Plant perceptions of plant growth-promoting Pseudomonas

Plant-associated Pseudomonas live as saprophytes and parasites on plant surfaces and inside plant tissues. Many plant-associated Pseudomonas promote plant growth by suppressing pathogenic micro-organisms, synthesizing growth-stimulating plant hormones and promoting increased plant disease resistance. Others inhibit plant growth and cause disease symptoms ranging from rot and necrosis through to developmental dystrophies such as galls. It is not easy to draw a clear distinction between pathogenic and plant growth-promoting Pseudomonas. They colonize the same ecological niches and possess similar mechanisms for plant colonization. Pathogenic, saprophytic and plant growth-promoting strains are often found within the same species, and the incidence and severity of Pseudomonas diseases are affected by environmental factors and host-specific interactions. Plants are faced with the challenge of how to recognize and exclude pathogens that pose a genuine threat, while tolerating more benign organisms. This review examines Pseudomonas from a plant perspective, focusing in particular on the question of how plants perceive and are affected by saprophytic and plant growth-promoting Pseudomonas (PGPP), in contrast to their interactions with plant pathogenic Pseudomonas. A better understanding of the molecular basis of plant-PGPP interactions and of the key differences between pathogens and PGPP will enable researchers to make more informed decisions in designing integrated disease-control strategies and in selecting, modifying and using PGPP for plant growth promotion, bioremediation and biocontrol.     

The indigenous Pseudomonas plasmid pQBR103 encodes plant-inducible genes, including three putative helicases

Plasmid pQBR103 ( approximately 400 kb) is representative of many self-transmissible, mercury resistant plasmids observed in the Pseudomonas community colonising the phytosphere of sugar beet. A promoter trapping strategy (IVET) was employed to identify pQBR103 genes showing elevated levels of expression on plant surfaces. Thirty-seven different plant-inducible gene fusions were isolated that were silent in laboratory media, but active in the plant environment. Three of the fusions were to DNA sequences whose protein products show significant homology to DNA-unwinding helicases. The three helicase-like genes, designated helA, helB and helC, are restricted to a defined group of related Pseudomonas plasmids. They are induced in both the root and shoot environments of sugar beet seedlings. Sequence analysis of the three plasmid-encoded helicase-like genes shows that they are phylogenetically distinct and likely to have independent evolutionary histories. The helA gene is predicted to encode a protein of 1121 amino acids, containing conserved domains found in the ultraviolet (UV) resistance helicase, UvrD. A helA knockout mutant was constructed and no phenotypic changes were found with plasmid-conferred UV resistance or plasmid conjugation. The other 34 fusions are unique with no homologues in the public gene databases, including the Pseudomonas genomes. These data demonstrate the presence of plant responsive genes in plasmid DNA comprising a component of the genomes of plant-associated bacteria.     

Antagonism of Lactic Acid Bacteria against Phytopathogenic Bacteria

A variety of lactic acid bacteria, isolated from plant surfaces and plant-associated products, were found to be antagonistic to test strains of the phytopathogens Xanthomonas campestris, Erwinia carotovora, and Pseudomonas syringae. Effective “in vitro” inhibition was found both on agar plates and in broth cultures. In pot trials, treatment of bean plants with a Lactobacillus plantarum strain before inoculation with P. syringae caused a significant reduction of the disease incidence.     

New insights on molecular regulation of biofilm formation in plant-associated bacteria

Biofilms are complex bacterial assemblages with a defined three-dimensional architecture, attached to solid surfaces, and surrounded by a self-produced matrix generally composed of exopolysaccharides, proteins, lipids and extracellular DNA. Biofilm formation has evolved as an adaptive strategy of bacteria to cope with harsh environmental conditions as well as to establish antagonistic or beneficial interactions with their host. Plant-associated bacteria attach and form biofilms on different tissues including leaves, stems,vasculature, seeds and roots. In this review, we examine the formation of biofilms from the plant-associated bacterial perspective and detail the recently-described mechanisms of genetic regulation used by these organisms to orchestrate biofilm formation on plant surfaces. In addition, we describe plant host signals that bacterial pathogens recognize to activate the transition from a planktonic lifestyle to multicellular behavior.     

Complete genome sequence of a beneficial plant root-associated bacterium, Pseudomonas brassicacearum

To shed light on the genetic equipment of the beneficial plant-associated bacterium Pseudomonas brassicacearum, we sequenced the whole genome of the strain NFM421. Its genome consists of one chromosome equipped with a repertoire of factors beneficial for plant growth. In addition, a complete type III secretion system and two complete type VI secretion systems were identified. We report here the first genome sequence of this species.     

