Phyllosphere

The above-ground parts of plants, including the stems, leaves, flower and fruits are normally colonized by a variety of bacteria, yeasts, and fungi. The aerial habitat colonized by these microbes is termed the phyllosphere, and the inhabitants are called epiphytes.

The phyllosphere can be further subdivided into the caulosphere (stems), phylloplane (leaves), anthosphere (flowers), and carposphere (fruits). Even though the Phyllosphere comprises stems, leaves, flower and fruits, most work on phyllosphere microbiology has focused on leaves, which is the most dominant aerial plant structure.

The microbial communities of leaves are diverse and include many different genera of bacteria, filamentous fungi, yeasts, algae, and, less frequently, protozoa and nematodes.

Filamentous fungi are considered transient inhabitants of leaf surfaces, with rapidly sporulating species and yeasts colonizing this habitat more actively.

Bacteria are  the most abundant inhabitants of the phyllosphere. Epiphytic bacterial populations differ in size among and within plants of the same species, and over time as well as over the growing season. These variations in population sizes are caused by the large fluctuations in the physical and nutritional conditions characteristic of the phyllosphere.

Factors influencing microbial growth and distribution on Phyllosphere

• Growth on the leaf surface is frequently limited by both water and nutrients, as well as exposure to high levels of ultraviolet (UV) radiation. 
• Variation in the physical and chemical landscape of the leaf itself and patchiness of nutrients on the leaf surface impose additional constraints for colonizing microorganisms.
• Leaf topography changes as the leaf ages, so that plant–microbial interactions are dependent on time as well as the specific characteristics of the host plant. 
• Abiotic stress including adverse environmental conditions also limit microbial growth on phyllosphere. Denser populations of bacteria are found in the grooves between plant cells, at the base of trichomes, and near leaf veins and stomatal openings. This is a protective measure from abiotic stress. 
• Moisture availability is a key limitation to microbial growth on the leaf surface. The cuticle prevents moisture from leaving the inside of the leaf and limits how much water remains on the leaf surface. To overcome this challenge, bacteria may form aggregates and biofilms, producing extracellular polymeric substances (EPS), which can resist desiccation. Some bacteria produce surfactants to increase the wettability of the leaf and lessen the ability of the cuticle to limit water accumulation.
• The leaf surface has a boundary layer with a microclimate that typically has higher humidity than the broader phyllosphere, and this reduces some of the desiccation pressure for microbiota that are in that layer. 

Microbial populations typically increase following precipitation, when there can be substantial changes in the diversity and composition of the phyllosphere microbiome. Microorganisms on the leaf surface are generally oligotrophs that can tolerate low-nutrient conditions or are microorganisms that can interact with the host plant to obtain more nutrients. Although cuticular waxes are generally resistant to chemical movement, some plant metabolites can move to the leaf surface, supporting microbial growth. These compounds may arrive on the leaf surface by excretion from leaf cells, or due to osmotic pressure when the leaf is wet. 

Plants also release volatile organic compounds, which support specific populations of microorganisms; for example, methylotrophic bacteria that metabolize plant-derived single-carbon compounds are abundant constituents of the phyllosphere of many plant species.

Phyllosphere microorganisms may obtain nitrogen from plant produced amino acids, which leach to the leaf surface or by inorganic forms of nitrogen. Nitrogen arrives on the leaf surface through atmospheric deposition. Ammonia is typically assimilated by the leaf microbiota, although chemoautotrophic ammonia oxidizers can also be present in the Phyllosphere. Oxidised nitrogen species (nitrate and nitrite) are water soluble, and their availability changes during rain events. 

Bacterial nitrogen fixation can occur on the leaf surface, likely in pockets of moisture because of the anaerobic constraints of the fixation process. Exposure to UV radiation poses a particular challenge to leaf epiphytes, and bacterial and fungal populations that have been isolated from the phyllosphere are typically more pigmented than those from soil. This pigmentation can increase survival in the phyllosphere environment. Sphingomonas is an abundantly occuring bacteria  in phyllosphere which produce yellow or orange pigments.

