Spatial and temporal variations in the microbiomes of different soil zones around clonal pedunculate oak trees (Quercus robur L.) out-planted as phytometers across grasslands in Europe

Soils harbor a huge diversity of microorganisms, which are dominated by bacteria and fungi. These soil microorganisms, collectively termed as the soil microbiome, are major contributors to soil biodiversity and play essential roles in soil functions (e.g. soil fertility and plant nutrition, organic matter degradation and nutrient cycling, and soil formation). Therefore, many studies in recent decades have explored soil microbial diversity in order to unravel driving forces of its variations. Hence, this thesis reports on spatial and temporal variations of the soil microbiome in response to site specificities, i.e. local climate as well as soil physico-chemistry, and host tree parameters. To avoid effects of intraspecific genetic variations, the pedunculate oak clone DF159 (Quercus robur L.) generated by the project TrophinOak-PhytOakmeter of the Soil Ecology Department at the Helmholtz Centre for Environmental Research (UFZ) was used as phytometer system. In the PhytOakmeter project of which this thesis is a part, saplings regenerated from microcuttings of DF159 were out-planted in grassland, forest and urban field sites in Central Germany and along a European North-South transect. The overall goal of the project is to analyze how the clone adapts to and performs under different regional climatic contexts and changing environment conditions. Pedunculate oak was chosen as a model tree species because it is engaged in highly complex and diverse multitrophic interactions, including soil microorganisms. Q. robur displays an endogenous rhythmic growth with alternating growing flushes in shoot and root, which can be repeated two to four times along a vegetation period. These alternating flushes have been shown to impact on variations of biological activities in soil zones close to the tree roots. Based on the above-described background, the current PhD study investigated changes in the soil microbial communities associated to the pedunculate oak phytometer outplanted in grassland sites at two different spatial scales: (1) the local scale by comparing the soil microbiomes associated to the phytometer in sites located within a close geographic space of Central Germany with similar climatic conditions; and (2) continental scale by making a similar comparison among sites along a European North-South transect, which encompasses a wide range of climatic and soil physico-chemical conditions. Moreover, temporal scale was considered, whereby the variability of the microbiomes intra-annually along a vegetation period was analyzed. Soil samples were taken not only in the tree root zone (RZ), i.e. soil zone containing living roots of the tree, but also in the tree root-free zone (RFZ), i.e. soil zone out of reach of any tree roots, but within the same field plot, to access also the local microbial pools. The analyses used a PCR-based Illumina MiSeq amplicon sequencing approach targeting bacteria and fungi, to assess their diversity, community structure and functionality after assignment of their OTUs to functional groups. In addition to Chapter 1, which introduces the whole work of this PhD research, the findings are presented within Chapters 2-4, of which two studies were already published in international peer-reviewed journals, while another study was published as a conference paper. The thesis is closed by the synopsis Chapter 5 that integrates discussion of all the publication chapters together with an outlook section. Chapter 2 “Tree root zone microbiome: exploring the magnitude of environmental conditions and host tree impact” published in Frontiers in Microbiology investigates the relative contribution of abiotic environmental and host tree parameters among four sites characterized by homogeneous climatic conditions in Central Germany, two years after the tree out-plant. We first compared at each field site the composition of the bacterial and fungal communities between the RZ of the oak clone, called PhytOakmeter in this chapter, and the tree RFZ. The chapter further evaluates the diversity and structure of the microbial communities within the tree RZ among the sites. The results revealed different microbial compositions between the tree RZ and RFZ, whereby the tree RZ-associated microbiome included numerous ectomycorrhizal fungi of the genera Hebeloma, Exophiala, Scleroderma, Tomentella, Trichophaea, and Tuber. This quick recruitment of specific beneficial microbial taxa from the local microbial pool seems to be among the tree