Genetic analysis of the aggA locus involved in agglutination and adherence of Pseudomonas putida, a beneficial fluorescent pseudomonad.

An isolate of Pseudomonas putida, which rapidly adheres to plant roots, is agglutinated by a glycoprotein from root surfaces. Agglutination is prevented and adherence to the root surface is diminished by Tn5 insertion in mutant 5123. Two cosmid clones from wild type P. putida and a 2.7-kbp EcoRI-HindIII subclone present in both cosmid clones restored agglutinable to wild type levels in transconjugants of the nonagglutinable (Agg-) Tn5 mutant 5123. These three clones increased agglutinability in transconjugants of the parental Agg+ isolate. The 2.7-kbp EcoRI-HindIII subclone restored adherence to bean root surfaces of 5123 to wild type levels in a short-term binding assay. Deletion analysis of the 2.7-kbp fragment indicated only 1.45 kbp was necessary for complementation of agglutinability in 5123. This sequence, termed the aggA locus, contains an open reading frame of 1,356 nucleotides encoding a predicted 50,509-Da protein. The distribution of the aggA locus in plant-associated bacteria, as detected through Southern hybridization, is limited to bacteria that express the agglutination phenotype.     

A vehicle for the introduction of transposons into plant-associated pseudomonads

A recombinant plasmid with wide-host-range transfer functions, narrow-host-range replication functions, and carrying a kanamycin-resistant transposon transferred kanamycin resistance to a number of plant-associated pseudomonads. Southern hybridization studies suggest that only a small portion of the plasmid, coinciding with the location of the transposon, is present in the kanamycin-resistant Pseudomonas derivatives. The plasmid sequences appear to be inserted at a number of different sites in the recipient genome. This plasmid can thus be used as a vehicle for the introduction of transposons into some plant-associated pseudomonads and should be useful in both genetic and ecological studies of these bacteria.     

Cyclic lipopeptide production by plant-associated Pseudomonas spp.: diversity, activity, biosynthesis, and regulation

Cyclic lipopeptides (CLPs) are versatile molecules produced by a variety of bacterial genera, including plant-associated Pseudomonas spp. CLPs are composed of a fatty acid tail linked to a short oligopeptide, which is cyclized to form a lactone ring between two amino acids in the peptide chain. CLPs are very diverse both structurally and in terms of their biological activity. The structural diversity is due to differences in the length and composition of the fatty acid tail and to variations in the number, type, and configuration of the amino acids in the peptide moiety. CLPs have received considerable attention for their antimicrobial, cytotoxic, and surfactant properties. For plant-pathogenic Pseudomonas spp., CLPs constitute important virulence factors, and pore formation, followed by cell lysis, is their main mode of action. For the antagonistic Pseudomonas sp., CLPs play a key role in antimicrobial activity, motility, and biofilm formation. CLPs are produced via nonribosomal synthesis on large, multifunctional peptide synthetases. Both the structural organization of the CLP synthetic templates and the presence of specific domains and signature sequences within peptide synthetase genes will be described for both pathogenic and antagonistic Pseudomonas spp. Finally, the role of various genes and regulatory mechanisms in CLP production by Pseudomonas spp., including two-component regulation and quorum sensing, will be discussed in detail.     

Ecomycins, unique antimycotics from Pseudomonas viridiflava

A novel family of peptide antimycotics, termed ecomycins, is described from Pseudomonas viridiflava , a plant-associated bacterium. Ecomycins B and C have molecular masses of 1153 and 1181. They contain equimolar amounts of a β hydroxyaspartic acid, homoserine, threonine, serine, alanine, glycine and one unknown amino acid. Fatty acids were detectable after hydrolysis, methylation and gas chromatography and mass spectroscopy. The ecomycins have significant bioactivities against a wide range of human and plant pathogenic fungi. The minimum inhibitory concentration values for ecomycin B were 4·0 μg ml⁻¹ against Cryptococcus neoformans and 31 μg ml⁻¹ against Candida albicans. Pseudomonas viridiflava also produces what appears to be syringotoxin, an antifungal lipopeptide previously described from Ps. syringae.     

Effects of plant genotype and growth stage on the structure of bacterial communities associated with potato (Solanum tuberosum L.)

