Biology subjects

Kuemmerli, R.

Publications and source records attributed to Kuemmerli, R..

At least 19 recordsLinked to original sources

bioRxiv2026

Low levels of metabolic auxotrophy among environmental Pseudomonas isolates

Auxotrophy, the inability of bacteria to synthesize one or multiple essential metabolites (e.g. amino acids, vitamins, metabolites) is thought to be common among bacteria. However, studies often rely either on bioinformatic tools to predict auxotrophies from genome data or on experiments with low numbers of strains. Here, we combine experimental and bioinformatic approaches to assess amino acid auxotrophy levels among 315 co-isolated natural Pseudomonas strains from pond and soil habitats. Both approaches revealed that Pseudomonas isolates are predominantly prototrophs. We identified one single histidine auxotroph and five non-specific auxotrophs featuring complex growth phenotypes incompatible with single amino acid auxotrophies. While different bioinformatic pipelines vary in the extent to which auxotrophy is over- or underestimated, none of the pipelines could resolve the basis of non-specific auxotrophies. Our analysis further revealed the existence of multiple alternative biosynthesis pathways for methionine, proline, and phenylalanine, with significant enrichments of specific pathways among soil or pond strains. We conclude that combining experiments with bioinformatics is a powerful approach to assess the metabolic potential of environmental bacteria. Moreover, taxa like Pseudomonas can be predominantly prototrophic possibly owing to their generalist lifestyle, thus calling for nuanced ecological concepts predicting auxotrophy levels based on lifestyle and habitat.

microbiology↗

bioRxiv2026

Tracking eco-evolutionary dynamics among lung pathobiome members from children with cystic fibrosis

Polymicrobial lung infections are common in individuals with cystic fibrosis (CF). Pathogen communities typically follow an ecological succession in which early colonizers such as Haemophilus influenzae and Staphylococcus aureus are later joined by Pseudomonas aeruginosa. Although adaptation to the lung environment is well described for single pathogens in adult people with CF, much less is known about the role of pathogen interactions and how changes in individual pathogens influence community dynamics at the early stages of disease. To address these questions, we combined genome sequencing with phenotypic screens and pathogen interaction assays using longitudinal clinical isolates (19 P. aeruginosa, 44 S. aureus, and 21 H. influenzae) collected from 23 children with CF (0-7.9 years of age) enrolled in the SCILD (Swiss CF Infant Lung Development) cohort. Our analysis revealed that early pathogen communities are characterized by a combination of strain turnover and persistence of isolates undergoing first steps of within-host evolution. Most notably, quorum-sensing-deficient P. aeruginosa variants repeatedly emerged, showing reduced protease production and diminished inhibition of S. aureus and H. influenzae. These changes indicate that P. aeruginosa becomes less antagonistic towards co-occurring pathogens, possibly promoting community stability. Together, our results show that ecological and evolutionary dynamics between pathobiome members may play an underappreciated role in shaping CF lung disease during early childhood.

evolutionary biology↗

bioRxiv2026

Siderophore-mediated inhibition of Legionella pneumophila by environmental Pseudomonas isolates

The genus Legionella comprises opportunistic pathogenic bacteria commonly found in natural and engineered water systems, where they interact with environmental microbes and protozoa, primarily in biofilms. Legionella pneumophila is the main causative agent of Legionnaires disease and is transmitted through inhalation of contaminated aerosols. Iron availability is a critical factor for L. pneumophila growth, persistence, and virulence, yet iron is often limited in aquatic environments. To overcome iron scarcity, many bacteria produce siderophores, secondary metabolites that scavenge ferric iron. Because siderophores are chemically diverse and species specific, they play a key role in inter-species competition and can withhold iron from competitors. Here, we investigated the effects of iron depletion and siderophore-mediated competition on L. pneumophila using commercial pyoverdines and extracellular metabolites from environmental Pseudomonas strains. Growth assays showed that L. pneumophila can grow under iron-limited conditions but with lag phases extended by more than 20 hours. Pyoverdines inhibited growth in a concentration-dependent manner, primarily increasing the time to mid-log phase (t_mid). Supernatants and crude pyoverdine extracts from siderophore-producing Pseudomonas strains caused the strongest inhibition, including lag-phase extensions of up to 55 hours or complete growth arrest. These results demonstrate that siderophore-producing bacteria can suppress L. pneumophila by limiting iron availability.

