Maria Faticov This thesis includes the following chapters: Spatial and temporal ecology of I Faticov, M., Abdelfattah, A., Hambäck, P., Roslin, T. and Tack, A. J. M. Different spatial structure of above- and belowground plant- oak-associated fungal associated fungal communities. Manuscript. Spatial and temporal ecology of oak-associated fungal communities communities II Faticov, M., Desprez-Loustau, M-L., Kiss, L., Massot, M., d'Arcier, J. F., Mutz, J., Németh, M. Z., Roslin, T. and Tack, A. J. M. 2020. Niche differentiation within a cryptic pathogen complex: Maria Faticov climatic drivers and hyperparasitism across spatial scales. Submitted manuscript.

III Faticov, M., Abdelfattah, A., Roslin, T., Vacher, C., Hambäck, P., F. Blanchet, G., Lindahl, B. D. and Tack, A. J. M. 2020. Climate warming dominates over plant genotype in shaping the seasonal trajectory of foliar fungal communities on oak. Submitted manuscript.

IV Faticov, M., Ekholm, A., Roslin, T. and Tack, A. J. M. 2020. Climate and host genotype jointly shape tree phenology, disease levels and insect attacks. Oikos 129: 391–401.

Maria Faticov Ecologist who is fascinated by fungi on trees.

ISBN 978-91-7911-412-1

Department of Ecology, Environment and Plant Sciences

Doctoral Thesis in Ecology and Evolution at Stockholm University, Sweden 2021

Spatial and temporal ecology of oak-associated fungal communities Maria Faticov Academic dissertation for the Degree of Doctor of Philosophy in Ecology and Evolution at Stockholm University to be publicly defended on Friday 16 April 2021 at 10.00 online via zoom, a link will be published a few days before the defence at https://www.su.se/deep/.

Abstract Plants host a large diversity of microorganisms, which includes fungi, bacteria and archaea. Among these, fungi are highly diverse, and known to play a vital role in plant health and in regulation of the essential ecosystem functions. Nevertheless, we still lack a comprehensive understanding of the forces structuring plant-associated fungal communities in space and time. The main aim of this thesis was to decipher the drivers of the spatial patterns and temporal dynamics of fungal communities on plants. To this aim, I focused on Quercus robur and its associated fungi. Using a combination of observational and experimental studies, I assessed i) the distribution and drivers of the above- and belowground fungal communities at the landscape scale; ii) the role of climatic and trophic factors in defining the niches of cryptic species within a pathogen complex on oak and iii) the relative importance of warming, plant genotype and their interaction in shaping oak phenology and the seasonal dynamics of the associated fungal and insect communities. I found that aboveground fungal communities were highly variable among leaves within a single tree, and that belowground fungal communities had a stronger spatial structure than aboveground fungi at the landscape scale. Yet, climate, tree phenology or the distribution of the host tree did not explain spatial patterns in the above- and belowground communities. When focusing on three cryptic powdery mildew species within a pathogen complex on oak, I demonstrated that the climatic dimension is more important than the species interaction dimension for niche differentiation of these cryptic pathogens. A field heating experiment showed strong seasonal change in the structure of the foliar fungal community, with experimental warming playing an important role in driving this change. This experiment also revealed that warming and plant genotype jointly shape plant phenology, disease levels and insect abundance across the growing season. In conclusion, my findings suggest that abiotic forces can override biotic forces in structuring spatial patterns and temporal dynamics of fungal communities associated with plants. The particularly strong impact of warmer temperatures on foliar fungi in some of my studies indicates that climate warming has the potential to structure foliar fungal communities, with important implications for plant health, interactions between plants and other organisms and ecosystem functions.

Keywords: abiotic and biotic forces, climate warming, community ecology, foliar fungal community, host genotype, plant microbiome, powdery mildew, seasonal dynamics, warming-by-genotype interaction, Quercus robur, soil fungal community, spatial patterns.

Stockholm 2021 http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-190746

ISBN 978-91-7911-412-1 ISBN 978-91-7911-413-8

Department of Ecology, Environment and Plant Sciences

Stockholm University, 106 91 Stockholm

SPATIAL AND TEMPORAL ECOLOGY OF OAK-ASSOCIATED FUNGAL COMMUNITIES

Maria Faticov

Spatial and temporal ecology of oak-associated fungal communities

Maria Faticov ©Maria Faticov, Stockholm University 2021

ISBN print 978-91-7911-412-1 ISBN PDF 978-91-7911-413-8

Cover and illustrations by Maria Faticov Drawing of powdery mildew on cover page, in Introduction (p. 10) and on cover of Chapter II by Emilia Regazzoni

Printed in Sweden by Universitetsservice US-AB, Stockholm 2021 To Igor, my best friend and deepest love.

List of chapters

This thesis is based on the following chapters, which are referred to in the text by their Roman numerals.

I Faticov, M., Abdelfattah, A., Hambäck, P., Roslin, T. and Tack, A. J. M. Different spatial structure of above- and belowground plant-associated fungal communities. Manuscript.

II Faticov, M., Desprez-Loustau, M-L., Kiss, L., Massot, M., d'Arcier, J. F., Mutz, J., Németh, M. Z., Roslin, T. and Tack, A. J. M. 2020. Niche differentiation within a cryptic pathogen complex: climatic drivers and hyperparasitism across spatial scales. Submitted manuscript.

III Faticov, M., Abdelfattah, A., Roslin, T., Vacher, C., Hambäck, P., Blanchet, G., Lindahl, B. D. and Tack, A. J. M. 2020. Climate warming dominates over plant genotype in shaping the seasonal trajectory of foliar fungal communities on oak. Submitted manuscript.

IV Faticov, M., Ekholm, A., Roslin, T. and Tack, A. J. M. 2020. Climate and host genotype jointly shape tree phenology, disease levels and insect attacks. Oikos 129: 391–401. Contributions

I II III IV Idea and design MF, AA, AT MF, AT, MLDL MF, AT, TR MF, AE, AT, TR Data collection MF, AA MF, AT, JM, LK, MF MF, AE MLDL, MN Molecular work MF, AA JdA, MM, MN MF, AA, BL and bioinformatics Analyses MF MF MF, GB MF Manuscript MF MF MF MF preparation Manuscript AA, AT, PH, AT, JM, LK, MM, AA, AT, BL, AE, AT, TR reviewing TR MLDL, MN, TR CV, PH, TR

MF = Maria Faticov AA = Ahmed Abdelfattah JM = Jessie Mutz AE = Adam Ekholm LK = Levente Kiss AT = Ayco Tack MLDL = Marie-Laure Desprez-Loustau BL = Björn Lindahl MM = Marie Massot CV = Corinne Vacher MN = Márk Németh GB = Guillaume Blanchet PH = Peter Hambäck JdA = Julie d'Arcier TR = Tomas Roslin

Main supervisor: Dr. Ayco Tack, Department of Ecology, Environment and Plant Sciences, Stockholm University, Sweden Co-supervisor: Professor Peter Hambäck, Department of Ecology, Environment and Plant Sciences, Stockholm University, Sweden Contents

