COMPARATIVE ECOLOGY OF WILD COLUMBIFORMES NATIVE TO EUROPE An analysis of movement behaviour, diet composition and haemosporidian infections Yvonne R. Schumm Dissertation presented for the degree of Doctor rerum naturalium JUSTUS LIEBIG UNIVERSITY GIESSEN Faculty of Biology and Chemistry Giessen | May 2022 DOCTORAL THESIS / DISSERTATION Comparative ecology of wild Columbiformes native to Europe - An analysis of movement behaviour, diet composition and haemosporidian infections SUPERVISOR / BETREUERIN Prof. Dr. Petra Quillfeldt DEAN / DEKAN Prof. Dr. Thomas Wilke REVIEWERS / GUTACHTER Prof. Dr. Petra Quillfeldt Behavioural Ecology & Ecophysiology Group, Department of Animal Ecology & Systematics, Justus Liebig University Prof. Dr. Volkmar Wolters Animal Ecology Group, Department of Animal Ecology & Systematics, Justus Liebig University DECLARATION / SELBSTSTÄNDIGKEITSERKLÄRUNG “I declare that I have completed this dissertation single-handedly without the unauthorized help of a second party and only with the assistance acknowledged therein. I have appropriately acknowledged and cited all text passages that are derived verbatim from or are based on the content of published work of others, and all information relating to verbal communications. I consent to the use of an anti-plagiarism software to check my thesis. I have abided by the principles of good scientific conduct laid down in the charter of the Justus Liebig University Giessen „Satzung der Justus-Liebig-Universität Gießen zur Sicherung guter wissenschaftlicher Praxis“ in carrying out the investigations described in the dissertation.” Giessen, May 2022 ________________________________ Yvonne R. Schummxxxxxxx CONTENTS ABSTRACT…………………………………………………………..…………………. I ZUSAMMENFASSUNG……………………………………..………………………… II SYNTHESIS 1 | General introduction…………………………………………...…………... 1 2 | Aims and structure of the thesis………………………….……..………... 10 3 | Chapter outline…………………………………………….………..……… 12 4 | General conclusions and future outlook………………….……..……….. 16 5 | References……………………………………………..….………...……… 21 CHAPTERS Chapter 1 | Feather stable isotopes (δ2Hf and δ 13Cf) identify the Sub-Saharan wintering grounds of turtle doves from Europe……..…...….. 30 Chapter 2 | Year‑round spatial distribution and migration phenology of a rapidly declining trans‑Saharan migrant - evidence of winter movements and breeding site fidelity in European turtle doves.………….……….…….. 46 Chapter 3 | Should I stay or should I fly? Migration phenology, individual- based migration decision and seasonal changes in foraging behaviour of Common Woodpigeons.……..………………………………….…………….. 64 Chapter 4 | Diet composition of wild columbiform birds: Next-generation sequencing of plant and metazoan DNA in faecal samples……………….. 82 Chapter 5 | Prevalence and genetic diversity of avian haemosporidian parasites in wild bird species of the order Columbiformes.….…………..….. 106 APPENDIX Supplementary material………………………………………….…...……….. 123 Curriculum vitae………………………………...……………….……...……… 131 List of publications……………………………………………….……...……... 132 Acknowledgements …………………………………….……....……….…….. 134 Illustration credits……………………………..…...……………..….………… 135 x ABSTRACT Behaviour encompasses all interactions of an animal with other organisms and with its environment. Studying animal behaviour can provide important information for species conservation and management. In this thesis, I aimed at closing fundamental knowledge gaps on different behavioural-ecological aspects of the three migratory species of Columbiformes (order of doves and pigeons) native to Europe. The species studied include one of the most rapidly declining breeding birds in Europe, the European Turtle Dove (Streptopelia turtur). As a Palearctic-African migratory species, it is the only long-distance migrant among the European native wild Columbiformes. On the contrary, Common Woodpigeons (Columba palumbus) and Stock Doves (Columba oenas) are both short-distance migrants. They belong to the most common European breeding birds or have a stable population trend, respectively. All three species are regularly hunted in several European countries. In my thesis, I examined the wintering areas, migration flyways and phenology of European Turtle Doves by means of analysing feather-isotope values (chapter 1) and satellite tracking data (chapter 2). My findings highlight the use of several migration flyways and suggest that individuals breeding in different parts of Europe may occupy separate African wintering grounds. Tracking of Common Woodpigeons with GPS-GSM transmitters (chapter 3) showed that individuals breeding in Germany are facultative partial migrants, which can switch their migratory strategy (resident vs. migrant) between years. Tracking data evaluated along with land cover data indicated that both European Turtle Doves and Common Woodpigeons adapt their foraging areas and distances to the availability of food resources (chapters 2 and 3). A comparative assessment of diet components, analysed from faecal samples through DNA metabarcoding (chapter 4), revealed a variation in the presence and frequency of occurrence of diet items between the three species. Identified diet components were mainly seed-bearing plants (Spermatopsida). Furthermore, comparisons with previous studies suggest distinct regional intraspecific differences in diet composition. An evaluation of haemosporidian parasite infections (Plasmodium sp., Haemoproteus sp. and Leucocytozoon sp.) from avian blood samples (chapter 5) demonstrated interspecific differences in lineage diversity, overall and genus-specific prevalence. The observed infection pattern supported the assumption that long-distance migrants harbour a higher parasite prevalence and diversity compared to resident or short-distance migratory species. Summarising, the findings of this thesis (i) enable a better interpretation of different behaviours observed in the Columbiformes, (ii) help to comprehend how behavioural habits are influenced by ecological causation and (iii) emphasize the distinct behavioural diversity and plasticity present among species as well as individuals. Furthermore, the results contribute to a greater understanding of the general and specific ecological requirements of the species of Columbiformes. This can help to optimise present or to plan effective future management and conservation strategies that reconcile the challenges of game bird management on the one hand and species conservation on the other hand. - I - ZUSAMMENFASSUNG Verhalten umfasst alle Interaktionen eines Tieres mit anderen Organismen und mit seiner Umwelt. Die Erforschung des Verhaltens einer Tierart kann wichtige Informationen für das Management und den Schutz einer Art liefern. Ziel meiner Dissertation war es, grundlegende Wissenslücken hinsichtlich verschiedener verhaltensökologischer Aspekte von drei in Europa einheimischen und migrierenden Wildtaubenarten (Ordnung Columbiformes) zu schließen. Zu den untersuchten Arten zählte die Europäische Turteltaube (Streptopelia turtur), die eine der am stärksten rückläufigen Vogelarten in Europa ist. Europäische Turteltauben, welche in Subsahara-Afrika überwintern, sind die einzigen Langstreckenzieher unter den in Europa einheimischen Taubenarten. Dahingegen sind Ringeltauben (Columba palumbus), die in Europa zu den häufigsten Brutvögeln gehören, und Hohltauben (Columba oenas), die einen stabilen Bestandstrend aufweisen, Kurzstreckenzieher. Alle drei Wildtaubenarten gehören in mehreren europäischen Ländern zu den legal jagdbaren Vogelarten. Im Rahmen der Dissertation untersuchte ich die Überwinterungsgebiete, Zugrouten und Migrationsphänologie von Turteltauben anhand einer Analyse von Isotopensignaturen von Federn (Kapitel 1) und Satellitenortungsdaten (Kapitel 2). Die Ergebnisse zeigten auf, dass unterschiedliche Individuen verschiedene Zugwege nutzen und dass möglicherweise eine räumliche Trennung in den afrikanischen Überwinterungsgebieten zwischen Individuen aus unterschiedlichen Brutgebieten existiert. Das Tracking mit GPS-GSM-Sendern von in Deutschland brütenden Ringeltauben (Kapitel 3) ergab, dass einige Individuen ihre Zugstrategie (Standvogel oder Zugvogel) von Jahr zu Jahr ändern. Eine Auswertung der Trackingdaten gemeinsam mit Daten zur Landbedeckung deutete daraufhin, dass sowohl Turteltauben als auch Ringeltauben Habitatnutzung und Flugdistanzen während der Nahrungssuche an die Verfügbarkeit von Nahrungsressourcen anpassen (Kapitel 2 und 3). Eine vergleichende Betrachtung der Nahrungszusammensetzung der drei Wildtaubenarten ergab, dass das Vorhandensein sowie die Häufigkeit von Nahrungsbestandteilen zwischen den drei Arten variierten (Kapitel 4). Für die Ermittlung der Nahrungsbestandteile wurden Kotproben mittels DNA-Metabarcoding analysiert. Bei den identifizierten Nahrungsbestandteilen handelte es sich hauptsächlich um Samenpflanzen (Spermatopsida). Vergleiche mit früheren Studien deuteten auf regionale Unterschiede innerhalb einer Taubenart hinsichtlich der Zusammensetzung der Nahrung hin. Eine Auswertung von Infektionen mit Hämosporidien (Gattungen: Plasmodium sp., Haemoproteus sp. und Leucocytozoon sp.) in Blutproben verschiedener Wildtaubenarten zeigte interspezifische Unterschiede in der Diversität genetischer Linien sowie der allgemeinen und gattungsspezifischen Prävalenz der Blutparasiten auf (Kapitel 5). Das beobachtete Infektionsmuster stützte die Annahme, dass Langstreckenzieher verglichen mit Kurzstreckenziehern oder Standvögeln eine höhere Parasitenprävalenz und -vielfalt aufweisen. - II - Zusammenfassungx Die Ergebnisse der vorliegenden Dissertation, im Allgemeinen und zusammenfassend betrachtet, (i) ermöglichen eine verbesserte Interpretation des bei den Taubenarten beobachteten Verhaltens, (ii) helfen zu verstehen, wie bestimmte Verhaltensweisen durch ökologische Ursachen beeinflusst werden, und (iii) verdeutlichen die ausgeprägte Vielfalt und Plastizität im Verhalten sowohl der jeweiligen Taubenarten als auch der Individuen. Darüber hinaus tragen die Ergebnisse zu einem besseren Verständnis der allgemeinen und spezifischen ökologischen Anforderungen der Wildtaubenarten bei. Dieses bessere Verständnis kann dazu beitragen, effektive Management- und Artenschutzstrategien zu planen oder bereits etablierte Maßnahmen anzupassen, welche sowohl die Anforderungen des Managements jagdbarer Arten als auch des Artenschutzes berücksichtigen. - III - x 1 | GENERAL INTRODUCTION Why, how, where and when do organisms move? Studying the movements of animals is often motivated by its importance for many ecological patterns. Migratory and foraging movements can affect ecological processes including the spread of diseases, inter- and intraspecific competition and the ability of species to cope with global as well as local environmental changes (Nathan et al. 2008; Spiegel et al. 2016; Miller et al. 2019; Shaw 2020). Besides migration and foraging strategies themselves, these ecological processes can lead to differences in individual fitness and subsequently can influence population dynamics (Palacín et al. 2016; Dadam et al. 2019). Birds, constituting the most diverse lineage of living tetrapod vertebrates, belong to the most mobile organisms. These ubiquitously distributed organisms include around 10,900 extant species, which exhibit an extraordinary diversity in ecological, morphological and behavioural traits, thus occupying a variety of different ecological niches (Winkler & Leisler 1985, Gill 1995; Prum et al. 2015; Stiels & Schidelko 2018; Gill et al. 2021). 