microbiology↗

bioRxiv2025

A combination of phenotypic responses and genetic adaptations enables Staphylococcus aureus to withstand inhibitory molecules secreted by Pseudomonas aeruginosa

Staphylococcus aureus and Pseudomonas aeruginosa frequently co-occur in infections, and there is evidence that their interactions can negatively affect disease outcomes. P. aeruginosa is known to be dominant, often compromising S. aureus through the secretion of inhibitory compounds. We previously demonstrated that S. aureus can become resistant to growth-inhibitory compounds during experimental evolution. While resistance arose rapidly, the underlying mechanisms were not obvious as only a few genetic mutations were associated with resistance, while ample phenotypic changes occurred. We thus hypothesize that resistance may result from a combination of phenotypic responses and genetic adaptation. Here, we tested this hypothesis using proteomics. We first focused on an evolved strain that acquired a single mutation in tcyA (encoding a transmembrane transporter unit) upon exposure to P. aeruginosa supernatant. We show that this mutation leads to a complete abolishment of transporter synthesis, which confers moderate protection against PQS and selenocystine, two toxic compounds produced by P. aeruginosa. However, this genetic effect was minor compared to the fundamental phenotypic changes observed at the proteome level when both ancestral and evolved S. aureus strains were exposed to P. aeruginosa supernatant. Major changes involved the downregulation of virulence factors, metabolic pathways, membrane transporters, and the upregulation of ROS scavengers and an efflux pump. Our results suggest that the observed multi-variate phenotypic response is a powerful adaptive strategy, offering instant protection against competitors in fluctuating environments and reducing the need for hard-wired genetic adaptations. ImportanceDifferent bacterial pathogens can co-occur in infections, where they interact with one another and influence disease severity. Previous research showed that pathogens can evolve and adapt to co-infecting species. Here, we show that evolution through genetic mutations and selection are not necessarily required to change pathogen behavior. Instead, we found that the human pathogen Staphylococcus aureus is able to plastically respond to the presence of Pseudomonas aeruginosa, a competing pathogen. Through proteomics and metabolomics, we demonstrate that S. aureus undergoes substantial proteomic alterations in response to P. aeruginosa by down-regulating virulence factor expression, changing metabolism, and mounting protective measures against toxic compounds. Our work highlights that pathogens possess sophisticated mechanisms to respond to competitors to secure growth and survival in polymicrobial infections. We predict such plastic responses to have significant impacts on infection outcomes.

microbiology↗

bioRxiv2024

Outer membrane changes enable evolutionary escape from bacterial predation

To combat antimicrobial resistant pathogens, natural predatory bacteria, like Bdellovibrio bacteriovorus, represent potential alternatives. B. bacteriovorus could be particularly potent as it kills a broad range of human bacterial pathogens, however, it remains unclear whether prey can evolve genetically-determined resistance against predation. Here, we show that the model bacterium Escherichia coli K-12 consistently evolves resistance against B. bacteriovorus during experimental evolution. Selection for resistance scaled positively with predation pressure and was widespread after two cycles of predator exposure. Like antibiotics, predation resistance was costly, manifesting in a trade-off between predation resistance and fitness in the absence of predators. Genetic analysis identified changes in outer membrane porin OmpF as common resistance mechanism, while a mutation in cell envelope lipopolysaccharide-modifying enzyme WaaF was rarer but also conferred predation resistance. Our study uncovers evolutionary and mechanistic aspects of prey escape from predation, generating important knowledge on predator-prey interactions and to advance sustainable treatments.

evolutionary biology↗

bioRxiv2024

Standardising a microbiome pipeline for body fluid identification from complex crime scene stains