Abstract ...... 4 1. Introduction ...... 6 1.1. Community ecology and forces structuring communities 6 1.2. Plant-associated microbial communities 7 1.3. Forces structuring fungal communities 8 1.4. Oak-associated fungi and insects as model organisms 11 2. Aims of the thesis...... 13 3. Methods ...... 15 3.1. Observational data 15 3.2. Heating experiment 16 3.3. Molecular methods 18 4. Insights ...... 20 4.1. Differential imprint of local environment and spatial processes on foliar and soil fungal communities (I) 20 4.2. Climate is more important than species interactions in defining the niches of cryptic powdery mildew species (II) 22 4.3. Climate warming and to lesser extent tree genotype shape foliar fungal communities (III) 23 4.4. Climate warming and tree genotype jointly affect tree phenology, fungal disease levels and insect density (IV) 24 5. Concluding remarks and future directions ...... 26 6. Svensk sammanfattning ...... 29 7. Acknowledgements ...... 31 8. References ...... 34 Kappa

Abstract

Plants host a diverse community of microorganisms, which includes fungi, bacteria and archaea. Among these, fungi are highly diverse, and known to play a vital role in plant health and in regulation of the essential ecosystem functions. Nevertheless, we still lack a comprehensive understanding of the forces structuring plant-associated fungal communities in space and time. The main aim of this thesis was to decipher the drivers of the spatial patterns and temporal dynamics of plant-associated fungal communities. To this aim, I focused on Quercus robur and its associated fungal communities. Using a combination of observational and experimental studies, I assessed i) the distribution and drivers of the above- and belowground fungal communities at the landscape scale; ii) the role of climatic and trophic factors in defining the niches of cryptic species within a pathogen complex on oak and iii) the relative importance of warming, plant genotype and their interaction in shaping oak phenology and the seasonal dynamics of the associated fungal and insect communities. I found that aboveground fungal communities were highly variable among leaves within a single tree, and that belowground fungal communities had a stronger spatial structure at the landscape scale than aboveground communities. Yet, climate, tree phenology or the distribution of the host tree did not explain spatial patterns in the above- and belowground communities. When focusing on three cryptic powdery mildew species within a pathogen complex on oak, I demonstrated that the climatic dimension is more important than the species interaction dimension for niche differentiation of these cryptic pathogens. A field heating experiment showed strong seasonal change in the structure of the foliar fungal community, with experimental warming playing an important role in driving this change. This experiment also revealed that warming and plant genotype jointly shape plant phenology, disease levels and insect abundance across the growing season. In conclusion, my findings suggest that abiotic forces can override biotic forces in structuring spatial patterns and temporal dynamics of fungal communities associated with plants. The particularly strong impact of warmer temperatures on foliar fungi in some of my studies indicates that climate warming has the potential to structure foliar fungal communities, with important implications for plant health, interactions between plants and other organisms and ecosystem functions.

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Keywords: abiotic and biotic forces, climate warming, community ecology, foliar fungal community, host genotype, plant microbiome, powdery mildew, seasonal dynamics, warming-by-genotype interaction, soil fungal community, spatial patterns.

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1. Introduction Ecology is the study of organisms and how they interact with the environment around them and with other organisms (Smith 1966). Understanding how these interactions play out in nature is more topical than ever, given that the abiotic environment is changing, and that the effects of this change on species are – to an unknown part – mediated by other abiotic and biotic forces (Walther 2010). In this thesis, I try to contribute to this understanding by looking at the relative importance of abiotic and biotic factors in structuring communities, specifically by focusing on microbial communities associated with plants.

1.1. Community ecology and forces structuring communities Community ecology aims to understand what abiotic and biotic forces determine the structure and dynamics of communities at different spatial and temporal scales. At the beginning, community ecology was a mostly a descriptive field, in which communities were classified based on species identities and their numbers (Elton 1927, Clements 1936). Scientists realised that species differed in their abundances among different locations, and these differences were often related to variation in environmental conditions. This led to the classification of communities according to the patterns of their composition and environmental preferences (Wallace 1876, Whittaker 1962). Many theoretical concepts, which emerged during the 20th century, shaped the field of modern community ecology. For example, in the early 20th century, Clements described communities as ‘superorganisms’ containing species that are not randomly structured and assembled in certain combinations (Clements 1916). By contrast, the large temporal and spatial variability in community composition, led Gleason to propose that different species respond to different climatic conditions and thus communities are not randomly structured (Gleason 1939). Niche theory, developed by Grinnell (i.e. climatic effects on species distributions) and Elton (the role of species interactions in niche differentiation), was formalised by Hutchinson with the notion of the fundamental and realised niche (Grinnell 1917, Elton 1927, Hutchinson 1959). Half a century later, Hubbell’s Neutral Theory proposed that all individuals are ecologically equivalent, and that community structures are independent of organism traits and governed by random events, such as dispersal colonization, extinction and drift (Hubbell 2001). While Neutral Theory received little empirical evidence, it often serves as a null model for

6 Kappa evaluation of the impact of evolutionary processes (e.g. adaptation and natural selection) and life-history traits in structuring communities. Several of the concepts discussed above, gave the foundation to modern theoretical frameworks, such as Metacommunity Theory (Leibold et al. 2004, Logue et al. 2011). Metacommunity Theory has been a particularly useful framework for integrating the abiotic environment, biotic interactions and dispersal events as drivers of communities across spatial scales. While a unifying theory of community ecology is still lacking, ecologists are in agreement that communities are often structured by abiotic, biotic and stochastic forces and that these forces can act either independently or interactively (Gravel et al. 2006, 2019, Weiher et al. 2011, Götzenberger et al. 2012). The relative importance of these processes in shaping communities of macroorganisms has received a lot of attention. However, how these forces affect microbial communities is largely unknown (Mony et al. 2020). My objective in the introduction is to review the abiotic and biotic forces that are of particular relevance for microbial communities associated with plants. First, I will briefly introduce fungal communities and the role they are playing in plant functioning and health. Second, I will present how fungal communities can vary in space and time. I will end the introduction by discussing the role of abiotic (i.e. climate) and biotic filters (i.e. plant characteristics, plant genotype and species interactions) in influencing plant fungal communities.

1.2. Plant-associated microbial communities Plant organs harbour many different microorganisms. Plant leaves and roots are no exception, and they interact with a multitude of microorganisms, including prokaryotes (bacteria and archaea), eukaryotes (fungi, oomycetes and nematodes) and viruses (Vorholt 2012, Bardgett and van der Putten 2014). These microorganisms form complex associations with plants and play important roles in promoting plant functioning and health. Among plant-associated microorganisms, fungi are one of the dominant groups. Plant-fungal encounters can be friendly, neutral or hostile. Some fungi can transform and translocate essential nutrients in the soil to make them available to plants (Gouda et al. 2018). Others protect the host against stresses, including drought and attack by pathogens and insects (Rodriguez et al. 2009, Peñuelas and Terradas 2014), whereas pathogenic fungi extract resources and weaken plant defences (Burdon and Thrall 2014). Fungi also contribute to the processes of leaf

7 Kappa senescence and decomposition (Voříšková and Baldrian 2013, Vacher et al. 2016). As such, fungal communities play an important role in regulating plant health, interactions between plants and other organisms and ecosystem functioning. The use of molecular methods to identify fungal taxa has taught us much about the distribution of fungi in natural world. At a biogeographic scale, fungi show some degree of habitat endemicity (Martiny et al. 2006) and also frequently vary in abundance and diversity over spatial scales ranging from millimetres to hundreds of kilometres (Fierer 2008, Hanson et al. 2012, Choudoir et al. 2018). At a given location, fungal communities can also dramatically change in their composition across the seasons and among years (Osono 2008, Jumpponen and Jones 2010, Martins et al. 2016). The knowledge of the spatial and temporal distribution of plant-associated fungi is fundamental to understanding of the ecological and evolutionary processes that shape plant-fungal interactions.