The effects of genotype, plant growth and experimental factors (soil and year) on potato-associated bacterial communities were studied. Cultivars Achirana Inta, Désirée, Merkur and transgenic Désirée line DL12 (containing T4 lysozyme gene) were assessed in two field experiments. Cross-comparisons between both experiments were made using Désirée plants. Culture-dependent and -independent approaches were used to demonstrate effects on total bacterial, actinobacterial and Pseudomonas communities in bulk and rhizosphere soils and endospheres. PCR-denaturing gradient gel electrophoresis fingerprints prepared with group-specific primers were analyzed using multivariate analyses and revealed that bacterial communities in Achirana Inta plants differed most from those of Désirée and Merkur. No significant effects were found between Désirée and DL12 lines. Plant growth stage strongly affected different plant-associated communities in both experiments. To investigate the effect of plant-associated communities on plant health, 800 isolates from rhizospheres and endospheres at the flowering stage were tested for suppression of Ralstonia solanacearum biovar 2 and/or Rhizoctonia solani AG3. A group of isolates closely resembling Lysobacter sp. dominated in young plants. Its prevalence was affected by plant growth stage and experiment rather than by plant genotype. It was concluded that plant growth stage overwhelmed any effect of plant genotype on the bacterial communities associated with potato.     

Phylogeny of HCN synthase-encoding hcnBC genes in biocontrol fluorescent pseudomonads and its relationship with host plant species and HCN synthesis ability

Hydrogen cyanide (HCN) is a broad-spectrum antimicrobial compound involved in biological control of root diseases by many plant-associated fluorescent pseudomonads. The HCN synthase is encoded by three biosynthetic genes (hcnA, hcnB, and hcnC), but little is known about the diversity of these genes in fluorescent Pseudomonas spp. and in other bacteria. Here, the partial hcnBC sequence was determined for a worldwide collection of biocontrol fluorescent Pseudomonas spp. Phylogenies based on hcnBC and deduced protein sequences revealed four main bacterial groups, but topological incongruences were found between hcnBC and rrs-based phylogenies, suggesting past lateral transfer of hcnBC among saprophytic root-colonizing pseudomonads. Three of the four groups included isolates from different countries and host plants. Yet, these groups corresponded to distinct, ecologically-adapted populations of HCN-producing biocontrol fluorescent pseudomonads, as indicated by high hcnBC distinctness ratio values and the differences in production levels of HCN in vitro found between groups. This is in accordance with previous results on catabolic properties and biocontrol abilities of these strains. HCN synthase gene diversity may thus reflect the adaptive radiation of HCN+ biocontrol fluorescent pseudomonads. Positive correlations were found between HCN production in vitro and plant protection in the cucumber/Pythium ultimum and tomato/Fusarium oxysporum f. sp. radicis-lycopersici pathosystems.     

The transfer RNA of certain Enterobacteriacae contain 2-methylthiozeatin riboside (ms2io6A) an isopentenyl adenosine derivative

Isopentenyl adenosine derivatives are always located adjacent to the 3' end of the anticodon in transfer RNA and have been implicated in certain biological functions. In the enteric bacterium, E. coli, the derivative is ms2i6A whereas in some plant associated bacteria the derivative is the hydroxylated form, ms2io6A. Anti-i6A immunoadsorbent chromatography has been employed to detect isopentenyl adenosine compounds. In the present study we show that the transfer RNA of three species of enteric bacteria, S. typhimurium, K. pneumoniae, and S. marcescens contains both ms2io6A and ms2i6A. Under the growth conditions utilized the ms2io6A is predominant. The presence of ms2io6A in Enterobacteriacae is particularly noteworthy since in previous work it has been found only in plant-associated species of bacteria.     

Plant associated protists—Untapped promising candidates for agrifood tools

The importance of host-associated microorganisms and their biotic interactions for plant health and performance has been increasingly acknowledged. Protists, main predators and regulators of bacteria and fungi, are abundant and ubiquitous eukaryotes in terrestrial ecosystems. Protists are considered to benefit plant health and performance, but the community structure and functions of plant-associated protists remain surprisingly underexplored. Harnessing plant-associated protists and other microbes can potentially enhance plant health and productivity and sustain healthy food and agriculture systems. In this review, we summarize the knowledge of multifunctionality of protists and their interactions with other microbes in plant hosts, and propose a future framework to study plant-associated protists and utilize protists as agrifood tools for benefiting agricultural production.     

