Development of a Pseudomonas-based biocontrol consortium with effective root colonization and extended beneficial side effects for plants under high-temperature stress.

The root microbiota plays a crucial role in plant performance. The use of microbial consortia is considered a very useful tool for studying microbial interactions in the rhizosphere of different agricultural crop plants. Thus, a consortium of 3 compatible beneficial rhizospheric Pseudomonas strains previously isolated from the avocado rhizosphere, was constructed. The consortium is composed of two compatible biocontrol P. chlororaphis strains (PCL1601 and PCL1606), and the biocontrol rhizobacterium Pseudomonas alcaligenes AVO110, which are all efficient root colonizers of avocado and tomato plants. These three strains were compatible with each other and reached stable levels both in liquid media and on plant roots. Bacterial strains were fluorescent tagged, and colonization-related traits were analyzed in vitro, revealing formation of mixed biofilm networks without exclusion of any of the strains. Additionally, bacterial colonization patterns compatible with the different strains were observed, with high survival traits on avocado and tomato roots. The bacteria composing the consortium shared the same root habitat and exhibited biocontrol activity against soil-borne fungal pathogens at similar levels to those displayed by the individual strains. As expected, because these strains were isolated from avocado roots, this Pseudomonas-based consortium had more stable bacterial counts on avocado roots than on tomato roots; however, inoculation of tomato roots with this consortium was shown to protect tomato plants under high-temperature stress. The results revealed that this consortium has side beneficial effect for tomato plants under high-temperature stress, thus improving the potential performance of the individual strains. We concluded that this rhizobacterial consortium do not improve the plant protection against soil-borne phytopathogenic fungi displayed by the single strains; however, its inoculation can show an specific improvement of plant performance on a horticultural non-host plant (such as tomato) when the plant was challenged by high temperature stress, thus extending the beneficial role of this bacterial consortium.

bacLIFE: a user-friendly computational workflow for genome analysis and prediction of lifestyle-associated genes in bacteria.

Bacteria have an extensive adaptive ability to live in close association with eukaryotic hosts, exhibiting detrimental, neutral or beneficial effects on host growth and health. However, the genes involved in niche adaptation are mostly unknown and their functions poorly characterized. Here, we present bacLIFE ( https://github.com/Carrion-lab/bacLIFE ) a streamlined computational workflow for genome annotation, large-scale comparative genomics, and prediction of lifestyle-associated genes (LAGs). As a proof of concept, we analyzed 16,846 genomes from the Burkholderia/Paraburkholderia and Pseudomonas genera, which led to the identification of hundreds of genes potentially associated with a plant pathogenic lifestyle. Site-directed mutagenesis of 14 of these predicted LAGs of unknown function, followed by plant bioassays, showed that 6 predicted LAGs are indeed involved in the phytopathogenic lifestyle of Burkholderia plantarii and Pseudomonas syringae pv. phaseolicola. These 6 LAGs encompassed a glycosyltransferase, extracellular binding proteins, homoserine dehydrogenases and hypothetical proteins. Collectively, our results highlight bacLIFE as an effective computational tool for prediction of LAGs and the generation of hypotheses for a better understanding of bacteria-host interactions.

Insecticidal features displayed by the beneficial rhizobacterium Pseudomonas chlororaphis PCL1606.

The biocontrol rhizobacterium Pseudomonas chlororaphis is one of the bacterial species of the P. fluorescens group where insecticide fit genes have been found. Fit toxin, supported with other antimicrobial compounds, gives the bacterial the ability to repel and to fight against eukaryotic organisms, such as nematodes and insect larvae, thus protecting the plant host and itself. Pseudomonas chlororaphis PCL1606 is an antagonistic rhizobacterium isolated from avocado roots and show efficient biocontrol against fungal soil-borne disease. The main antimicrobial compound produced by P. chlororaphis PCL606 is 2-hexyl-5-propyl resorcinol (HPR), which plays a crucial role in effective biocontrol against fungal pathogens. Further analysis of the P. chlororaphis PCL1606 genome showed the presence of hydrogen cyanide (HCN), pyrrolnitrin (PRN), and homologous fit genes. To test the insecticidal activity and to determine the bases for such activity, single and double mutants on the biosynthetic genes of these four compounds were tested in a Galleria mellonella larval model using inoculation by injection. The results revealed that Fit toxin and HPR in combination are involved in the insecticide phenotype of P. chlororaphis PCL1606, and additional compounds such as HCN and PRN could be considered supporting compounds.