BackgroundRecent advances in next-generation sequencing have opened up new possibilities for utilizing the human microbiome in various fields, including forensics. Researchers have capitalized on the site-specific microbial communities found in different parts of the body to identify body fluids from biological evidence. Despite promising results, microbiome-based methods have not yet been fully integrated into forensic practice due to the lack of standardized protocols and systematic testing of methods on forensically relevant samples. Our study addresses critical decisions in establishing these protocols, focusing on bioinformatics choices and the use of machine learning to present microbiome results in court for forensically relevant and challenging samples. ResultsWe propose using Operational Taxonomic Units (OTUs) for read data processing and creating heterogeneous training datasets for training a random forest classifier. Our classifier incorporates six forensically relevant classes: saliva, semen, hand skin, penile skin, urine, and vaginal/menstrual fluid. Across these classes, our classifier achieved a high weighted average F1 score of 0.89. Systematic testing on mixed-source samples and underwear revealed reliable detection of at least one component of the mixture and the identification of vaginal fluid from underwear substrates. Additionally, when investigating the sexually shared microbiome (sexome) of heterosexual couples, our classifier shows promising results for the inference of sexual activity. ConclusionIn our study, we recommend the use of a novel random forest classifier trained on a heterogenous dataset for obtaining predictions from samples mimicking forensic evidence. We also highlight the potential of the sexome for assessing the nature of sexual activities in forensic investigations, while delineating areas that warrant further research. Furthermore, we underscore key considerations when presenting machine learning results for classifying mixed-source samples.

bioinformatics↗

bioRxiv2024

Pyoverdine-antibiotic combination treatment: its efficacy and effects on resistance evolution in Escherichia coli

Antibiotic resistance is a growing concern for global health, demanding innovative and effective strategies to combat pathogenic bacteria. Pyoverdines, iron-chelating siderophores produced by environmental Pseudomonas spp., present a novel promising approach to induce growth arrest in pathogens through iron starvation. While we have previously demonstrated the efficacy of pyoverdines as antibacterials, our understanding of how these molecules interact with antibiotics and impact resistance evolution remains unknown. Here, we investigate the propensity of different Escherichia coli variants to evolve resistance against pyoverdine, the cephalosporin antibiotic ceftazidime, and their combination. We found that strong resistance against ceftazidime and weak resistance against pyoverdine evolved in the wildtype E. coli strain under single and combination treatment. Ceftazidime resistance was linked to mutations in outer membrane porin genes (envZ and ompF), whereas pyoverdine resistance was associated with mutations in the oligopeptide permease (opp) operon. In contrast, resistance phenotypes were attenuated under combination treatment, particularly in an E. coli strain carrying a costly multicopy plasmid. Altogether, our results show that pyoverdine as an antibacterial is particularly potent and evolutionarily robust against plasmid-carrying E. coli strains, presumably because iron starvation compromises both cellular metabolism and plasmid replication.

microbiology↗

bioRxiv2024

A new protocol for multispecies bacterial infections in zebrafish and their monitoring through automated image analysis

The zebrafish Danio rerio has become a popular model host to explore disease pathology caused by infectious agents. A main advantage is its transparency at an early age, which enables live imaging of infection dynamics. While multispecies infections are common in patients, the zebrafish model is rarely used to study them, although the model would be ideal for investigating pathogen-pathogen and pathogen-host interactions. This may be due to the absence of an established multispecies infection protocol for a defined organ and the lack of suitable image analysis pipelines for automated image processing. To address these issues, we developed a protocol for establishing and tracking single and multispecies bacterial infections in the inner ear structure (otic vesicle) of the zebrafish by imaging. Subsequently, we generated an image analysis pipeline that involved deep learning for the automated segmentation of the otic vesicle, and scripts for quantifying pathogen frequencies through fluorescence intensity measures. We used Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae, three of the difficult-to-treat ESKAPE pathogens, to show that our infection protocol and image analysis pipeline work both for single pathogens and pairwise pathogen combinations. Thus, our protocols provide a comprehensive toolbox for studying single and multispecies infections in real-time in zebrafish.

microbiology↗

bioRxiv2023

From sequence to ecology: siderophore-receptor coevolution algorithm predicts bacterial interactions in complex communities

