Urban greenspace types and climate factors jointly drive the microbial community structure and co-occurrence network.

The benefits of urban green space are socially widely recognized as a direct link between plant-microbe interactions and the maintenance of biodiversity, community stability, and ecosystem functioning. Nevertheless, there is a lack of knowledge about the factors influencing microbial communities in urban green spaces, especially those related to phyllosphere epiphytes and stem epiphytes. In this study, we analyzed the microbial community assembly in leaf and stem bark samples collected from Square, Road, Campus, and Park. Illumina sequecing of 16S amplicons was performed to characterize microbial diversity and composition. The α-diversity was significantly higher in the bark epiphytic community, compared to the phyllosphere. Moreover, urban greenspaces'type altered the way communities gathered. The main soil and air properties factors of the urban greenhouse (e.g. soil temperature, atmospheric moisture, air temperature) were shaping the characteristics of bacterial communities on the leaf surface and bark epiphytic. In addition, in the co-occurrence network analysis, keystone taxa were not mostly observed in abundant species, which may be necessary to maintain ecosystem functions. Finally, our findings provide a deeper understanding of the ecological dynamics and microbial interactions within plant phyllosphere and stem epiphytes microbiomes.

Host specificity of plant-associated bacteria is negatively associated with genome size and host abundance along a latitudinal gradient.

Host specialization plays a critical role in the ecology and evolution of plant-microbe symbiosis. Theory predicts that host specialization is associated with microbial genome streamlining and is influenced by the abundance of host species, both of which can vary across latitudes, leading to a latitudinal gradient in host specificity. Here, we quantified the host specificity and composition of plant-bacteria symbioses on leaves across 329 tree species spanning a latitudinal gradient. Our analysis revealed a predominance of host-specialized leaf bacteria. The degree of host specificity was negatively correlated with bacterial genome size and the local abundance of host plants. Additionally, we found an increased host specificity at lower latitudes, aligning with the high prevalence of small bacterial genomes and rare host species in the tropics. These findings underscore the importance of genome streamlining and host abundance in the evolution of host specificity in plant-associated bacteria along the latitudinal gradient.

Relationships between Phyllosphere Bacterial Communities and Leaf Functional Traits in a Temperate Forest.

As a vital component of biodiversity, phyllosphere bacteria in forest canopy play a critical role in maintaining plant health and influencing the global biogeochemical cycle. There is limited research on the community structure of phyllosphere bacteria in natural forests, which creates a gap in our understanding of whether and/or how phyllosphere bacteria are connected to leaf traits of their host. In this study, we investigated the bacterial diversity and composition of the canopy leaves of six dominant tree species in deciduous broad-leaved forests in northeastern China, using high-throughput sequencing. We then compare the differences in phyllosphere bacterial community structure and functional genes of dominant tree species. Fourteen key leaf functional traits of their host trees were also measured according to standard protocols to investigate the relationships between bacterial community composition and leaf functional traits. Our result suggested that tree species with closer evolutionary distances had similar phyllosphere microbial alpha diversity. The dominant phyla of phyllosphere bacteria were Proteobacteria, Actinobacteria, and Firmicutes. For these six tree species, the functional genes of phyllosphere bacteria were mainly involved in amino acid metabolism and carbohydrate metabolism processes. The redundancy and envfit analysis results showed that the functional traits relating to plant nutrient acquisition and resistance to diseases and pests (such as leaf area, isotope carbon content, and copper content) were the main factors influencing the community structure of phyllosphere bacteria. This study highlights the key role of plant interspecific genetic relationships and plant attributes in shaping phyllosphere bacterial diversity.

Diversity and biogeography of plant phyllosphere bacteria are governed by latitude-dependent mechanisms.