strategies to acclimate to local site conditions. However, the overall tree contribution to shape soil microbial communities was lower than the impact of abiotic environmental parameters. The results revealed also a similar level of microbial diversity within the tree RZ among the sites for both the bacteria and fungi, an outcome attributed to the homogeneous climatic conditions within the sites and the common genetic identity of the host trees. In contrast, structure of the microbial communities was site-specific. Chapter 3 “Balance between geographic, soil, and host tree parameters to shape soil microbiomes associated to clonal oak varies across soil zones along a European North-South transect” published in Environmental Microbiology, also examines the relative impact of geographic, soil physico-chemical, and pedunculate oak clone parameters on the variability of the soil microbiome, but at a larger spatial scale from Lapinjärvi (Finland) to Bordeaux (Southwestern France), which is characterized by a broad range of geographic and soil physico-chemical conditions. In addition to the tree RFZ total microbiome and the tree RZ total microbiome, this chapter introduces a new sub-microbiome called tree RZ affine microbiome. The latter was defined as a subset of the RZ bacteria and fungi, significantly enriched in this zone compared to the tree RFZ. The results demonstrated an interplay among abiotic environmental and host tree parameters in shaping bacterial and fungal communities of the tree RZ along the European transect. These parameters showed a descending order of magnitude of their impact on the tree RZ total microbiome: geographic > soil physico-chemical > host tree parameters. However, for the variability of the RZ affine microbiome alone, the impact of the abiotic environmental parameters decreased, while the tree influence was strongly increased, particularly for fungi. Another important result was the highest proportion of the tree RZ affine microbial OTUs shared among all four sites, which was here designated as the tree “core” microbiome. These bacteria and fungi with significant affinity to the host tree, and shared by all the sites because of their ability to cope with diverging environmental conditions across the transect, may be playing a crucial role in supporting the wide distribution of Q. robur across Europe. Interestingly, we found no members of the RZ affine microbiome to be exclusive of only one particular site. Chapter 4 “Temporal changes and alternating host tree root and shoot growth affect soil microbiomes” published in Proceedings as conference paper after “The 1st International Electronic Conference on Microbiology”, considers a temporal scale, and here the variability of the tree RZ and RFZ total microbiomes was analyzed along a vegetation period in two sites of Central Germany. The soil was sampled at different time points coinciding with the tree alternating root and shoot growth, and the fall senescence that concludes the vegetation period. The results show a directional change over time along a vegetation period for the bacterial communities. However, the fungal communities did not show such temporal changes; they rather displayed a fine spatial scale partitioning closely linked to host plant individuals. In addition to the effect of temporal succession, deeper analyses of the generated data set will enable us to specify the impact of the alternating root and shoot growth characteristic of the tree endogenous rhythmic growth in the near future. These further analyses will include for example zooming in the tree RZ affine microbiome and in individual microbial functional groups. The results presented in this thesis evidence the quick impact of pedunculate oak tree clone on the soil microbiome within a two-year time span after the tree out-plant. Also, to different extents, geographic, soil physico-chemical, and host tree concurrently shape the soil bacterial and fungal communities. This thesis shows different spatial and temporal responses to the abiotic environmental and tree parameters between the soil bacterial and fungal communities. The use of tree clonal phytometer to study the tree-related parameters on soil microbiomes was proved to be a promising tool, to unravel the hierarchy of different abiotic and biotic factors in shaping the soil microbiome associated to long live trees. Finally, this work represents a first step toward establishing a long term monitoring of the dynamics of soil microbiomes associated to trees, as a strategy to unravel how these microorganisms participate to the long term acclimation of these long live plants to diverse and changing environments

