Poly-γ-Glutamic Acids Contribute to Biofilm Formation and Plant Root Colonization in Selected Environmental Isolates of Bacillus subtilis

The soil ubiquitous Bacillus subtilis is known to promote plant growth and protect plants against disease. These characteristics make B. subtilis highly relevant in an agricultural perspective, fueling the interest in studying B. subtilis-plant interactions. Here, we employ an experimental evolution approach to explore adaptation of B. subtilis to Arabidopsis thaliana roots. B. subtilis rapidly adapts to the plant root environment, as evidenced by improved root colonizers observed already after 12 consecutive transfers between seedlings in a hydroponic setup. Further phenotypic characterization of evolved isolates from transfer 30 revealed that increased root colonization was associated with robust biofilm formation in response to the plant polysaccharide xylan. Additionally, several evolved isolates across independent populations were impaired in motility, a redundant trait in the selective environment. Interestingly, two evolved isolates outcompeted the ancestor during competition on the root but suffered a fitness disadvantage in non-selective environment, demonstrating an evolutionary cost of adaptation to the plant root. Finally, increased root colonization by a selected evolved isolate was also demonstrated in the presence of resident soil microbes. Our findings provide novel insights into how a well-known PGPR rapidly adapts to an ecologically relevant environment and reveal evolutionary consequences that are fundamental to consider when evolving strains for biocontrol purposes.

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Cyclic di-AMP Acts as an Extracellular Signal That ImpactsBacillus subtilisBiofilm Formation and Plant Attachment

mBio ◽

10.1128/mbio.00341-18 ◽

2018 ◽

Vol 9 (2) ◽

Cited By ~ 26

Author(s):

Loni Townsley ◽

Sarah M. Yannarell ◽

Tuanh Ngoc Huynh ◽

Joshua J. Woodward ◽

Elizabeth A. Shank

Keyword(s):

Gene Expression ◽

Bacillus Subtilis ◽

Intracellular Signaling ◽

Biofilm Formation ◽

Plant Root ◽

Second Messenger ◽

Pathogen Resistance ◽

Plant Roots ◽

Extracellular Signal ◽

The Impact

ABSTRACTThere is a growing appreciation for the impact that bacteria have on higher organisms. Plant roots often harbor beneficial microbes, such as the Gram-positive rhizobacteriumBacillus subtilis, that influence their growth and susceptibility to disease. The ability to form surface-attached microbial communities called biofilms is crucial for the ability ofB. subtilisto adhere to and protect plant roots. In this study, strains harboring deletions of theB. subtilisgenes known to synthesize and degrade the second messenger cyclic di-adenylate monophosphate (c-di-AMP) were examined for their involvement in biofilm formation and plant attachment. We found that intracellular production of c-di-AMP impacts colony biofilm architecture, biofilm gene expression, and plant attachment inB. subtilis. We also show thatB. subtilissecretes c-di-AMP and that putative c-di-AMP transporters impact biofilm formation and plant root colonization. Taken together, our data describe a new role for c-di-AMP as a chemical signal that affects important cellular processes in the environmentally and agriculturally important soil bacteriumB. subtilis. These results suggest that the “intracellular” signaling molecule c-di-AMP may also play a previously unappreciated role in interbacterial cell-cell communication within plant microbiomes.IMPORTANCEPlants harbor bacterial communities on their roots that can significantly impact their growth and pathogen resistance. In most cases, however, the signals that mediate host-microbe and microbe-microbe interactions within these communities are unknown. A detailed understanding of these interaction mechanisms could facilitate the manipulation of these communities for agricultural or environmental purposes.Bacillus subtilisis a plant-growth-promoting bacterium that adheres to roots by forming biofilms. We therefore began by exploring signals that might impact its biofilm formation. We found thatB. subtilissecretes c-di-AMP and that the ability to produce, degrade, or transport cyclic di-adenylate monophosphate (c-di-AMP; a common bacterial second messenger) affectsB. subtilisbiofilm gene expression and plant attachment. To our knowledge, this is the first demonstration of c-di-AMP impacting a mutualist host-microbe association and suggests that c-di-AMP may function as a previously unappreciated extracellular signal able to mediate interactions within plant microbiomes.