Interplay between rhizospheric Pseudomonas chlororaphis strains lays the basis for beneficial bacterial consortia.

Pseudomonas chlororaphis (Pc) representatives are found as part of the rhizosphere-associated microbiome, and different rhizospheric Pc strains frequently perform beneficial activities for the plant. In this study we described the interactions between the rhizospheric Pc strains PCL1601, PCL1606 and PCL1607 with a focus on their effects on root performance. Differences among the three rhizospheric Pc strains selected were first observed in phylogenetic studies and confirmed by genome analysis, which showed variation in the presence of genes related to antifungal compounds or siderophore production, among others. Observation of the interactions among these strains under lab conditions revealed that PCL1606 has a better adaptation to environments rich in nutrients, and forms biofilms. Interaction experiments on plant roots confirmed the role of the different phenotypes in their lifestyle. The PCL1606 strain was the best adapted to the habitat of avocado roots, and PCL1607 was the least, and disappeared from the plant root scenario after a few days of interaction. These results confirm that 2 out 3 rhizospheric Pc strains were fully compatible (PCL1601 and PCL1606), efficiently colonizing avocado roots and showing biocontrol activity against the fungal pathogen Rosellinia necatrix. The third strain (PCL1607) has colonizing abilities when it is alone on the root but displayed difficulties under the competition scenario, and did not cause deleterious effects on the other Pc competitors when they were present. These results suggest that strains PCL1601 and PCL1606 are very well adapted to the avocado root environment and could constitute a basis for constructing a more complex beneficial microbial synthetic community associated with avocado plant roots.

Mitigation of Pseudomonas syringae virulence by signal inactivation.

Pseudomonas syringae is an important plant pathogen of many valuable crops worldwide, with more than 60 identified pathovars. The phytotoxins produced by these organisms were related to the severity of the damage caused to the plant. An emerging strategy to treat bacterial infections relies on interference with their signaling systems. In this study, we investigated P. syringae pv. syringae, which produces the virulence factor mangotoxin that causes bacterial apical necrosis on mango leaves. A previously unknown signaling molecule named leudiazen was identified, determined to be unstable and volatile, and responsible for mangotoxin production. A strategy using potassium permanganate, compatible with organic farming, was developed to degrade leudiazen and thus to attenuate the pathogenicity of P. syringae pv. syringae.

The Rhizobacterium Pseudomonas alcaligenes AVO110 Induces the Expression of Biofilm-Related Genes in Response to Rosellinia necatrix Exudates.

The rhizobacterium Pseudomonas alcaligenes AVO110 exhibits antagonism toward the phytopathogenic fungus Rosellinia necatrix. This strain efficiently colonizes R. necatrix hyphae and is able to feed on their exudates. Here, we report the complete genome sequence of P. alcaligenes AVO110. The phylogeny of all available P. alcaligenes genomes separates environmental isolates, including AVO110, from those obtained from infected human blood and oyster tissues, which cluster together with Pseudomonas otitidis. Core and pan-genome analyses showed that P. alcaligenes strains encode highly heterogenic gene pools, with the AVO110 genome encoding the largest and most exclusive variable region (~1.6 Mb, 1795 genes). The AVO110 singletons include a wide repertoire of genes related to biofilm formation, several of which are transcriptionally modulated by R. necatrix exudates. One of these genes (cmpA) encodes a GGDEF/EAL domain protein specific to Pseudomonas spp. strains isolated primarily from the rhizosphere of diverse plants, but also from soil and water samples. We also show that CmpA has a role in biofilm formation and that the integrity of its EAL domain is involved in this function. This study contributes to a better understanding of the niche-specific adaptations and lifestyles of P. alcaligenes, including the mycophagous behavior of strain AVO110.

Beyond the Wall: Exopolysaccharides in the Biofilm Lifestyle of Pathogenic and Beneficial Plant-Associated Pseudomonas.

