Inner Plant Values: Colonization, Diversity and Benefits from Endophytic Bacteria
Plants host a considerable number of bacteria inside their tissues. The importance of these endophytic bacteria in plant growth, development and resistance to biotic and abiotic stresses has been increasingly recognized. The recent advent of high throughput sequencing has greatly enhanced our understanding of endophytic bacterial communities. Plant roots robustly screen the soil bacteria via rhizosphere and rhizoplane, which results in a bacterial community dominated by Proteobacteria, Actinobacteria and to a lesser extent Bacteroidetes and Firmicutes, while being depleted of Acidobacteria. Metagenomic and genomic analysis of bacterial communities and model strains, respectively, have revealed many traits potentially involved in endophytic bacterial interactions with plants. Many of these traits have been confirmed in studies on function-defective mutants. Motility, cell-wall degradation products, and reactive oxygen species scavenging seem to be crucial factors for successful endophytic colonization and establishment. Although endophytic bacterial infections generate fewer responses of the plant immune system than pathogens, they endow plant priming conditions which elicit a faster and stronger defense once pathogens attack. Due to their plant growth-promoting traits, endophytic bacteria are being widely explored for their use in the improvement of plant performance.
Biodiversity of Endophytic Bacteria
The current technologies available for next generation sequencing (also known as high-throughput sequencing) allow the remarkable insights into bacterial composition and diversity. It has been shown that the plant interior harbors bacterial community with a much lower abundance and diversity compared with rhizosphere. The taxonomic structure of bacterial communities in plant endophytic compartment is dominated by phyla Proteobacteria and Actinobacteria. Firmicutes, Bacteroidetes, Gemmatimonadetes, Verrucomicrobia, Planctomycetes, Fusobacteria and some the other bacterial phyla may also present but in lower abundances. In contrast, Archaea and Acidobacteria are totally depleted from these habitats. Given that many plant growth promoting bacteria (PGPB) belong to these taxa and have been reported to be competent colonizers of plant tissues, endophytic bacteria holds great potentials as targets in screening studies aiming at the isolation of beneficial bacteria for plant growth or disease control in agricultural systems.
Our previous 16S rRNA Illumina sequencing of the archaeal and bacterial communities in bulk soil, ectrozhizosphere, and endophytic roots of 10-day’s wheat seedlings clearly shows the gradient selection of bacteria from soil to root endophytic compartment. As shown, Archaea and Acidobacteria greatly decreased in proportion from soil to extorhizosphere, and further decreased from ectorhizosphere to endophytic root compartment, which results in the eventual depletion from inside wheat roots. On the opposite, Proteobacteria, especially the γ-Proteobacteria and β-Proteobacteria gradually increased in relative abundance from soil to root endosphere (>70%). The γ-Proteobacteria are mainly composed of Pseudomonas and Enterobacteria-realted OTUs.
Biodiversity of Endophytic Bacteria
The current technologies available for next generation sequencing (also known as high-throughput sequencing) allow the remarkable insights into bacterial composition and diversity. It has been shown that the plant interior harbors bacterial community with a much lower abundance and diversity compared with rhizosphere. The taxonomic structure of bacterial communities in plant endophytic compartment is dominated by phyla Proteobacteria and Actinobacteria. Firmicutes, Bacteroidetes, Gemmatimonadetes, Verrucomicrobia, Planctomycetes, Fusobacteria and some the other bacterial phyla may also present but in lower abundances. In contrast, Archaea and Acidobacteria are totally depleted from these habitats. Given that many plant growth promoting bacteria (PGPB) belong to these taxa and have been reported to be competent colonizers of plant tissues, endophytic bacteria holds great potentials as targets in screening studies aiming at the isolation of beneficial bacteria for plant growth or disease control in agricultural systems.