1.3. Forces structuring fungal communities 1.3.1. Spatial and temporal variation in fungal communities Fungal communities associated with plants are often not randomly distributed. A handful of studies, looking at the spatial distribution of aboveground fungi showed that foliar fungal dispersal is rather common over both short and long distances (Peay et al. 2010, Barge et al. 2019, I). As for belowground fungi, recent evidence shows spatial clustering of soil fungal communities at scales from square millimetres to hundreds of meters (Ettema and Wardle 2002, Vannette et al. 2016). The observed clustering can be explained by dispersal limitation or by small-scale differences in environmental conditions. In addition to the spatial variation, foliar and soil fungal communities can also vary temporally, for example across the season, although less is known about the drivers of this variation. Seasonal variation in communities has been attributed to fungal life cycles, physical and chemical changes associated with leaf ageing, and temporal changes in climatic conditions (Osono 2008, Jumpponen and Jones 2010, Martins et al. 2016, Fort et al. 2016). While foliage of evergreen trees remains active throughout the season, deciduous trees produce their leaves after winter and lose them in autumn. The development of the fungal community often follows tree phenology, with community structure showing rapid turnover when leaves mature and senesce (Peñuelas et al. 2012, Lebeis 2015, III). Spring and autumn phenology of trees are partly under genetic control, which means that

8 Kappa trees within the same population can differ in the timing of bud burst and leaf senescence (Forrest and Miller-Rushing 2010, Evans et al. 2016, IV). These differences in spring and autumn phenology among trees can also explain the variation in the structure and dynamics of their fungal communities.

1.3.2. Climate Several studies have shown that foliar and soil fungal communities vary along latitudinal and elevational gradients (Zimmerman and Vitousek 2012, Coince et al. 2014, Siles and Margesin 2016), which are often used as proxies for temperature gradients. In such observational studies, other environmental factors, such as precipitation and soil characteristics, can confound the effect of temperature. While disentangling the effect of different environmental factors can be difficult in observational studies, multifactorial experiments can help us to assess the relative importance of different climatic factors and inform us about the potential consequences of environmental change for fungal communities. While the impact of temperature on soil and root microbial communities has received a lot of attention (Frey et al. 2008, Solly et al. 2017, Romero-Olivares et al. 2017), few experiments have manipulated air temperature to assess the effect of warming on foliar fungal communities. The available experimental evidence indicates that foliar fungi are influenced by elevated temperatures, with warming changing community composition and decreasing fungal richness (Bálint et al. 2015, III). A better understanding of the effects of climate warming on fungal communities is crucial for predicting the consequences of spatial and temporal variation in climate, and of climate change, for plant health and ecosystem functions.

1.3.3. Plant genotype and genotype-by-environment interactions In addition to spatial variation in climate, genetic variation among plants can also influence the colonization and establishment of the associated fungal communities, which has been studied within the framework of community genetics (Gaylord et al. 1996, Whitham et al. 2006, Zytynska et al. 2011). Specifically, differences in physical and chemical characteristics of leaf traits among genotypes may result in differential colonization of tree genotypes by fungi, which may lead to variation in fungal community composition (Hoffman and Arnold 2008, Bálint et al. 2013, Lamit et al. 2015). Several experimental studies have shown that tree genotype plays a larger role than climate in shaping foliar fungal communities. However, tree

9 Kappa genotypes in these studies are often collected from a large geographic area, whereas the experiments are usually conducted in a single environment. Given this mismatch between the scale from which tree genotypes are collected and the experimental setting, there is a high probability that the importance of tree genetic variation in shaping fungal community composition is overestimated. When tree genotypes in the study were reflective of the genetic variation at the landscape scale (Bálint et al. 2015, III), tree genotype was shown to play a minor role compared to elevated temperatures in shaping foliar fungal communities. The effect of tree genotype on the foliar fungal community may also depend on climate (Singh et al. 2018, Li et al. 2018). For example, an interactive effect between tree genotype and warming indicates that warming influences the fungal community, but differently for each tree genotype.

1.3.4. Natural enemies While climate and host genetic identity were show to play important roles in driving the structure of fungal communities, the impact of biotic interactions in governing fungi remains unclear. Specifically, the distribution of fungi can be explained by their interactions with natural enemies. For example, in nature plant pathogens are themselves a target for other fungi, which are often referred to as hyperparasites. Hyperparasites can reduce pathogen population growth, reproduction and overwintering success, and consequently affect disease dynamics (Parratt and Laine 2016). Despite the use of hyperparasites as biological control measures in agricultural systems, the role of hyperparasites in shaping the distribution of plant pathogens in the wild has rarely been investigated (but see e.g. Tollenaere et al. 2014 and II).

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1.4. Oak-associated fungi and insects as model organisms My thesis is based on a specific model system: the oak Quercus robur and its associated fungal and insect communities. Below, I provide information on each organism or group of organisms studied in the thesis.

1.4.1. Oaks

The pedunculate oak (Quercus robur) is a long-lived deciduous tree, which is widely distributed across Europe and reaches its northernmost limit in Sweden, and in southern parts of Norway and Finland (Lindbladh and Foster 2010). Oaks provide habitat for a wide range of generalist and specialist fungi and plant-feeding insects. For example, oak leaves are often attacked by specialised fungal pathogen from the genus Erysiphe and sap sucking insect, aphid Tuberculatus annulatus (Marçais and Desprez-Loustau 2014, Avila et al. 2014, Gross et al. 2021).

1.4.2 Fungal communities Like the foliage of any tree, the leaves of the pedunculate oak host many fungal species, both on their surface (epiphytes) and within and between cells (endophytes) (Jumpponen and Jones 2010, Cordier et al. 2012). Oak leaves are dominated by the fungal phylum Ascomycota, with the genera Aureobasidium, Cladosporium and Taphrina being particularly abundant. While Cryptococcus, Filobasidium and Sporobolomyces are the most abundant genera from the phylum Basidiomycota. These fungal genera play important roles in processes like nutrient cycling and the decomposition of senescent leaves and young litter (Cordier et al. 2012, Fort et al. 2016, Vacher et al. 2016). Oak leaves are often attacked by fungal pathogens, among which powdery mildew species from the genus Erysiphe are particularly well studied (Box 1) (Desprez-Loustau et al. 2011, Marçais and Desprez-Loustau 2014). The fungal community in oak roots and soil contains, among others, several ectomycorrhizal (e.g. Scleroderma, Tuber, Thelephora and Cenococcum), putative pathogenic (e.g. Cylindrocarpon and Armillaria) and saprotrophic genera (e.g. Alternaria and Epicoccum). 11 Kappa

