Biology subjects

Keel, C.

Publications and source records attributed to Keel, C..

11 recordsLinked to original sources

bioRxiv2026

The insect- and plant-associated lifestyles of Pseudomonas protegens CHA0 are preserved following serial passage through insect larvae

The plant-beneficial bacterium Pseudomonas protegens CHA0 (CHA0) is widely studied for the biological control of soil-borne plant diseases. Beyond its root-colonising capabilities, CHA0 can also infect and kill insect larvae and thus exhibits a multi-host lifestyle shared with other plant- and insect-colonising bacteria. To better understand the robustness of this multi-host lifestyle, we subjected CHA0 to ten consecutive passages through larvae of the pest insect Plutella xylostella via repeated cycles of insect colonisation and killing forcing it into an insect-only lifestyle. Overall, serial passaging did not result in consistent changes in insect killing speed, larval or root colonisation, plant protection efficiency, microbial antagonism or in vitro growth. This suggests that its multi-host lifestyle was conserved following serial passage. Nonetheless, a few independently passaged lines showed an increase in larval killing speed, which in one case might be linked to choline uptake. To disentangle changes specific to the insect host from those arising due to the experimental system itself, we conducted parallel serial passages through the same system while omitting the insect host. In some of these lines, exposure to the background of the system led to changes in microbial antagonism and in in vitro growth, which likely are associated with mutations in regions encoding for regulatory systems. Our findings indicate that P. protegens CHA0 remains phenotypically stable in complex environments such as an insect host, suggesting that the multi-host lifestyle might also be conserved when applied in the field and supporting CHA0s potential for reliable biocontrol performance against both plant diseases and insect pests. Author summaryControlling insect pests with living organisms, known as biological control, offers an environmentally friendly alternative to chemical pesticides. The plant-beneficial bacterium Pseudomonas protegens CHA0 is a promising biocontrol candidate that not only colonizes plant roots but also infects and kills certain insect larvae. This ability to colonize different hosts appears to be a conserved trait also observed in other bacteria. To better understand the robustness of this multi-host lifestyle, we repeatedly exposed CHA0 to larvae of the insect pest Plutella xylostella and assessed the resulting physiological and genetic changes. Surprisingly, after ten cycles, CHA0 largely retained its insect-killing and plant-protective traits. Although a few populations showed minor changes, including slightly faster insect killing and traits associated with aspects of the experimental system, these changes were limited in scope. Overall, our findings suggest that P. protegens CHA0 does not change rapidly in complex environments such as an insect host, supporting its potential for reliable biocontrol performance in the field.

microbiology↗

bioRxiv2025

Exploring novel inducers of phage tail-like particle expression using a fluorescent reporter bacterium.

R-tailocins are phage tail-like particles produced by diverse bacterial species, playing crucial roles in interbacterial competition and microbial community dynamics. While their production is typically triggered by DNA damage through the bacterial SOS response, the full range of potential inducers remains largely unexplored. This study aimed at identifying novel chemical compounds capable of triggering R-tailocin gene expression. A fluorescent reporter bacterial strain was employed to monitor R-tailocin gene cluster activity, and over 1,500 compounds were screened using Biolog Phenotype MicroArray plates. This screen identified several previously unreported inducers of R-tailocin expression. Among the strongest inducers identified were pipemidic acid and hydroxyurea, although their induction levels did not exceed those observed with the commonly used inducer mitomycin C. The identified compounds exhibited distinct temporal induction patterns, suggesting variation in their underlying regulatory mechanisms. Several also displayed clear dose-response patterns, providing further insights into their potency and potential modes of action. Notably, R-tailocin induction did not necessarily lead to complete population lysis, as a subset of cells consistently survived the induction process. Collectively, these findings expand the current understanding of R-tailocin induction and production in bacterial populations, uncovering novel chemical triggers and revealing diverse expression dynamics. These insights offer potential implications for microbial community ecology and the development of targeted antimicrobial strategies.

microbiology↗

bioRxiv2025

A LexA-like repressor and global H-NS-like regulators enable the fine-tuning of R-tailocin expression in environmental Pseudomonas