The formation of biofilms results from a multicellular mode of growth, in which bacteria remain enwrapped by an extracellular matrix of their own production. Many different bacteria form biofilms, but among the most studied species are those that belong to the Pseudomonas genus due to the metabolic versatility, ubiquity, and ecological significance of members of this group of microorganisms. Within the Pseudomonas genus, biofilm studies have mainly focused on the opportunistic human pathogen Pseudomonas aeruginosa due to its clinical importance. The extracellular matrix of P. aeruginosa is mainly composed of exopolysaccharides, which have been shown to be important for the biofilm architecture and pathogenic features of this bacterium. Notably, some of the exopolysaccharides recurrently used by P. aeruginosa during biofilm formation, such as the alginate and polysaccharide synthesis loci (Psl) polysaccharides, are also used by pathogenic and beneficial plant-associated Pseudomonas during their interaction with plants. Interestingly, their functions are multifaceted and seem to be highly dependent on the bacterial lifestyle and genetic context of production. This paper reviews the functions and significance of the exopolysaccharides produced by plant-associated Pseudomonas, particularly the alginate, Psl, and cellulose polysaccharides, focusing on their equivalents produced in P. aeruginosa within the context of pathogenic and beneficial interactions.

Role of extracellular matrix components in the formation of biofilms and their contribution to the biocontrol activity of Pseudomonas chlororaphis PCL1606.

Pseudomonas chlororaphis PCL1606 (PcPCL1606) displays plant-colonizing features and exhibits antagonistic traits against soil-borne phytopathogenic fungi. Biofilm formation could be relevant for the PcPCL1606 lifestyle, and in this study the role of some putative extracellular matrix components (EMC; Fap-like fibre, alginate and Psl-like polysaccharides) in the biofilm architecture and biocontrol activity of this bacterium were determined. EMC such as the Fap-like fibre and alginate polysaccharide play secondary roles in biofilm formation in PcPCL1606, because they are not fundamental to its biofilm architecture in flow cell chamber, but synergistically they have shown to favour bacterial competition during biofilm formation. Conversely, studies on Psl-like polysaccharide have revealed that it may contain mannose, and that it is strongly involved in the PcPCL1606 biofilm architecture and niche competition. Furthermore, the Fap-like fibre and Psl-like exopolysaccharide play roles in early surface attachment and contribute to biocontrol activity against the white root rot disease caused by Rosellinia necatrix in avocado plants. These results constitute the first report regarding the study of the extracellular matrix of the PcPCL1606 strain and highlight the importance of a putative Fap-like fibre and Psl-like exopolysaccharide produced by PcPCL1606 in the biofilm formation process and interactions with the host plant root.

A Large Tn7-like Transposon Confers Hyper-Resistance to Copper in Pseudomonas syringae pv. syringae.

Copper resistance mechanisms provide an important adaptive advantage to plant pathogenic bacteria under exposure to copper treatments. Copper resistance determinants have been described in Pseudomonas syringae pv. syringae (Pss) strains isolated from mango intimately associated with 62 kb plasmids belonging to the pPT23A family (PFP). It has been previously described that the indiscriminate use of copper-based compounds promotes the selection of copper resistant bacterial strains and constitutes a selective pressure in the evolution of copper resistance determinants. Hence, we have explored in this study the copper resistance evolution and the distribution of specific genetic determinants in two different Pss mango populations isolated from the same geographical regions, mainly from southern Spain with an average of 20 years of difference. The total content of plasmids, in particular the 62 kb plasmids, and the number of copper resistant Pss strains were maintained at similar levels over the time. Interestingly, the phylogenetic analysis indicated the presence of a phylogenetic subgroup (PSG) in the Pss mango phylotype, mostly composed of the recent Pss population analyzed in this study that was strongly associated with a hyper-resistant phenotype to copper. Genome sequencing of two selected Pss strains from this PSG revealed the presence of a large Tn7-like transposon of chromosomal location, which harbored putative copper and arsenic resistance genes (COARS Tn7-like). Transformation of the copper sensitive Pss UMAF0158 strain with some putative copper resistance genes and RT-qPCR experiments brought into light the role of COARS Tn7-like transposon in the hyper-resistant phenotype to copper in Pss.IMPORTANCECopper compounds have traditionally been used as standard bactericides in agriculture in the past few decades. However, the extensive use of copper has fostered the evolution of bacterial copper resistance mechanisms. Pseudomonas syringae is a plant pathogenic bacterium used worldwide as a model to study plant-pathogen interactions. The adaption of P. syringae to plant surface environment is the most important step prior to an infection. In this scenario, copper resistance mechanisms could play a key role in improving its epiphytic survival. In this work, a novel Tn7-like transposon of chromosomal location was detected in P. syringae pv. syringae strains isolated from mango. This transposon conferred the highest resistance to copper sulfate described to date for this bacterial phytopathogen. Understanding in depth the copper resistance mechanisms and their evolution are important steps to the agricultural industry to get a better improvement of disease management strategies.