The Nature and Composition of the Phyllosphere Microbiome

Phyllosphere microbiota represent a diverse array of microorganisms, but they are typically dominated by bacteria. Phyllosphere bacterial assemblages are generally less species rich than the rhizosphere or soil. Alpha proteobacteria are particularly well represented on the leaf surface, and these bacteria play many ecological roles. Gamma proteobacteria have also commonly been reported in surveys of phyllosphere bacterial community composition.

Proteobacteria are metabolically diverse, and the phyllosphere bacteria that carry out methyltrophy, nitrification, nitrogen fixation, or anoxygenic photosynthesis are typically representatives of this phylum. 

Bacteroidetes and Actinobacteria are generally the next most dominant bacterial lineages in phyllosphere communities. Bacteroidetes in the phyllosphere tend to be from families such as the Cytophagaceae or Chitinophagaceae. These organisms are often aerobic and pigmented, suggesting that they are well adapted to the leaf surface. 

The phylum Actinobacteria includes members that are plant pathogens, nitrogen-fixing symbionts, and fungal antagonists, as well as decomposers. Many of these roles have not been explored in the phyllosphere environment, but the Actinobacteria Corynebacterium has been used as a foliar-applied plant growth promoter (Nitrogen fixer). 

The fungal community is composed of organisms with a wide variety of ecological roles whose population sizes fluctuate in distinct seasonal trends based on the growing season and, ultimately, leaf senescence.

Moulds belonging to the Ascomycota are often the dominant fungi on the leaf surface before senescence. Other important fungi are yeasts belonging to the Ascomycota and Basidiomycota. 

Following leaf senescence, the fungal microbiome becomes dominated by filamentous fungi. Plant senescence is the process of aging in plants. Plants have both stress-induced and age-related developmental aging. Chlorophyll degradation during leaf senescence reveals the carotenoids, and is the cause of autumn leaf color in deciduous trees.

Interactions Between the Leaf Microbiome and the Plant Host

1. The Role of Phyllosphere Microorganisms in Plant Nutrient Acquisition

In tropical environments, nitrogen fixation occurs in the phyllosphere, potentially because higher moisture availability at the leaf surface allow nitrogen-fixing bacteria to be active. In areas subject to pollution from elevated levels of nitrogen, phyllosphere microorganisms may play a protective role by oxidising ammonia to nitrate through nitrification.

2. The Influence of the Phyllosphere Microbiome on Host Stress Tolerance

Biofilms in the phyllosphere resist desiccation. For example, Pseudomonas putida biofilms grown on phyllosphere retained their morphology better. Pseudomonas spp. are often  dominant constituents of the phyllosphere suggesting that naturally occurring biofilms may limit the loss of water and also protect from exposure to UV radiation. Pigmentation of bacteria and production of EPS can also provide some UV protection to the plant host.

Phyllosphere bacteria associated with aquatic plants can oxidise arsenite, preventing its accumulation and reducing toxicity and the phyllosphere microbiome may be a major contributor to aquatic arsenic cycling. Similarly, bacteria in the terrestrial phyllosphere may remediate airborne pollutants. e.g. Pseudomonas strains can accumulate phenol on the leaves of bean plants at concentrations 10-fold higher than in the ambient air, and use that phenol metabolically as a source of both energy and carbon. This provides some protection to the plant from airborne phenolics.

3. Interactions Between the Phyllosphere Microbiome and Plant Hormones

Many of the microorganisms that have been isolated from the phyllosphere have the ability to synthesise IAA, and promote plant-growth.

4. The Role of the Phyllosphere Microbiome in Mediating Plant–Pathogen Interactions

Biological control can be accomplished through; the induction of a plant immune response by non-pathogenic microorganisms, direct competition between non-pathogenic microorganisms and pathogens, or through the production of antibiotics. Competitive exclusion of pathogens by the broader phyllosphere community plays an important role in plant pathogen resistance.

✓ For example, Sphingomonas strains limit the plant pathogen P. syringae in Arabidopsis, while Methylobacterium strains do not, likely because Sphingomonas is a direct competitor with P. syringae for glucose, fructose, and sucrose, none of which are metabolised by Methylobacterium.