Biology subjects

Kuipers, O. P.

Publications and source records attributed to Kuipers, O. P..

6 recordsLinked to original sources

bioRxiv2025

Pyocyanin produced by Pseudomonas aeruginosa Creates Legacy Effects That Boost Antibiotic Resistance Evolution in Enterococci

Polymicrobial infections are small communities of multiple interacting bacterial species. Interactions among constituent species may modify the growth of community members in the presence of antibiotics, for example via degradation of the antibiotic or induction of specific resistance mechanisms. However, for most polymicrobial infections the nature of such interactions is opaque, while they may affect both treatment efficacy and the evolution of antibiotic resistance. Here, we describe that past interaction of enterococci with Pseudomonas aeruginosa creates legacy effects that substantially alter their antibiotic tolerance and resistance evolution. Specifically, we find that the temporary exposure to pyocyanin, a secondary metabolite produced by P. aeruginosa, increases the efflux in enterococci. These tolerance legacy effects promote the evolution of antibiotic resistance of enterococci. This work shows that transient interactions in polymicrobial communities can alter the evolutionary fate of community members.

microbiology↗

bioRxiv2022

Brevicidine, a bacterial non-ribosomally produced cyclic antimicrobial lipopeptide with a unique modus operandi

Due to the accelerated appearance of antibiotic-resistant (AMR) pathogens in clinical infections, new first-in-class antibiotics, operating via novel modes of action, are desperately-needed. Brevicidine, a bacterial non-ribosomally produced cyclic lipopeptide, has shown potent and selective antimicrobial activity against Gram-negative pathogens. However, before our investigations, little was known about how brevicidine exerts its potent bactericidal effect against Gram-negative pathogens. In this study, we find that brevicidine has potent antimicrobial activity against AMR Enterobacteriaceae pathogens, with a MIC value ranging between 0.5M (0.8mg/L) and 2M (3.0mg/L). In addition, brevicidine showed potent anti-biofilm activity against the Enterobacteriaceae pathogens, with same 100% inhibition and 100% eradication concentration of 4M (6.1mg/L). Further mechanistic studies showed that brevicidine exerts its potent bactericidal activity via interacting with lipopolysaccharide in the outer membrane, targeting phosphatidylglycerol and cardiolipin in the inner membrane, and dissipating the proton motive force of bacteria. This results in metabolic perturbation, including inhibition of adenosine triphosphate synthesis, inhibits the dehydrogenation of nicotinamide adenine dinucleotides, accumulation of reactive oxygen species in bacteria, and inhibition of protein synthesis. Lastly, brevicidine showed a good therapeutic effect in a mouse peritonitis-sepsis model. Our findings pave the way for further research on clinical applications of brevicidine, to combat the prevalent infections caused by AMR Gram-negative pathogens worldwide.

microbiology↗

bioRxiv2021

Nisin- and ripcin-derived hybrid lanthipeptides display selective antimicrobial activity against Staphylococcus aureus

Lanthipeptides are (methyl)lanthionine ring-containing ribosomally synthesized and post-translationally modified peptides (RiPPs). Many lanthipeptides show strong antimicrobial activity against bacterial pathogens, including antibiotic-resistant bacterial pathogens. The group of disulfide bond-containing antimicrobial peptides (AMPs) is well known in nature and forms a rich source of templates for the production of novel peptides with corresponding (methyl)lanthionine analogues instead of disulfides. Here, we show that novel macrocyclic lanthipeptides (termed thanacin and ripcin) can be synthesized using the known antimicrobials thanatin and rip-thanatin as templates. Notably, the synthesized nisin(1-20)-ripcin hybrid lanthipeptides (ripcin B-G) showed selective antimicrobial activity against S. aureus, including an antibiotic-resistant MRSA strain. Interestingly, ripcin B-G, which are hybrid peptides of nisin(1-20) and ripcin, respectively, that are each inactive against Gram-negative pathogens, showed substantial antimicrobial activity against the tested Gram-negative pathogens. Moreover, ripcin B-G was highly resistant against the nisin resistance protein (NSR; a protease could cleave nisin and strongly reduce its activity), opposed to nisin itself. Mode of action studies show that ripcin C exerts its antimicrobial activity against Gram-positive pathogens by binding to the cell wall synthesis precursor lipid II and thereafter arrests cell growth. In addition, ripcin C exerts its antimicrobial activity against Gram-negative pathogens by binding to LPS and the cell wall synthesis precursor lipid II. This study provides an example of converting disulfide bond-based AMPs into (methyl)lanthionine-based macrocyclic hybrid lanthipeptides and can yield antimicrobial peptides with selective antimicrobial activity against S. aureus. O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY For Table of Contents Use Only C_FIG_DISPLAY

synthetic biology↗

bioRxiv2021

Brevibacillin 2V, a novel antimicrobial lipopeptide with an exceptional low hemolytic activity

