streptomycetes grow apically, where they synthesize a glycan that is exposed at the outer surface, but how it gets there is unknown. Here we show that deposition of the apical glycan at the cell surface depends on two key enzymes, the endoglucanase CslZ and the lytic polysaccharide monooxygenase LpmP. Activity of these enzymes allows localized remodeling and degradation of the PG, and we propose that this facilitates passage of the glycan. The absence of both enzymes not only prevents morphological development, but also sensitizes strains to lysozyme. Given that lytic polysaccharide monooxygenases are commonly found in microbes, this newly identified biological role in cell-wall remodelling may be widespread.

microbiology↗

bioRxiv2020

RRE-Finder: A Genome-Mining Tool for Class-Independent RiPP Discovery

Nearly half of the classes of natural products known as ribosomally synthesized and post-translationally modified peptides (RiPPs) are reliant on a protein domain called the RiPP recognition element (RRE) for peptide maturation. The RRE binds specifically to a linear precursor peptide and directs the post-translational modification enzymes to their substrate. Given its prevalence across various types of RiPP biosynthetic gene clusters (BGCs), the RRE could theoretically be used as a bioinformatic handle to identify novel classes of RiPPs. In addition, due to the high affinity and specificity of most RRE:precursor peptide complexes, a thorough understanding of the RRE domain could be exploited for biotechnological applications. However, sequence divergence of the RRE domain across RiPP classes has precluded automated identification of RREs based solely on sequence similarity. Here, we introduce RRE-Finder, a novel tool for identifying RRE domains with high sensitivity. RRE-Finder can be used in "precision" mode to confidently identify RREs in a class-specific manner or in "exploratory" mode, which was designed to assist in the discovery of novel RiPP classes. RRE-Finder operating in precision mode on the UniProtKB protein database retrieved over 30,000 high-confidence RREs spanning all characterized RRE-dependent RiPP classes, as well as several yet-uncharacterized RiPP, putatively novel gene cluster architectures that will require future experimental work. Finally, RRE-Finder was used in precision mode to explore a possible evolutionary origin of the RRE domain. Altogether, RRE-Finder provides a powerful new method to probe RiPP biosynthetic diversity and delivers a rich dataset of RRE sequences that will provide a foundation for deeper biochemical studies into this intriguing and versatile protein domain.

bioinformatics↗

bioRxiv2019

Genome rearrangements and megaplasmid loss in the filamentous bacterium Kitasatospora viridifaciens are associated with protoplast formation and regeneration

Filamentous Actinobacteria are multicellular bacteria with linear replicons. Kitasatospora viridifaciens DSM 40239 contains a linear 7.8 Mb chromosome and an autonomously replicating plasmid KVP1 of 1.7 Mb. Here we show that lysozyme-induced protoplast formation of the multinucleated mycelium of K. viridifaciens drives morphological diversity. Characterization and sequencing of an individual revertant colony that had lost the ability to differentiate revealed that the strain had not only lost most of KVP1 but also carried lesions in the right arm of the chromosome. Strikingly, the lesion sites were preceded by insertion sequence elements, suggesting that the rearrangements may have been caused by replicative transposition and homologous recombination between both replicons. These data indicate that protoplast formation is a stressful process that can lead to profound genetic changes.\n\nRepositoriesGenomic sequence data for strain B3.1 has been deposited in the NCBI SRA database under accession code SAMN11514356.

microbiology↗

bioRxiv2018

High-resolution analysis of the peptidoglycan composition in Streptomyces coelicolor

The bacterial cell wall maintains cell shape and protects against bursting by the turgor. A major constituent of the cell wall is peptidoglycan (PG), which is continuously modified to allow cell growth and differentiation through the concerted activity of biosynthetic and hydrolytic enzymes. Streptomycetes are Gram-positive bacteria with a complex multicellular life style alternating between mycelial growth and the formation of reproductive spores. This involves cell-wall remodeling at apical sites of the hyphae during cell elongation and autolytic degradation of the vegetative mycelium during the onset of development and antibiotic production. Here, we show that there are distinct differences in the cross-linking and maturation of the PG between exponentially growing vegetative hyphae and the aerial hyphae that undergo sporulation. LC-MS/MS analysis identified over 80 different muropeptides, revealing that major PG hydrolysis takes place over the course of mycelial growth. Half of the dimers lack one of the disaccharide units in transition-phase cells, most likely due to autolytic activity. De-acetylation of MurNAc to MurN was particularly pronounced in spores, suggesting that MurN plays a role in spore development. Taken together, our work highlights dynamic and growth phase-dependent construction and remodeling of PG in Streptomyces.\n\nIMPORTANCEStreptomycetes are bacteria with a complex lifestyle, which are model organisms for bacterial multicellularity. From a single spore a large multigenomic, multicellular mycelium is formed, which differentiates to form spores. Programmed cell death is an important event during the onset of morphological differentiation. In this work we provide new insights into the changes in the peptidoglycan architecture over time, highlighting changes over the course of development and between growing mycelia and spores. This revealed dynamic changes in the peptidoglycan when the mycelia age, showing extensive PG hydrolysis and in particular an increase in the proportion of 3-3-cross-links. Additionally, we identified a muropeptide that is highly abundant specifically in spores, which may relate to dormancy and germination.

biochemistry↗

bioRxiv2018

Classification of the gifted natural product producer Streptomyces roseofaciens sp. nov. by polyphasic taxonomy

