A better understanding of soil fungal communities is very useful in revealing the effects of an agroforestry system and would also help us to understand the fungi-mediated effects of agricultural practices on the processes of soil nutrient cycling and crop productivity. Compared to conventional monoculture farming, agroforestry systems have obvious advantages in improving land use efficiency and maintaining soil physicochemical properties, reducing losses of water, soil material, organic matter, and nutrients, as well as ensuring the stability of yields. In this study, we attempted to investigate the impact of a mulberry/alfalfa intercropping system on the soil physicochemical properties and the rhizosphere fungal characteristics (such as the diversity and structure of the fungal community), and to analyze possible correlations among the planting pattern, the soil physicochemical factors, and the fungal community structure. In the intercropping and monoculture systems, we determined the soil physicochemical properties using chemical analysis and the fungal community structure with MiSeq sequencing of the fungal ITS1 region. The results showed that intercropping significantly improved the soil physicochemical properties of alfalfa (total nitrogen, alkaline hydrolysable nitrogen, available potassium, and total carbon contents). Sequencing results showed that the dominant taxonomic groups were Ascomycota, Basidiomycota, and Mucoromycota. Intercropping increased the fungal richness of mulberry and alfalfa rhizosphere soils and improved the fungal diversity of mulberry. The diversity and structure of the fungal community were predominantly influenced by both the planting pattern and soil environmental factors (total nitrogen, total phosphate, and total carbon). Variance partitioning analysis showed that the planting pattern explained 25.9% of the variation of the fungal community structure, and soil environmental factors explained 63.1% of the variation. Planting patterns and soil physicochemical properties conjointly resulted in changes of the soil fungal community structure in proportion.

Soil methanogenic microorganisms are one of the primary methane-producing microbes in wetlands. However, we still poorly understand the community characteristic and metabolic patterns of these microorganisms according to vegetation type and seasonal changes. Therefore, to better elucidate the effects of the vegetation type and seasonal factors on the methanogenic community structure and metabolic patterns, we detected the characteristics of the soil methanogenic mcrA gene from three types of natural wetlands in different seasons in the Xiaoxing'an Mountain region, China. The results indicated that the distribution of Methanobacteriaceae (hydrogenotrophic methanogens) was higher in winter, while Methanosarcinaceae and Methanosaetaceae accounted for a higher proportion in summer. Hydrogenotrophic methanogenesis was the dominant trophic pattern in each wetland. The results of principal coordinate analysis and cluster analysis showed that the vegetation type considerably influenced the methanogenic community composition. The methanogenic community structure in the Betula platyphylla–Larix gmelinii wetland was relatively different from the structure of the other two wetland types. Indicator species analysis further demonstrated that the corresponding species of indicator operational taxonomic units from the Alnus sibirica wetland and the Betula ovalifolia wetland were similar. Network analysis showed that cooperative and competitive relationships exist both within and between the same or different trophic methanogens. The core methanogens with higher abundance in each wetland were conducive to the adaptation to environmental disturbances. This information is crucial for the assessment of metabolic patterns of soil methanogenic archaea and future fluxes in the wetlands of the Xiaoxing'an Mountain region given their vulnerability.

Introduction: As an ephemeral and oligotrophic environment, the phyllosphere harbors many highly diverse microorganisms. Importantly, it is known that their colonization of plant leaf surfaces is considerably influenced by a few abiotic factors related to climatic conditions. Yet how the dynamics of phyllosphere bacterial community assembly are shaped by detailed climatological elements, such as various bioclimatic variables, remains poorly understood.Methods: Using high-throughput 16S rRNA gene amplicon sequencing technology, we analyzed the bacterial communities inhabiting the leaf surfaces of an oilseed tree, yellowhorn (Xanthoceras sorbifolium), grown at four sites (Yinchuan, Otogqianqi, Tongliao, and Zhangwu) whose climatic status differs in northern China.Results and Discussion: We found that the yellowhorn phyllosphere’s bacterial community was generally dominated by four phyla: Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes. Nevertheless, bacterial community composition differed significantly among the four sampled site regions, indicating the possible impact of climatological factors upon the phyllosphere microbiome. Interestingly, we also noted that the α-diversities of phyllosphere microbiota showed strong positive or negative correlation with 13 bioclimatic factors (including 7 precipitation factors and 6 temperature factors). Furthermore, the relative abundances of 55 amplicon sequence variants (ASVs), including three ASVs representing two keystone taxa (the genera Curtobacterium and Streptomyces), exhibited significant yet contrary responses to the precipitation and temperature climatic variables. That pattern was consistent with all ASVs’ trends of possessing opposite correlations to those two parameter classes. In addition, the total number of links and nodes, which conveys community network complexity, increased with rising values of most temperature variables. Besides that, remarkably positive relevance was found between average clustering coefficient and most precipitation variables. Altogether, these results suggest the yellowhorn phyllosphere bacterial community is capable of responding to variation in rainfall and temperature regimes in distinctive ways.