Predicting and managing the structure and function of plant microbiomes requires quantitative understanding of community assembly and predictive models of spatial distributions at broad geographic scales. Here, we quantified the relative contribution of abiotic and biotic factors to the assembly of phyllosphere bacterial communities, and developed spatial distribution models for keystone bacterial taxa along a latitudinal gradient, by analyzing 16S rRNA gene sequences from 1453 leaf samples taken from 329 plant species in China. We demonstrated a latitudinal gradient in phyllosphere bacterial diversity and community composition, which was mostly explained by climate and host plant factors. We found that host-related factors were increasingly important in explaining bacterial assembly at higher latitudes while nonhost factors including abiotic environments, spatial proximity and plant neighbors were more important at lower latitudes. We further showed that local plant-bacteria associations were interconnected by hub bacteria taxa to form metacommunity-level networks, and the spatial distribution of these hub taxa was controlled by hosts and spatial factors with varying importance across latitudes. For the first time, we documented a latitude-dependent importance in the driving factors of phyllosphere bacteria assembly and distribution, serving as a baseline for predicting future changes in plant phyllosphere microbiomes under global change and human activities.

Contribution of tree species to the co-occurrence network of the leaf phyllosphere and soil bacterial community in the subtropical forests.

The underlying mechanisms of the interactions between bacterial communities and tree species are still unknown, primarily attributed to a focus on the soil system while ignoring the leaf phyllosphere, which is a complex and diverse ecosystem that supports microbial diversity in the forest ecosystem. To gain insights into the mechanisms, the effects of seven common subtropical tree species, involving Pinus massoniana Lamb., Mytilaria laosensis Lecomte., Ilex chinensis Sims., Michelia macclurei Dandy., Liquidambar formosana Hance., Quercus acutissima Carruth., and Betula luminifera H.Winkler on the bacterial communities were investigated in the leaf phyllosphere and soil systems. We found that the bacterial community was dominated by Proteobacteria in the leaf phyllosphere (63.2-84.7%), and was dominated by Proteobacteria (34.3-45.0%) and Acidobacteria (32.5-40.6%) in soil. Mycorrhizal types and leaf phenology had no apparent effects on the bacterial abundance in the bacterial diversity in the leaf phyllosphere and soil. The bacterial community composition was greatly influenced by tree species in the leaf phyllosphere rather than in soil, with soil parameters (soil pH and C/N) and litter N identified as the most important factors. Ectomycorrhizal trees exerted positive effects on the complexity of the bacterial community at the expense of decreasing the robustness of the soil bacterial community, potentially threatening ecosystem stability. Evergreen trees decreased the network robustness of bacterial community by 21.9% higher than this of deciduous trees in the leaf phyllosphere. Similarly, evergreen trees decreased soil bacterial abundance by 50.8% and network robustness by 8.0% compared to deciduous trees, indicating the adverse impacts of leaf phenology on the bacterial stability in both leaf and soil. Overall, our results highlight the need for studies of leaf-associated bacteria to comprehensively understand the potential effects of tree species on microbial diversity and stability in subtropical forests.

Regulation of gut bacteria in silkworm (Bombyx mori) after exposure to endogenous cadmium-polluted mulberry leaves.

Soil cadmium (Cd) pollution presents a severe pollution burden to flora and fauna due to its non-degradability and transferability. The Cd in the soil is stressing the silkworm (Bombyx mori) out through a soil-mulberry-silkworm system. The gut microbiota of B.mori are reported to shape host health. However, earlier research had not reported the effect of endogenous Cd-polluted mulberry leaves on the gut microbiota of B.mori. In the current research, we compared the phyllosphere bacteria of endogenous Cd-polluted mulberry leaves at different concentrations. The investigation of the gut bacteria of B.mori fed with the mulberry leaves was done to evaluate the impact of endogenous Cd- polluted mulberry leaves on the gut bacteria of the silkworm. The results revealed a dramatic change in the gut bacteria of B.mori whereas, the changes in the phyllosphere bacteria of mulberry leaves in response to an increased Cd concentration were insignificant. It also increased the α-diversity and altered the gut bacterial community structure of B. mori. A significant change in the abundance of dominant phyla of gut bacteria of B.mori was recorded. At the genus level, the abundance of Enterococcus, Brachybacterium and Brevibacterium group related to disease resistance, and the abundance of Sphingomonas, Glutamicibacter and Thermus related to metal detoxification was significantly increased after Cd exposure. Meanwhile, there was a significant decrease in the abundance of the pathogenic bacteria Serratia and Enterobacter. The results demonstrated that endogenous Cd-polluted mulberry leaves caused perturbations in the gut bacterial composition of B.mori, which may driven by Cd content rather than phyllosphere bacteria. A significant variation in the specific bacterial community indicated the adaptation of B. mori gut for its role in heavy metal detoxification and immune function regulation. The results of this study help to understand the bacterial community associated with endogenous Cd-polluted resistance in the gut of B.mori, which proves to be a novel addition in describing its response in activating the detoxification mechanism and promoting its growth and development. This research work will help to explore the other mechanisms and microbiota associated with the adaptations to mitigate the Cd pollution problems.

