Biology subjects

Krismer, B.

Publications and source records attributed to Krismer, B..

7 recordsLinked to original sources

bioRxiv2025

Genetic modification of intractable staphylococcal clones by heat-shock facilitated phage transduction

Increasing recognition of commensal bacteria as a requirement for microbiome integrity and pathogen exclusion puts urgency to the molecular characterization of commensal interactions. However, many commensals cannot be transformed with available methodology because of potent restriction barriers. We developed a new method for the introduction of plasmid DNA into otherwise intractable non-Staphylococcus aureus (NAS) staphylococci, important commensal members of human nose and skin microbiomes, by phage transduction. We demonstrate that a precise pulse of recipient bacteria with elevated temperatures prior to exposure to transducing phages renders NAS isolates effectively and transiently amenable to transduction. Transduction of NAS mutants lacking restriction-modification (RM) systems did not profit from a heat shock indicating that it is the transient deactivation of RM enzymes that permit transduction. Our method also facilitated the transduction of other Bacillota from the genera Bacillus and Listeria indicating that it will support research on different bacterial groups from diverse ecosystems. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/647181v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@233348org.highwire.dtl.DTLVardef@fbf57corg.highwire.dtl.DTLVardef@1b1abadorg.highwire.dtl.DTLVardef@17e283f_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2024

Multiple regulators control the biosynthesis of brasilicardin in Nocardia terpenica

Brasilicardin A, BraA, is a secondary metabolite produced by the bacterium Nocardia terpenica, and a promising drug due to its potent immunosuppressive activity and low cytotoxicity. Currently, a semisynthetic approach confers production of a complete compound but suffers from insufficient heterologous biosynthesis of BraA intermediates used in the chemical semi-synthesis steps leading to only lab scale quantities of the compound. A better understanding of the involved gene expression regulatory pathways within the brasilicardin biosynthetic gene cluster, Bra-BGC, is a prerequisite to further improve production titers. However, the transcriptional regulation of the Bra-BGC has only been superficially analyzed, till now. In this study, we comprehensively analyze the functions of several unstudied transcriptional regulators, KstR, SdpR and OmpR, encoded within the close vicinity of the Bra-BGC, and delve into the role of the previously described cluster-situated activator Bra12. We present, that Bra12 and the novel regulator SdpR, bind several DNA sequences located in the promoter regions of the genes essential for BraA biosynthesis. Subsequently, we demonstrate the complex regulatory network through which both regulators are capable of controlling activity of those gene promoters and thus gene expression in Bra-BGC. Furthermore, using the heterologous producer strain Amycolatopsis japonicum, we present, that Bra12 and SdpR regulators play opposite roles in brasilicardin congener biosynthesis. Finally, we propose a comprehensive model of multilevel gene expression regulation in Bra-BGC and propose the roles of locally encoded transcriptional regulators.

molecular biology↗

bioRxiv2024

Probiotic Lactobacilli activate Formyl-Peptide Receptor 2

Changes in the composition of the human microbiota can negatively impact human health. Probiotic bacteria like many lactobacilli help prevent or repair dysbiosis but it is largely unclear which molecules of these bacteria mediate the probiotic effects. Given the extensive crosstalk between the immune system and microbiome members, we investigated whether lactobacilli activate the formyl-peptide receptor 2 (FPR2), a pattern recognition receptor that is expressed on the surface of intestinal epithelial cells and known to promote wound healing and immune homeostasis. Probiotic strains of Lacticaseibacillus paracasei, Lactiplantibacillus plantarum, and Lacticaseibacillus rhamnosus were isolated from probiotic compounds and sequenced. Calcium influx experiments in FPR1 or FPR2 overexpressing HL60 cells, and primary human neutrophils, along with pharmacological inhibition of FPR2, revealed that culture filtrates of the isolated lactobacilli strongly activate FPR2, promote killing of the methicillin resistant S. aureus USA300 and induce neutrophil chemotaxis. Pretreatment of culture filtrates with proteinase K reduced FPR2 activity, indicating that the FPR2 ligands are peptides. In silico analysis of the amphipathic properties of the signal peptides of lactic acid bacteria identified selected signal peptides of L. plantarum with the ability to predominantly activate FPR2 in vitro. Thereby, via targeted activation of FPR2, peptides released by some lactobacilli are likely to positively influence the outcome of inflammatory gut diseases and could be used to treat inflammatory diseases.