Our previous 16S rRNA Illumina sequencing of the archaeal and bacterial communities in bulk soil, ectrozhizosphere, and endophytic roots of 10-day’s wheat seedlings clearly shows the gradient selection of bacteria from soil to root endophytic compartment. As shown, Archaea and Acidobacteria greatly decreased in proportion from soil to extorhizosphere, and further decreased from ectorhizosphere to endophytic root compartment, which results in the eventual depletion from inside wheat roots. On the opposite, Proteobacteria, especially the γ-Proteobacteria and β-Proteobacteria gradually increased in relative abundance from soil to root endosphere (>70%). The γ-Proteobacteria are mainly composed of Pseudomonas and Enterobacteria-realted OTUs.
Figure 1 Gradient selection of bacteria from soil to ecotorhizosphere and further to inside roots of wheat seedlings. These results were obtained by 16S rRNA Illumina deep sequencing (n=3). The red and yellow dashed lines on the pie graphs respectively highlight the gradient increase of Proteobacteria, and the depletion of both the Archaea and Acidobacteria from the inside wheat roots.
Figure 2 Circular map of a beneficial microbe's genome. The distribution of the circle from the outermost to the center is, (i) scale marks of the genome; (ii) protein-coding genes on the forward strand; (iii) protein-coding genes on the reverse strand; (iv) tRNA (black) and rRNA (red) on the forward strand; (v) tRNA (black) and rRNA (red) genes on the reverse strand; (vi) GC content; (vii) GC skew. Protein-coding genes are color coded according to their COG categories.
Factors Driving Endophytic Bacterial Communities
The endophytic bacterial communities may vary according to plant species, plant genotypes, plant organs, plant developmental stage, growing seasons (for trees), geographical location (field conditions), soil type, host plant nutrient status and cultivation practices. Characteristics of the soil and plant host seem to be the main drivers in shaping the endophytic microbiome; however, the plant associated factors may have a stronger influence on this selection. Besides the taxonomy-based approaches, function-based metagenomic analysis can reveal the functional potentials of endophytic communities. Studies on the functional changes of endophytic communities have been performed to a lesser extent than the phylogeny based analysis.
Distributions of Endophytic Bacteria and Colonization Patterns
Bacterial colonization patterns in plant endophytic compartment have thus far been mainly studied on grass plants (e.g., rice and kallar grass) using cultivated model stains. One of the most popular approaches for such evaluation include fluorescence in situ hybridization and genetically engineered bacteria strains tagged with reporter genes (e.g. gfp or gus) combined with microscopy have been used to enumerate and visualize colonization of endophytic bacteria in plant tissues. Endophytic bacteria are typically detected on outer cell layers, root cortex, phloem and xylem in the apoplast as well as intracellularly. Lateral root emergence sites are usually hot spots for bacterial colonization. Emerging lateral roots break through the epidermis, cortex, endodermis, casparian strip (band around endodermis) and pericycle thereby naturally forming a ‘freeway’ for bacteria to infect at these sites. From there, bacteria can further enter the phloem and xylem vessels that transport photosynthates (phloem), nutrients and water (xylem). Bacteria colonizing inside the root conductive tissues can be further transported to shoots and leaves.
Traits for Successful Translocation, Invasion and Colonization
To successfully colonize in the endophytic environment of plants, endophytic bacteria are then equipped with some necessary traits. Motility, chemotaxis, production of cell-wall degrading products and lipopolysaccharide formation are among the observed traits for bacteria to infect and adapt inside plants. Comparative genomic or metagenomic analysis together with mutational studies has confirmed the importance of these traits.
Bacterial Endophytes Circumvent Host Plant Defense
To avoid antagonistic effects, unlike the phytopathogens, the endophytic bacteria generally don’t elicit significant immune response such as the production of pathogenesis-related proteins in plants. One of the important cell surface components is the bacterial protein secretion systems (SS) which are large protein complexes that transverse the cell envelope and contains a channel mediating the translocation of proteins or protein-DNA complexes. Eight (Type I SS~ Type VI SS and Sec, Tat) and six (Sec, Tat, secA2, Sortase, Injectosome and Type VII SS) different protein SS have been described for Gram-negative and Gram-positive bacteria, respectively. Among the SS, T3SS and T4SS are pivotal for pathogens to deliver effector proteins into plants, which can induce plant MTI defense. However, the endophytic bacteria don’t seem to elicit significant plant immune defense as T3SS and T4SS may be either absent or present in small numbers. Colonization of endophytic bacteria also elicits an oxidative burst in the rice and traditional Chinese medicine plant of Atractylodes lancea. To detoxify the ROS produced by plant, the endophytic bacteria may resort to the ROS scavenging enzymes for help. A high number and diversity of genes encoding enzymes involved in ROS scavenging such as superoxide dismutase (SOD) and glutathione reductase (GR) are presented in the metagenome of the endophytic bacterial communities in rice roots.