Box 1. Oak powdery mildew species and their antagonists In Europe, Quercus robur is a host to three specialist powdery mildew species from the genus Erysiphe: E. hypophylla, E. alphitoides and E. quercicola. The three species cannot be distinguished in the field, and microscopic differences are minor. The two species, E. alphitoides and E. quercicola, are most commonly found on the upper leaf surface (but occasionally also found on the lower leaf surface), whereas E. hypophylla is mostly restricted to the lower surface. The mycelia of these fungal pathogens grows on the leaf surface, and only their feeding organs (haustoria) penetrate the epidermal cells (Bushnell 1972). Infection of oaks in spring starts through sexual spores (ascospores) that are released by overwintering fruiting bodies (chasmothecia) or by mycelia that overwinter in the buds (Desprez-Loustau et al. 2010). During the growing season, powdery mildew produces asexual spores (conidia). Powdery mildew has multiple generations during the growing season. The mycelium and asexual spores colour the oak leaves white, which is especially apparent in summer when the pathogen is most abundant. There is also a fourth powdery mildew species, Phyllactinia guttatta, but it is relatively rare (Desprez-Loustau et al. 2011). Powdery mildew species are frequently attacked by fungal hyperparasites from the genus Ampelomyces. Hyperparasites grow within the mycelium of the powdery mildew and disperse via conidia (Kiss 1998, Kiss et al. 2004, Németh et al. 2019). When the conidium of the hyperparasite lands on the powdery mildew mycelium, it will penetrate the host mycelium. Asexual fruiting bodies of Ampelomyces, called pycnidia, are produced in the hyphae, conidiophores and immature chasmothecia of their fungal hosts. Several studies have revealed that Ampelomyces is a genetically diverse group, with four major clades (putative species) (Kiss 1997, Liang et al. 2007, Park et al. 2010)

1.4.3. Aphids Among the many plant-feeding insects associated with oak, the aphid T. annulatus has received relatively little attention. T. annulatus has multiple asexual generations during the growing season, allowing for rapid population growth. Aphid densities usually peak in July, and after that, densities decline (Silva- Bohorquez 1987). Aphids overwinter as eggs and hatch at the start of the next growing season.

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2. Aims of the thesis

The overarching aim of my thesis was to identify and quantify the forces structuring fungal communities of oak (Quercus robur) in space and time (Figure 1).

In chapter I, I investigated the local and spatial factors that shape the fungal communities in the leaves and soil, and whether there was a relationship between below- and aboveground fungal communities (Figure 1a, c).

In chapter II, I investigated the role of climate, host characteristics and natural enemies in defining the niches of three cryptic species within the oak powdery mildew complex at multiple spatial scales (Figure 1a, c).

In chapter III, I used a two-factorial heating experiment to disentangle the role of warming, tree genotype and their interaction in shaping foliar fungal communities across the full growing season (Figure 1b, c).

In chapter IV, I assessed the impact of warming, tree genotype and their interaction on the spring and autumn phenology of oak trees, the seasonal dynamics of a fungal pathogen, and the density of a sap-sucking insect (Figure 1b, c).

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Figure 1. Conceptual framework of my PhD thesis. Chapter I and II focused on spatial patterns of fungal communities (Panel a), whereas in chapter III I also investigated their temporal dynamics (Panel b). I used both observational and experimental approaches to determine what factors drive the observed spatial and temporal patterns. For this, I focused my studies on the role of climate, host genotype, host characteristics and species interactions in influencing the spatial and temporal ecology of fungal communities on oak (I, II, III, IV; Panel c). Note that several factors may interact, and, in my thesis, I mainly focussed on the interactive effect of warming and host genotype (III, IV).

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3. Methods

To answer the questions posed in my thesis, I used a combination of observational studies to examine the distribution and potential forces structuring foliar and soil fungal communities in nature. I also used experimental approach to pinpoint causal relationships and assess the relative importance of different abiotic and biotic forces in shaping fungal communities. I utilized molecular methods, i.e. metabarcoding, to uncover the diversity and composition of foliar and soil fungi.

3.1. Observational data

In chapter I, I investigated the distribution and potential drivers of the fungal community associated with oak leaves and soil under natural conditions. The study was conducted on the island Wattkast in southwestern Finland. To investigate the spatial structure of fungal communities, I sampled leaves and soil of Quercus robur across the island and described foliar and soil community composition using DNA metabarcoding. I characterised microclimatic conditions around sampled trees by measuring temperature and relative humidity in the tree canopy and temperature in the soil. I also scored leaf discoloration, to understand the impact of tree autumn phenology on foliar fungal communities. In chapter II, I investigated the distribution of cryptic pathogen species within the oak powdery mildew complex and identified the role of climate, host characteristics and species interactions in shaping the coexistence of the powdery mildew species. To tease apart the role of each factor in defining the niches of the cryptic species, I used well-replicated sampling at the continental (Europe), national (France, Sweden) and landscape scales (the island Wattkast in Finland) (Figure 1a). I used molecular methods to distinguish species within the cryptic species complex and to identify the presence of the fungal hyperparasite (Ampelomyces) on powdery mildew (Box 1). To relate the distribution of the cryptic powdery mildew species and Ampelomyces to the spatial variation in climate in Europe, Sweden and France, I extracted temperature and precipitation data from WorldClim database. At the continental scale, I took advantage of a previous sampling campaign of cryptic powdery mildew species on oak trees throughout Europe (Desprez-Loustau et al. 2018) and used the same samples to detect the presence of Ampelomyces (Box 1). At the national scale, I assessed how climate, oak phenology and the hyperparasite influenced the distribution of the cryptic

15 Kappa powdery mildew species. In Sweden, I used leaf samples collected by schoolchildren as part of citizen science project throughout the range of Q. robur, and scored tree autumn phenology for a subset of sampled trees. In France, I used the data from a previous multi-year survey of cryptic mildew species on oak samplings, to investigate the role of climate in explaining the distribution of the hyperparasite. Finally, at the landscape scale, I investigated the impact of climate and oak phenology on cryptic powdery mildew species (E. alphitoides and E. hypophylla) and their hyperparasite on the island Wattkast. Since E. alphitoides is the only powdery mildew species that occurs on the upper leaf surface, we estimated its disease intensity visually by recording the infection percentage on the upper leaf surface in several consecutive years. To identify the other cryptic species, present within this landscape, E. hypophylla, we used molecular tools. I measured microclimatic data at the tree level using dataloggers, thereby recording temperature and relative humidity in the canopy for several years.

3.2. Heating experiment

During my PhD, I conducted a two-factorial field-heating experiment, to investigate the mechanisms and relative importance of different abiotic and biotic forces in shaping fungal communities (Figure 2). More specifically, I used small oak trees (ca. 1.2 m in height) that were grafted from five large mother trees (henceforth referred to as ‘genotypes’). The mother trees were randomly selected from the island Wattkast. For the experiment, six cages were built using wooden frames, and covered with mesh net to exclude insect predators. We increased the temperature in three of the cages by ca. 2°C above ambient temperature. The heating lasted for half a year, starting several weeks before bud burst and continuing until leaf drop. We used three ceramic heaters (2000 W, 240 V) placed at 120-degree angles to each other (Kimball 2005), in each of the three heated cages, to maintain elevated temperatures. The experiment was run in a pasture close to the city of Uppsala (north of Stockholm) (Figure 2). Within each of the six cages, we had trees belonging to the five genotypes (Figure 2).