A novel verticillate strain of streptomycetes, Streptomyces strain MBT76T, was isolated from the QinLing mountains, which harbours more than 40 biosynthetic gene clusters for natural products. Here we present full taxonomic classification of strain MBT76T, and show that it has chemotaxonomic, genomic and morphological properties consistent with its classification in the genus Streptomyces. Strain MBT76T is part of the cluster of Streptoverticillates, a group within the genus Streptomyces that has characteristic whorl-forming spores produced in chains along the lateral wall of the hyphae. Multi-locus sequence analysis based on five housekeeping gene alleles showed that MBT76T is closely related to Streptomyces hiroshimensis. Average Nucleotide Identification (ANI) and Genome to Genome Distance Calculation (GGDC) of the genomes of strain MBT76T and S. hiroshimensis separated them into distinct species. Strain MBT76T represents a novel species of the genus Streptomyces for which we propose the name Streptomyces roseofaciens sp. nov. The type strain is MBT76T (=NCCB 100637T =DSM 106196T). The whole genome of MBT76T has 7974 predicted open reading frames and a total genome size of 8.64 Mb. Further genomic analysis showed that verticillate streptomycetes lack the sporulation gene ssgE, and our data suggest that this is a useful genetic marker for the spore-chain morphology of the verticillates.

microbiology↗

bioRxiv2017

Inter- and intracellular colonization of Arabidopsis roots by endophytic actinobacteria and the impact of plant hormones on their antimicrobial activity

Many actinobacteria live in close association with eukaryotes like fungi, insects, animals and plants. Plant-associated actinobacteria display (endo)symbiotic, saprophytic or pathogenic life styles, and can make up a substantial part of the endophytic community. Here, we characterised endophytic actinobacteria isolated from root tissue of Arabidopsis thaliana (Arabidopsis) plants grown in soil from a natural ecosystem. Many of these actinobacteria belong to the family of Streptomycetaceae with Streptomyces olivochromogenes and Streptomyces clavifer as well represented species. When seeds of Arabidopsis were inoculated with spores of Streptomyces strain coa1, which shows high similarity to S. olivochromogenes, roots were colonised intercellularly and, unexpectedly, also intracellularly. Subsequent exposure of endophytic isolates to plant hormones typically found in root and shoot tissues of Arabidopsis led to altered antibiotic production against Escherichia coli and Bacillus subtilis. Taken together, our work reveals remarkable colonization patterns of endophytic streptomycetes with specific traits that may allow a competitive advantage inside root tissue.

microbiology↗

bioRxiv2017

YlmD and YlmE are required for correct sporulation-specific cell division in Streptomyces coelicolor A3(2)

Cell division during the reproductive phase of the Streptomyces life-cycle requires tight coordination between synchronous formation of multiple septa and DNA segregation. One remarkable difference with most other bacterial systems is that cell division in Streptomyces is positively controlled by the recruitment of FtsZ by SsgB. Here we show that deletion of ylmD (SCO2081) or ylmE (SCO2080), which lie in operon with ftsZ in the dcw cluster of actinomycetes, has major consequences for sporulation-specific cell division in Streptomyces coelicolor. Electron and fluorescence microscopy demonstrated that ylmE mutants have a highly aberrant phenotype with defective septum synthesis, and produce very few spores with low viability and high heat sensitivity. FtsZ-ring formation was also highly disturbed in ylmE mutants. Deletion of ylmD had a far less severe effect on sporulation. Interestingly, the additional deletion of ylmD restored sporulation to the ylmE null mutant. YlmD and YlmE are not part of the divisome, but instead localize diffusely in aerial hyphae, with differential intensity throughout the sporogenic part of the hyphae. Taken together, our work reveals a function for YlmD and YlmE in the control of sporulation-specific cell division in S. coelicolor, whereby the presence of YlmD alone results in major developmental defects.

microbiology↗

bioRxiv2017

Micro-encapsulation extends mycelial viability of Streptomyces lividans 66 and increases enzyme production

BackgroundFilamentous bacteria of the genus Streptomyces produce a large arsenal of industrially relevant antibiotics and enzymes. The industrial production of these molecules occurs in large fermenters, where many streptomycetes form dense mycelial networks called pellets. Pellets are characterized by slow growth and inefficient nutrient transfer and therefore regarded as undesirable from the perspective of productivity. Although non-pelleting strains have increased growth rates, their morphology also leads to a dramatic increase in the viscosity of the culture broth, which negatively impacts the process dynamics.\n\nResultsHere, we applied immobilization of Streptomyces lividans 66 using alginate as semi-solid matrix. This alginate-mediated micro-encapsulation increased the production of the extracellular enzyme tyrosinase more than three-fold. The increased production was accompanied by extended viability of the mycelium and a dramatic reduction in the release of intracellular proteins into the culture broth.\n\nConclusionsOur data demonstrate the utility of microencapsulation as a powerful technique to achieve higher yields and lower downstream-processing costs of streptomycetes.

microbiology↗

bioRxiv2017

Morphology-driven downscaling of Streptomyces lividans to micro-cultivation

Actinobacteria are prolific producers of secondary metabolites and industrially relevant enzymes. Growth of these mycelial microorganisms in small culture volumes is challenging due to their complex morphology. Since morphology and production are typically linked, scaling down culture volumes requires better control over morphogenesis. In larger scale platforms, ranging from shake flasks to bioreactors, the hydrodynamics play an important role in shaping the morphology and determining product formation. Here, we report on the effects of agitation on the mycelial morphology of Streptomyces lividans grown in microtitre plates (MTP). Our work shows that at the proper agitation rates cultures can be scaled down to volumes as small as 100 l while maintaining the same morphology as seen in larger scale platforms. Using image analysis we compared the morphologies of the cultures; when agitated at 1400 rpm the mycelial morphology in microcultures approached that obtained in shake flasks, while product formation was also maintained. Our study shows that the morphology of actinobacteria in microcultures can be controlled in a similar manner as in larger scale cultures by carefully controlling the mixing rate. This could facilitate high-throughput screening and upscaling.

bioengineering↗