Biological role of EPS from Pseudomonas syringae pv. syringae UMAF0158 extracellular matrix, focusing on a Psl-like polysaccharide.

Pseudomonas syringae is a phytopathogenic model bacterium that is used worldwide to study plant-bacteria interactions and biofilm formation in association with a plant host. Within this species, the syringae pathovar is the most studied due to its wide host range, affecting both, woody and herbaceous plants. In particular, Pseudomonas syringae pv. syringae (Pss) has been previously described as the causal agent of bacterial apical necrosis on mango trees. Pss exhibits major epiphytic traits and virulence factors that improve its epiphytic survival and pathogenicity in mango trees. The cellulose exopolysaccharide has been described as a key component in the development of the biofilm lifestyle of the P. syringae pv. syringae UMAF0158 strain (PssUMAF0158). PssUMAF0158 contains two additional genomic regions that putatively encode for exopolysaccharides such as alginate and a Psl-like polysaccharide. To date, the Psl polysaccharide has only been studied in Pseudomonas aeruginosa, in which it plays an important role during biofilm development. However, its function in plant-associated bacteria is still unknown. To understand how these exopolysaccharides contribute to the biofilm matrix of PssUMAF0158, knockout mutants of genes encoding these putative exopolysaccharides were constructed. Flow-cell chamber experiments revealed that cellulose and the Psl-like polysaccharide constitute a basic scaffold for biofilm architecture in this bacterium. Curiously, the Psl-like polysaccharide of PssUMAF0158 plays a role in virulence similar to what has been described for cellulose. Finally, the impaired swarming motility of the Psl-like exopolysaccharide mutant suggests that this exopolysaccharide may play a role in the motility of PssUMAF0158 over the mango plant surface.

Aer Receptors Influence the Pseudomonas chlororaphis PCL1606 Lifestyle.

Pseudomonas chlororaphis PCL1606 (PcPCL1606) is a rhizobacterium isolated from avocado roots, which is a favorable niche for its development. This strain extensively interacts with plant roots and surrounding microbes and is considered a biocontrol rhizobacterium. Genome sequencing has shown the presence of thirty-one potential methyl-accepting chemotaxis proteins (MCPs). Among these MCPs, two candidates are putative functional aerotaxis receptors, encoded at locus PCL1606_41090 (aer1-1) and locus PLC1606_20530 (aer1-2), that are homologous to the Aer receptor of Pseudomonas aeruginosa strain PaO1. Single- and double-deletion mutants in one or both genes have led to motility deficiencies in oxygen-rich areas, particularly reduced swimming motility compared with that of wildtype PcPCL1606. No differences in swarming tests were detected, and less adhesion by the aer double mutant was observed. However, the single and double mutants on avocado plant roots showed delayed biocontrol ability. During the first days of the biocontrol experiment, the aer-defective mutants also showed delayed root colonization. The current research characterizes the presence of aer transductors on P. chlororaphis. Thus, the functions of the PCL1606_41090 and PCL1606_20530 loci, corresponding to genes aer1-1 and aer1-2, respectively, are elucidated.

Soil Application of a Formulated Biocontrol Rhizobacterium, Pseudomonas chlororaphis PCL1606, Induces Soil Suppressiveness by Impacting Specific Microbial Communities.