Bacterial non-ribosomally produced peptides (NRPs) form a rich source of antibiotics, including more than twenty of these antibiotics that are used in the clinic, such as penicillin G, colistin, vancomycin, and chloramphenicol. Here we report the identification, purification, and characterization of a novel NRP, i.e., brevibacillin 2V, from Brevibacillus laterosporus DSM 25. Brevibacillin 2V has strong antimicrobial activity against Gram-positive bacterial pathogens, including difficult-to-treat antibiotic-resistant Enterococcus faecium, Enterococcus faecalis, and Staphylococcus aureus. Notably, brevibacillin 2V has a much lower hemolytic activity to eukaryotic cells than previously reported NRPs of the lipo-tridecapeptide family, including other brevibacillins, which makes it a promising candidate for antibiotic development. In addition, our results demonstrate that brevibacillins display synergistic action with established antibiotics against Gram-negative bacterial pathogens. Thus, we identified and characterized a novel and promising antibiotic candidate (brevibacillin 2V) with a low hemolytic activity, which can be used either on its own or as a template for further total synthesis and modification.

microbiology↗

bioRxiv2018

Boosting heterologous protein production yield by adjusting global nitrogen and carbon metabolic regulatory networks in Bacillus subtilis

Bacillus subtilis is extensively applied as a microorganism for the high-level production of heterologous proteins. Traditional strategies for increasing the productivity of this microbial cell factory generally focused on the targeted modification of rate-limiting components or steps. However, the longstanding problems of limited productivity of the expression host, metabolic burden and non-optimal nutrient intake, have not yet been solved to achieve production strain improvements. To tackle this problem, we systematically rewired the regulatory networks of the global nitrogen and carbon metabolism by random mutagenesis of the pleiotropic transcriptional regulators CodY and CcpA, to allow for optimal nutrient intake, translating into significantly higher heterologous protein production yields. Using a {beta}-galactosidase expression and screening system and consecutive rounds of mutagenesis, we identified mutant variants of both CcpA and CodY that in conjunction increased production levels up to 290%. RNA-Seq and electrophoretic gel mobility shift analyses showed that amino acid substitutions within the DNA-binding domains altered the overall binding specificity and regulatory activity of the two transcription factors. Consequently, fine-tuning of the central metabolic pathways allowed for enhanced protein production levels. The improved cell factory capacity was further demonstrated by the successfully increased overexpression of GFP, xylanase and a peptidase in the double mutant strain.\n\nHighlightsO_LIThe global transcription machinery engineering (gTME) technique was applied to build mutational libraries of the pleiotropic regulators CodY and CcpA in Bacillus subtilis\nC_LIO_LISpecific point mutations within the DNA-binding domains of CodY and CcpA elicited alterations of the binding specificity and regulatory activity\nC_LIO_LIChanges in the transcriptome evoked the reprogramming of networks that gear the carbon and nitrogen metabolism\nC_LIO_LIThe rewired metabolic networks provided a higher building block capacity for heterologous protein production by adjusting the nutrient uptake and channeling its utilization for protein overexpression\nC_LI

bioengineering↗

bioRxiv2017

Unleashing natural competence in Lactococcus lactis by induction of the competence regulator ComX

In biotechnological work horses like Streptococcus thermophilus and Bacillus subtilis natural competence can be induced, which facilitates genetic manipulation of these microbes. However, in strains of the important dairy starter Lactococcus lactis natural competence has not been established to date. However, in silico analysis of complete genome sequences of 43 L. lactis strains revealed complete late-competence gene-sets in 2 L. lactis subspecies cremoris strains (KW2 and KW10) and 8 L. lactis subspecies lactis strains, including the model strain IL1403 and the plant-derived strain KF147. The remainder of the strains, including all dairy isolates, displayed genomic decay in one or more of the late competence genes. Nisin-controlled expression of the competence regulator comX in L. lactis subsp. lactis KF147 resulted in the induction of expression of the canonical competence regulon, and elicited a state of natural competence in this strain. By contrast, comX expression in L. lactis NZ9000, predicted to encode an incomplete competence gene-set, failed to induce natural competence. Moreover, mutagenesis of the comEA-EC operon in strain KF147, abolished the comX driven natural competence, underpinning the involvement of the competence machinery. Finally, introduction of nisin-inducible comX expression into nisRK-harboring derivatives of strains IL1403 and KW2 allowed the induction of natural competence also in these strains, expanding this phenotype to other L. lactis strains of both subspecies.\n\nSignificance statementSpecific bacterial species are able to enter a state of natural competence in which DNA is taken up from the environment, allowing the introduction of novel traits. Strains of the species Lactococcus lactis are very important starter cultures for the fermentation of milk in the cheese production process, where these bacteria contribute to the flavor and texture of the end-product. The activation of natural competence in this industrially relevant organism can accelerate research aiming to understand industrially relevant traits of these bacteria, and can facilitate engineering strategies to harness the natural biodiversity of the species in optimized starter strains.

microbiology↗