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In chapter III, I was interested in the role of climate warming and tree genotype in shaping foliar fungal communities across the full growing season. To answer the questions of the study, I collected leaves from each tree in the experiment at the start, middle and end of the growing season and used DNA metabarcoding to characterise foliar fungal diversity and community composition during the season. In chapter IV, I focused on the role of climate warming and tree genotype in impacting spring and autumn phenology of the trees, the disease levels of one of the powdery mildew species, E. alphitoides, and the density of the aphid species T. annulatus. I started by scoring spring phenology of each tree in the experiment in the early season and continued with scoring autumn phenology in the late season. More specifically, I measured chlorophyll concentration, scored leaf discoloration and estimated the percentage of leaves that dropped from each tree. To detect the impact of warming and tree genotype on powdery mildew disease levels and insect density, I recorded the incidence and severity of the powdery, as well as the number of the aphids, on each tree.

Figure 2. In chapters III and IV, I investigated the effect of warming and tree genetic variation on foliar fungal communities, plant phenology and plant attackers. The photo to the left shows the setting of the heating experiment, where oak trees in cages 1, 3 and 6 were exposed to heating, while trees in cages 2, 4 and 5 were under ambient temperatures. The photo to the right shows young leaves of an oak tree in the experiment.

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3.3. Molecular methods 3.3.1. Metabarcoding

Research on fungal community ecology has long been hampered by the fact that fungi are harder to identify than plants or animals. The fact that fungal species could not readily be identified and quantified has prevented many ecologists from describing the patterns in fugal communities, let alone to apply ecological theories. The development of the field of fungal community ecology became possible only after the rise of molecular methods, and in particular metabarcoding (Peay et al. 2008, Nilsson et al. 2019). Fungal diversity can now be studied by sequencing genetic markers such as the internal transcribed spacer (ITS) region, also known as the primary fungal DNA barcode (Schoch et al. 2012). Since the ITS region is relatively long (500-700 bases), most studies use subregions within the ITS, either ITS1 or ITS2, which are shorter in length (e.g. 150-400 bases) (Nilsson et al. 2019). In my studies, I targeted the ITS2 subregion of fungi, as the use of this region results in less taxonomic bias than ITS1 (Tedersoo et al. 2015). Typically, a metabarcoding study includes several steps, that is, sample collection, extraction of the DNA from the collected samples, polymerase chain reaction (PCR) with target specific primers, DNA sequencing, sequence processing (e.g. clustering and cleaning) and data analysis. In Box 2, I focus on the methods that I used to characterise fungal communities in chapters I and III.

3.3.2. Identification of cryptic powdery mildew species and their hyperparasite The nucleotide diversity of the ITS region has also proven to be very useful for species identification within the powdery mildew species complex (Takamatsu et al. 2007, Mougou et al. 2008, Desprez-Loustau et al. 2018). The molecular identification of Erysiphe species is based on polymorphisms in the ITS region, as developed by Mougou et al. (2008). This protocol was followed in chapter II, in order to identify closely-related species within the oak powdery mildew species complex. In brief, DNA was extracted from the leaf discs, followed by PCR amplification using the ITS1 subregion and o-micro-rev primers (Heuser and Zimmer 2002). Sequencing was done by Sanger technology. Sequences were aligned with BioEdit and Erysiphe species were identified using six fixed single nucleotide polymorphisms (SNPs) (Takamatsu et al. 2007, Mougou et al. 2008).

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Box 2. Mains steps in fungal metabarcoding For my projects, I sequenced both leaves and soil fungi (1). For DNA extraction step (2) leaves and soil samples were grinded first. During PCR step (3), I used target specific primers, fITS7 and ITS4, with added tags and handles. After PCR amplification, I pooled my samples and sent them for sequencing (4). In chapter I of my thesis, sequencing was done using the Illumina platform, while in chapter III, samples were sequenced on the PacBio platform. Both methods were shown to have its advantages and disadvantages. For example, Illumina platform offers full ITS2 fungal sequences for most taxa and allows for high sequencing depth, however the large length variation of the fungal ITS region makes this sequencing platform to over-represent shorter fragments. PacBio, on the other hand, produces much longer reads than Illumina, but the sequencing depth (i.e., the number of sequences produced per sample) is lower compared to Illumina (Castaño et al. 2020). After sequencing, the received reads were cleaned, aligned and merged. Clustering of sequences into putative species operational taxonomic units (OTUs) is the most common approach in fungal metabarcoding (5). The ITS sequence clustering is done using different similarity thresholds, often between 97% and 98.5%. Once, the threshold is chosen and sequences are clustered in OTUS, the taxonomic annotation of OTUs is done through reference databases (6). In my studies, to assign taxonomy to the OTUs, I used the BLAST search tool for sequence comparison and the fungal reference database UNITE for taxonomic annotation (Pruitt et al. 2007, Abarenkov et al. 2010).

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We used the same leaves to detect the hyperparasite. The first primers for hyperparasite, Ampelomyces quisqualis, amplification were developed by Kiss et al. (2011), using the universal fungal primer ITS1F as a forward and a degenerate primer as a reverse, in a region showing high polymorphism between Ampelomyces quisqualis and related species. However, the first trials showed that this primer pair lacked specificity, since sequences closely matching related species were also amplified. Therefore, we used a slightly modified version of the method developed by Tollenaere et al. (2014) for hyperparasite identification in my studies.

3.3.3. Statistical approaches When modelling the univariate responses (e.g. fungal species richness or diversity), I used the framework of linear mixed models. Linear mixed models are an extension of simple linear models that allows modelling both fixed and random effects. When modelling binary data or proportions (e.g. presence-absences of species or disease levels), I frequently used generalized linear mixed models (or GLMMs), which allow response variables from different distributions, such as binary responses. For community data, for example, data containing presence-absences or abundances of each species in each sample, I used multivariate analysis, i.e. principal component analysis, redundancy analysis and Mantel test.

4. Insights 4.1. Differential imprint of local environment and spatial processes on foliar and soil fungal communities (I)

Molecular studies have demonstrated that fungal communities can be highly variable at the microscale and show distinct biogeographical patterns at the global scale (Peay et al. 2008, Zimmerman and Vitousek 2012, Cordier et al. 2012, Tedersoo et al. 2014). So far, the majority of such studies had focused on either foliar or soil fungal communities, and on mixed plant communities rather than single plant species (Mummey and Rillig 2008, Porras-Alfaro et al. 2011). Thus, a major question is how foliar and soil microbial communities are distributed within and among plant individuals of the same species and whether above- and belowground microbial communities respond to similar or distinct environmental forces. In chapter I, I sampled leaves and soil associated with a patchily distributed

20 Kappa oak species to disentangle the effect of local environment and spatial processes on the foliar and soil communities. I showed that the foliar fungal community was highly variable among leaves within trees, with only a minor part of the variation found among trees (Figure 3). The soil community was spatially clustered up to 50 m, whereas I found only weak evidence for spatial structure in the foliar community. Interestingly, microclimatic factors, oak autumn phenology and the spatial connectivity of oak trees did not explain the distribution of foliar and soil fungi. I also found that fungal communities in the leaves and soil were highly distinct, and there was no link between them. The findings of this chapter have several important implications, of which I would like to emphasize two. First, the large variation in foliar fungal community composition within trees suggests that we may need to shift our Figure 3. In chapter I, I demonstrate how foliar focus to finer spatial scales when community structure differs within and among describing the spatial structure of foliar trees in the landscape. The stacked bar charts fungi in nature. With most of the illustrate the percentage of variation in foliar fungal richness, evenness and community variation in foliar fungal communities composition found among leaves compared to found within individual tree canopies, between trees. fungi might primarily respond to differences in leaf surface temperature and humidity or the differences in chemical characteristics among leaves (Gripenberg 2007, Gripenberg et al. 2008, Cordier et al. 2012, Leff et al. 2015). Second, the finding of a stronger spatial structure of soil community than foliar community, suggests that soil fungi can be dispersal limited. However, abiotic and biotic forces that we did not explore in this study, e.g. species interactions or stochastic processes could, also drive the observed spatial patterning in the soil.