1.1 The order Columbiformes Pigeons and doves (Columbiformes) are one of the oldest and most diverse extant lineages of birds (Soares et al. 2016). In many cultures, pigeons and doves can be found as a symbol of gentleness, love, sacrifice, grace and peace, especially in religions and belief systems. Nevertheless, they currently are one of the most threatened avian orders. Of the 354 extant species of Columbiformes worldwide, 72 species (20%) are threatened with extinction and 47 species (13%) are categorized as ‘near threatened’ (Wood & Cerbini 2021). Despite this, the order has not received a level of attention proportionate to the number of threatened species, in particular considering the vital ecological roles of many Columbiformes, e.g. as keystone species in tropical forests, seed dispersers, or important prey items (Gibbs et al. 2001; Kissling et al. 2009; Santos et al. 2019; Panter & Amar 2021; Wood & Cerbini 2021). Notwithstanding their relatively conserved anatomy and morphology, Columbiformes display a wide range of variations in their ecological adaptations (Soares et al. 2016). Except for the polar regions, pigeons and doves are widespread over all continents, albeit in varying numbers of species and individuals. The greatest species richness occurs in the tropical zones, particularly in the Southeast Asian and Oceanic regions (Rösler 1996; Gibbs et al. 2001). 1.2 Species of the order Columbiformes native to Europe In contrast to the species-rich tropics, in Europe, besides the rock pigeon and domesticated feral pigeon (Columba livia and C. livia f. domestica, Gmelin 1789, respectively) as well as a few endemic species on the Canary Island, only four species of native wild Columbiformes occur (Gibbs et al. 2001; Gill et al. 2021). These are the Common Woodpigeon (Columba palumbus, Linnaeus 1758), the Eurasian Collared Dove (Streptopelia decaocto, Frivaldszky 1838), the European Turtle Dove (Streptopelia turtur, Linnaeus 1758) and the Stock Dove (Columba oenas, Linnaeus 1758). - 1 - General introductionx All four species are listed in Annex II of the Council Directive 79/409/EEC and therefore can be legally hunted. However, the species-specific protection status, hunting effort and regulations differ between European countries. This thesis has not considered the Eurasian Collared Dove as it only spread from its original distribution range in the Oriental region over the European continent throughout the 20th century (Gibbs et al. 2001; Rocha-Camarero & de Trucios 2002). In addition, it is a resident species in Europe, compared to the three other species, known to perform migratory movements (Gibbs et al. 2001; Baptista et al. 2015). Therefore, my thesis focuses on the remaining three species of wild Columbiformes native to Europe, namely the Common Woodpigeon, European Turtle Dove and Stock Dove. These species are briefly introduced in the following sections. 1.2.1 The Common Woodpigeon - From forest dweller to city resident The Common Woodpigeon (henceforth Woodpigeon) is widely distributed throughout Europe (Fig. 1). With a population size of around 41 to 57 million mature individuals, they are one of the most common breeding bird species in Europe (BirdLife International 2021). Notwithstanding its great abundance and despite being considered by some as an agricultural pest, which causes damage to different crops, and the most important European game bird species (Murton 1960; Slater et al. 2001; Höfle et al. 2004; Butkauskas et al. 2013), the Woodpigeon receives relatively little attention in general as well as in scientific research (König et al. 2015). It was once a typical woodland species, which inhabited deciduous, mixed and coniferous forests, but an increase in population size since the mid-20th century has resulted in an expansion in its breeding habitats to suburban and urban areas (Tomiałojć 1976; Slater et al. 2001; Bea et al. 2011). Thus, the Woodpigeon is classified as synanthropic species nowadays (Kurucz et al. 2021). Possibly due to their high adaptability Woodpigeons now occur in almost all natural and man-altered habitats and their European population continues to increase further (Gibbs et al. 2001; Bea et al. 2011; Sakhvon & Kövér 2020; PECBMS 2022). Figure 1 Maps of breeding and non-breeding areas, i.e. wintering areas, of Common Woodpigeons, European Turtle Doves and Stock Doves. Maps adapted from BirdLife International (2022). Orange arrows depict a simplified course of migration directions from breeding to wintering areas (adapted from Möckel 1988 for Stock Doves; Cohou et al. 2007 for Common Woodpigeons, Marx et al. 2016 for European Turtle Doves). - 2 - x 1.2.2 The European Turtle Dove - One of Europe’s most rapidly declining species Given its rather vast breeding range (Fig. 1), it can be assumed that the European Turtle Dove (henceforth Turtle Dove) occurs in multiple habitats and landscapes. Often the Turtle Dove is associated predominantly with farmland. However, research indicates that it occupies a more diverse range of habitats, often combining open ground areas, such as arable land or grassland, with hedgerows, trees, or small woodlands, i.e. an ecotone between agricultural areas and forest (reviewed in Carboneras et al. 2022). The Turtle Dove, a trans-Saharan migrant, is one of many migratory landbirds within the Afro-Palaearctic system that is declining (Carboneras et al. 2022). The European population has declined by almost 80% since 1980. Consequently, the species has been classified as globally threatened (‘vulnerable’; BirdLife International 2021). Nevertheless, the severity of the population decline varies between countries (Fig. 2). Despite the ubiquity of decline, the detailed causes remain uncertain, in particular, because certain pressures, such as food availability or predation pressure, affect different stages of the Turtle Dove life cycle, making it difficult to estimate their relative contribution to the overall decline (de Vries et al. 2022). The main causes suggested to contribute to the ongoing population decline are habitat loss and deterioration at breeding and wintering grounds associated with changes in agricultural practices, illegal killing and unsustainable levels of legal hunting (Fig. 2; Fisher et al. 2018; Lormée et al. 2020; Moreno-Zarate et al. 2021; Carboneras et al. 2022). Figure 2 Long-term breeding population trends of Turtle Doves per European country for which sufficient data is available. The population trend is reflected by the numbers given in the respective circle (‘-’ corresponds to decline [%] and ‘+’ to growth [%]). Yellow colour marks EU countries in which Turtle Doves can be legally hunted or in which hunting is generally permitted, but a temporary hunting ban exists (for hunting seasons 2020/21 and/or 2021/22). Modified figure according to FACE (2020). - 3 - General introductionx 1.2.3 The Stock Dove – The cavity-nesting columbiform species The Stock Dove occurs almost all over Europe (Fig. 1) and inhabits mainly agricultural land with plenty of trees, woodland edges and broad-leaved forests, especially old forest stands (Möckel 1988; Gibbs et al. 2001; Kosiński et al. 2011). In contrast to most other wild doves and pigeons, which usually construct an open nest by accumulating some twigs to a platform nest, Stock Doves are cavity-breeders (Möckel 1988; Gibbs et al. 2001). Therefore, the occurrence and density of Stock Doves depend on the number of available trees with suitable nest holes, such as holes excavated by Black Woodpeckers (Drycopus martius, Linnaeus 1758; Kosiński et al. 2011). Also, artificial nest boxes are accepted as nesting opportunities by Stock Doves (Möckel 1988; Møller et al. 2016). The European Stock Dove population declined at the beginning of the 20th century. This decline was attributed mainly to a lack of adequate breeding sites due to changes and modernisation in forestry management, such as the increase of spruces or loss of old broad- leaved forest stands (Hillerich 1984; Möckel 1988). Since around 1970, after increased protection efforts, the population has been stable and has even increased locally (Möckel 1988; Møller et al. 2016; BirdLife International 2022). Nowadays, Stock Doves are classified as ‘least concern’ in Europe and Germany (Ryslavy et al. 2020; BirdLife International 2021). 1.3 The relevance of behavioural studies in species conservation and management With animal species increasingly facing threats from changing climate, diseases, habitat loss and other pressures from human activities, accelerated rates of species decline and extinction risk are observed (Swift et al. 2018; Soroye et al. 2020; Sattar et al. 2021). Thus, for effective conservation and management, it is important to have accurate information about the status and dynamics of natural populations (Swift et al. 2018). The study of animal behaviour can provide a valuable contribution to solving conservation and management problems (Sutherland 1998; Buchholz 2007). Attempts to relate the behaviour of animals to conservation are increasing in the academic literature (Caro & Sherman 2013). To be more precise, the behaviour of an animal is relevant to conservation biology as behaviour can affect the persistence of species, e.g. by dispersal and settlement decisions or learned foraging techniques (Reed 2002). Moreover, it is stated that the study of animal behaviour is crucial for solving issues of coexistence between wild animals and humans (Blackwell et al. 2018), especially in the case of legally huntable species. Although the Turtle Dove is a rapidly declining species and the Woodpigeon is one of the most common species in Europe, large gaps in our knowledge regarding different behavioural aspects of these and related columbiform species exist. For instance, little is known about the migration patterns of the Turtle Dove (Marx et al. 2016), the dietary habits of the Stock Dove and Woodpigeon (Dunn et al. 2018), or the effects of diseases on mortality and fitness of columbiform host species (Fisher et al. 2018). In the presented thesis, I aimed to obtain a comparative and integrated picture of three behavioural-ecological