Pseudomonas fluorescens 2P24 is a rhizosphere bacterium that protects many crop plants against soil-borne diseases caused by phytopathogens. The PcoI/PcoR quorum-sensing (QS) system and polyketide antibiotic 2,4-diacetylphloroglucinol (2,4-DAPG) are particularly relevant to the strain9s biocontrol potential. In this study, we investigated the effects of the c-di-GMP on the biocontrol activity of strain 2P24. The expression of the Escherichia coli diguanylate cyclase (YedQ) and phosphodiesterase (YhjH) in P. fluorescens 2P24 significantly increased and decreased the cellular concentration of c-di-GMP, respectively. The production of the QS signal N-acyl homoserine lactones (AHLs) and 2,4-DAPG were negatively regulated by c-di-GMP in 2P24. The regulatory proteins RsmA and RsmE were positively regulated by c-di-GMP. Genomic analysis revealed that 2P24 has 23 predicted proteins that contain c-di-GMP synthesizing or degrading domains. Among these proteins, C0J56_12915, C0J56_13325, and C0J56_27925 contributed to the production of c-di-GMP, and were also involved in the regulation of the QS signal and antibiotic 2,4-DAPG production in P. fluorescens. Overexpression of C0J56_12915, C0J56_13325, and C0J56_27925 in 2P24 impaired its root colonization and biocontrol activities. Taken together, these results demonstrated that c-di-GMP played an important role in fine-tuning of the biocontrol traits of P. fluorescens. Importance In various bacteria, the bacterial second messenger c-di-GMP influences a wide range of cellular processes. However, the function of c-di-GMP on biocontrol traits in the plant-beneficial rhizobacteria remains largely unclear. The present work shows that the QS system and polyketide antibiotic 2,4-DAPG production are regulated by c-di-GMP through RsmA and RsmE proteins in P. fluorescens 2P24. The diguanylate cyclases (DGCs) C0J56_12915, C0J56_13325, and C0J56_27925 are specially involved in regulating the biocontrol traits of 2P24. Our work also demonstrated a connection between the Gac/Rsm cascade and the c-di-GMP signaling pathway in P. fluorescens.

Background: Pseudomonas fluorescens 2P24 is a rhizosphere bacterium that produces 2,4-diacetyphloroglucinol (2,4-DAPG) as the decisive secondary metabolite to suppress soilborne plant diseases. The biosynthesis of 2,4-DAPG is strictly regulated by the RsmA family proteins RsmA and RsmE. However, mutation of both of rsmA and rsmE genes results in reduced bacterial growth. Results: In this study, we showed that overproduction of 2,4-DAPG in the rsmA rsmE double mutant influenced the growth of strain 2P24. This delay of growth could be partially reversal when the phlD gene was deleted or overexpression of the phlG gene encoding the 2,4-DAPG hydrolase in the rsmA rsmE double mutant. RNA-seq analysis of the rsmA rsmE double mutant revealed that a substantial portion of the P. fluorescens genome was regulated by RsmA family proteins. These genes are involved in the regulation of 2,4-DAPG production, cell motility, carbon metabolism, and type six secretion system. Conclusions: These results suggest that RsmA and RsmE are the important regulators of genes involved in the plant-associated strain 2P24 ecologic fitness and operate a sophisticated mechanism for fine-tuning the concentration of 2,4-DAPG in the cells.

The PcoI/PcoR quorum-sensing system of Pseudomonas fluorescens 2P24 is important for its effective colonization in the plant rhizosphere. Many regulatory elements appear to directly or indirectly influence the QS system.

A Gram-stain-negative aerobic bacterium, strain 11K1^T, was isolated from a rhizosphere soil of broad bean collected from Qujing, Yunnan, PR China and characterized by using polyphasic taxonomy. The bacterial cells of strain 11K1^Twere rod-shaped, motile by two polar flagella and positive for oxidase and catalase. Results of phylogenetic analysis based on 16S rRNA gene sequences revealed that the strain had the highest similarities toPseudomonas thivervalensisDSM 13194^T(99.52 %),Pseudomonas liniCFBP 5737^T(99.45 %),Pseudomonas chlororaphissubsp.chlororaphis NBRC 3904^T(99.31 %),Pseudomonas kilonensisDSM 13647^T(99.25 %) andPseudomonas brassicacearumJCM11938^T(99.24 %). Multilocus sequence analysis using the 16S rRNA,gyrB,rpoBandrpoDgene sequences demonstrated that strain 11K1^Twas a member of thePseudomonas corrugatasubgroup within thePseudomonas fluorescenslineage, but was distant from all closely related species. The average nucleotide identity andin silicoDNA–DNA hybridization values were lower than recommended thresholds of 95 and 70 %, respectively, for species delineation. The major isoprenoid quinone of strain 11K1^Twas ubiquinone (Q-9) and the major cellular fatty acids were C_16 : 0, summed feature 3 (C_16 : 1 ω7c/C_16 : 1 ω6c), summed feature 8 (C_18 : 1 ω7c/C_18 : 1 ω6c) and C_17 : 0cyclo. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, aminophospholipid and two unidentified lipids. Based on the results of phenotypic characterization, phylogenetic analysis and genome comparison, strain 11K1^Trepresents a novel species of the genusPseudomonas, for which the namePseudomonas viciaesp. nov. is proposed. The type strain is 11K1^T(=GDMCC 1.1743^T=KACC 21650^T).

The GacS/GacA two-component system is essential for virulence in many plant pathogenic bacteria, and thus represents a promising anti-virulence target. In the present study, we isolated and screened rhizobacteria that were capable of inhibiting the expression of the gacS gene in the phytopathogenic bacterium Pseudomonas syringae pv. tomato (Pto) DC3000. One candidate inhibitor bacterium, BR3 was obtained and identified as a Bacillus sp. strain based on 16s rRNA gene sequence analysis. Besides the gacS gene, the GacA-dependent small RNA genes rsmZ and rsmY were repressed transcriptionally when DC3000 was treated with an extract from strain BR3. Importantly, the extract also influenced bacterial motility, the expression of type three secretion system effector AvrPto, and the plant hypersensitive response triggered by strain DC3000. The results suggested that the extract from strain BR3 might offer an alternative method to control bacterial diseases in plants by targeting the GacS/GacA system.