Predicting bacterial social interactions from genome sequences is notoriously difficult. Here, we developed bioinformatic tools to predict whether secreted iron-scavenging siderophores stimulate or inhibit the growth of community members. Siderophores are chemically diverse and can be stimulatory or inhibitory depending on whether bacteria possess or lack corresponding uptake receptors. We focused on 1928 representative Pseudomonas genomes and developed a co-evolution algorithm to match all encoded siderophore synthetases to corresponding receptor gene groups with >90% accuracy based on experimental validation. We derived community-level iron interaction networks to show that selection for siderophore-mediated interactions differs across habitats and lifestyles. Specifically, dense networks of siderophore sharing and competition were observed among environmental (soil/water/plant) strains and non-pathogenic species, while only fragmented networks occurred among human-derived strains and pathogenic species. Altogether, our sequence-to-ecology approach empowers the analyses of social interactions among thousands of bacterial strains and uncovers ways for targeted intervention to microbial communities. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/565711v3_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@60d907org.highwire.dtl.DTLVardef@48756eorg.highwire.dtl.DTLVardef@115aac1org.highwire.dtl.DTLVardef@17d76e5_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2023

Space and epigenetic inheritance determine inter-individual differences in siderophore gene expression in bacterial colonies

Heterogeneity in gene expression among cells in clonal groups is common in bacteria. Albeit ubiquitous, it often remains unclear what the sources of variation are and whether variation has functional significance. Here, we tracked the expression of genes involved in the synthesis of iron-chelating siderophores (pyoverdine and pyochelin) in individual cells of the bacterium Pseudomonas aeruginosa during colony growth on surfaces using time-lapse fluorescence microscopy, to explore potential sources and functions of cellular heterogeneity. Regarding sources, we found that the physical position of cells within the colony and epigenetic gene expression inheritance from mother to daughter cells significantly contributed to cellular heterogeneity. In contrast, cell pole age and cellular lifespan had no effect. Regarding functions, our results indicate that cells optimize their siderophore investment strategies (pyoverdine vs. pyochelin) along a gradient from the centre to the edge of the colony. Moreover, we found evidence that cell lineages with above-average siderophore investment increase the fitness of cell lineages with below-average investment through cooperative sharing of secreted siderophores. Altogether, our study highlights that single-cell experiments combined with automated image and cell-tracking analyses can identify sources of heterogeneity and yield adaptive explanations for gene expression variation among clonal bacterial cells.

microbiology↗

bioRxiv2023

Antimicrobial activity of iron-depriving pyoverdines against human opportunistic pathogens

The global rise of antibiotic resistance calls for new drugs against bacterial pathogens. A common approach is to search for natural compounds deployed by microbes to inhibit competitors. Here we show that the iron chelating pyoverdines, siderophores produced by environmental Pseudomonas spp., have strong antibacterial properties by inducing iron starvation and growth arrest in pathogens. A screen of 320 natural Pseudomonas isolates used against 12 human pathogens uncovered several pyoverdines with particularly high antibacterial properties and distinct chemical characteristics. The most potent pyoverdine effectively reduced growth of the pathogens Acinetobacter baumannii, Klebsiella pneumoniae and Staphylococcus aureus in a concentration- and iron-dependent manner. Pyoverdine increased survival of infected Galleria mellonella host larvae, and showed low toxicity for the host, mammalian cell lines, and erythrocytes. Furthermore, experimental evolution combined with whole-genome sequencing revealed reduced potentials for resistance evolution compared to an antibiotic. Thus, pyoverdines from environmental strains have the potential to become a new class of sustainable antibacterials against specific human pathogens.

microbiology↗

bioRxiv2023

Predicting bacterial interaction outcomes from monoculture growth and supernatant assays

How to derive principles of community dynamics and stability is a central question in microbial ecology. Bottom-up experiments, in which a small number of bacterial species are mixed, have become popular to address it. However, experimental setups are typically limited because co-culture experiments are labor-intensive and species are difficult to distinguish. Here, we use a 4-species bacterial community to show that information from monoculture growth and inhibitory effects induced by secreted compounds can be combined to predict the competitive rank order in the community. Specifically, integrative monoculture growth parameters allow building a preliminary competitive rank order, which is then adjusted using inhibitory effects from supernatant assays. While our procedure worked for two different media, we observed differences in species rank orders between media. We then parameterized computer simulations with our empirical data to show that higher-order species interactions largely follow the dynamics predicted from pairwise interactions with one important exception. The impact of inhibitory compounds was reduced in higher-order communities because their negative effects were spread across multiple target species. Altogether, we formulated three simple rules of how monoculture growth and supernatant assay data can be combined to establish a competitive species rank order in an experimental 4-species community.