aspects of the three aforementioned species of Columbiformes: movement, dietary and disease ecology. - 4 - x 1.3.1 Movement ecology – Migration and foraging behaviour The ability to fly makes birds one of the most mobile animals. Their movements occur across a wide range of spatiotemporal scales, extending from daily foraging movements, one-way dispersal movements to seasonal migratory movements (Thorup et al. 2017; Shaw et al. 2020). Migration can be defined as the regular movement of a population to and from a specific region, resulting in distantly separated breeding and wintering grounds, and is characterised by movement patterns that vary within a year but usually not between years (Teitelbaum & Mueller 2019). Many avian migrants demonstrate partial migratory behaviour, i.e. just a fraction of the population migrates whereas the remaining individuals reside in a single habitat for the entire year (Chapman et al. 2011). Factors that influence population size dynamics include events that occur over the full annual cycle and thus, in the case of migrants, events that occur at different localities. Identifying the relative importance of effects, carryover effects and relationships among events at different annual stages (breeding, wintering and migration) is fundamental to the effective conservation and management of species (Robinson et al. 2010; Sedinger et al. 2011; Calderòn et al. 2019). However, these data are rare for most species, often due to the inability to track individuals throughout their annual cycle, resulting in a lack of knowledge about where birds breed and winter (Robinson et al. 2010; González et al. 2020). Traditionally, mark-recapture techniques, such as marking individuals with leg rings and subsequent sighting or recapture, were used to acquire knowledge on migration flyways (Robinson et al. 2010; Negrier et al. 2020). Also, ratios of isotopes in tissue, genetic markers (reviewed in Robinson et al. 2010) or radar systems (Nilsson et al. 2018; Cui et al. 2020) were used to monitor migratory movements. Rapid advancements in tracking technologies, such as radiotelemetry or satellite transmitter systems, enabled the collection of positioning information on a wide variety of species remotely. With this progress, the movement of animals can be tracked at various spatial and temporal scales over long periods, often without the need for recapture (Robinson et al. 2010; Yoda 2019; Mitchell et al. 2019; Jetz et al. 2022). The diverse tracking techniques can be used to study various aspects of migration, e.g. orientation, navigation, phenology, flyways, stopover site selection, migratory connectivity and occurrence of niche tracking behaviour (Robinson et al. 2010; Illan et al. 2020). For example, flyways of Turtle Doves were initially identified based on ring-recoveries and sightings, suggesting three flyways between Europe and Sub-Saharan Africa: a Western, a Central and an Eastern one (Fig. 1; Marx et al. 2016). Later on, individual Turtle Doves were equipped with Argos satellite transmitters at their breeding grounds in France to analyse flyways, stopover and African wintering sites in the Sub-Saharan region in more detail (Lormee et al. 2016). Stock Doves and Woodpigeons are partial migrants. Both species can be roughly divided into individuals from Central and Eastern Europe, which are expected to perform short- distance migration within Europe, and individuals from Western Europe, which are supposed to be residents (Fig. 1; von Blotzheim & Bauer 1994). For instance, a tracking study of - 5 - General introductionx Woodpigeons equipped with Argos transmitters during the wintering period in Portugal showed that these individuals migrate to breeding grounds in Poland or Switzerland (Cohou 2011). Improving and overcoming deficiencies, like low resolution and accuracy, modern technology, such as high-resolution GPS tracking, provide the level of detail needed to identify and characterise fine-scale space use and movements. The high-frequency movement data enables to link movements with behaviours and thus gain greater insight into the behavioural ecology of species, such as foraging behaviour, representing one of the most important animal behavioural components (Yoda 2019; McDuie et al. 2019; Van Donk et al. 2020). For example, a radiotracking study suggested a memory-based strategy with a flocking mechanism to be the best fit for explaining observed foraging patterns in Woodpigeons (Kułakowska et al. 2014). Further studies like this may allow to precisely predict spatial and temporal characteristics of foraging habits of avian species like Woodpigeons, taking into account their excellent memory, ability to fly long foraging distances and distinctive flocking behaviour. A precise knowledge of foraging behaviour can be used to assess aspects relevant to species conservation, e.g. predictions of the exposure to pesticides based on known foraging characteristics (Kułakowska et al. 2014; de Montaigu & Goulson 2022). As another example, with leg-ring radio-tag attachments it was shown that in the first weeks after fledging, Turtle Doves forage mainly in the immediate vicinity (≈ 300 m) of their nest, highlighting food availability close to nest areas as a crucial factor for survival in juveniles (Dunn et al. 2016). 1.3.2 Dietary ecology - Reconstruction of diet composition Worldwide, avian communities inhabiting agroecosystems are threatened as a consequence of agricultural intensification (Crisol-Martínez et al. 2016). In Europe, the mid-20th-century agricultural intensification and the past and ongoing changes in farming practices are expected to have contributed to the decline of several species of farmland birds. The declines are often associated with decreased weed seed availability from farmlands (Gutiérrez-Galán & Alonso 2016; Negrier et al. 2020). Precise knowledge of the diet of a species might be of special interest for designing sound conservation and management strategies (Valentini et al. 2009), such as planning grazing schemes that increase the abundance of relevant plant species to improve food availability (Gutiérrez-Galán & Alonso 2016). Traditionally, dietary analyses have been conducted mainly by direct observation of foraging behaviour or microhistological analysis of stomach or gut contents and faecal samples. These traditional techniques are often hampered by the difficulty of continuous observation and decomposition by the digestion process impeding identification, respectively. The development of non-invasive molecular techniques has allowed a more precise detection of DNA remains and higher taxonomic resolution of diet items in faecal samples (Valentini et al. 2009; Ando et al. 2013; Crisol-Martínez et al. 2016). The molecular analyses of faeces, i.e. molecular scatology, can be used to reveal significant differences in dietary composition and overlap (potential competition) between bird species (Crisol-Martínez et al. 2016; Dunn et al. 2018; Swift et al. 2018). - 6 - x With the development of DNA barcoding and next-generation sequencing (NGS) techniques, faecal analysis using high-throughput sequencing (HTC) is nowadays a widespread, non-invasive method to study animal diets (Fig. 3; Pompanon et al. 2012; Ando et al. 2016). Generally, most species of Columbiformes are considered generalists with broad foraging ranges, often altering their diet components according to the availability of specific food resources (Ando et al. 2016). For instance, Woodpigeons adapt their diet and foraging behaviour due to synurbization by exploiting supplementary feeding at garden bird feeders (Luniak et al. 2004; Dunn et al. 2018). Turtle Doves and Stock Doves feed predominantly on seeds, whereas Woodpigeons also consume leaves, fruits and other plant matter. Animal material, e.g. small invertebrates, is rarely a component of the diet of the columbiform birds (Murton et al. 1964; Browne & Aebischer 2003; Dunn et al. 2018; Negrier et al. 2020). However, precise data on the diet of Columbiformes are fragmentary and limited (Mansouri et al. 2019; Negrier et al. 2020). Previous studies have demonstrated that the diet of the Turtle Dove changed from mainly non-cultivated (natural) arable plants in the 1960s to seeds of cultivated plants, such as wheat and oilseed rape, in the 1990s (Murton et al. 1964; Browne & Aebischer 2003). The dietary switch may be associated with the reduction in food availability during key periods of the breeding season when seeding natural arable plants have become scarce as a result of agricultural changes (Browne & Aebischer 2004; Dunn et al. 2018). The lack of natural arable plant species potentially deteriorates the condition of nestling Turtle Doves (Dunn et al. 2018). Moreover, it was hypothesised that the reduced availability of suitable food, both spatially and temporally, contributes to the decline, by causing Turtle Doves to cease breeding earlier and thus produce fewer offspring due to a reduced number of nesting attempts (Browne & Aebischer 2004; Browne et al. 2005). Furthermore, Turtle Doves might be energetically challenged due to the reduction of both diet quality and food availability (Dunn & Morris 2012). This stress could cause weakened immune defense, which in turn could increase susceptibility to infections (Altizer et al. 2006). Figure 3 Flowchart diagram showing the main steps (simplified) of a non-invasive, molecular approach for assessing diet composition using faecal samples. Figure modified according to Valentini et al. (2009) and Valkiūnas & Atkinson (2020). - 7 - General introductionx 1.3.3 Disease ecology - Avian haemosporidian parasites Endoparasitic infections are a potential stressor for wild birds. Endoparasites, which can be classified roughly into gastrointestinal parasites and blood parasites, can have effects on their avian hosts ranging from minor to fatal diseases (Kumar et al. 2018; Zahan et al. 2018; Attia & Salem 2022). In recent decades, changing environments, climate change, globalization, human and animal movements and other drivers contributed to the spread of parasites and the emergence of novel pathogens (Rosenberg et al. 2019; Rush et al. 2021). Species of Columbiformes infested with endoparasites are often assumed responsible for the spread of parasites to other avian species (Omer et al. 2015; Attia & Salem 2022). Among others, they can act as a reservoir host for the flagellated protozoan parasite Trichomonas gallinae, causing avian trichomonosis (Marx et al. 2017; Martínez-Herrero et al. 2020) or as a reservoir of various avian malaria and malaria-like parasites (Zahan et al. 2018). Nowadays, it is recognised that avian haemosporidian parasites, the causative agents of avian malaria and related diseases, are among the most pathogenic organisms. These diseases can be responsible for mortality, population