High diversity and low specificity of fungi associated with seedless epiphytic plants

Epiphytes, which grow on other plants for support, make up a large portion of Earth's plant diversity. Like other plants, their surfaces and interiors are colonized by diverse assemblages of fungi that can benefit their hosts by increasing tolerance for abiotic stressors and resistance to disease or harm them as pathogens. Fungal communities associated with epiphytic plants and the processes that structure these communities are poorly known. To address this, we sampled seven epiphytic seedless plant taxa in a Costa Rican rainforest and examined the effects of host identity and microhabitat on external and endophytic fungal communities. We found low host specificity for both external and endophytic fungi and weak differentiation between epiphytic and neighboring epilithic plant hosts. High turnover in fungi within and between hosts and habitats reveals that epiphytic plant-associated fungal communities are highly diverse and suggests that they are structured by stochastic processes.     

Evidence for phage-mediated gene transfer among Pseudomonas aeruginosa strains on the phylloplane.

As the use of genetically engineered microorganisms for agricultural tasks becomes more frequent, the ability of bacteria to exchange genetic material in the agricultural setting must be assessed. Transduction (bacterial virus-mediated horizontal gene transfer) is a potentially important mechanism of gene transfer in natural environments. This study investigated the potential of plant leaves to act as surfaces on which transduction can take place among microorganisms. Pseudomonas aeruginosa and its generalized transducing bacteriophage F116 were used as a model system. The application of P. aeruginosa lysogens of F116 to plant leaves resulted in genetic exchange among donor and recipient organisms resident on the same plant. Transduction was also observed when these bacterial strains were inoculated onto adjacent plants and contact was made possible through high-density planting.     

Bacterial endophytes from seeds of Norway spruce (Picea abies L. Karst)

Endophytic bacteria from wooden plants and especially seed-associated endophytes are not well studied. Fresh seeds collected from four Norway spruce trees (Picea abies) from different locations in the Slovene subalpine region were surface-sterilised and dissected into a seed coat, embryo and endosperm. The presence of endophytes was detected by culturing methods and by direct amplification of the eubacterial 16S rDNA gene. Both approaches identified bacteria from genera Pseudomonas and Rahnella in the Norway spruce seeds. Both are known plant-associated bacteria with growth-promoting properties and biological control potential. We suggest that plant seeds could serve as a vector for transmission of beneficial bacteria.     

Microbial antibiotics take the lead in the fight against plant pathogens

The plant microbiome is essential for plant fitness and health. Antibiotics produced by plant-associated bacteria have been shown to play an important role in protecting plant hosts against phytopathogens. Here, we highlight the strong biotechnological potential of (i) antibiotic producing plant-associated bacteria as biocontrol agents and (ii) the heterologous expression of antibiotic biosynthetic gene clusters in non-pathogenic plant-associated bacteria. We also provide the complete list of the active substances based on bacteria, fungi, and viruses currently approved or pending approval in the European Union, as an indication of the significant emergence and biotechnological applicability of biopesticides. Further progress in this field of research will enable the development of novel biopesticides for the biocontrol of agricultural pests.     

Mobile genetic elements in the genome of the beneficial rhizobacterium Pseudomonas fluorescens Pf-5

Background  

Pseudomonas fluorescens Pf-5 is a plant-associated bacterium that inhabits the rhizosphere of a wide variety of plant species and and produces secondary metabolites suppressive of fungal and oomycete plant pathogens. The Pf-5 genome is rich in features consistent with its commensal lifestyle, and its sequence has revealed attributes associated with the strain's ability to compete and survive in the dynamic and microbiologically complex rhizosphere habitat. In this study, we analyzed mobile genetic elements of the Pf-5 genome in an effort to identify determinants that might contribute to Pf-5's ability to adapt to changing environmental conditions and/or colonize new ecological niches.     

Symbiotic microorganisms, a key for ecological success and protection of plants

Plant-associated microbial diversity encompasses symbionts, protecting their host against various aggressions. Mycorrhizal and rhizospheric microorganisms buffer effects of soil toxic compounds and soil-borne pathogens. Endophytic bacteria and fungi, some of which are vertically inherited through seeds, take part in plant protection by acting directly on aggressive factors (mainly pathogens and herbivores) or by enhancing plant responses. Plant protective microbial symbionts determine the ecological success of plants; they drastically modify plant communities and related trophic webs. This review suggests approaches to improve the inventory of diversity and functions of in situ plant-associated microorganisms.