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Surfactin production is not essential for pellicle and root-associated biofilm development of Bacillus subtilis

10.1101/865345 ◽

2019 ◽

Author(s):

Maude Thérien ◽

Heiko T. Kiesewalter ◽

Emile Auria ◽

Vincent Charron-Lamoureux ◽

Mario Wibowo ◽

...

Keyword(s):

Bacillus Subtilis ◽

Biofilm Formation ◽

Plant Root ◽

Model Systems ◽

Colony Development ◽

Laboratory Model ◽

Pellicle Formation ◽

Surfactin Production ◽

Biofilm Development ◽

The Impact

AbstractSecondary metabolites have an important impact on the biocontrol potential of soil-derived microbes. In addition, various microbe-produced chemicals have been suggested to impact the development and phenotypic differentiation of bacteria, including biofilms. The non-ribosomal synthesized lipopeptide of Bacillus subtilis, surfactin, has been described to impact the plant promoting capacity of the bacterium. Here, we investigated the impact of surfactin production on biofilm formation of B. subtilis using the laboratory model systems; pellicle formation at the air-medium interface and architecturally complex colony development, in addition to plant root-associated biofilms. We found that the production of surfactin by B. subtilis is not essential for pellicle biofilm formation neither in the well-studied strain, NCIB 3610, nor in the newly isolated environmental strains, but lack of surfactin reduces colony expansion. Further, plant root colonization was comparable both in the presence or absence of surfactin synthesis. Our results suggest that surfactin-related biocontrol and plant promotion in B. subtilis strains are independent of biofilm formation.

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Isolation and characterization of Bacillus subtilis strain 1-L-29, an endophytic bacteria from Camellia oleifera with antimicrobial activity and efficient plant-root colonization

PLoS ONE ◽

10.1371/journal.pone.0232096 ◽

2020 ◽

Vol 15 (4) ◽

pp. e0232096 ◽

Cited By ~ 1

Author(s):

Jin-Xin Xu ◽

Zi-Yang Li ◽

Xing Lv ◽

Hua Yan ◽

Guo-Ying Zhou ◽

...

Keyword(s):

Antimicrobial Activity ◽

Bacillus Subtilis ◽

Endophytic Bacteria ◽

Root Colonization ◽

Plant Root ◽

Subtilis Strain ◽

Camellia Oleifera ◽

Isolation And Characterization

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Diversification of B. subtilis during experimental evolution on A. thaliana leads to synergism in root colonization of evolved subpopulations

10.1101/2021.03.06.434191 ◽

2021 ◽

Author(s):

Christopher Blake ◽

Mathilde Nordgaard Christensen ◽

Gergely Maróti ◽

Ákos T. Kovács

Keyword(s):

Arabidopsis Thaliana ◽

Bacillus Subtilis ◽

Plant Growth ◽

Plant Species ◽

Experimental Evolution ◽

Root Colonization ◽

Plant Root ◽

Plant Roots ◽

Stable Strategy ◽

Pellicle Formation

SummaryThe soil bacterium Bacillus subtilis is known to suppress pathogens as well as promote plant growth. However, in order to fully exploit the potential as natural fertilizer, we need a better understanding of the interactions between B. subtilis and plants. Here, B. subtilis was examined for root colonization through experimental evolution on Arabidopsis thaliana. The populations evolved rapidly, improved in root colonization and diversified into three distinct morphotypes. In order to better understand the adaptation that had taken place, single evolved isolates from the final transfer were randomly selected for further characterization, revealing changes in growth and pellicle formation in medium supplemented with plant polysaccharides. Intriguingly, certain evolved isolates showed improved root colonization only on the plant species they evolved on, but not on another plant species, namely tomato, suggesting A. thaliana specific adaption paths. Finally, synergism in plant root colonization was observed for a mix of all three morphotypes, as the mix performed better than the sum of its constituents in monoculture. Our results suggest, that genetic diversification occurs in an ecological relevant setting on plant roots and proves to be a stable strategy for root colonization.Significance StatementUnderstanding how plant-growth-promoting rhizobacteria colonize plant roots is crucial to fully utilize their potential for agricultural applications. Here, we utilized experimental evolution of the PGPR Bacillus subtilis on Arabidopsis thaliana to study root colonization. We revealed that evolving populations rapidly improve in root colonization and diversify into distinct morphotypes. Notably, improved root colonization by evolved isolates was observed on A. thaliana, not on tomato. Moreover, isolates of distinct morphotypes interacted during root colonization and the mixture of morphotypes showed higher productivity then predicted. These findings suggest that genetic diversification might be a stable strategy to maximize root colonization.