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4.2. Climate is more important than species interactions in defining the niches of cryptic powdery mildew species (II)

In chapter II, I narrowed my focus from the broad fungal community to a specific group of fungi, namely oak pathogens from the genus Erysiphe. As mentioned earlier, there are three closely related and nearly indistinguishable species within the oak powdery mildew species complex. The cryptic nature of these oak powdery mildew species raises an intriguing question: How can species that are nearly indistinguishable coexist? In this chapter, I explored different dimensions of the ecological niche that might explain their coexistence at different spatial scales. The ecological niche of cryptic species can differ due to different interactions with hosts and natural enemies, but also due to different responses of cryptic species to climatic conditions (Grinnell 1917, Elton 1927, Gravel et al. 2019). Knowledge of the drivers of the distribution of cryptic species within pathogen complexes is crucial for predicting the spread of existing and emerging diseases in a changing climate (Chenuil et al. 2019, Byrne et al. 2019). Previously, Desprez-Loustau et al. (2018) and Marçais et al. (2017) demonstrated that cryptic powdery mildew species differed in their response to climate at the continental and national scale, in France. In Chapter II, I showed that at the national scale, in Sweden, two cryptic powdery mildew species, E. alphitoides and E. hypohylla, were unaffected by climate and oak autumn phenology, and were attacked to a similar degree by the hyperparasite. When scaling down to the landscape level, I found that one of the powdery mildew species, E. alphitoides, but not E. hypophylla, was negatively affected by temperature in two out of four years. At the same time, I found no evidence of niche differentiation when looking at host characteristics (autumn phenology) and hyperparasitism. While we lack comparable data on the relationship between hyperparasitism and cryptic species, some studies have looked at the degree of specialization of Ampelomyces on powdery mildew strains and showed that Ampelomyces collected from a single powdery mildew species can infect other mildew species, indicating that Ampelomyces largely lacks host specificity (Kiss et al. 2004, Sucharzewska et al. 2011, Tollenaere et al. 2014). Overall, I showed that species belonging to the same cryptic pathogen complex show some degree of climatic niche separation. I also found no evidence of niche differentiation when targeting host tree autumn phenology and hyperparasitism. Obviously, both climatic and trophic dimensions of the niches are

22 Kappa highly complex and other drivers not investigated in this study may be important for niche separation of the cryptic powdery mildew species.

4.3. Climate warming and to lesser extent tree genotype shape foliar fungal communities (III) In addition to climatic factors and species interactions (I, II), the distribution and diversity of fungal communities can be influenced by plant species identity and plant genetic variation. A number of studies have demonstrated an important role of plant intraspecific genetic variation in structuring natural communities, including fungi (Martins et al. 2016, Wagner et al. 2016, Hamonts et al. 2018). The importance of plant genotype might also depend on local environmental conditions such as temperature (Bálint et al. 2015, Randriamanana et al. 2015). Importantly, the role of plant genotype and temperature in influencing foliar fungal communities may shift across the growing season, thus shaping the seasonal community dynamics. In chapter III, I compared the relative importance of warming and plant genotype in influencing the foliar fungal community across the full growing season (Figure 2). I demonstrated a strong seasonal change in foliar fungal community composition, with the seasonal change largely explained by a shift in fungal functional guilds. For example, basidiomycetous yeasts gradually replaced ascomycetous endophytes and pathogens towards the end of the growing season. I also showed that the seasonal dynamics of foliar fungal communities was mainly affected by warming and only to a minor degree by tree genotype and warming- by-genotype interactions (Figure 4). (a) (b) (c)

Figure 4. In chapter III, I demonstrate the role of warming and plant genotype in shaping the foliar fungal community across the full growing season. This figure shows dissimilarities in fungal community composition under elevated and ambient temperatures in early (Panel a), mid- and late season (Panels b and c, respectively), revealed by Principal Coordinates Analysis (PCoA, also known as metric multidimensional scaling).

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Overall, I provided experimental insights into the impact of warming and plant genetic variation on the ecology and evolution of leaf fungal communities and their hosts under a changing climate. From a more applied perspective, deciphering the interactions between plant genotypes and foliar fungal communities are prerequisites for managing these interactions, for instance, in terms of disease resistance and plant health.

4.4. Climate warming and tree genotype jointly affect tree phenology, fungal disease levels and insect density (IV) Climate warming has been shown to advance the timing of spring phenology of many plant species (Fu et al. 2015, Vitasse et al. 2018, Roslin et al. 2021). At the same time, genotypes of the same plant species can differ strongly in their phenology (Crawley and Akhteruzzaman 1988, Lutter et al. 2016). Yet, we lack an understanding of how climate warming and intraspecific genetic variation jointly shape the phenology of the host plant and the dynamics of plant attackers. In chapter IV, I used a heating experiment to investigate the role of climate warming and tree genotype in shaping host phenology, in both spring and autumn, as well as the population dynamics of the fungal pathogen Erysiphe alphitoides and the aphid Tuberculatus annulatus. I found that warming advanced spring phenology and delayed autumn phenology. The delay in leaf senescence under elevated temperatures matches the findings of other observational and experimental studies (Vitasse et al. 2009, Liu et al. 2016). Importantly, while warming advanced bud burst and delayed leaf senescence, it also extended the leaf longevity. Thus, the changes in the timing of the spring bud burst and autumn leaf senescence can have significant implications for the length of the growing season, and hence for the exposure of plants to early autumn frost (Liu et al. 2018) or changes in the carbon uptake (Keenan et al. 2014). Interestingly, for spring phenology, the response to warming differed among the tree genotypes at some of the dates. In autumn, the timing of leaf senescence was mainly affected by tree genotype, whereas warming substantially delayed leaf senescence later in the season (Figure 5a). The strong effect of tree genotype and genotype-by-warming interactions on the timing of spring and autumn phenology indicates a potential for natural selection to act on oak phenology, which may lead to adaptation of oak trees to climate change (Vitasse et al. 2013, Hänninen 2016). As for the plant attackers, trees exposed to elevated temperatures had similar

24 Kappa disease pressure during the onset of the growing season, but during the peak of the epidemic, disease levels were influenced by a weak interactive effect of temperature and tree genotype (Figure 5b). Aphid density was higher in the heated cages, and also differed among trees genotypes at the end of the growing season (Figure 5c). Several complex mechanisms can explain the observed effect of warming and tree genotype on plant attackers in my study: warming can either directly affect fungal and insect physiology and phenology or indirectly. For example, the warming-induced changes in plant traits consequently influence the performance of plant attackers (Dostálek et al. 2020). Taken together, the findings of this study highlight that climate change and tree genetic variation may influence the timing of spring and autumn phenology. These findings also imply that climate change and tree genetic identity can jointly affect the interactions of trees with their natural attackers. Thus, to predict how climate change will affect the phenology of the host plant and the dynamics of diseases and herbivores at higher trophic levels, it is important to take into account both the independent and interactive effects of temperature and plant genetic variation.