microbiology↗

bioRxiv2024

Siderophore piracy enables the nasal commensal Staphylococcus lugdunensis to antagonize the pathogen Staphylococcus aureus

Bacterial pathogens such as Staphylococcus aureus colonize body surfaces of part of the human population, which represents a critical risk factor for skin disorders and invasive infections. However, such pathogens do not belong to the human core microbiomes - beneficial commensal bacteria can often prevent the persistence of certain pathogens - using molecular strategies that are only superficially understood. We recently reported that the commensal bacterium Staphylococcus lugdunensis produces the novel antibiotic lugdunin, which eradicates S. aureus from nasal microbiomes of hospitalized patients. However, it has remained unclear if S. lugdunensis may affect S. aureus carriage in the general population and how S. lugdunensis carriage could be promoted to enhance its S. aureus-eliminating capacity. We could cultivate S. lugdunensis from the noses of 6.3% of healthy human volunteers. In addition, S. lugdunensis DNA could be identified in metagenomes of many culture-negative nasal samples indicating that cultivation success depends on a specific bacterial threshold density. Healthy S. lugdunensis carriers had a 5.2-fold lower propensity to be colonized by S. aureus indicating that lugdunin can eliminate S. aureus also in healthy humans. S. lugdunensis-positive microbiomes were dominated by either Staphylococcus epidermidis, Corynebacterium species, or Dolosigranulum pigrum. These and further bacterial commensals, whose abundance was positively associated with S. lugdunensis, promoted S. lugdunensis growth in co-culture. Such mutualistic interactions depended on the production of iron-scavenging siderophores by supporting commensals, and on the capacity of S. lugdunensis to import siderophores. These findings underscore the importance of microbiome homeostasis for eliminating pathogen colonization. Elucidating mechanisms that drive microbiome interactions will become crucial for microbiome-precision editing approaches.

microbiology↗

bioRxiv2024

Nasal commensals reduce Staphylococcus aureus proliferation by restricting siderophore availability

BackgroundThe human microbiome is critically associated with human health and disease. One aspect of this is that antibiotic-resistant opportunistic bacterial pathogens such as methicillin-resistant Staphylococcus aureus can reside within the nasal microbiota which increases the risk of infections. Epidemiological studies of the nasal microbiome have revealed positive and negative correlations between non-pathogenic species and S. aureus, but the underlying molecular mechanisms remain poorly understood. The nasal cavity is iron-limited and bacteria are known to produce iron-scavenging siderophores to proliferate in such environments. Siderophores are public goods that can be consumed by all members of a bacterial community. Accordingly, siderophores are known to mediate bacterial competition and collaboration but their role in the nasal microbiome is unknown. ResultsHere we show that siderophore acquisition is crucial for S. aureus nasal colonization in vivo. We screened 94 nasal bacterial strains from seven genera for their capacity to produce siderophores as well as to consume the siderophores produced by S. aureus. We found that 80% of the strains engaged in siderophoremediated interactions with S. aureus. Non-pathogenic corynebacterial species were found to be prominent consumers of S. aureus siderophores. In co-culture experiments, consumption of siderophores by competitors reduced S. aureus growth in an iron-dependent fashion. ConclusionsOur data show a wide network of siderophore mediated interactions between the species of the human nasal microbiome and provide mechanistic evidence for inter-species competition and collaboration impacting pathogen proliferation. This opens avenues for designing nasal probiotics to displace S. aureus from the nasal cavity of humans.