To successfully colonize in the endophytic environment of plants, endophytic bacteria are then equipped with some necessary traits. Motility, chemotaxis, production of cell-wall degrading products and lipopolysaccharide formation are among the observed traits for bacteria to infect and adapt inside plants. Comparative genomic or metagenomic analysis together with mutational studies has confirmed the importance of these traits.
Bacterial Endophytes Circumvent Host Plant Defense
To avoid antagonistic effects, unlike the phytopathogens, the endophytic bacteria generally don’t elicit significant immune response such as the production of pathogenesis-related proteins in plants. One of the important cell surface components is the bacterial protein secretion systems (SS) which are large protein complexes that transverse the cell envelope and contains a channel mediating the translocation of proteins or protein-DNA complexes. Eight (Type I SS~ Type VI SS and Sec, Tat) and six (Sec, Tat, secA2, Sortase, Injectosome and Type VII SS) different protein SS have been described for Gram-negative and Gram-positive bacteria, respectively. Among the SS, T3SS and T4SS are pivotal for pathogens to deliver effector proteins into plants, which can induce plant MTI defense. However, the endophytic bacteria don’t seem to elicit significant plant immune defense as T3SS and T4SS may be either absent or present in small numbers. Colonization of endophytic bacteria also elicits an oxidative burst in the rice and traditional Chinese medicine plant of Atractylodes lancea. To detoxify the ROS produced by plant, the endophytic bacteria may resort to the ROS scavenging enzymes for help. A high number and diversity of genes encoding enzymes involved in ROS scavenging such as superoxide dismutase (SOD) and glutathione reductase (GR) are presented in the metagenome of the endophytic bacterial communities in rice roots.
Plant Growth Promoting Traits
The elucidation of the mechanics of plant growth promoting traits (PGPTs) will facilitate the development of potent biofertilizer and lead to a more sustainable agriculture. The endophytic bacterial community and even the single endophytic bacterial strain may have multiple PGPTs. The growth stimulation by endophytic bacteria can be the consequence of phytohormone production, induction of plant primed conditions, suppression of phytopathogens and improvement of plant nutrition. The figure below summarizes the proposed PGPTs of endophytic bacteria but which may not be exhausted.
The elucidation of the mechanics of plant growth promoting traits (PGPTs) will facilitate the development of potent biofertilizer and lead to a more sustainable agriculture. The endophytic bacterial community and even the single endophytic bacterial strain may have multiple PGPTs. The growth stimulation by endophytic bacteria can be the consequence of phytohormone production, induction of plant primed conditions, suppression of phytopathogens and improvement of plant nutrition. The figure below summarizes the proposed PGPTs of endophytic bacteria but which may not be exhausted.
Figure 3 Schematic representation summarizing proposed plant growth promoting traits (PGPTs) of endophytic bacteria. Endophytic bacteria may promote plant biomass by providing solubilizing phosphate, assimilable N to plants and suppress the ethylene synthesis in plant. The indirect plant growth promoting are mainly related to biocontrol especially in the plant root areas, which is mediated by the production of antimicrobial agents, siderophore production, competition for nutrients and the induction of plant defense. The arrows denote the plant-endophytic bacteria interactions and the symbol ‘⊥’ indicates inhibition. Abbreviations: QS, quorum sensing; IAA, indoleacetic acid; ACC, 1 aminocyclopropane-1-carboxylic acid; GAS, the gibberellins; CK, cytokinin, EPS, extracellular polymeric substance; LPS, lipopolysaccharide; αkb, α-ketobutyrate.