The Effects of Phyllosphere Bacteria on Plant Physiology and Growth of Soybean Infected with Pseudomonas syringae.

Phyllosphere bacteria are an important determinant of plant growth and resistance to pathogens. However, the efficacy of phyllosphere bacteria in regulating infection of Pseudomonas syringae pv. glycinea (Psg) and its influence on soybean growth and physiology is unknown. In a greenhouse study, we assessed the influence of a phyllosphere bacterial consortium (BC) of 13 species isolated from field-grown soybean leaves on uninfected and deliberately Psg infected soybean plants. We measured Psg density on infected leaves with and without the application of the BC. The BC application resulted in a significant reduction in Psg cells. We also measured plant biomass, nodule mass and number, gas exchange, and leaf chlorophyll and nitrogen in four treatment groups: control plants, plants with a BC and no infection (BC), plants with BC and infected with Psg (BC + Psg), and plants infected with Psg alone. For all variables, plants infected with Psg alone showed significant reduction in measured variables compared to both BC treatments. Therefore, the bacterial consortium was effective in controlling the negative effects of Psg on growth and physiology. The BC treatment sometimes resulted in increases in measured variables such as plant biomass, nodule numbers, and leaf chlorophyll as compared to control and BC + Psg treatments. Overall, the positive influence of BC treatment on plant growth and physiology highlights its potential applications to increase crop yield and control bacterial pathogens.

Community Structure of Phyllosphere Bacteria in Different Cultivars of Fingered Citron (Citrus medica 'Fingered') and Their Correlations With Fragrance.

In recent years, plant metabolomics and microbiome studies have suggested that the synthesis and secretion of plant secondary metabolites are affected by microbial-host symbiotic interactions. In this study, six varieties of fingered citron (Citrus medica 'Fingered') are sampled to study their phyllosphere bacterial communities and volatile organic compounds (VOCs). High-throughput sequencing is used to sequence the V5-V7 region of the 16S rRNA of the fingered citron phyllosphere bacteria, and the results showed that Proteobacteria, Actinobacteria, Firmicutes, and Bacteroidetes were the dominant bacterial phylum in the phyllosphere of fingered citron. There were significant differences in the phyllosphere bacteria community between XiuZhen and the remaining five varieties. The relative abundance of Actinomycetospora was highest in XiuZhen, and Halomonas, Methylobacterium, Nocardioides, and Pseudokineococcus were also dominant. Among the remaining varieties, Halomonas was the genus with the highest relative abundance, while the relative abundances of all the other genera were low. Headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS) were used to analyze and identify the aroma compounds of six different fingered citron, and a total of 76 aroma compounds were detected in six varieties. Pinene, geraniol, and linalool were found to be the primary VOCs that affect the aroma of fingered citron based on relative odor activity value. The correlation analysis showed 55 positive and 60 negative correlations between the phyllosphere bacterial flora and aroma compounds of fingered citron. The top 10 genera in the relative abundance were all significantly associated with aroma compounds. This study provides deep insight into the relation between bacteria and VOCs of fingered citron, and this may better explain the complexity of the analysis of bacterial and metabolic interactions.

Environmental Factors Influencing Phyllosphere Bacterial Communities in Giant Pandas' Staple Food Bamboos.