WRKY transcription factors have been implicated in the regulation of disease resistance associated with plant immune responses, which has crucial implications for defense responses against stress in plants. The role played by the PsnWRKY70 gene of Populus ( Populus simonii × P. nigra ) in triggering the mechanism between the phyllosphere microbiome and plant defense against foliar pathogens remains unclear. Molecular ecological network analysis demonstrated that the stability and complexity of the phyllosphere bacterial community of Populus were influenced by Alternaria alternata infection. Specifically, compared to the wild-type line, the PsnWRKY70 -overexpressing (OE) line had a higher average clustering coefficient and modularity. Furthermore, metabolomic analysis revealed that 19 differential metabolites were significantly enriched in the leaves of the OE line. Among these metabolites, coumarin compounds, such as fraxetin-8-O-glucoside (fraxin) and scopoletin-7-O-glucoside (scopolin), significantly promoted the proliferation of the genera Methylobacterium and Achromobacter with resistance to A. alternata . Additionally, these genera also served as connectors in the molecular ecological network of the phyllosphere microbiome of the OE line. Thus, we concluded that the PsnWRKY70 gene enhanced the stability, complexity, and core taxa cooperation of the phyllosphere microbial network in Populus and regulated the biosynthesis of fraxin and scopolin to recruit beneficial bacteria controlling A. alternata infection. These findings provide valuable insights into the ability of resistant plant genotypes to drive the assembly of the phyllosphere microbiome, advancing our understanding of defense against pathogens using the biocontrol phyllosphere microbial community. IMPORTANCE: Poplar leaf blight caused by Alternaria alternata , a common disease in Northeast China, can cause abnormal abscission of poplar leaves and even lead to plant death in severe cases. WRKY transcription factors have been implicated in the regulation of disease resistance associated with plant immune responses to secondary metabolism via a complicated gene network. However, little is known about how the metabolites regulated by the PsnWRKY70 gene trigger changes in the phyllosphere microbiome, leading to increased resistance to foliar pathogens. Here, the PsnWRKY70 overexpressing line of Populus ( Populus simonii × P. nigra ) exhibited increased coumarin synthesis in the leaves, triggering changes in microbial species central in phyllosphere microbial networks and leading to increased resistance to A. alternata infection. This study provides insights into the role of the PsnWRKY70 gene in triggering the resistance mechanism to A . alternata in Populus .

Microorganisms colonizing plant surfaces and internal tissues may possess beneficial functions in promoting plant growth and health. However, information on the microbiome of bark tissues of woody plants remains limited, especially regarding the endophytic and epiphytic bacterial microbiota of the bark of Populus. To overcome this limitation, we established a workflow to quantify the composition and diversity of the endophytic and epiphytic bacterial microbiota colonizing the bark of Populus trichocarpa. Briefly, the epidermis of the stem of P. trichocarpa was repeatedly wiped with a cotton ball dipped in 0.1% Tween 20 to acquire epiphytic bacterial samples. The stripped epidermis was sterilized and then repeatedly frozen and crushed using liquid nitrogen and a bead beater, respectively, to collect endophytic bacterial samples. Genomic DNA was extracted from the cotton-adhered epiphytic bacterial communities and the crushed bark of P. trichocarpa, and underwent ploymerase chain reaction (PCR) amplification with primers targeting the hypervariable V5-V7 and V4 regions of the bacterial 16S rRNA gene. Three replicates of the obtained PCR products of each endophytic or epiphytic sample were mixed in equal concentrations to build the amplicon library, which was then sequenced. The obtained sequences were analyzed by sequence splicing, quality filtering, chimera removal, and taxonomic annotations. In summary, we established a reliable and efficient workflow from sample collection to data analysis for determining the endophytic and epiphytic bacterial microbiomes of the bark of P. trichocarpa through 16S rRNA gene profiling. Together with the methods for exploring microbiota colonizing barks established in the previous study, our methodology may serve as a blueprint for designing protocols for investigating the bark microbiome of other woody plant species, particularly economically important trees of forests.