Biocontrol bacteria can be used for plant protection against some plant diseases. Pseudomonas chlororaphis PCL1606 (PcPCL1606) is a model bacterium isolated from the avocado rhizosphere with strong antifungal antagonism mediated by the production of 2-hexyl, 5-propil resorcinol (HPR). Additionally, PcPCL1606 has biological control against different soil-borne fungal pathogens, including the causal agent of the white root rot of many woody crops and avocado in the Mediterranean area, Rosellinia necatrix. The objective of this study was to assess whether the semicommercial application of PcPCL1606 to soil can potentially affect avocado soil and rhizosphere microbial communities and their activities in natural conditions and under R. necatrix infection. To test the putative effects of PcPCL1606 on soil eukaryotic and prokaryotic communities, a formulated PcPCL1606 was prepared and applied to the soil of avocado plants growing in mesocosm experiments, and the communities were analyzed by using 16S/ITS metagenomics. PcPCL1606 survived until the end of the experiments. The effect of PcPCL1606 application on prokaryotic communities in soil and rhizosphere samples from natural soil was not detectable, and very minor changes were observed in eukaryotic communities. In the infested soils, the presence of R. necatrix strongly impacted the soil and rhizosphere microbial communities. However, after PcPCL1606 was applied to soil infested with R. necatrix, the prokaryotic community reacted by increasing the relative abundance of few families with protective features against fungal soilborne pathogens and organic matter decomposition (Chitinophagaceae, Cytophagaceae), but no new prokaryotic families were detected. The treatment of PcPCL1606 impacted the fungal profile, which strongly reduced the presence of R. necatrix in avocado soil and rhizosphere, minimizing its effect on the rest of the microbial communities. The bacterial treatment of formulated PcPCL1606 on avocado soils infested with R. necatrix resulted in biological control of the pathogen. This suppressiveness phenotype was analyzed, and PcPCL1606 has a key role in suppressiveness induction; in addition, this phenotype was strongly dependent on the production of HPR.

Pseudomonas syringae pv. syringae Associated With Mango Trees, a Particular Pathogen Within the "Hodgepodge" of the Pseudomonas syringae Complex.

The Pseudomonas syringae complex comprises different genetic groups that include strains from both agricultural and environmental habitats. This complex group has been used for decades as a "hodgepodge," including many taxonomically related species. More than 60 pathovars of P. syringae have been described based on distinct host ranges and disease symptoms they cause. These pathovars cause disease relying on an array of virulence mechanisms. However, P. syringae pv. syringae (Pss) is the most polyphagous bacterium in the P. syringae complex, based on its wide host range, that primarily affects woody and herbaceous host plants. In early 1990s, bacterial apical necrosis (BAN) of mango trees, a critical disease elicited by Pss in Southern Spain was described for the first time. Pss exhibits important epiphytic traits and virulence factors, which may promote its survival and pathogenicity in mango trees and in other plant hosts. Over more than two decades, Pss strains isolated from mango trees have been comprehensively investigated to elucidate the mechanisms that governs their epiphytic and pathogenic lifestyles. In particular, the vast majority of Pss strains isolated from mango trees produce an antimetabolite toxin, called mangotoxin, whose leading role in virulence has been clearly demonstrated. Moreover, phenotypic, genetic and phylogenetic approaches support that Pss strains producers of BAN symptoms on mango trees all belong to a single phylotype within phylogroup 2, are adapted to the mango host, and produce mangotoxin. Remarkably, a genome sequencing project of the Pss model strain UMAF0158 revealed the presence of other factors that may play major roles in its different lifestyles, such as the presence of two different type III secretion systems, two type VI secretion systems and an operon for cellulose biosynthesis. The role of cellulose in increasing mango leaf colonization and biofilm formation, and impairing virulence of Pss, suggests that cellulose may play a pivotal role with regards to the balance of its different lifestyles. In addition, 62-kb plasmids belonging to the pPT23A-family of plasmids (PFPs) have been strongly associated with Pss strains that inhabit mango trees. Further, complete sequence and comparative genomic analyses revealed major roles of PFPs in detoxification of copper compounds and ultraviolet radiation resistance, both improving the epiphytic lifestyle of Pss on mango surfaces. Hence, in this review we summarize the research that has been conducted on Pss by our research group to elucidate the molecular mechanisms that underpin the epiphytic and pathogenic lifestyle on mango trees. Finally, future directions in this particular plant-pathogen story are discussed.