declines and even extinctions of poultry and wild birds (Palinauskas et al. 2020). They can cause serious infections in avian hosts with symptoms such as anaemia, weight loss, anorexia, arthritis, hepatosplenomegaly, reduced strength in flight and have long-term negative effects on their reproductive system (Ciloglu et al. 2019). Avian haemosporidian parasites (Apicomplexa) of the genera Plasmodium, Haemoproteus and Leucocytozoon have been documented in a great variety of avian host species all over the world and are one of the most prevalent and diverse groups of avian parasites (Atkinson 1999; Valkiūnas 2005; Galen et al. 2018; Ciloglu et al. 2019; de Angeli Dutra et al. 2021). The life cycles of these vector-borne parasites are obligately heteroxenous, developing in two groups of hosts: vertebrates (here: birds) and vectors (depending on the parasite genus different blood-sucking dipterans). The vector inoculates sporozoites, the infective stage of the parasites, into birds while blood-feeding (Fig. 4). The sexual reproduction process occurs in the vector and thus the birds are intermediate hosts and the dipterans are definitive hosts (Fig. 4; Valkiūnas 2005; Valkiūnas & Atkinson 2020). Generally, three measures are most commonly considered analysing host-parasite interactions: (i) prevalence refers to the proportion of individuals of a population or group that are infected, (ii) intensity or parasitemia is a measure of how many parasites of one species infect a host individual and (iii) richness is the number of parasite species found in an individual host, a group, or a host species (Herrera & Nunn 2019). While microscopy was the method to identify blood parasites with morphological data before 2000, recent molecular studies have recognised a huge genetic diversity of avian haemosporidian parasites with at least 260 species and over 3600 lineages (Harl et al. 2020; Mandal 2021). - 8 - x Also, it is known that the avian life-history strategy influences haemosporidian prevalence and richness (Dunn & Outlaw 2019; Fecchio et al. 2021). For example, it is expected that migratory birds might have more prevalent haemosporidian infections and higher parasite richness because they move between breeding and wintering areas and use stopover sites. At these different locations they may encounter a much greater variety of haemosporidian parasites and vectors. In comparison, residents encounter parasites and their vectors in only one ecosystem (Ciloglu et al. 2019; de Angeli Dutra et al. 2021; Fecchio et al. 2021). Even though Plasmodium parasites are among the best-studied pathogens, there is still a large gap in our understanding of their diversity, specificity, virulence and development in vertebrate hosts. Even less is known about the other genera Leucocytozoon and Haemoproteus (Palinauskas et al. 2020). By sampling wild birds, it is possible to define composition up to lineage level and prevalence of haemosporidian parasites in different bird species using a combination of microscopy and molecular tools (Ciloglu et al. 2019; Palinauskas et al. 2020). Accurate detection of avian haemosporidian parasites includes a thorough sampling of avian host species, in particular columbiform species (Zahan et al. 2018). The accurate detection is required to understand host-parasite interactions, effects of coinfections (e.g. with Trichomonas gallinae), true diversity and epidemiology of infections and to develop control strategies against these diseases (Ciloglu et al. 2019; Dunn & Outlaw 2019; Thomas et al. 2022). Figure 4 General representation of the life cycle of avian haemosporidian parasites. Figure adapted from Atkinson (1999), Mata (2012) and Valkiūnas & Atkinson (2020). - 9 - Aims and structure of the thesisx 2 | AIMS AND STRUCTURE OF THE THESIS The main objective of this thesis was to improve our knowledge about several behavioural- ecological aspects of different migratory species of Columbiformes native to Europe by a comparative approach. Thus, the studies included intend to complement missing basic and fundamental information on the species' ecology. The knowledge acquired through the studies can potentially be used to identify prospective protection strategies, considering interests in both management of game bird species and species conservation. In addition, the data gathered can help to estimate the influence of the different aspects analysed on the population dynamics of the columbiform species. The included aspects can be classified into three broad research topics (Fig. 5 and 6):  movement ecology, encompassing migration and foraging behaviour (chapters 1 to 3)  dietary ecology, focussing on analysing diet compositions (chapter 4)  disease ecology, in particular, avian haemosporidian parasites (chapter 5) While chapters 1 to 3 focus on a single species each (chapters 1 and 2: Turtle Dove, chapter 3: Woodpigeon), chapters 4 and 5 are community analyses, including several species of wild Columbiformes. Due to the pluralist approach in terms of investigated behavioural- ecological aspects and applied methods (Fig. 6), the present cumulative thesis is structured in five publications, each presented as an individual chapter, with the following specific aims: CHAPTER 1 | Feather stable isotopes (δ2H and δ13f Cf) identify the Sub-Saharan wintering grounds of turtle doves from Europe  Determine the non-breeding, i.e. wintering, regions of the only long-distance migratory species among the Columbiformes native to Europe, namely Turtle Doves  Analyse whether wintering grounds differ for individuals using different flyways to estimate the migratory connectivity of Turtle Doves across Europe CHAPTER 2 | Year‑round spatial distribution and migration phenology of a rapidly declining trans‑Saharan migrant - evidence of winter movements and breeding site fidelity in European turtle doves  Identify migration flyways and phenology as well as wintering areas of Turtle Doves following different flyways on a finer spatial and temporal scale  Characterise habitat parameters at breeding and wintering sites  Check for the occurrence of niche tracking or niche switching behaviour with regard to environmental parameters - 10 - x CHAPTER 3 | Should I stay or should I fly? Migration phenology, individual-based migration decision and seasonal changes in foraging behaviour of Common Woodpigeons  Investigate the migration and foraging behaviour of Woodpigeons from different breeding regions  Evaluate within-population migratory dimorphism  Describe habitat characteristics and choice based on land cover data CHAPTER 4 | Diet composition of wild columbiform birds: Next-generation sequencing of plant and metazoan DNA in faecal samples  Demonstrate how plant and metazoan diet components can be identified based on a non-invasive approach  Provide a detailed reconstruction of the food items ingested by species from the order Columbiformes  Determine the potential dietary overlap between the included bird species CHAPTER 5 | Prevalence and genetic diversity of avian haemosporidian parasites in wild bird species of the order Columbiformes  Examine the overall and species-specific prevalence, diversity and parasitemia of avian haemosporidian parasites in wild columbiform bird species  Check for potential differences between host species, taking into account the variable behavioural characteristics of each species  Evaluate the possible negative effects of haemosporidian parasites on host species Figure 5 Schematic overview of relevant connections and interactions, represented by arrows, between the three broad research topics included in the presented cumulative thesis (dietary ecology: chapter 4, disease ecology: chapter 5 and movement ecology: chapters 1 to 3). - 11 - Chapter outlinex 3 | CHAPTER OUTLINE The thesis compromises five studies that provide insight in the above-mentioned main objective, separated into multiple specific aims, presented in the chapters accordingly. CHAPTER 1 | Feather stable isotopes (δ2Hf and δ 13Cf) identify the Sub-Saharan wintering grounds of turtle doves from Europe [PUBLISHED] Outline – In this publication, analyses of feather-isotope signatures were used to identify the African wintering origins of Turtle Doves. Samples of tenth primary feathers (n = 181), known to grow during the wintering period, have been collected during the breeding and migration period or originated from museum collections. By assigning stable hydrogen (δ2Hf) and carbon (δ13Cf) isotope values of feathers to multi-isotopic landscapes (‘isoscapes’) of precipitation stable hydrogen (δ2Hp) and theoretical vegetation stable carbon (δ13C) in Africa, we presented a first-order estimation of wintering regions of sampled Turtle Doves. Furthermore, it was assessed how these likely wintering quarters, located in the Western and Central Sub-Sahara, overlap with present hunting and protected areas. The use of samples collected at different sites across Europe allowed checking for flyway-specific wintering regions. The probabilistic assignments did not indicate a marked difference between the wintering origin of individuals following the Western (samples from Spain and France) vs. the Central/Eastern flyway (samples from Greece, Malta, Italy and Bulgaria). However, comparisons of the raw δ2Hf and δ13Cf values revealed differences, suggesting that individuals potentially have spent the winter in separate regions, had different diets, or occupied different habitats. The advantages and disadvantages of stable isotope analyses, as applied in the study, were discussed. This may provide thought-provoking impulses for research on other long-distance migratory species, in particular for those too small to be equipped with currently available tracking devices. Contributions – Shared lead author (with M. Marx), including manuscript writing and editing; corresponding author; partial data and statistical analyses of stable isotope data and spatial dataset of protected and hunting areas (shared mainly with M. Marx and K.J. Kardynal) CHAPTER 2 | Year‑round spatial distribution and migration phenology of a rapidly declining trans‑Saharan migrant - evidence of winter movements and breeding site fidelity in European turtle doves [PUBLISHED] Outline – This publication focused on the evaluation of satellite tracking data of Turtle Doves to increase our knowledge on migration flyways, migration timing, stopovers, European breeding and African wintering sites. Individuals have been equipped with Argos satellite tags, providing location fixes based on Doppler calculations, during spring migration on Malta (n = 8) and the breeding season in Germany (n = 8). Tracking data combined with environmental habitat parameters showed that environmental parameters are more uniform at the wintering grounds compared to the breeding grounds. This suggests that Turtle Doves - 