Temporal Changes and Alternating Host Tree Root and Shoot Growth Affect Soil Microbiomes

Patterns of trees’ endogenous rhythmic growth (ERG) and paralleled C allocation shift between root and shoot systems have been studied, but there is still a need to understand their impact in shaping soil microbiomes. Moreover, the impact of plants on soil microbial communities can be modulated or overweighed by time-induced plant and/or seasonal changes. Thus, we intended to analyze the structure of soil microbiomes as response to simultaneous alternated host tree root and shoot flushes and time-induced changes within one vegetation period at two sites in Central Germany. In this study, we utilized oak phytometers (Quercus robur L., clone DF159) as host trees, and made use of their ERG, whereby consecutive root and shoot flushes make a complete growth cycle. We studied two complete growth cycles during the same vegetation period, performed a non-destructive soil sampling and applied high-throughput amplicon sequencing of the bacterial 16S gene and the fungal ITS2 region. As C allocation shifts between the tree root and shoot, released root exudates and consequently the nutrient availability alternate for soil microorganisms. We therefore anticipated different microbial communities in the host tree root zone along the growth cycles until autumnal leaf senescence. In our results, the bacterial community exhibited a directional change over time along the vegetation period. In contrast, the fungal community appeared sample specific. Our findings enlarge the current understanding of the temporal microbial assembly in the host tree root zone

Tree Root Zone Microbiome: Exploring the Magnitude of Environmental Conditions and Host Tree Impact

Tree roots attract their associated microbial partners from the local soil community. Accordingly, tree root-associated microbial communities are shaped by both the host tree and local environmental variables. To rationally compare the magnitude of environmental conditions and host tree impact, the “PhytOakmeter” project planted clonal oak saplings (Quercus robur L., clone DF159) as phytometers into different field sites that are within a close geographic space across the Central German lowland region. The PhytOakmeters were produced via micro-propagation to maintain their genetic identity. The current study analyzed the microbial communities in the PhytOakmeter root zone vs. the tree root-free zone of soil two years after out-planting the trees. Soil DNA was extracted, 16S and ITS2 genes were respectively amplified for bacteria and fungi, and sequenced using Illumina MiSeq technology. The obtained microbial communities were analyzed in relation to soil chemistry and weather data as environmental conditions, and the host tree growth. Although microbial diversity in soils of the tree root zone was similar among the field sites, the community structure was site-specific. Likewise, within respective sites, the microbial diversity between PhytOakmeter root and root-free zones was comparable. The number of microbial species exclusive to either zone, however, was higher in the host tree root zone than in the tree root-free zone. PhytOakmeter “core” and “site-specific” microbiomes were identified and attributed to the host tree selection effect and/or to the ambient conditions of the sites, respectively. The identified PhytOakmeter root zone-associated microbiome predominantly included ectomycorrhizal fungi, yeasts and saprotrophs. Soil pH, soil organic matter, and soil temperature were significantly correlated with the microbial diversity and/or community structure. Although the host tree contributed to shape the soil microbial communities, its effect was surpassed by the impact of environmental factors. The current study helps to understand site-specific microbe recruitment processes by young host trees

Balance between geographic, soil, and host tree parameters to shape soil microbiomes associated to clonal oak varies across soil zones along a European North–South transect

Tree root-associated microbiomes are shaped by geographic, soil physico-chemical, and host tree parameters. However, their respective impacts on microbiome variations in soils across larger spatial scales remain weakly studied. We out-planted saplings of oak clone DF159 (Quercus robur L.) as phytometer in four grassland field sites along a European North–South transect. After four years, we first compared the soil microbiomes of the tree root zone (RZ) and the tree root-free zone (RFZ). Then, we separately considered the total microbiomes of both zones, besides the microbiome with significant affinity to the RZ and compared their variability along the transect. Variations within the microbiome of the tree RFZ were shaped by geographic and soil physico-chemical changes, whereby bacteria responded more than fungi. Variations within both microbiomes of the tree RZ depended on the host tree and abiotic parameters. Based on perMANOVA and Mantel correlation tests, impacts of site specificities and geographic distance strongly decreased for the tree RZ affine microbiome. This pattern was more pronounced for fungi than bacteria. Shaping the microbiome of the soil zones in root proximity might be a mechanism mediating the acclimation of oaks to a wide range of environmental conditions across geographic regions

Temporal Changes and Alternating Host Tree Root and Shoot Growth Affect Soil Microbiomes