The extracellular matrix protects Bacillus subtilis colonies from Pseudomonas invasion and modulates plant co-colonization.

Bacteria of the genera Pseudomonas and Bacillus can promote plant growth and protect plants from pathogens. However, the interactions between these plant-beneficial bacteria are understudied. Here, we explore the interaction between Bacillus subtilis 3610 and Pseudomonas chlororaphis PCL1606. We show that the extracellular matrix protects B. subtilis colonies from infiltration by P. chlororaphis. The absence of extracellular matrix results in increased fluidity and loss of structure of the B. subtilis colony. The P. chlororaphis type VI secretion system (T6SS) is activated upon contact with B. subtilis cells, and stimulates B. subtilis sporulation. Furthermore, we find that B. subtilis sporulation observed prior to direct contact with P. chlororaphis is mediated by histidine kinases KinA and KinB. Finally, we demonstrate the importance of the extracellular matrix and the T6SS in modulating the coexistence of the two species on melon plant leaves and seeds.

Fitness Features Involved in the Biocontrol Interaction of Pseudomonas chlororaphis With Host Plants: The Case Study of PcPCL1606.

The goal of this mini review is to summarize the relevant contribution of some beneficial traits to the behavior of the species Pseudomonas chlororaphis, and using that information, to give a practical point of view using the model biocontrol strain P. chlororaphis PCL1606 (PcPCL1606). Among the group of plant-beneficial rhizobacteria, P. chlororaphis has emerged as a plant- and soil-related bacterium that is mainly known because of its biological control of phytopathogenic fungi. Many traits have been reported to be crucial during the multitrophic interaction involving the plant, the fungal pathogen and the soil environment. To explore the different biocontrol-related traits, the biocontrol rhizobacterium PcPCL1606 has been used as a model in recent studies. This bacterium is antagonistic to many phytopathogenic fungi and displays effective biocontrol against fungal phytopathogens. Antagonistic and biocontrol activities are directly related to the production of the compound 2-hexyl, 5-propyl resorcinol (HPR), despite the production of other antifungal compounds. Furthermore, PcPCL1606 has displayed additional traits regarding its fitness in soil and plant root environments such as soil survival, efficient plant root colonization, cell-to-cell interaction or promotion of plant growth.

The Compound 2-Hexyl, 5-Propyl Resorcinol Has a Key Role in Biofilm Formation by the Biocontrol Rhizobacterium Pseudomonas chlororaphis PCL1606.

The production of the compound 2-hexyl-5-propyl resorcinol (HPR) by the biocontrol rhizobacterium Pseudomonas chlororaphis PCL1606 (PcPCL1606) is crucial for fungal antagonism and biocontrol activity that protects plants against the phytopathogenic fungus Rosellinia necatrix. The production of HPR is also involved in avocado root colonization during the biocontrol process. This pleiotrophic response prompted us to study the potential role of HPR production in biofilm formation. The swimming motility of PcPLL1606 is enhanced by the disruption of HPR production. Mutants impaired in HPR production, revealed that adhesion, colony morphology, and typical air-liquid interphase pellicles were all dependent on HPR production. The role of HPR production in biofilm architecture was also analyzed in flow chamber experiments. These experiments revealed that the HPR mutant cells had less tight unions than those producing HPR, suggesting an involvement of HPR in the production of the biofilm matrix.

Pantoea agglomerans as a New Etiological Agent of a Bacterial Necrotic Disease of Mango Trees.