The giant panda has developed a series of evolutionary strategies to adapt to a bamboo diet. The abundance and diversity of the phyllosphere microbiome change dramatically depending on the season, host species, location, etc., which may, in turn, affect the growth and health of host plants. However, few studies have investigated the factors that influence phyllosphere bacteria in bamboo, a staple food source of the giant panda. Amplicon sequencing of the 16S rRNA gene was used to explore the abundance and diversity of phyllosphere bacteria in three bamboo species (Arundinaria spanostachya, Yushania lineolate, and Fargesia ferax) over different seasons (spring vs. autumn), elevation, distance from water, etc., in Liziping National Nature Reserve (Liziping NR), China. And whole-genome shotgun sequencing uncovered the differences in biological functions (KEGG and Carbohydrate-Active enzymes functions) of A. spanostachya phyllosphere bacteria between spring and autumn. The results showed that the abundance and diversity of F. ferax phyllosphere bacteria were greater than that of the other two bamboo species in both seasons. And three kinds of bamboo phyllosphere bacteria in autumn were significantly higher than in spring. The season was a more important factor than host bamboo species in determining the community structure of phyllosphere bacteria based on the (un)weighted UniFrac distance matrix. The composition, diversity, and community structure of phyllosphere bacteria in bamboo were primarily affected by the season, species, altitude, tree layer, and shrub layer. Different bacterial communities perform different functions in different bamboo species, and long-term low temperatures may shape more varied and complex KEGG and Carbohydrate-Active enzymes functions in spring. Our study presented a deeper understanding of factors influencing the bacterial community in the bamboo phyllosphere. These integrated results offer an original insight into bamboo, which can provide a reference for the restoration and management of giant panda bamboo food resources in the Xiaoxiangling mountains.

Leaf metabolic influence of glyphosate and nanotubes on the Arabidopsis thaliana phyllosphere.

Chemical exposure can indirectly affect leaf microbiota communities, but the mechanism driving this phenomenon remains largely unknown. Results revealed that the co-exposure of glyphosate and multi-carbon nanotubes (CNTs) caused a synergistic inhibitory effect on the growth and metabolism of Arabidopsis thaliana shoots. However, only a slight inhibitory effect was induced by nanotubes or glyphosate alone at the tested concentrations. Several intermediate metabolites of nitrogen metabolism and fatty acid synthesis pathways were upregulated under the combined treatment, which increased the amount of energy required to alleviate the disruption caused by the combined treatment. Additionally, compared with the two individual treatments, the glyphosate/nanotube combination treatment induced greater fluctuations in the phyllosphere bacterial community members with low abundance (relative abundance (RA) <1%) at both the family and genus levels, and among these bacteria some plant growth promotion and nutrient supplement related bacteria were markable increased. Strikingly, strong correlations between phyllosphere bacterial diversity and metabolites suggested a potential role of leaf metabolism, particularly nitrogen and carbohydrate metabolism, in restricting the range of leaf microbial taxa. These correlations between phyllosphere bacterial diversity and leaf metabolism will improve our understanding of plant-microbe interactions and the extent of their drivers of variation and the underlying causes of variability in bacterial community composition.

Extract from phyllosphere bacteria with antibiofilm and quorum quenching activity to control several fish pathogenic bacteria.

OBJECTIVE: The objective of this research were to screen quorum quenching activity compound from phyllosphere bacteria as well as antibiofilm activity against several fish pathogen bacteria such as Aeromonas hydrophila, Streptococcus agalactiae, and Vibrio harveyi. RESULTS: We found eight phyllosphere bacteria isolates with potential quorum quenching activity to inhibit Chromobacterium violaceum as indicator bacteria. Crude extracts (20 mg/mL) showed various antibiofilm activity against fish pathogenic bacteria used in this study. Isolate JB 17B showed the highest activity to inhibit biofilm formation of A. hydrophila and V. harveyi, meanwhile isolate JB 3B showed the highest activity to inhibit biofilm of S. agalactiae. From destruction assay, isolate JB 8F showed the highest activity to disrupt biofilm of A. hydrophila isolate JB 20B showed the highest activity to disrupt biofilm of V. harveyi, isolate JB 17B also showed the highest activity to disrupt biofilm of S. agalactiae.