Bacteria rely on an arsenal of weapons to challenge their opponents in highly competitive environments. To specifically counter closely related bacteria, specialized weapons with a narrow activity spectrum are deployed, particularly contractile phage tail-like particles or R-tailocins. Their production leads to the lysis of the producing cells, indicating that their expression must be carefully orchestrated so that only a small percentage of cells produce R-tailocins for the benefit of the entire population. In this study, we set out to better understand how the production of these phage tail-like weapons is regulated in environmental pseudomonads using the competitive plant root colonizer and environmental model Pseudomonas protegens CHA0. Using an RNA sequencing (RNA-seq) approach, we found that genes involved in DNA repair, particularly the SOS response program, are upregulated following exposure of the pseudomonad to the DNA-damaging agents mitomycin C and hydrogen peroxide, while genes involved in cell division and primary metabolism are downregulated. The R-tailocin and prophage gene clusters were also upregulated in response to these DNA damaging agents. By combining reverse genetics, transcriptional reporters and chromatin immunoprecipitation sequencing (ChIP-seq), we show that the R-tailocin locus-specific LexA-like regulator PrtR1 represses R-tailocin gene expression by binding directly to the promoter region of the cluster, while the histone-like nucleoid structuring (H-NS) proteins MvaT and MvaV act as master regulators that indirectly regulate R-tailocin cluster expression. Our results suggest that at least these three regulators operate in concert to ensure tight control of R-tailocin expression and cell lytic release in environmental Pseudomonas strains. Author SummaryTo face their opponents in highly competitive environments, bacteria rely on an arsenal of weapons. To specifically target closely related bacterial strains, specialized weapons with a narrow activity spectrum are produced, notably contractile phage tail-like particles such as R-tailocins. Since production of these particles leads to the lysis and death of the producing cell, their expression must be tightly regulated. Here, we set out to better understand how the expression of these phage tail-like weapons is regulated in environmental bacteria using the competitive plant root colonizer and environmental model Pseudomonas protegens CHA0. We found that following DNA-damaging stress, which activates DNA repair systems and downregulates primary metabolism, the expression of gene clusters encoding viral-like particles such as the R-tailocins is upregulated. We identified cluster-specific and global regulators that are either directly or indirectly involved in the control of R-tailocin cluster expression. Our results contribute to a better understanding of the regulatory mechanisms of viral-like particle production in environmental Pseudomonas bacteria.

microbiology↗

bioRxiv2024

Single nucleotide switches confer bacteriophage resistance to Pseudomonas protegens

Phage therapy offers a promising strategy against bacterial pathogens in medicine and agriculture, but the rise of phage-resistant bacteria presents a significant challenge to its sustainability. Here, we used an environmental model bacterium, Pseudomonas protegens CHA0, to investigate phage resistance mechanisms in laboratory conditions through genomic analysis of four phage-resistant variants (C2, C4, C17, C18). Whole-genome sequencing revealed frequent deletions, insertions, and single nucleotide substitutions, particularly in genes encoding enzymes involved in cell surface modifications. The T428P mutation in AlgC, a phosphoglucomutase, and the P229T substitution in YkcC, a glycosyltransferase, each conferred resistance by potentially altering phage receptor accessibility while preserving bacterial fitness. These findings suggest that subtle mutations in surface-modifying enzymes enable phage resistance by modifying phage-receptor accessibility while maintaining bacterial growth, highlighting adaptive mechanisms in bacterial phage interactions. Further structural analysis is needed to fully understand these mutation-driven resistance mechanisms.

microbiology↗

bioRxiv2024

Priority effects of heritable seed-borne bacteria drive early assembly of the wheat rhizosphere microbiome

Microbial communities play a critical role in supporting plant health and productivity, making the ability to obtain reproducible plant-associated microbiomes an essential asset for experimentally testing hypotheses related to microbiome manipulation and fundamental principles governing community dynamics. We used a sequential propagation strategy to generate a complex and reproducible wheat rhizosphere microbiome (RhizCom) that was shaped by host selection and periodic habitat resetting. Heritable seed-borne rhizosphere bacteria (SbRB) emerged as the dominant microbiome source after coalescing with the soil community, driven by priority effects and efficient niche exploitation during early habitat development. Functional analyses revealed that niche partitioning through the ability of SbRB to degrade specific saccharides and niche facilitation contributed to the assembly of the RhizCom. Our results advance our understanding of the principles governing microbial community dynamics in early plant development and provide strategies for future microbiome manipulation aimed at improving crop productivity and health.

microbiology↗

bioRxiv2024

Natural soil suppressiveness against soilborne phytopathogens extends to the control of insect pest