Patterns of trees’ endogenous rhythmic growth (ERG) and paralleled C allocation shift between root and shoot systems have been studied, but there is still a need to understand their impact in shaping soil microbiomes. Moreover, the impact of plants on soil microbial communities can be modulated or overweighed by time-induced plant and/or seasonal changes. Thus, we intended to analyze the structure of soil microbiomes as response to simultaneous alternated host tree root and shoot flushes and time-induced changes within one vegetation period at two sites in Central Germany. In this study, we utilized oak phytometers (Quercus robur L., clone DF159) as host trees, and made use of their ERG, whereby consecutive root and shoot flushes make a complete growth cycle. We studied two complete growth cycles during the same vegetation period, performed a non-destructive soil sampling and applied high-throughput amplicon sequencing of the bacterial 16S gene and the fungal ITS2 region. As C allocation shifts between the tree root and shoot, released root exudates and consequently the nutrient availability alternate for soil microorganisms. We therefore anticipated different microbial communities in the host tree root zone along the growth cycles until autumnal leaf senescence. In our results, the bacterial community exhibited a directional change over time along the vegetation period. In contrast, the fungal community appeared sample specific. Our findings enlarge the current understanding of the temporal microbial assembly in the host tree root zone

Temporal Changes and Alternating Host Tree Root and Shoot Growth Affect Soil Microbiomes

Patterns of trees’ endogenous rhythmic growth (ERG) and paralleled C allocation shift between root and shoot systems have been studied, but there is still a need to understand their impact in shaping soil microbiomes. Moreover, the impact of plants on soil microbial communities can be modulated or overweighed by time-induced plant and/or seasonal changes. Thus, we intended to analyze the structure of soil microbiomes as response to simultaneous alternated host tree root and shoot flushes and time-induced changes within one vegetation period at two sites in Central Germany. In this study, we utilized oak phytometers (Quercus robur L., clone DF159) as host trees, and made use of their ERG, whereby consecutive root and shoot flushes make a complete growth cycle. We studied two complete growth cycles during the same vegetation period, performed a non-destructive soil sampling and applied high-throughput amplicon sequencing of the bacterial 16S gene and the fungal ITS2 region. As C allocation shifts between the tree root and shoot, released root exudates and consequently the nutrient availability alternate for soil microorganisms. We therefore anticipated different microbial communities in the host tree root zone along the growth cycles until autumnal leaf senescence. In our results, the bacterial community exhibited a directional change over time along the vegetation period. In contrast, the fungal community appeared sample specific. Our findings enlarge the current understanding of the temporal microbial assembly in the host tree root zone

Innovate side event : learning journeys of smallholder finance and innovation adoption : MEDA innovate learning series

Analysis of market segments and their characteristics can help define action areas for organizations that offer financial products and services to smallholders. Some parameters are: subsistence smallholder farmers; those with more diversified incomes; commercial farmers in low value markets; those in higher value markets; and / or commercial farmers in tightly coordinated value chains. The presentation provides examples of this customer-centric approach. Overall, the project aims to assess potential of non-traditional financial services to enable large scale adoption of agricultural innovations among men and women smallholder farmers in South Asia, South America and East Africa, and to inform policymaking and programing on agricultural development

Balance between geographic, soil, and host tree parameters to shape soil microbiomes associated to clonal oak varies across soil zones along a European North–South transect

Tree root-associated microbiomes are shaped by geographic, soil physico-chemical, and host tree parameters. However, their respective impacts on microbiome variations in soils across larger spatial scales remain weakly studied. We out-planted saplings of oak clone DF159 (Quercus robur L.) as phytometer in four grassland field sites along a European North–South transect. After four years, we first compared the soil microbiomes of the tree root zone (RZ) and the tree root-free zone (RFZ). Then, we separately considered the total microbiomes of both zones, besides the microbiome with significant affinity to the RZ and compared their variability along the transect. Variations within the microbiome of the tree RFZ were shaped by geographic and soil physico-chemical changes, whereby bacteria responded more than fungi. Variations within both microbiomes of the tree RZ depended on the host tree and abiotic parameters. Based on perMANOVA and Mantel correlation tests, impacts of site specificities and geographic distance strongly decreased for the tree RZ affine microbiome. This pattern was more pronounced for fungi than bacteria. Shaping the microbiome of the soil zones in root proximity might be a mechanism mediating the acclimation of oaks to a wide range of environmental conditions across geographic regions