Bacterial apical necrosis of mango trees, a disease elicited by Pseudomonas syringae pv. syringae, is a primary limiting factor of mango crop production in the Mediterranean region. In this study, a collection of bacterial isolates associated with necrotic symptoms in mango trees similar to those produced by bacterial apical necrosis disease were isolated over five consecutive years in orchards from the Canary Islands. The bacterial isolates were characterized and identified as Pantoea agglomerans. Pathogenicity tests conducted on onion bulbs and mango plants confirmed that P. agglomerans strains isolated from mango trees are a new etiological agent of a bacterial necrotic disease in the Canary Islands. Pathogenicity plasmids of the pPATH family have been previously reported in P. agglomerans. The majority of putatively pathogenic (n = 23) and pathogenic (n = 4) P. agglomerans strains isolated from mango trees harbored four plasmids, one of which was close in size to the 135-kb pPATH pathogenicity plasmid. The analysis of the presence of two major genes in pPATH plasmids (repA and hrpJ) was undertaken in P. agglomerans strains isolated from mango trees. The hrpJ gene was detected in the 140-kb plasmid of pathogenic P. agglomerans strains isolated from mango trees but it showed differences in nucleotide sequences compared with other pathogenic strains. In contrast, the repA gene was not detected in any of the putatively pathogenic and pathogenic P. agglomerans strains isolated from mango trees. Finally, genetic characterization and phylogenetic analysis using the hrpJ gene and the housekeeping genes gyrB and rpoB showed that almost all P. agglomerans strains that were putatively pathogenic and pathogenic on mango trees clustered together, forming a differentiated phylogroup with respect to the other pathogenic P. agglomerans strains described from other hosts.

Response of the Biocontrol Agent Pseudomonas pseudoalcaligenes AVO110 to Rosellinia necatrix Exudate.

The rhizobacterium Pseudomonas pseudoalcaligenes AVO110, isolated by the enrichment of competitive avocado root tip colonizers, controls avocado white root rot disease caused by Rosellinia necatrix Here, we applied signature-tagged mutagenesis (STM) during the growth and survival of AVO110 in fungal exudate-containing medium with the goal of identifying the molecular mechanisms linked to the interaction of this bacterium with R. necatrix A total of 26 STM mutants outcompeted by the parental strain in fungal exudate, but not in rich medium, were selected and named growth-attenuated mutants (GAMs). Twenty-one genes were identified as being required for this bacterial-fungal interaction, including membrane transporters, transcriptional regulators, and genes related to the metabolism of hydrocarbons, amino acids, fatty acids, and aromatic compounds. The bacterial traits identified here that are involved in the colonization of fungal hyphae include proteins involved in membrane maintenance (a dynamin-like protein and ColS) or cyclic-di-GMP signaling and chemotaxis. In addition, genes encoding a DNA helicase (recB) and a regulator of alginate production (algQ) were identified as being required for efficient colonization of the avocado rhizosphere.IMPORTANCE Diseases associated with fungal root invasion cause a significant loss of fruit tree production worldwide. The bacterium Pseudomonas pseudoalcaligenes AVO110 controls avocado white root rot disease caused by Rosellinia necatrix by using mechanisms involving competition for nutrients and niches. Here, a functional genomics approach was conducted to identify the bacterial traits involved in the interaction with this fungal pathogen. Our results contribute to a better understanding of the multitrophic interactions established among bacterial biocontrol agents, the plant rhizosphere, and the mycelia of soilborne pathogens.

Diversity of phytobeneficial traits revealed by whole-genome analysis of worldwide-isolated phenazine-producing Pseudomonas spp.

Plant-beneficial Pseudomonas spp. competitively colonize the rhizosphere and display plant-growth promotion and/or disease-suppression activities. Some strains within the P. fluorescens species complex produce phenazine derivatives, such as phenazine-1-carboxylic acid. These antimicrobial compounds are broadly inhibitory to numerous soil-dwelling plant pathogens and play a role in the ecological competence of phenazine-producing Pseudomonas spp. We assembled a collection encompassing 63 strains representative of the worldwide diversity of plant-beneficial phenazine-producing Pseudomonas spp. In this study, we report the sequencing of 58 complete genomes using PacBio RS II sequencing technology. Distributed among four subgroups within the P. fluorescens species complex, the diversity of our collection is reflected by the large pangenome which accounts for 25 413 protein-coding genes. We identified genes and clusters encoding for numerous phytobeneficial traits, including antibiotics, siderophores and cyclic lipopeptides biosynthesis, some of which were previously unknown in these microorganisms. Finally, we gained insight into the evolutionary history of the phenazine biosynthetic operon. Given its diverse genomic context, it is likely that this operon was relocated several times during Pseudomonas evolution. Our findings acknowledge the tremendous diversity of plant-beneficial phenazine-producing Pseudomonas spp., paving the way for comparative analyses to identify new genetic determinants involved in biocontrol, plant-growth promotion and rhizosphere competence.