12 - x might be even more vulnerable to future changes in their winter ranges than in their breeding ranges. ‘Niche tracking’ behaviour, i.e. following environmental conditions of similar type throughout the year, was only observed regarding night-time temperatures. Analysis of land cover data on breeding grounds demonstrated the use of a wide range of forest and agricultural landscapes. Home range size at breeding grounds increased with an increasing proportion of agricultural areas. This is probably due to Turtle Doves being forced to fly large distances in intensively farmed areas to reach good quality food resources. Year-round tracking, partly for consecutive years, of individuals revealed behavioural characteristics relevant for planning conservation measures, e.g. breeding site fidelity, prolonged stopovers in Europe, use of multiple wintering sites, evidence for loop migration and a curtailed breeding season. The gathered data were particularly needed for individuals migrating along the Central and Eastern flyway, as no tracking data from there had been analysed before this study. Contributions – Lead author; partial material collection and bird handling in the field (catching, sampling and tagging); molecular work (sex determination); data and statistical analyses (environmental habitat parameters, tracking and land cover data) CHAPTER 3 | Should I stay or should I fly? Migration phenology, individual-based migration decision and seasonal changes in foraging behaviour of Common Woodpigeons [PUBLISHED] Outline – This manuscript dealt with foraging and migratory movements over the annual cycle of Woodpigeons. The analyses were based on ring-recovery and two types of satellite tracking data (Argos and GPS-GSM/GPRS). Individuals (n = 29) with breeding areas located in Germany and Portugal, mainly caught in an urban environment, were equipped with GPS-GSM/GPRS transmitters. Woodpigeons from Lisbon (Portugal) stayed almost exclusively within a large wooded park within the city. Contrary, Woodpigeons from Giessen (Germany) regularly left the city area to forage on surrounding farmland. In general, home ranges were larger for Woodpigeons in Giessen compared to individuals in Lisbon. The results of ringing and tracking data showed a migratory dimorphism: all Woodpigeons breeding in Portugal and the majority of individuals with breeding sites in Germany were residents, but other individuals from Germany were migrants. These migrants wintered either in Germany, but away from their breeding sites, or followed the European sector of the East Atlantic flyway to winter mainly in France and less frequently in Spain, Portugal, Belgium, Netherlands and Denmark. Using the tracking data, it was possible to observe behavioural characteristics on an individual level, revealing a low wintering site fidelity, use of multiple winter sites during one season and switches of migratory strategies (resident vs. migrant) between years. This study, the first to analyse satellite tracking data of Woodpigeons from consecutive years, emphasises their behavioural plasticity. Studying species with such pronounced variation in migratory behaviour might be of value to investigate the effects of ongoing climate change and increasing urbanisation on migratory decisions. - 13 - Chapter outlinex Contributions – Lead author; funding acquisition; partial material collection and bird handling in the field (catching, sampling and tagging); molecular work (sex determination); data analyses (ring-recovery, tracking and land cover data) CHAPTER 4 | Diet composition of wild columbiform birds: Next-generation sequencing of plant and metazoan DNA in faecal samples [IN PREPARATION] Outline – In this study, the focus was on a detailed analysis of the diet compositions of different species of Columbiformes by applying next-generation sequencing (NGS) technology as a tool for diet reconstruction through DNA metabarcoding. The use of primer pairs targeting plant nuclear DNA and metazoan DNA, isolated from collected faecal samples (n = 139), provided a complete picture of the food items ingested. Assessing the dietary overlap between the included columbiform species (Turtle Doves, Stock Doves and Woodpigeons) revealed variability in consumed plant taxa (a diverse range of Spermatopsida) among the species. Woodpigeons and Stock Doves showed the highest dietary overlap. Non-metric Multidimensional Scaling was used to visualize the detected differences in plant diet compositions. Animal prey, i.e. metazoan DNA, was present only very rarely. Plant taxa observed in this study, but previously not listed as known food items in other studies, and an evaluation of the proportion of wild vs. cultivated plant species give indications for dietary changes due to urbanization and agricultural intensification. Especially for declining Turtle Doves, identified regional variations may be relevant for the implementation of proposed conservation options, e.g. tailored seed mixtures. The sharp decline of many avian species over the past decades calls for a more thorough knowledge of their dietary requirements. Our study supports the non-invasive approach applied as an accurate method for diet analyses that could be used in other species as well. Contributions – Lead author; organisation of faecal sample collection with cooperation partners; partial material collection in the field; molecular work (DNA isolation and preparation of NGS libraries); partial data and statistical analyses (shared mainly with J.F. Masello) CHAPTER 5 | Prevalence and genetic diversity of avian haemosporidian parasites in wild bird species of the order Columbiformes [PUBLISHED] Outline – Here, the prevalence, diversity and parasitemia of haemosporidian parasites in Columbiformes were presented. Blood samples (n = 259) of different species, predominantly Turtle Dove, Stock Dove and Woodpigeon, were collected in seven countries between 2013 and 2019. Haemosporidian infections were determined through nested PCR, revealing an overall prevalence of 42%. More precisely, the majority of individuals (34%) harboured a single haemosporidian lineage (Leucocytozoon: 15%, Haemoproteus: 15%, Plasmodium: 4%) and 7% had mixed infections. Stock Doves had a lower overall prevalence (4%) than Turtle Doves (49%) and Woodpigeons (62%), potentially because they might be shielded better from dipteran vectors as cavity-nesters. - 14 - x Overall, 15 distinct mitochondrial cytochrome b lineages were detected, including five newly discovered ones. A second PCR assay (One-step multiplex) was applied for improved detection of mixed Plasmodium/Haemoproteus infections. However, we showed that this assay displays infections of the subgenera H. (Haemoproteus) at the expected band height of Plasmodium infections. Thus, this PCR assay is rather not advisable to examine orders prone to H. (Haemoproteus) infections. Despite a relatively high observed prevalence and diversity, the parasitemia (obtained by microscopic examination of blood smears) was rather low for most samples, indicating mainly chronic instead of acute infections. The low parasitemia suggests the contribution of haemosporidian infections to population dynamics of sampled hosts might be rather insignificant. Obtained data can constitute an important reference to monitor future changes in parasite ranges and diversity expected as a consequence of climate change. Contributions – Lead author; partial material collection in the field (blood sampling); examination of blood smears; molecular work (DNA isolation and PCR assays); phylogenetic (medium joining networks and phylogenetic tree reconstruction) and statistical analyses of datasets resulting from Sanger sequencing Figure 6 Overview of thesis structure, depicting the three broad research topics with the respective species studied and applied methods (simplified) for each chapter. - 15 - General conclusions and future outlookx 4 | GENERAL CONCLUSIONS AND FUTURE OUTLOOK 4.1 General conclusions The presented cumulative thesis consists of five chapters focusing on behavioural-ecological aspects in the fields of movement, dietary and disease ecology. All chapters together contribute to form a more complete picture of the behaviours exhibited by the studied columbiform species. In particular, because the different fields of ecology are connected and interact with each other (Fig. 5). The presented findings result from the combination of studying single species (chapters 1 to 3) and comparative approaches (chapters 4 and 5; Fig. 6). Given the fact that many migratory birds spend most of the year away from their breeding grounds and face seasonally specific threats and limitations with various carryover effects, it is important to consider data from both the breeding and the non-breeding season, i.e. across the entire annual cycle (Hostetler et al. 2015; Briedis et al. 2018; Swift et al. 2020). Studying the migratory movements of Turtle Doves (chapters 1 and 2) and Woodpigeons (chapter 3) has contributed to a better understanding of the year-round distribution of these two species. The main results obtained under this thematic aspect of my thesis point out that:  Turtle Doves from the European population migrate along different migration flyways from their European breeding grounds to their African wintering grounds, located in the Western and Central Sub-Sahara. Moreover, we could show that they partly wintered farther south than 10°N (chapters 1 and 2), which is often given as a limit in literature. Consequently, also areas farther south than 10°N should be considered as potential wintering areas.  Turtle Doves following the different flyways are likely to occupy different regions and habitats during the wintering period (chapters 1 and 2). This potentially exposes them to different and variable levels of threats during this period in the life cycle, which might be reflected in varying levels of observed population declines across the European breeding grounds (see de Vries et al. 2022).  In Woodpigeons, which were breeding in Portugal, only resident behaviour was observed. Whereas, the individual-based migratory decision of Woodpigeons with breeding sites in Germany switched between resident and migrant between years. Therefore, they can be classified as facultative partial migrants (chapter 3). The annually changing numbers of migrants and residents may provide an explanation for the observed fluctuating numbers of Woodpigeons in their wintering areas (e.g. Lormée & Aubry 2018).  