Since the 1980s, soils in a 22-km2 area near Lake Neuchatel in Switzerland have been recognized for their innate ability to suppress the black root rot plant disease. Their efficacy against insect pests has not been studied. We demonstrate that natural soil suppressiveness also protects plants from the leaf-feeding pest insect Oulema melanopus. Plants grown in the most suppressive soil have a reduced stress response to Oulema feeding, reflected by dampened levels of herbivore defense-related phytohormones and benzoxazinoids, and enhanced salicylate levels in plants without the insect indicate defense-priming. The rhizosphere microbiome network of the suppressive soils was highly tolerant to the destabilizing impact of insect exposure. The presence of plant-beneficial bacteria in the suppressive soils along with priming conferred plant resistance to the insect pest, manifesting also in the onset of insect microbiome dysbiosis. This intricate soil-plant-insect feedback extends natural soil suppressiveness from soilborne diseases to insect pests.

plant biology↗

bioRxiv2024

Fragmented micro-growth habitats present opportunities for alternative competitive outcomes

Bacteria in nature often proliferate in highly patchy environments, such as soil pores, particles, plant roots or leaves. The resulting spatial fragmentation leads to cells being constrained to smaller habitats, shared with potentially fewer other species. The effects of microhabitats on the emergence of bacterial interspecific interactions are poorly understood, but potentially important for the maintenance of diversity at a larger scale. To study this more in-depth, we contrasted paired species-growth in picoliter droplets at low population census with that in large (macro) population liquid suspended cultures. Four interaction scenarios were imposed by using different bacterial strain combinations and media: substrate competition, substrate independence, growth inhibition, and cell killing by tailocins. In contrast to macro-level culturing, we observed that fragmented growth in picoliter droplets in all cases yielded more variable outcomes, and even reversing the macro-level assumed interaction type in a small proportion of droplet habitats. Timelapse imaging and mathematical simulations indicated that the variable and alternative interaction outcomes are a consequence of founder cell phenotypic variation and small founder population sizes. Simulations further suggested that increased growth kinetic variation may be a crucial selectable property for slower-growing bacterial species to survive competition. Our results thus demonstrate how microhabitat fragmentation enables the proliferation of alternative interaction trajectories and contributes to the maintenance of higher species diversity under substrate competition.

microbiology↗

bioRxiv2023

Molecular and evolutionary basis of O-antigenic polysaccharide driven phage sensitivity in environmental pseudomonads

Pseudomonas protegens CHA0, a bacterial strain able to suppress plant pathogens as well as efficiently kill lepidopteran pest insects, has been studied as biocontrol agent to prevent ensuing agricultural damage. However, the success of this method is dependent on the efficient plant colonization by the bacterial inoculant while it faces competition from the resident microbiota as well as predators such as bacteriophages. One of these naturally occurring phages, {Phi}GP100, was found to drastically reduce the abundance of CHA0 once inoculated into plant microcosms, resulting in the loss of plant protection against a phytopathogen. Here, we investigated the molecular determinants implicated in the interaction between CHA0 and the phage {Phi}GP100 using a high-density transposon-sequencing approach. We show that lipopolysaccharide cell surface decorations, specifically the longer OBC3-type O-antigenic polysaccharide (O-PS, O-antigen) of the two dominant O-PS of CHA0 is essential for the attachment and infection of {Phi}GP100. Moreover, when exploring the distribution of the OBC3 cluster in bacterial genomes, we identified several parts of this gene cluster that are conserved in phylogenetically distant bacteria. Through heterologous complementation, we integrated an OBC3-type gene copy from a phylogenetically distant bacterium and were able to restore the phage sensitivity of a CHA0 mutant which lacked the ability to form long O-PS. Finally, we evidence that the OBC3 gene cluster of CHA0 displays a high genomic plasticity and likely underwent several horizontal acquisitions and genomic rearrangements. Collectively, this study underlines the complexity of phage-bacteria interaction and the multifunctional aspect of bacterial cell surface decorations. ImportanceThe application of plant-beneficial microorganisms to protect crop plants is a promising alternative to the usage of chemicals. However, biocontrol research often faces difficulties to implement this approach due to the inconsistency of the bacterial inoculant to establish itself within the root microbiome. Beneficial bacterial inoculants can be decimated by the presence of their natural predators, notably bacteriophages (also-called phages). Thus, it is important to gain knowledge regarding the mechanisms behind phage-bacteria interaction to overcome this challenge. Here, we evidence that the major long O-antigenic polysaccharide (O-PS, O-antigen) of the widely used model plant-beneficial bacterium Pseudomonas protegens CHA0 is the receptor of its natural predator, phage {Phi}GP100. We examined the distribution of the gene cluster directing the synthesis of this O-PS and identified signatures of horizontal gene acquisitions. Altogether, our study highlights the importance of bacterial cell surface structure variation in the complex interplay between phages and their Pseudomonas hosts.