Plant resistance and leaf chemical characteristic jointly shape phyllosphere bacterial community.

Phyllosphere bacteria have an important role in plant growth and resistance to pathogen infection and are partially influenced by plant genotype and leaf environment. How plant resistance to pathogens and leaf chemical characteristics shape the phyllosphere bacterial communities is unclear. In this study, the phyllosphere bacterial communities of maize hybrids with various resistance to Setosphaeria turcica were compared using the high-throughput sequencing and large-scale culturing methods. The results showed that Shannon and Simpson indices of phyllosphere bacterial communities were markedly higher in the highly resistant hybrid (HR) compared with the susceptible one. Hierarchical clustering analysis, unweighted UniFrac principal component analysis (PCoA) and the analysis of similarities (ANOSIM) demonstrated that the phyllosphere bacterial communities were significantly distinct between resistant and susceptible hybrids. The redundancy analysis (RDA) demonstrated that leaf chemical characteristics, including nitrogen and phosphorus concentration, and disease resistance play an important role in shaping the phyllosphere bacterial community. Linear discriminant effect size (LEfSe) analysis indicated that Bacillus, Pseudomonas and Tumebacillus were the biomarker species in the phyllosphere of HR. Biocontrol bacteria against S. turcica (such as Pseudomonas and Bacillus) were isolated from the phyllosphere of HR by large-scale culturing. The work contributes to understanding of the phyllosphere bacterial community assembly and provides a new clue to screening for strong biocontrol bacteria from HR and to facilitating future breeding efforts for enhancing disease resistance.

Mitigation of drought stress in rice crop with plant growth-promoting abiotic stress-tolerant rice phyllosphere bacteria.

In the search of effective drought-alleviating and growth-promoting phyllosphere bacteria, a total of 44 bacterial isolates were isolated from the leaf surface of drought-tolerant rice varieties, Mattaikar, Nootripattu, Anna R(4), and PMK3, and screened for their abiotic stress tolerance by subjecting their growth medium to temperature, salinity, and osmotic stress. Only eight isolates were found to grow and proliferate under different abiotic stress conditions. These isolates were identified using 16S ribosomal DNA gene sequence and submitted to the NCBI database. All the bacterial isolates were identified as Bacillus sp., except PB24, which was identified as Staphylococcus sp., and these isolates were further screened for plant growth-promoting (PGP) traits such as IAA production, GA production, ACC deaminase activity, and exopolysaccharide production under three different osmotic stress conditions adjusted using polyethylene glycol (PEG 6000). Additionally, mineral solubilization was measured under the normal condition. Bacillus endophyticus PB3, Bacillus altitudinis PB46, and Bacillus megaterium PB50 were found to have multifarious PGP traits. Consecutively, the performance of an individual strain to improve the plant growth was investigated under the osmotic stress (25% PEG 6000) and nonstress condition by inoculating them into rice seeds using hydroponics culture. Furthermore, the drought-alleviating potency of bacterial strains was assessed in the rice plants using pot experiment (-1.2 MPa) through bacterial foliar application during the reproductive stage. Finally, as a result of seed inoculation and foliar spray, the application of B. megaterium PB50 significantly improved the plant growth under osmotic stress, protected plants from physical drought through stomatal closure, and improved carotenoid, total soluble sugars, and total protein content. Metabolites of PB50 were profiled under both stress and nonstress conditions using gas chromatography-mass spectroscopy.

Dynamics Relationship of Phyllosphere and Rhizosphere Bacterial Communities During the Development of Bothriochloa ischaemum in Copper Tailings.

