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Transcriptome profiling of Variovorax paradoxus EPS under different growth conditions reveals regulatory and structural novelty in biofilm formation.

ABSTRACT: We used transcriptome analysis by paired-end strand-specific RNA-seq to evaluate the specific changes in gene expression associated with the transition to static biofilm growth in the rhizosphere plant growth-promoting bacterium Variovorax paradoxus EPS. Triplicate biological samples of exponential growth, stationary phase and static biofilm samples were examined. DESeq2 and Rockhopper were used to identify robust and widespread shifts in gene expression specific to each growth phase. We identified 1711 protein-coding genes (28%) using DESeq2 that had altered expression greater than twofold specifically in biofilms compared to exponential growth. Fewer genes were specifically differentially expressed in stationary-phase culture (757, 12%). A small set of genes (103/6020) were differentially expressed in opposing fashions in biofilm and stationary phase, indicating potentially substantial shifts in phenotype. Gene-ontology analysis showed that the only class of genes specifically upregulated in biofilms was associated with nutrient transport, highlighting the importance of nutrient uptake in the biofilm. The biofilm-specific genes did not overlap substantially with the loci identified by mutagenesis studies, although some were present in both sets. The most highly upregulated biofilm-specific gene is predicted to be a part of the RNA degradosome, which indicates that RNA stability is used to regulate the biofilm phenotype. Two small putative proteins, Varpa_0407 and Varpa_3832, are highly expressed specifically in biofilms and are predicted to be secreted DNA-binding proteins, which may stabilize extracellular DNA as a component of the biofilm matrix. An flp/tad type-IV pilus locus (Varpa_5148-60) is strongly downregulated specifically in biofilms, in contrast with results from other systems for these pili. Mutagenesis confirms that this locus is important in surface motility rather than biofilm formation. These experimental results suggest that V. paradoxus EPS biofilms have substantial regulatory and structural novelty.

SUBMITTER: Fredendall RJ

PROVIDER: S-EPMC7494200 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Transcriptome profiling of <i>Variovorax paradoxus</i> EPS under different growth conditions reveals regulatory and structural novelty in biofilm formation.

Fredendall Richard J RJ Stone Jenny L JL Pehl Michael J MJ Orwin Paul M PM

Access microbiology 20200403 6

We used transcriptome analysis by paired-end strand-specific RNA-seq to evaluate the specific changes in gene expression associated with the transition to static biofilm growth in the rhizosphere plant growth-promoting bacterium <i>Variovorax paradoxus</i> EPS. Triplicate biological samples of exponential growth, stationary phase and static biofilm samples were examined. DESeq2 and Rockhopper were used to identify robust and widespread shifts in gene expression specific to each growth phase. We ...[more]

PMID: 32974586

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Project description:BackgroundVariovorax paradoxus is an aerobic soil bacterium frequently associated with important biodegradative processes in nature. Our group has cultivated a mucoid strain of Variovorax paradoxus for study as a model of bacterial development and response to environmental conditions. Colonies of this organism vary widely in appearance depending on agar plate type.ResultsSurface motility was observed on minimal defined agar plates with 0.5% agarose, similar in nature to swarming motility identified in Pseudomonas aeruginosa PAO1. We examined this motility under several culture conditions, including inhibition of flagellar motility using Congo Red. We demonstrated that the presence of a wetting agent, mineral, and nutrient content of the media altered the swarming phenotype. We also demonstrated that the wetting agent reduces the surface tension of the agar. We were able to directly observe the presence of the wetting agent in the presence and absence of Congo Red, and found that incubation in a humidified chamber inhibited the production of wetting agent, and also slowed the progression of the swarming colony. We observed that swarming was related to both carbon and nitrogen sources, as well as mineral salts base. The phosphate concentration of the mineral base was critical for growth and swarming on glucose, but not succinate. Swarming on other carbon sources was generally only observed using M9 salts mineral base. Rapid swarming was observed on malic acid, d-sorbitol, casamino acids, and succinate. Swarming at a lower but still detectable rate was observed on glucose and sucrose, with weak swarming on maltose. Nitrogen source tests using succinate as carbon source demonstrated two distinct forms of swarming, with very different macroscopic swarm characteristics. Rapid swarming was observed when ammonium ion was provided as nitrogen source, as well as when histidine, tryptophan, or glycine was provided. Slower swarming was observed with methionine, arginine, or tyrosine. Large effects of mineral content on swarming were seen with tyrosine and methionine as nitrogen sources. Biofilms form readily under various culture circumstances, and show wide variance in structure under different conditions. The amount of biofilm as measured by crystal violet retention was dependent on carbon source, but not nitrogen source. Filamentous growth in the biofilm depends on shear stress, and is enhanced by continuous input of nutrients in chemostat culture.ConclusionOur studies have established that the beta-proteobacterium Variovorax paradoxus displays a number of distinct physiologies when grown on surfaces, indicative of a complex response to several growth parameters. We have identified a number of factors that drive sessile and motile surface phenotypes. This work forms a basis for future studies using this genetically tractable soil bacterium to study the regulation of microbial development on surfaces.

Dataset Information

Transcriptome profiling of Variovorax paradoxus EPS under different growth conditions reveals regulatory and structural novelty in biofilm formation.

Publications

Transcriptome profiling of <i>Variovorax paradoxus</i> EPS under different growth conditions reveals regulatory and structural novelty in biofilm formation.

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