microbiology↗

bioRxiv2022

Horizontal transfer of bacteriocin biosynthesis genes requires metabolic adaptation to improve compound production and cellular fitness

Biosynthetic gene clusters (BGCs) encoding the production of bacteriocins are widespread amongst bacterial isolates and are important genetic determinants of competitive fitness within a given habitat. Staphylococci produce a tremendous diversity of compounds and the corresponding BGCs are frequently associated with mobile genetic elements, suggesting gain and loss of biosynthetic capacity. Pharmaceutical biology has shown that compound production in heterologous hosts is often challenging and many BGC recipients produce initially low compound amounts or show reduced growth rates. To assess whether transfer of BGCs between closely related S. aureus strains can be instantly effective or requires elaborate metabolic adaptation, we investigated the intra species transfer of a BGC encoding the ribosomally synthesized and post-translationally modified peptide (RiPP) micrococcin P1 (MP1). We found that acquisition of the BGC by S. aureus RN4220 enabled immediate MP1 production but also imposed a metabolic burden, which was relieved after prolonged cultivation by adaptive mutation. We used a multiomics approach to study this phenomenon and found adaptive evolution to select for strains with increased activity of the tricarboxylic acid cycle (TCA), which enhanced metabolic fitness and levels of compound production. Metabolome analysis revealed increases of central metabolites including citrate and -ketoglutarate in the adapted strain, suggesting metabolic adaptation to overcome the BGC-associated growth defects. Our results indicate that BCG acquisition requires genetic and metabolic predispositions allowing the integration of bacteriocin production into the cellular metabolism. Inappropriate metabolic characteristics of recipients can entail physiological burdens, negatively impacting the competitive fitness of recipients within natural bacterial communities. ImportanceHuman microbiomes are critically associated with human health and disease. Importantly, pathogenic bacteria can hide in human associated communities and can cause disease when the composition of the community becomes dysbalanced. Bacteriocin producing commensals are able to displace pathogens from microbial communities, suggesting that their targeted introduction in human microbiomes might prevent pathogen colonisation and infection. However, in view of future probiotic approaches, strains are needed that produce high levels of bioactive compounds and retain cellular fitness within mixed bacterial communities. Our work offers insights into the metabolic burdens associated with the production of the bacteriocin micrococcin P1 and highlights evolutionary strategies that increase cellular fitness in the context of production. Most likely metabolic adaptations are broadly relevant for bacteriocin producers and need to be considered for the future development of effective microbiome editing strategies.

microbiology↗

bioRxiv2020

Decoding and blocking the bacterial lipid flippase MprF yields a novel anti-virulence strategy

The pandemic of antibiotic resistance represents a major human health threat demanding new antimicrobial strategies. MprF is the synthase and flippase of the phospholipid lysyl-phosphatidylglycerol that increases virulence and resistance of methicillin-resistant Staphylococcus aureus (MRSA) and other pathogens to cationic host defense peptides and antibiotics. With the aim to design MprF inhibitors that could sensitize MRSA to both, human antimicrobials and antibiotics and support the clearance of staphylococcal infections with minimal selection pressure, we developed MprF-targeting monoclonal antibodies, which bound and blocked the MprF flippase subunit. Antibody M-C7.1 targeted a specific loop in the flippase domain that proved to be exposed at both sides of the bacterial membrane, thereby enhancing the mechanistic understanding into bacterial lipid translocation. M-C7.1 rendered MRSA susceptible to host antimicrobial peptides and antibiotics such as daptomycin. Moreover, it impaired MRSA survival in human phagocytes, which recommends MprF inhibitors for new anti-MRSA approaches. MprF-directed monoclonal antibodies provide a proof of concept for development of precisely targeted anti-virulence approaches, which block bacterial antimicrobial resistance mechanisms.

microbiology↗