microbiology↗

bioRxiv2023

Identification of the conserved iol gene cluster involved in rhizosphere competence in Pseudomonas

The Pseudomonas genus has shown great potential as a sustainable solution to support agriculture through its plant-growth promoting and biocontrol activities. However, their efficacy as bioinoculants is limited by unpredictable colonization in natural conditions. Our study identifies the iol locus, a gene cluster in Pseudomonas involved in inositol catabolism, as a feature enriched among superior root colonizers in natural soil. Further characterization revealed that the iol locus increases competitiveness by inducing swimming motility and fluorescent siderophore production in response to inositol, a plant-derived compound. Public data analyses indicate that the iol locus is broadly conserved in the Pseudomonas genus and linked to diverse host-microbe interactions. Our findings suggest the iol locus as a potential target for developing more effective bioinoculants, given its conservation and association with diverse host-microbe interactions.

microbiology↗

bioRxiv2021

Much more than a toxin: hydrogen cyanide is a volatile modulator of bacterial behaviour

Bacterial volatile compounds have emerged as important chemical messengers between bacteria themselves as well as in their interactions with other organisms. One of the earliest examples of bioactive volatiles emitted by bacteria is hydrogen cyanide (HCN), which was long considered a mere respiratory toxin conferring competitive advantage to cyanogenic strains. Using cyanide-deficient mutants in two Pseudomonas strains and global transcriptome analysis, we demonstrate that the impact of HCN is much more global than previously thought. We first observed that the lack of cyanogenesis in emitting strains led to massive transcriptome reprogramming affecting diverse traits such as motility and biofilm formation (respectively inhibited vs. promoted by HCN), or the production of siderophores, phenazines and other antimicrobial compounds (repressed by HCN). We then exposed non-cyanogenic strains to biogenically emitted HCN from neighboring cells and observed similar transcriptome modulations and phenotypic changes, suggesting that HCN not only acts endogenously but also exogenously, remotely manipulating important traits involved in competition and virulence, e.g. siderophore production, in other organisms. Cyanogenesis in Pseudomonas has long been known to play a role in both the virulence of opportunistic pathogens and the efficient biocontrol activity of plant-beneficial strains, however this impact was so far thought to occur solely through the inhibition of respiration. We demonstrate here new ecological roles for a small and fast-diffusing volatile compound, which opens novel avenues in our understanding of and ability to interfere with important processes taking place in pathogenic and beneficial Pseudomonas strains. ImportanceBacteria communicate by exchanging chemical signals, some of which are volatile and can remotely reach other organisms. Hydrogen cyanide (HCN) was one of the first volatiles discovered to severely impact exposed organisms by inhibiting their respiration. Using HCN-deficient mutants in two Pseudomonas strains, we demonstrate that HCNs impact goes beyond the sole inhibition of respiration and affects both emitting and receiving bacteria in a global way, modulating their motility, biofilm formation and production of antimicrobial compounds. Our data suggest that bacteria could use HCN not only to control their own cellular functions, but also to remotely influence the behavior of other bacteria sharing the same environment. Since HCN emission occurs in both clinically and environmentally relevant Pseudomonas, these findings are important to better understand or even modulate the expression of bacterial traits involved in both virulence of opportunistic pathogens and in biocontrol efficacy of plant-beneficial strains.

microbiology↗

bioRxiv2020

Spatially Restricted Immune Responses Allow for Root Meristematic Activity During Bacterial Colonisation

Plants circumscribe microbe-associated molecular pattern (MAMP)-triggered immune responses to weak points of the roots. This spatially restricted immunity was suggested to avoid constitutive responses to rhizosphere microbiota. To demonstrate its relevance, we combined cell-type specific expression of the plant flagellin receptor (FLS2) with fluorescent defence markers and mapped immune competency at cellular resolution. Our analysis distinguishes cell-autonomous and non-cell autonomous responses and reveals lignification to be tissue-independent, contrasting cell-type specific suberisation. Importantly, our analysis divides the non-responsive meristem into a central zone refractory to FLS2 expression, and a cortex that becomes highly sensitised by FLS2 expression, causing meristem collapse upon MAMP exposure. Meristematic epidermal expression generates super-competent lines that detect native bacterial flagellin and bypass the absence of response to commensals, providing a powerful tool for studying root immunity. Our precise manipulations and read-outs demonstrate incompatibility of meristematic activity and defence and the importance of cell-resolved studies of plant immunity.

plant biology↗