Tree Root Zone Microbiome: Exploring the Magnitude of Environmental Conditions and Host Tree Impact

Tree roots attract their associated microbial partners from the local soil community. Accordingly, tree root-associated microbial communities are shaped by both the host tree and local environmental variables. To rationally compare the magnitude of environmental conditions and host tree impact, the “PhytOakmeter” project planted clonal oak saplings (Quercus robur L., clone DF159) as phytometers into different field sites that are within a close geographic space across the Central German lowland region. The PhytOakmeters were produced via micro-propagation to maintain their genetic identity. The current study analyzed the microbial communities in the PhytOakmeter root zone vs. the tree root-free zone of soil two years after out-planting the trees. Soil DNA was extracted, 16S and ITS2 genes were respectively amplified for bacteria and fungi, and sequenced using Illumina MiSeq technology. The obtained microbial communities were analyzed in relation to soil chemistry and weather data as environmental conditions, and the host tree growth. Although microbial diversity in soils of the tree root zone was similar among the field sites, the community structure was site-specific. Likewise, within respective sites, the microbial diversity between PhytOakmeter root and root-free zones was comparable. The number of microbial species exclusive to either zone, however, was higher in the host tree root zone than in the tree root-free zone. PhytOakmeter “core” and “site-specific” microbiomes were identified and attributed to the host tree selection effect and/or to the ambient conditions of the sites, respectively. The identified PhytOakmeter root zone-associated microbiome predominantly included ectomycorrhizal fungi, yeasts and saprotrophs. Soil pH, soil organic matter, and soil temperature were significantly correlated with the microbial diversity and/or community structure. Although the host tree contributed to shape the soil microbial communities, its effect was surpassed by the impact of environmental factors. The current study helps to understand site-specific microbe recruitment processes by young host trees

Tree Root Zone Microbiome: Exploring the Magnitude of Environmental Conditions and Host Tree Impact

Tree roots attract their associated microbial partners from the local soil community. Accordingly, tree root-associated microbial communities are shaped by both the host tree and local environmental variables. To rationally compare the magnitude of environmental conditions and host tree impact, the “PhytOakmeter” project planted clonal oak saplings (Quercus robur L., clone DF159) as phytometers into different field sites that are within a close geographic space across the Central German lowland region. The PhytOakmeters were produced via micro-propagation to maintain their genetic identity. The current study analyzed the microbial communities in the PhytOakmeter root zone vs. the tree root-free zone of soil two years after out-planting the trees. Soil DNA was extracted, 16S and ITS2 genes were respectively amplified for bacteria and fungi, and sequenced using Illumina MiSeq technology. The obtained microbial communities were analyzed in relation to soil chemistry and weather data as environmental conditions, and the host tree growth. Although microbial diversity in soils of the tree root zone was similar among the field sites, the community structure was site-specific. Likewise, within respective sites, the microbial diversity between PhytOakmeter root and root-free zones was comparable. The number of microbial species exclusive to either zone, however, was higher in the host tree root zone than in the tree root-free zone. PhytOakmeter “core” and “site-specific” microbiomes were identified and attributed to the host tree selection effect and/or to the ambient conditions of the sites, respectively. The identified PhytOakmeter root zone-associated microbiome predominantly included ectomycorrhizal fungi, yeasts and saprotrophs. Soil pH, soil organic matter, and soil temperature were significantly correlated with the microbial diversity and/or community structure. Although the host tree contributed to shape the soil microbial communities, its effect was surpassed by the impact of environmental factors. The current study helps to understand site-specific microbe recruitment processes by young host trees