Woodpigeons show pronounced behavioural plasticity in terms of migration tactics (chapter 3). Firstly, this supports the hypothesis that migratory movements in partial migrants are not solely a genetically fixed behaviour (Lundblad & Conway 2020). Secondly, given that Woodpigeons are nowadays one of the most common bird species in many European cities, our results support the general postulation that behavioural plasticity plays an influential role in successful urban colonization processes in birds (Chyb et al. 2021). - 16 - x  Both migratory Woodpigeons and Turtle Doves show a high breeding site fidelity, i.e. return to the same breeding areas, compared to a lower wintering site fidelity. Also similar in both species is the use of multiple distinct wintering sites during one wintering season by the majority of individuals (chapters 2 and 3). Probably these winter movements are linked to the temporally fluctuating availability of food resources. The dietary and foraging ecology of a species are closely linked to each other, e.g. food availability exerts major influences on foraging distances (Burke & Montevecchi 2009). Therefore, dietary studies can give insights into foraging behaviour and vice versa (Gaglio et al. 2016). Considering the results of the molecular diet analyses (chapter 4) and the studies on foraging behaviour based on tracking data (chapters 2 and 3) together, the following conclusions can be drawn:  Turtle Doves and Woodpigeons seem to adapt their foraging areas and distances in breeding areas to the availability of food resources (chapters 2 and 3, respectively). This, on the one hand, might highlight an opportunistic or rather plastic nature in both species, but on the other hand, indicates that suitable food resources might not always be available near nesting sites. This forces individuals to forage over longer distances, which might be energetically costly and therefore potentially affect the species negatively (Browne & Aebischer 2001; Masden et al. 2010; Trevail et al. 2019).  The DNA data from faecal samples showed some differences between the columbiform species in terms of consumed plant species (chapter 4). Woodpigeons also consumed plants growing mainly in urban environments, whereas this could not be detected as distinctly in Turtle Doves and Stock Doves. This is also reflected in the tracking studies, demonstrating that much of the foraging of Woodpigeons takes place in urban areas, whereas Turtle Doves very rarely visit urban sites (chapters 2 and 3), highlighting the successful adaptation of Woodpigeons to urban environments.  In general, only certain species, characterized by a combination of specific traits and by certain niche requirements, seem to be capable of coping with the environmental alterations that are imposed by urbanisation (Jokimäki et al. 2016; Patankar et al. 2021). The adapted behaviour in terms of urbanisation of Woodpigeons could be one of the reasons why population numbers of Woodpigeons are not declining despite the continuous increase of urban settlements, which generally constitute a major threat to biodiversity (Seto et al. 2012; Patankar et al. 2021). In contrast, the Turtle Dove has a strong declining population trend in Europe, potentially indicating that this species copes less well with the ongoing urbanisation and associated changes (but see Eddajjani et al. 2022).  Obtained results point out distinct plasticity in foraging habits, in particular for ‘urban’ Woodpigeons (chapter 3). This is likely an adaptation to the seasonally changing productivity of exploited foraging sites. Moreover, out of the three columbiform species, Woodpigeons had the highest overall diversity in consumed plant families (chapter 4). - 17 - General conclusions and future outlookx Many birds living in urban environments adapt their food resources and show a loss in migratory behaviour since there is enough food available in urban areas to support them through the winter (Méndez et al. 2020; Patankar et al. 2021). This could also be the case in Woodpigeons, as most individuals tagged in an urban area in Germany were residents and especially during winter rarely left the city (chapter 4). Contrary, Turtle Doves and Stock Doves, which typically do not exploit food resources in urban areas, are forced to migrate to find adequate and sufficient food during winter. Migration plays a significant role in the ecology and evolution of host species and consequently their parasites (Ishtiaq & Renner 2020). Moreover, migrants are important agents for the distribution of parasites, particularly those lacking free-living stages, and thus disperse only with their hosts and vectors, such as avian haemosporidian parasites (Ricklefs et al. 2017). By analysing avian haemosporidian parasites in columbiform species with different migration strategies (chapters 2, 3 and 5), the results presented in my thesis could:  Contribute to the debate whether migratory avian species are more often infected by haemosporidian parasites (e.g. Soares et al. 2019; Ciloglu et al. 2020; de Angeli Dutra et al. 2021). Our results support the hypothesis that long-distance migrants harbour a higher diversity of haemosporidian parasites than residents or short-distance migrants. In terms of prevalence, this pattern was only evident for Plasmodium and Haemoproteus infections. Contrary, Leucocytozoon infections were most prevalent in partial migrants, namely Woodpigeons (chapters 3 and 5). The detected variation in patterns by parasite genus along with methodological issues raised in the publication emphasises that an accurate detection of avian haemosporidians is crucial for correctly investigating host-parasite interactions and true parasite diversity (Ciloglu et al. 2019).  Assess the potential impact of blood parasites as a threat to declining Turtle Doves (Fisher et al. 2018). Due to the rather low observed parasitemia, we deem the contribution of haemosporidian infections to the decline of this species to be rather insignificant at the moment (chapter 5). However, expected future changes in parasite transmission areas, distribution and diversity associated with global change can pose an upcoming risk (Ishtiaq et al. 2021). My results provide reference information to monitor future changes. Beyond the contribution to a better general understanding of different behavioural aspects of the included columbiform species, the studies assembled in this thesis provide some insight to draw prospective directive lines for conservation purposes for declining Turtle Doves and management of Woodpigeons:  For Turtle Doves we demonstrated that prolonged stopovers during autumn migration in Europe overlap with the time of legal hunting activities (chapter 2). The sustainability of Turtle Dove hunting in Europe has been discussed during the last several years due to severe population declines (Thomaidis et al. 2022). - 18 - x Turtle Dove individuals migrating along the Western flyway mainly pass European countries with temporary hunting bans (Fig. 2). However, hunting is still allowed in some countries along the Central and Eastern flyways. The sustainability of harvest and effectiveness of hunting regulations along the Central and Eastern flyway should be re- assessed, similarly as done along the Western flyway (Lormée et al. 2020; Delibes-Mateos et al. 2021; Moreno-Zarate et al. 2021), to develop sustainable hunting concepts.  In Germany, Woodpigeons can usually be hunted during the winter month (November to February; JagdzeitV 1977). Our results demonstrated that Woodpigeons, which breed in urban areas, rarely leave the urban areas during these months (chapter 3). Therefore, shot Woodpigeons are likely mainly local individuals breeding outside the urban area or migrants from more northern European countries, which winter in Germany. Thus, hunting probably has relatively little impact on resident urban Woodpigeons. However, it must be noted that our study represents the situation of only one city in Germany. Woodpigeons in other cities might behave differently, e.g. depending on the type of habitats surrounding the city or food availability within the city.  In line with Dunn et al. (2021) and Chiatante et al. (2021) we discovered that the home range size of Turtle Doves in breeding areas seems to depend on the availability of food resources connected to certain habitat types. In our case, a higher proportion of agricultural areas led to an increase in home range size (chapter 2). Also, the dietary shift from wild plant species to cultivated ones (chapter 4) was already observed in other breeding areas and has been proposed to negatively influence Turtle Doves (e.g. Browne & Aebischer 2004; Dunn et al. 2015, 2018). Both, increased home range size and dietary shift, reinforce the importance of the close proximity of suitable nesting sites and feeding areas with accessible seeds throughout the entire breeding season (Browne et al. 2004; Dias et al. 2013; Dunn et al. 2018). To ensure abundant and adequate food resources at the breeding areas, managed feeding areas sown with tailored seed mixtures according to regionally preferred wild plant species could prove to be a successful conservation measure.  Finally, the obtained results add to the body of research, which highlights the variability of habitats, both breeding and foraging habitats, and food resources exploited by Turtle Doves across European breeding sites. This indicates that the Turtle Dove is not a specialist species, but rather a plastic one, able to adapt to a variety of environmental conditions, i.e. variable habitat compositions. Thus, the ‘optimal’ Turtle Dove habitat might look very different across regions. This must be considered when planning conservation measures, as it cannot be assumed that a particular measure will be equally effective and useful across different breeding regions, e.g. Western versus Eastern Europe. Therefore, conservation measures should be tailored to regional conditions. - 19 - General conclusions and future outlookx 4.2 Future outlook Not all pertinent questions dealing with behavioural aspects of the included species of Columbiformes and, moreover, the resulting consequences of specific behaviours for populations dynamics, could be covered and answered comprehensively within the scope of my thesis. Some of the included analyses could be examined in more depth with a better data basis. For instance, the analyses of habitat requirements would benefit from an improved understanding on the relationships between specific landscape components and bird abundance (cf. Saâd et al. 2021). This could be achieved if more detailed information were available on landscape elements, such as the presence of small-scale elements like hedgerows or field margins, and on land management practices, e.g. the use of agrochemicals (cf. Marx & Quillfeldt 2018). A thorough identification of the landscape elements that are crucial in explaining abundance patterns can be helpful to promote specifically these elements through measures and thus enhance potential habitats. In the particular case of Turtle Doves, a closer look at habitat requirements in their wintering areas is needed, as