microbiology↗

bioRxiv2021

Rapid and strain-specific resistance evolution of Staphylococcus aureus against inhibitory molecules secreted by Pseudomonas aeruginosa

Pseudomonas aeruginosa and Staphylococcus aureus frequently occur together in polymicrobial infections, and there is evidence that their interactions negatively affect disease outcome in patients. At the molecular level, interactions between the two bacterial species are well-described, with P. aeruginosa usually being the dominant species suppressing S. aureus through a variety of inhibitory molecules. However, in chronic infections the two species interact over prolonged periods of time, and S. aureus might be able to evolve resistance against inhibitory molecules deployed by P. aeruginosa. Here, we used experimental evolution to test this hypothesis by exposing three different S. aureus strains (Cowan I, 6850, JE2) to the growth-inhibitory supernatant of P. aeruginosa PAO1 over 30 days. Prior to evolution, we found that S. aureus strains were inhibited by secreted compounds regulatorily controlled by the PQS quorum-sensing system of P. aeruginosa. Following evolution, S. aureus strains were no longer inhibited: we observed that phenotypic adaptations were strain-specific and involved the up-regulation of virulence traits, such as staphyloxanthin production and the formation of small colony variants. At the genetic level, mutations in membrane transporters were the most frequent evolutionary targets. Our work indicates that adaptations of S. aureus to co-infecting pathogens occurs rapidly and involves both virulence traits and membrane transporters involved in drug resistance. Thus, pathogen evolution could promote species co-existence, complicate treatment options and therefore worsen disease outcome.

microbiology↗

bioRxiv2021

Enforced specialization fosters mutual cheating and not division of labour in the bacterium Pseudomonas aeruginosa

A common way for bacteria to cooperate is via the secretion of beneficial public goods (proteases, siderophores, biosurfactants) that can be shared among individuals in a group. Bacteria often simultaneously deploy multiple public goods with complementary functions. This raises the question whether natural selection could favour division of labour where subpopulations or species specialise in the production of a single public good, whilst sharing the complementary goods at the group level. Here we use an experimental system, where we genetically enforce specialization in the bacterium Pseudomonas aeruginosa with regard to the production of its two siderophores, pyochelin and pyoverdine, and explore the conditions under which specialization can lead to division of labour. When growing pyochelin and pyoverdine specialists at different mixing ratios in various iron limited environments, we found that specialists could only successfully complement each other in environments with moderate iron limitation and grow as good as the generalist wildtype but not better. Under more stringent iron limitation, the dynamics in specialist communities was characterized by mutual cheating and with higher proportions of pyochelin producers greatly compromising group productivity. Nonetheless, specialist communities remained stable through negative frequency-dependent selection. Our work shows that specialization in a bacterial community can be spurred by mutual cheating and does not necessarily result in beneficial division of labour. We propose that natural selection might favour fine-tuned regulatory mechanisms in generalists over division of labour because the former enables generalists to remain flexible and adequately adjust public good investments in fluctuating environments.

evolutionary biology↗

bioRxiv2021

Siderophores drive invasion dynamics in bacterial communities through their dual role as public good versus public bad

Microbial invasions can compromise ecosystem services and spur dysbiosis and disease in hosts. Nevertheless, the mechanisms determining invasion outcomes often remain unclear. Here, we examine the role of iron-scavenging siderophores in driving invasions of Pseudomonas aeruginosa into resident communities of environmental pseudomonads. Siderophores can be "public goods" by delivering iron to individuals possessing matching receptors; but they can also be "public bads" by withholding iron from competitors lacking these receptors. Accordingly, siderophores should either promote or impede invasion, depending on their dual effects on invader and resident growth. Using supernatant feeding and invasion assays, we show that invasion success indeed decreased when the invader was inhibited (public bad) rather than stimulated (public good) by the residents siderophores. Conversely, invasion success often increased when the invader could use its siderophore to inhibit the residents. Our findings identify siderophores as a major driver of invasion dynamics in bacterial communities.