Figure 5. In chapter IV, I show the independent and interactive effects of warming and plant genotype on oak autumn phenology (a), powdery mildew disease levels (b) and aphid density (c). When lines cross or differ significantly in slopes it indicates an interactive effect: in this case, I show that warming may have a different effect on disease levels and insect density on different tree genotypes.

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5. Concluding remarks and future directions In my thesis, I systematically dissect the imprint of different abiotic and biotic forces on the spatial patterns and temporal dynamics of foliar and soil fungal communities associated with oak. Through my work, I demonstrated that foliar and soil fungal communities associated with the same host plant can be structured by different community assembly processes (Figure 6a). When focusing on a complex of cryptic powdery mildew species, I showed that that the climatic dimension is more important than the species interaction dimension for niche differentiation of these fungal pathogens (Figure 6b). When experimentally teasing apart the role of climate and tree genotype in influencing the temporal dynamics of foliar fungal communities, I demonstrated a strong effect of warming and a weak effect of tree genotype in shaping the seasonal trajectory of foliar fungi (Figure 6c). In terms of host responses to changing climate, I showed a strong effect of tree genotype and genotype-by-warming interactions on the timing of spring and autumn phenology (Figure 6d), indicating a potential for natural selection to act on oak phenology, which may lead to adaptation of oak trees to climate change. Taken together, my findings suggest that abiotic forces, in particular warmer temperatures, can prevail over biotic forces in structuring fungal communities on trees. While my thesis provides insights into the forces shaping tree-associated fungal communities, it also leaves many questions open for future work. Several future directions are identified in the individual chapters, but I would here like to emphasize three topics that I personally find particularly interesting and novel, and may consider exploring in future. As a part of chapter II, I investigated – among many other things – the role of hyperparasites in defining the niches of cryptic powdery mildew species. While my study suggests that the hyperparasite plays no role in niche differentiation of the cryptic powdery mildew species, the hyperparasite may still be important for the dynamics of the powdery mildew species. Maybe the powdery mildew is absent from some regions, because the hyperparasite thrives there, and readily leads its host to extinction? Does the hyperparasite have the potential to regulate the density of the powdery mildew on some tree individuals or under some environmental conditions? To answer these questions, we need experiments. In particular, I would be interested in an experimental study that investigates the role of hyperparasitism in shaping the population dynamics of oak powdery mildew species among neighbouring trees.

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While I demonstrate in my thesis that foliar fungal communities can be affected by a range of biotic and abiotic drivers (I, II, III), it remains unclear what role interactions between microorganisms play in fungal community structure and how these microbe-microbe interactions are affected by climate warming. To be able to answer these questions, we first need to identify the most important microbes and microbe-microbe interactions within natural fungal communities. We may use network analysis to identify microbial “hubs”— highly connected microorganisms in interaction networks, which exert a considerable influence on microbiome structure and functioning irrespective of their abundance across space and time. Once we have identified these ecologically relevant “hub” microbes, we can conduct experiments to investigate the relative importance of abiotic and biotic factors in shaping fungal communities in the presence and absence of “hub” fungi. Such experiment would shed light on the relevance of keystone taxa for the composition and functioning of the microbiome, and subsequently their role in orchestrating plant–fungal interactions. Fungal communities on oak leaves share their habitat with a diverse insect community, including aphids, caterpillars, leaf miners and gallers, as well as natural enemies such as parasitoids. It turns out that not only plants are full of microorganisms, but so are insects. In insects, we refer to the microorganisms as endosymbionts. During my PhD, I have tried to characterise the diversity of endosymbionts in the insect community, and to identify the drivers of these insect- associated microbial communities. Given that insects can acquire their microorganisms from the host plant they feed on, changes in the plant microbiome can potentially alter the insect microbiome and consequently the performance of plant-feeding insects. Yet, in the end, time proved short. At the time of writing, the DNA sequences from the endosymbionts are still hidden in a series of DNA libraries submitted for sequencing.

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Figure 6. A schematic depiction of the main conclusions of the thesis. (a) The aboveground fungal community was highly variable among leaves within trees, whereas the belowground fungal community was spatially clustered at distances up to 50 meters. In this observational study, I found no imprint of temperature, relative humidity and autumn phenology on the above- and belowground fungal communities. (b) When focusing on a cryptic pathogen complex on oak, I found some degree of climatic niche separation, while autumn phenology and hyperparasitism did not contribute to niche differentiation. (c) Fungal species richness increased and community composition shifted during the season. Experimental climate warming had a major impact on the seasonal trajectory of the foliar fungal community, whereas tree genotype explained little of the variation in foliar fungal community composition. (d) When investigating the effect of experimental warming and tree genotype on tree phenology, I detected a strong interactive effect (indicated here with a cross) between warming and tree genotype on spring phenology, and a strong independent effect of warming on autumn phenology. Solid brown arrows represent significant relationships, while dashed brown arrows indicate non-significant relationships.

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6. Svensk sammanfattning

Växter är värdar för artrika mikroorganismsamhällen som inkluderar svampar (eng: fungi), bakterier och arkéer. Svampar är en mycket artrik grupp inom dessa samhällen och är kända för att spela en avgörande roll för växters hälsa samt för att reglera viktiga ekosystemfunktioner. Trots detta saknar vi fortfarande övergripande kunskaper om vilka faktorer som strukturerar växtassocierade svampsamhällen i tid och rum. Det övergripande målet med denna avhandling var att komma fram till vilka faktorer som påverkar rumsliga mönster och dynamik över tid hos växtassocierade svampsamhällen. I detta syfte fokuserade jag på trädarten ek (Quercus robur) och dess svampsamhällen. Genom att använda en kombination av observations- och experimentstudier utvärderade) utbredningen jag i av ekassocierade svampsamhällen i landskapet, ovan och under jord, samt vilka faktorer som påverkar dessa utbredningsmönster; ii) klimatets samt olika trofinivåers roll i att definiera olika kryptiska arters nischer inom ett svampartkomplex associerat med ek samt iii) den relativa betydelsen av uppvärmning, växtindividens genotyp och interaktionen där emellan för att förklara ekens fenologi och den säsongsberoende dynamiken hos svamp- och insektssamhällen associerade med ek. Genom dessa studier fann jag att svampsamhällena ovan jord varierade stort mellan olika löv på enskilda träd. Vidare fann jag skillnader i utbredningsmönster hos svampsamhällen ovan och under jord, samt att svampsamhällena under jord var mer tydligt rumsligt strukturerade än de ovan jord. Trots dessa skillnader fann jag att varken klimat eller värdträdens fenologi och utbredningsmönster kunde förklara rumsliga mönster hos de olika svampsamhällena. I studien där jag fokuserade på tre kryptiska arter av mjöldagg (eng: powdery mildew) inom ett artkomplex av för eken sjukdomsframkallande arter, kunde jag visa att klimatfaktorer var viktigare än interaktionerna mellan arterna för att definiera deras olika nischer. I en experimentell fältstudie där jag på konstgjord väg värmde upp vissa ekindivider kunde jag visa att lövverkens svampsamhällen uppvisade stora skillnader över säsongerna samt att dessa skillnader påverkades av den experimentella uppvärmningen. Detta experiment visade även att uppvärmning och växtindividers genotyp gemensamt påverkar ekens fenologi samt förekomsten av sjukdomar och insektsangrepp över växtsäsongen.