their sensitivity to changes in their wintering quarters has already been suggested (Eraud et al. 2009). However, a significant knowledge gap on threats in the wintering areas remains (Fisher et al. 2018). To be able to draw more direct causal links in future research, it should be striven to assess more concrete connections between different behavioural aspects (Fig. 5). For example, assessing if the availability of specific food resources influences the migratory decision in partial migrants. Another research question could be whether individuals weakened by a suboptimal diet are more likely to be affected negatively by pathogens. During research for this thesis, I attempted to equip Stock Doves with transmitters, but tracking them was not successful due to different reasons, such as the breaking off of transmitter antennas or low recapture success. Future studies should take a step forward by expanding applied tracking methods, such as using transmitters with GPS accuracy on all species. Moreover, they should include individuals of all columbiform species, using the same location to ascertain habitat segregation or potential competition on particular resources, e.g. nesting sites or certain food resources (cf. Benghedier et al. 2020; Floigl et al. 2022; Squalli et al. 2022). It should be noted that the presented studies only cover small sub-areas of the complete distribution ranges of the respective species and that different behavioural patterns should be expected in other areas, especially since our studies have already determined regional differences. For example, there are hardly any studies from the Eastern part of the breeding distribution range (Eastern Europe and parts of Asia) of Turtle Doves. To recapitulate, the outcomes of the studies incorporated in my thesis show how research into different behavioural aspects not only provides insights into the general biology of species and species communities but can also provide information relevant for appropriate management and conservation actions (cf. Hays et al. 2019; Saâd et al. 2021). However, to actually help declining species in practice, it will be particularly important to close the prevailing science-practice implementation gap in the future. - 20 - x 5 | REFERENCES Altizer S, Dobson A, Hosseini P, Hudson P, Pascual, Rohani P (2006) Seasonality and the dynamics of infectious diseases. Ecology Letters 9:467-484. 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DOI: 10.15312/EurasianJVetSci.2018.189 - 28 - x - 29 - CHAPTER 1 FEATHER STABLE ISOTOPES (δ2H AND δ13f Cf) IDENTIFY THE SUB-SAHARAN WINTERING GROUNDS OF TURTLE DOVES FROM EUROPE Melanie Marx*, Yvonne R. Schumm*, Kevin J. Kardynal, Keith A. Hobson, Gregorio Rocha, Pavel Zehtindjiev, Dimitris Bakaloudis, Benjamin Metzger, Jacopo G. Cecere, Fernando Spina, Marco Cianchetti-Benedetti, Sylke Frahnert, Christian C. Voigt, Hervé Lormée, Cyril Eraud, Petra Quillfeldt * These authors contributed equally to this work European Journal of Wildlife Research (2022) 68: 21. DOI: 10.1007/s10344-022-01567-w - 30 - x - 31 - Chapter 1 | Wintering grounds of Turtle Dovesx - 32 - x - 33 - Chapter 1 | Wintering grounds of Turtle Dovesx - 34 - x - 35 - Chapter 1 | Wintering grounds of Turtle Dovesx - 36 - x - 37 - Chapter 1 | Wintering grounds of Turtle Dovesx - 38 - x - 39 - Chapter 1 | Wintering grounds of Turtle Dovesx - 40 - x - 41 - Chapter 1 | Wintering grounds of Turtle Dovesx - 42 - x - 43 - Chapter 1 | Wintering grounds of Turtle Dovesx - 44 - x - 45 - x CHAPTER 2 YEAR-ROUND SPATIAL DISTRIBUTION AND MIGRATION PHENOLOGY OF A RAPIDLY DECLINING TRANS-SAHARAN MIGRANT – EVIDENCE OF WINTER MOVEMENTS AND BREEDING SITE FIDELITY IN EUROPEAN TURTLE DOVES Yvonne R. Schumm, Benjamin Metzger, Eric Neuling, Martin Austad, Nicholas Galea, Nicholas Barbara, Petra Quillfeldt Behavioral Ecology and Sociobiology (2021) 75: 152. DOI: 10.1007/s00265-021-03082-5 - 46 - x - 47 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 48 - x - 49 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 50 - x - 51 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 52 - x - 53 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 54 - x - 55 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 56 - x - 57 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 58 - x - 59 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 60 - x - 61 - Chapter 2 | Migration and habitat use of Turtle Dovesx - 62 - x - 63 - CHAPTER 3 SHOULD I STAY OR SHOULD I FLY? MIGRATION PHENOLOGY, INDIVIDUAL-BASED MIGRATION DECISION AND SEASONAL CHANGES IN FORAGING BEHAVIOUR OF COMMON WOODPIGEONS Yvonne R. Schumm, Juan F. Masello, Valerie Cohou, Philippe Mourguiart, Benjamin Metzger, Sascha Rösner, Petra Quillfeldt The Science of Nature (2022) 109: 44. DOI: 10.1007/s00114-022-01812-x - 64 - x - 65 - Chapter 3 | Migration and foraging of Woodpigeonsx - 66 - x - 67 - Chapter 3 | Migration and foraging of Woodpigeonsx - 68 - x - 69 - Chapter 3 | Migration and foraging of Woodpigeonsx - 70 - x - 71 - Chapter 3 | Migration and foraging of Woodpigeonsx - 72 - x - 73 - Chapter 3 | Migration and foraging of Woodpigeonsx - 74 - x - 75 - Chapter 3 | Migration and foraging of Woodpigeonsx - 76 - x - 77 - Chapter 3 | Migration and foraging of Woodpigeonsx - 78 - x - 79 - Chapter 3 | Migration and foraging of Woodpigeonsx - 80 - x - 81 - CHAPTER 4 DIET COMPOSITION OF WILD COLUMBIFORM BIRDS: NEXT-GENERATION SEQUENCING OF PLANT AND METAZOAN DNA IN FAECAL SAMPLES Yvonne R. Schumm, Juan F. Masello, Jennifer Vreugdenhil-Rowlands, Dominik Fischer, Klaus Hillerich, Petra Quillfeldt Manuscript prepared for submission - 82 - x Diet composition of wild columbiform birds: next-generation sequencing of plant and metazoan DNA in faecal samples Yvonne R. Schumm1#, Juan F. Masello1, Jennifer Vreugdenhil-Rowlands2, Dominik Fischer3, Klaus Hillerich4, Petra Quillfeldt1 1 Department of Animal Ecology & Systematics, Justus Liebig University, Heinrich-Buff-Ring 26-32, 35392 Giessen, Germany 2 Lewestraat 52, 4481 BE Kloetinge, Netherlands 3 Clinic for Birds, Reptiles, Amphibians and Fish, Veterinary Faculty, Justus Liebig University Giessen, Frankfurter Strasse 114, 35392 Giessen, Germany; Present address: Zoo Wuppertal, Hubertusallee 30, 42117 Wuppertal, Germany 4 Röntgenstraße 7, 64823 Groß-Umstadt, Germany # Corresponding author: Yvonne.R.Schumm@bio.uni-giessen.de - 83 - Chapter 4 | Diet analysesx Abstract Next-generation sequencing technology has become a powerful and non-invasive tool for diet reconstruction through DNA metabarcoding. Accurate knowledge of species’ diets is fundamental to understand their ecological requirements. Here we applied next-generation sequencing and DNA metabarcoding on faecal samples of European Turtle Doves Streptopelia turtur (n = 19), Stock Doves Columba oenas (n = 71) and Common Woodpigeons C. palumbus (n = 49) to investigate their dietary composition. By applying primer pairs targeting both the ITS2 region of plant nuclear DNA and the mitochondrial COI region of metazoan DNA, we provide a complete picture of the food ingested and estimate the dietary overlap between the columbiform species. Animal DNA was present very rarely, and a diverse range of plants from the class Spermatopsida dominated the diet, with Asteraceae, Brassicaceae, Cucurbitaceae, Fabaceae and Poaceae as the most frequently represented families. Generally, we detected a variability between species but also among individual samples. Plant species already known from previous studies, mainly visual analyses, could be confirmed for our individuals sampled in Germany and the Netherlands. However, the molecular approach also revealed new plant taxa. Although most of the plant species were categorised as ‘wild’, the majority of cultivated plants were present with higher frequencies of occurrence, suggesting that cultivated food items likely constitute an important part of the Columbiformes’ diet. For Turtle Doves a comparison with previous studies suggested regional differences and that food items (historically) considered as important part of their diet, were missing in our samples. This indicates that regional variations as well as historic and current data on diet should be considered to plan tailored seed mixtures, which are currently proposed as an important conservation measure for rapidly declining Turtle Doves. Keywords Columbiformes; Common Woodpigeon; DNA metabarcoding; European Turtle Dove; high- throughput sequencing; molecular diet analysis; non-invasive sampling; Stock Dove Introduction Analyses of diet are important to understand the feeding ecology and habitat requirements of animals as well as to manage and protect species (Oehm et al., 2011; Gong et al., 2019). Conventional methods of dietary studies rely on visually identifying diet components during foraging (behavioural observations) or within stomachs, guts or faeces (morphological classification). These techniques often suffer from misidentification of similar-looking prey items, underrepresentation of soft-bodied or small components, and low taxonomic resolution due to observation distance or digestion stage. Furthermore, analyses of the stomach and crop - 84 - x content are invasive, as dead individuals are needed or regurgitation needs to be forced (Jordan, 2005; Oehm et al., 2011; Bowser et al., 2013; Gong et al., 2019). Biochemical and molecular methods like fatty acids, protein electrophoretic, stable isotope analyses and DNA metabarcoding have been adapted to be used in diet studies (Pompanon et al., 2012). The application of DNA metabarcoding in diet studies has increased considerably with the advent of next-generation sequencing (NGS) technology (Pompanon et al., 2012). With NGS it is now possible to identify (rare) prey items of multiple samples up to species level in a single sequencing run while maintaining the ability to trace back each prey to the sample of origin (Bowser et al., 2013; Gong et al., 2019). NGS technology is increasingly used for dietary analyses across a variety of animal taxa (arthropods: Krey et al., 2020; fish: Chow et al., 2019; mammals: Buglione et al., 2020; birds: Dunn et al., 2018, Kleinschmidt et al., 2019) with faeces being the most popular sample type (Alberdi et al., 2018). Up to now, detailed information about the range and composition of the diets of many free-ranging wild animals is still extremely limited, and often, only a generalized approximation of food items consumed is known. Accurate and comprehensive knowledge of a species’ diet is fundamental to understand its ecological requirements (Wood, 1954; Newton, 1998; Jordan, 2005). This is, for example, essential for appreciating resource partitioning between species and competition for food. Furthermore, knowledge of the feeding habits and ecology of a species is important to evaluate how food availability