ecology↗

bioRxiv2021

Pseudomonas aeruginosa reaches collective decisions via transient segregation of quorum sensing activities across cells

Bacteria engage in a cell-to-cell communication process called quorum sensing (QS) to coordinate expression of cooperative exoproducts at the group level. While population-level QS-responses are well studied, we know little about commitments of single cells to QS. Here, we use flow cytometry to track the investment of Pseudomonas aeruginosa individuals into their intertwined Las and Rhl QS-systems. Using fluorescent reporters, we show that QS gene expression (signal synthase, receptor and exoproduct) was heterogenous and followed a gradual instead of a sharp temporal induction pattern. The simultaneous monitoring of two QS genes revealed that cells transiently segregate into low receptor (lasR) expressers that fully commit to QS, and high receptor expressers that delay QS commitment. Our mathematical model shows that such gene expression segregation could mechanistically be spurred by transcription factor limitation. In evolutionary terms, temporal segregation could serve as a QS-brake to allow for a bet-hedging strategy in unpredictable environments.

microbiology↗

bioRxiv2021

From heterogeneity to homogeneity: coordination of siderophore gene expression among clonal cells of the bacterium Pseudomonas aeruginosa

There has been great progress in understanding how bacterial groups coordinate social actions, such as biofilm formation, swarming and public-goods secretion. Less clear, however, is whether the seemingly coordinated responses observed at the group level actually mirror what individual cells do. Here, we use a microscopy approach to simultaneously quantify the investment of individual cells of the bacterium Pseudomonas aeruginosa into two public goods, the siderophores pyochelin and pyoverdine. Using gene expression as a proxy for investment, we initially observed no coordination but high heterogeneity and bimodality in siderophore gene expression across cells. With increasing cell density, gene expression became homogenized across cells, accompanied by a shift from pyochelin to pyoverdine expression. We found positive correlations in the expression of pyochelin and pyoverdine genes across cells, and show that cell-to-cell variation is driven by differences in cellular metabolic states. We propose a model explaining how variation in internal iron stocks can spur initial erratic gene expression, while siderophore-mediated signalling and intra-cellular feedbacks later on can induce highly coordinated gene expression and synchronized shifts from pyochelin to pyoverdine. Our work provides new insights into bacterial collective decision-making processes and reveals a three-phase chronobiological siderophore investment cycle in P. aeruginosa.

microbiology↗

bioRxiv2021

Ecology drives the evolution of diverse siderophore-production strategies in the opportunistic human pathogen Pseudomonas aeruginosa

Bacteria often cooperate by secreting molecules that can be shared as public goods between cells. Because the production of public goods is subject to cheating by mutants that exploit the good without contributing to it, there has been great interest in elucidating the evolutionary forces that maintain cooperation. However, little is known on how bacterial cooperation evolves under conditions where cheating is unlikely of importance. Here we use experimental evolution to follow changes in the production of a model public good, the iron-scavenging siderophore pyoverdine, of the bacterium Pseudomonas aeruginosa. After 1200 generations of evolution in nine different environments, we observed that cheaters only reached high frequency in liquid medium with low iron availability. Conversely, when adding iron to reduce the cost of producing pyoverdine, we observed selection for pyoverdine hyper-producers. Similarly, hyper-producers also spread in populations evolved in highly viscous media, where relatedness between interacting individuals is higher. Whole-genome sequencing of evolved clones revealed that hyper-production is associated with mutations/deletions in genes encoding quorum-sensing communication systems, while cheater clones had mutations in the iron-starvation sigma factor or in pyoverdine biosynthesis genes. Our findings demonstrate that bacterial social traits can evolve rapidly in divergent directions, with particularly strong selection for increased levels of cooperation occurring in environments where individual dispersal is reduced, as predicted by social evolution theory. Moreover, we establish a regulatory link between pyoverdine production and quorum-sensing, showing that increased cooperation at one trait (pyoverdine) can be associated with the loss (quorum-sensing) of another social trait.

evolutionary biology↗