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Biofilm hydrophobicity in environmental isolates of Bacillus subtilis

10.1101/2021.04.29.441976 ◽

2021 ◽

Author(s):

Margarita Kalamara ◽

James C. Abbott ◽

Cait E. MacPhee ◽

Nicola. R. Stanley-Wall

Keyword(s):

Extracellular Matrix ◽

Bacillus Subtilis ◽

Glutamic Acid ◽

Biofilm Formation ◽

Environmental Isolates ◽

Diverse Range ◽

European Nucleotide Archive ◽

Universal Feature ◽

The Matrix ◽

Highly Hydrophobic

AbstractBiofilms are communities of bacteria that are attached to a surface and surrounded by an extracellular matrix. The extracellular matrix protects the community from stressors in the environment, making biofilms robust. The Gram-positive soil bacterium Bacillus subtilis, particularly the isolate NCIB 3610, is widely used as a model for studying biofilm formation. B. subtilis NCIB 3610 forms colony biofilms that are architecturally complex and highly hydrophobic. The hydrophobicity is linked, in part, to the localisation of the protein BslA at the surface of the biofilm, which provides the community with increased resistance to biocides. As most of our knowledge about B. subtilis biofilm formation comes from one isolate, it is unclear if biofilm hydrophobicity is a widely distributed feature of the species. To address this knowledge gap, we collated a library of B. subtilis soil isolates and acquired their whole genome sequences. We used our new isolates to examine biofilm hydrophobicity and found that, although BslA is encoded and produced by all isolates in our collection, hydrophobicity is not a universal feature of B. subtilis colony biofilms. To test whether the matrix exopolymer poly γ-glutamic acid could be masking hydrophobicity in our hydrophilic isolates, we constructed deletion mutants and found, contrary to our hypothesis, that the presence of poly γ-glutamic acid was not the reason behind the observed hydrophilicity. This study highlights the natural variation in the properties of biofilms formed by different isolates and the importance of using a more diverse range of isolates as representatives of a species.RepositoriesRaw sequence reads and annotated assemblies have been submitted to the European Nucleotide Archive under accession PRJEB43128.

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Antagonist Bacillus subtilis HJ5 controls Verticillium wilt of cotton by root colonization and biofilm formation

Biology and Fertility of Soils ◽

10.1007/s00374-012-0718-x ◽

2012 ◽

Vol 49 (3) ◽

pp. 295-303 ◽

Cited By ~ 48

Author(s):

Shuqing Li ◽

Nan Zhang ◽

Zhenhua Zhang ◽

Jia Luo ◽

Biao Shen ◽

...

Keyword(s):

Bacillus Subtilis ◽

Verticillium Wilt ◽

Biofilm Formation ◽

Root Colonization

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Faculty Opinions recommendation of LapF, the second largest Pseudomonas putida protein, contributes to plant root colonization and determines biofilm architecture.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.3744960.3848054 ◽

2010 ◽

Author(s):

Ben Lugtenberg

Keyword(s):

Pseudomonas Putida ◽

Root Colonization ◽

Plant Root ◽

Biofilm Architecture

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Physiology of guanosine-based second messenger signaling in Bacillus subtilis

Biological Chemistry ◽

10.1515/hsz-2020-0241 ◽

2020 ◽

Vol 401 (12) ◽

pp. 1307-1322

Author(s):

Gert Bange ◽

Patricia Bedrunka

Keyword(s):

Bacillus Subtilis ◽

Biofilm Formation ◽

Second Messengers ◽

Model Organism ◽

Physiological Regulation ◽

Open Questions ◽

Key Players ◽

Stress Signals ◽

Second Messenger Signaling ◽

Synthesis And Degradation

AbstractThe guanosine-based second messengers (p)ppGpp and c-di-GMP are key players of the physiological regulation of the Gram-positive model organism Bacillus subtilis. Their regulatory spectrum ranges from key metabolic processes over motility to biofilm formation. Here we review our mechanistic knowledge on their synthesis and degradation in response to environmental and stress signals as well as what is known on their cellular effectors and targets. Moreover, we discuss open questions and our gaps in knowledge on these two important second messengers.

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