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Sammanfattningsvis tyder mina resultat på att abiotiska faktorer kan dominera över biotiska faktorer när det gäller att strukturera rumsliga mönster och temporal dynamik hos växtassocierade svampsamhällen. Den starka påverkan varmare temperaturer hade på lövverkens svampar indikerar att den globala uppvärmningen har potential att omstrukturera dessa svampsamhällen, vilket i sin tur kan få viktiga konsekvenser för ekars hälsa, för interaktioner mellan ekar och andra organismer samt för olika ekosystemfunktioner.

Ordförklaringar Abiotiska och biotiska faktorer – organismer påverkas av olika faktorer i miljöerna där de lever. Dessa miljöfaktorer kan vara antingen abiotiska (icke levande) eller biotiska (levande). Exempel på abiotiska faktorerna är: solljus, fuktighet, jordmån, temperatur, vind och bränder. Biotiska faktorer är andra levande organismer och interaktionerna dem emellan. Arkéer – encelliga organismer som utgör en av de tre grundläggande indelningarna (domänerna) av levande organismer på Jorden. De andra två domänerna utgörs av bakterier och eukaryoter (ex. svampar, växter och djur). Artkomplex – en grupp närbesläktade organismer som är så lika att det uppstår oklarheter när det kommer till att artbegränsa dem. Fenologi – tidpunkten för olika händelser i en arts livscykel, t.ex. lövknoppning, blomning eller fågelmigration. Genotyp – en individs specifika genetiska egenskaper. Kryptiska arter – arter som är så lika varandra att de inte går att skilja utan genetiska analyser. Mjökdagg – sjukdom på ek förorsakad av en svampparasit. Svamparnas mycel och sporer färgar ekens löv vita vilket syns tydligt om sommaren när parasiten är som vanligast. Nisch – en arts nisch är dess roll i det organismsamhälle den lever i. Nischen begränsas av många faktorer, dels av vilka miljöfaktorer arten kan tolerera men också genom interaktioner med andra arter. Trofisk nivå – den position en art intar i en näringskedja.

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7. Acknowledgements

Thank you to all the trees and their tiny microorganisms. The last four and a half years has been a great, challenging at times, but such a great journey. During this journey, I was extremely lucky to meet many extraordinary colleagues, collaborators and co-authors. I will most probably forget someone and for that, I am sorry, but you should know that I am grateful for each conversation, collaboration and discussion I had with all of you.

To people in Stockholm,

Ayco, I was so lucky to have you as a supervisor! Thank you for introducing me into the wonderful world of microbes and giving the most intriguing PhD project (not that I knew it from the start ;)). I have learnt a ton, or actually a megaton, from you during these years and for that I am grateful. I really hope that at least a tiny bit of your beautiful writing style, time management skills and fascination in natural enemies, I will carry with me for a long time. P.S. I hope you caught my indent joke 

Big thanks goes also to my co-supervisor, Peter. Thank you for always being available for spontaneous discussions. I really appreciate all the instant feedback and spot on comments you gave me on my projects and manuscripts. Pil, I will be eternally grateful for all the support during the past years. I am so happy we became friends and I look forward to all the adventures we will share. I hope you know that you are always close to my heart 

Ditte and Laura, well what is there to say? Your friendship has carried me through PhD. I always enjoyed our numerous scientific/R/political/personal discussions. I admire you for being such wonderful persons and scientists. I am grateful for all the fun moments we had together (some documented on photos that we will never share ;)). My only wish is to be your friend forever  Daan, I am happy I got to know you (really amazed by the coolest hobbies you have).

Line, I am lucky I became friends with you. I would like to thank you for all the time we spent together, including the discussions of global environmental issues and runs in the forest! I am sure you will be (are) an extraordinary scientist.

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Beate, my dear friend, you were an amazing office buddy: it was pretty lonely in the office without you this past year. I think we finally have all the time we want to catch up 

Ahmed, thank you so much for all the bioinfo help, knowledge you shared with me and being always ready to answer my stupid questions over WhatsApp. It was so great to have a microbial enthusiast in our group. Oak-coffee-anemone group: Alvaro, Beyene, Biruk and Emilia. Thank you for being great colleagues and all the fun we had shooting each other in different games.

And to all current and past members of Ecofika group: Matilda (thank you a lot for helping out with Swedish translation), Elsa, Sonia and Alicia (it’s such a pleasure knowing you, thank you for great time we spent together). Irena, Xuyue, David, Magda (thank you so much for the help with the lab and nice company). Ove, Kristoffer and Tanja for the great discussions. Lastly, I want to thank Johan and Sarahi for reviewing my thesis. Amanda and Peter B., I am glad I had the opportunity to learn about science communication from you - it was a great pleasure. Oskar, thank you for an awesome company in a PhD council board. Andreas N. for the great time we spent at Tovetorp and all the help with the molecular work. Andrea C., Aleksandra, Jörg, Leonard, Fede, Julie, Laura S., Robert, Maria S., Serena, Sven, Dimi and Ben thank you all for being so awesome PhD colleagues! PhD would be a very different experience without you all.

Thank you for the administrative staff at DEEP, particularly huge thanks goes to Erica and Kristina.

To people in Uppsala, a town that has been my go-to place for many months during the PhD, Adam, thank you for being such a great colleague. It was a pleasure to run the warming experiment with you. Tomas, I greatly appreciate all the time you spent reading and commenting on my manuscripts and project plans. I always admired your insightful and wise conceptual suggestions.

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To people in Kyiv,

Friends:

Julie (Volyanskaya), Tetyana (Mamula), Nastya (Buchkova), Tanya (Betsa) and Ketrin (Kotenok), our friendship means a lot to me, no matter how far away from each other we live! Friends will be friends! Thank you for being there for me during all these years and even showing some interest in the ‘boring’ life of a biologist, who is weird enough to find fascination in microorganisms on plants.

Family: My parents, I am so grateful you taught to be free, to be free in making the choices I wanted and to love with all my might. Мамочка и папочка, я очень вам благодарна за огромную поддержку. Именно благодаря вам, я знаю, что такое любовь и как это любить безоговорочно. Все мои успехи, в них огромная ваша заслуга. Я очень по вам скучаю. My brother, I will be eternally grateful to my parents for bringing you in my life. Even though we have a big age difference and I missed most of your teenage years (phew, lucky me), when I moved out, you are always in my thoughts. Тимур, я тебя так сильно при сильно люблю!

Igor, here it would be appropriate to say that “the best feelings in the world are the ones there are no words to describe” and I cannot agree more. You have been my greatest cheerleader during these past years and I only hope that once I could be as much support to you as you have been to me. You helped me in so many ways that even A5 will not be enough to list them all, but most importantly, you never said no to my crazy ideas (like sleeping in hammocks in the middle of the Amazonian jungle ). You are probably the only husband (of a biologist) in the world, who knows all the species of leaf miners and galls on oak. Crazy how great you are in finding the right words and warmest hugs to calm down your emotional wife. I want to thank you for always believing in me and for taking care of our favourite new member, Altai, while I was busy finishing up. I am grateful I can share my life with you and looking forward to new adventures it will bring us. «Я не маг, чтоб подглядывать в завтра, Но в мире, где светят и любят, Будущее - будто деталька, Что стрелку сдвигает секундную»

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8. References

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