Copper mining and the byproducts associated with the industry have led to serious pollution in the Loess Plateau of China. There is a potential in improving the ecological restoration efficiency of such degraded land through combining microbial and plant remediation approaches. However, the community structure and function of phyllosphere and rhizosphere microorganisms and their response to plant development in copper tailings dams are poorly understood. This study investigated the impact of the phyllosphere and rhizosphere microbial communities on Bothriochloa ischaemum during three distinct plant development stages: seedling, tiller, and mature. The relative species abundance and Shannon index of bacterial communities of the rhizosphere during the seedling and tiller stages were distinct from that in the mature stage. Dominant bacteria at the level of phyla, such as Proteobacteria, Cyanobacteria, Actinobacteria, and Bacteroidetes, followed distinct patterns associated with plant development in the phyllosphere, but the predominant bacteria were similar in the rhizosphere. Redundancy analysis showed that aboveground total nitrogen and the carbon and nitrogen ratio of this plant species significantly affected phyllosphere bacterial community structure, whereas soil water content, soil nutrients, electrical conductivity, and salinity significantly affected rhizosphere bacterial community structure. Moreover, keystone phyllosphere and rhizosphere bacterial species differed significantly. This study sheds new light on understanding the dynamic relationship of phyllosphere and rhizosphere bacterial communities during plant development in copper tailings. These results are beneficial to the development and utilization of beneficial microbial communities at different stages of development, which might help to reclaim and stabilize tailings more effectively.

Abundance of human pathogen genes in the phyllosphere of four landscape plants.

The surface of leaf, also known as phyllosphere, harbors diverse microbial communities which include both beneficial microorganisms promoting plants growth and harmful microorganisms, such as plant pathogens and human pathogens. Several studies have investigated the interaction between plants and human pathogens, while few works have focused on the quantitative analysis of pathogenic bacteria. On the basis of real-time polymerase chain reaction (qPCR), this study aimed to evaluate the abundance of following genes: the nuc and pvl of Staphylococcus aureus, the lytA and psaA of Streptococcus pneumoniae, and the ttr and invA of Salmonella enterica in the phyllosphere of four landscape plants (Nandina domestica, Rhododendron pulchrum, Photinia serrulata, and Cinnamomum camphora) growing in two habitats. Our results indicated that the relative abundance of pathogenic genes in the phyllosphere ranged from 10-9 to 10-6. The specific genes of S. aureus, S. pneumoniae and S. enterica in landscape plants were pvl, lytA and ttr, respectively. The two pathogenic genes of S. pneumoniae and the 16S rRNA gene were mainly affected by habitats, host species, and habitats-species interaction. Moreover, for the abundance of lytA and 16S rRNA, results showed that plants present in roadside with traffic pollution were relatively higher than that of campus with less pollution. The N. domestica and C. camphora were recommended for planting along the roadsides due to lower abundance of pathogenic genes. However, we have observed no significant difference in the abundance of pathogenic genes among four plants in the campus. Thereby, this study provided a valuable reference for selecting landscape plants in view of human health.

High-yield production of indole-3-acetic acid by Enterobacter sp. DMKU-RP206, a rice phyllosphere bacterium that possesses plant growth-promoting traits.

Enterobacter sp. DMKU-RP206 was isolated from rice leaves in Thailand and identified by the 16S rRNA gene and multilocus sequence (gyrB, rpoB, atpD, and infB genes) analysis. The bacterium was assessed on plant growth-promoting traits including indole-3-acetic acid (IAA) production. Phosphate solubilization, ammonia production, and antagonism to fungal plant pathogens, as well as siderophore production, were shown by this bacterium. However, only IAA production was focused on. The production of IAA by Enterobacter sp. DMKU-RP206 was optimized by statistical methods. A Box-Behnken design was used for the investigation of interactions among the basic influencing factors and for the optimization of IAA production. The results showed that l-tryptophan had a significant importance in terms of IAA production. Enterobacter sp. DMKU-RP206 produced a higher amount of IAA than previously reported for the genus Enterobacter. 0.85% of lactose as a carbon source, 1.3% of yeast extract as a nitrogen source, 1.1% of l-tryptophan as a precursor, 0.4% of NaCl, an initial pH of 5.8, an incubation temperature at 30 °C, and a shaking speed of 200 rpm were found to be the optimum conditions for IAA production. In addition, IAA production was performed to scale up IAA production, and the highest amount, 5561.7 mg l-1, was obtained. This study reported a 13.4-fold improvement in IAA production by Enterobacter sp. DMKU-RP206.