can affect its population status and to identify key resources for designing conservation strategies (Valentini, 2008; Gutiérrez-Galán et al., 2017). In general, the Turtle Dove is considered an obligate granivore, whereas the other two columbiform species have more generalist diets, including green plant material, fruits and invertebrates, especially if seed availability is low (Murton et al., 1964; Möckel, 1988; Gutiérrez- Galán et al., 2017; Negrier et al., 2021). The present study aimed to update and improve our knowledge on the diet composition of three species of Columbiformes, sampled in Germany and the Netherlands, representing regions where their feeding ecology was little studied in recent years. NGS technology was used to generate a diet reconstruction through DNA metabarcoding based on faecal samples, which in turn was used to compare the diet of the three species to assess their dietary overlap. Material & Methods Faecal sample collection and DNA isolation Faecal samples (n = 139) were collected from three species Common Woodpigeon (n = 49), European Turtle Dove (n = 19), and Stock Dove (n = 71) at different sampling sites in Germany and the Netherlands (Fig. S1, Table S1). Birds were caught using mist nets, trapping cages, clap nets or in the case of some Stock Doves traps installed to their artificial nest boxes. Faecal samples were collected either opportunistically from the bird during handling or from the inside - 85 - Chapter 4 | Diet analysesx of clean bird bags within which the birds were temporarily held. Some faecal samples of Woodpigeons were collected as fresh droppings of active nests or roosting sites (n = 26) or from transport containers of individuals brought to the clinic for birds by the public (‘Vetmed’, n = 19). Some individuals were caught at temporarily baited sites with seeds used to lure individuals (Table S1). Thus, we expected a small amount of baited seeds (Table S2) to be present in faecal if individuals were using baited sites (cf. Dunn et al. 2018). Sampled nestlings were at least one week or older to ensure they did not receive crop milk only (Glutz von Blotzheim & Bauer, 1987). All faecal samples were stored dark and frozen at −20°C. Prior to DNA isolation, 180-200 µg of each sample were weighed. If less material was available, the entire sample was used (minimum: 21 µg). DNA was extracted using the QIAamp ® Fast DNA Stool Kit Mini (QIAGEN GmbH, Germany) with the following modifications to the manufacturer's instructions: 2-3 bashing beads (ZR Bashing BeadTM 2.0 mm, Zymo Research, USA) were added to ensure proper homogenization using the Disruptor Genie™ (Scientific Industries SI™). Incubation with Buffer AL and proteinase K was increased from 10 to 30 min. Two negative extraction controls, i.e. empty Eppendorf tubes, were run along with the faecal samples during isolation and through the entire process. DNA concentration was determined with a NanoDrop2000c UV-Vis Spectrophotometer (NanoDrop Technologies, USA) and samples were diluted to 20 ng/µl if the DNA concentration was higher than 100 ng/µl. Construction of sequencing library A sequencing library (NGS library) was constructed with two consecutive PCR reactions, first, an amplicon PCR followed by an indexing PCR. Initial tests (Supplementary material A1) resulted in the following amplicon PCRs: We used primers UniPlantF and UniPlantR amplifying a 187 to 380 bp region encompassing the second internal transcribed spacer of nuclear ribosomal DNA (ITS2) of plant nuclear DNA (Moorhouse-Gann et al., 2018). The primer pair mICOIintF/dgHCO-2198 (Meyer, 2003, Leray et al., 2013) was used to amplify a fragment of approx. 300 bp of the highly variable mitochondrial cytochrome c oxidase subunit 1 (COI) region of metazoan DNA (Supplementary material A2). All used primers had Illumina overhang adapters attached (P5 for forward primers: 5’-TCGTCGGCAGCGTCAGATGTGTATAAGAGACAG-3’ and P7 for reverse primers: 5’-GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAG-3’). PCR runs included PCR grade water as negative control and negative extraction controls, as well as positive controls (DNA isolated directly from plants or Gastropoda). PCR amplicons were visualised using QIAxcel Advanced (QIAGEN) high-resolution capillary gel electrophoresis. A 5 µl aliquot of the amplicon PCR products was purified using an Illustra™ ExoproStar 1-Step Kit for enzymatic PCR clean-up (GE Healthcare, UK) according to the manufacturer’s protocol. After this purification, an index PCR was performed in order to individually mark each - 86 - x PCR product with specific Illumina indices added to the P5 and P7 sequencing adapters (Supplementary material A2). Index PCR products were purified and normalised with a SequalPrep™ Normalization Plate Kit (Thermo Fisher Scientific, USA) and 2 µl of each normalised and individually tagged sample were pooled to finalise the NGS library. In total, 136 and 104 samples were successfully amplified with the metazoan and plant primers, respectively, and sent for sequencing (Table S1). The library was sequenced using 250‑bp paired‑end reads on a MiSeq desktop sequencer (Illumina) at SEQ-IT GmbH & Co. KG, Kaiserslautern, Germany. Bioinformatics analyses of sequences from faecal samples To transform the raw Illumina sequence data into a list of MOTUs (molecular operational taxonomic units) with assigned taxonomy, a custom workflow (Masello et al., 2021; for detailed steps see Supplementary material A3) in GALAXY (https://www.computational.bio.uni- giessen.de/galaxy; Afgan et al., 2018) was used. Subsequently, MOTUs that corresponded to regular fieldwork contaminants in faecal samples (bacteria, soil fungi, bird DNA) were manually discarded (Kleinschmidt et al., 2019). As short fragments are less likely to contain reliable taxonomic information (Deagle et al., 2009), sequences with a length of less than 100 bp and a BLASTn assignment match of less than 98% were discarded. MOTUs were assigned to the lowest shared taxonomic level (Kleinschmidt et al., 2019; Table S3). Those that could not be determined at least at family level were excluded. Further filter steps were performed to obtain reliable data, i.e. avoid contamination and false positives (Crisol-Martínez et al., 2016): MOTUs were accepted only if they contained a minimum of five sequences or accounted for > 1% of the maximum total of hits per columbiform species. For each MOTU, we identified the highest read number within the negative samples and removed this MOTU from any sample where the read number was below this threshold. Statistical analysis All statistical analyses were carried out in R v.4.0.4 (R Core Team, 2021). For dietary overlap analyses and statistical analyses, we used the presence or absence data of each MOTU or respective genus or family. The frequency of occurrence ‘FOO’ per single species was calculated as FOO% = (n/t)*100, where ‘n’ is the number of samples, in which the MOTU was detected, and t is the total number of samples (Table 1). Since the data are qualitative data, ꭓ² tests (pairwise for species) were performed to compare frequencies of plant families and genera between columbiform species (Table S4, S5). Furthermore, we tested for differences in diet species composition at family and genus level with permutation tests in the R package ‘VEGAN’ (Oksanen et al., 2009). Non-metric Multidimensional Scaling (NMDS, function metaMDS) was used to visualize species differences in diet compositions. NMDS uses rank orders to collapse information from multiple - 87 - Chapter 4 | Diet analysesx dimensions into usually two dimensions to facilitate visualization as well as interpretation and is generally considered the most robust unconstrained ordination method in community ecology (Faith et al., 1987; Minchin, 1987). The function metaMDS allowed us to investigate the agreement between the two-dimension configuration and the original configuration through a stress parameter (if the stress value < 0.1 the agreement is very good, < 0.2 is a good representation). For this analysis at family level samples containing only a single plant family (n = 8) were discarded. All stress values in the present tests were < 0.25 at family and < 0.19 at genus level. We performed Permutational Multivariate Analysis of Variance Using Distance Matrices (PERMANOVA) with the function adonis and checked for the multivariate homogeneity of group dispersions (variances) with the function betadisper. To assess the dietary overlap of each species pair according to the presence/absence data at family and genus level of valid MOTUs, we calculated Pianka's measure of overlap Ojk (Pianka, 1986) in the R package ‘SPAA’ (Zhang, 2016) using the niche.overlap function. To evaluate differences between the species in the proportion of wild to cultivated plant species we categorised the MOTUs in five broad categories according to their likely source (Table S3; Dunn et al., 2018): ‘fed’ (seeds likely to be offered at bird feeders), ‘cultivated’ (crop plants as well as those widely cultivated as components of seed mixes sown to provide seed for wild birds), ‘natural’ (wild plant species), ‘tree’, and ‘brassica’ (all MOTUs of the family Brassicaceae). ‘Brassica’ was considered as a separate category as the family of Brassicaceae includes plants used to provision birds, as well as cultivated and several naturally occurring wild species (Dunn et al., 2018). If a species and respective genus occurred both as separate MOTUs in one species, e.g. Achillea sp. and Achillea millefolium, they were combined for categorisation. Results Diet composition – metazoan DNA Apart from the consumption of a few insects (9 samples, Table S3), only one valid metazoan prey MOTU was present in the faecal sample of one Stock Dove. This was DNA of a common earthworm Lumbricus terrestris (Table S3). Due to the low presence of animal prey in our samples, the further statistical evaluation refers only to the plant components found in the faecal samples. - 88 - x Table 1 Presence of valid MOTUs (molecular operational taxonomic units) of Spermatopsida in the diet of Common Woodpigeons Columba palumbus (WP), European Turtle Doves Streptopelia turtur (TD), and Stock Doves C. oenas (SD) with the respective frequency of occurrence (FOO%) of each MOTU. Order Family Genus Species MOTU Common FOO % name WP TD SD Apiales Apiaceae Heracleum - Heracleum sp hogweeds - 5.6 - Araliaceae Hedera - Hedera sp ivies 6.3 - 1.9 Asterales Asteraceae Achillea - Achillea sp yarrows 15.6 - - A. millefolium A. millefolium common 15.6 - 3.7 yarrow Artemisia A. vulgaris A. vulgaris common 6.3 - 5.6 mugwort Bellis B. perennis B. perennis common 3.1 - - daisy Carthamus C. tinctorius C. tinctorius safflower - - 3.7 Cichorium - Cichorium sp chicories - - 3.7 Cirsum - Cirsum sp thistles - 11.1 - Crepis C. capillaris C. capillaris smooth 3.1 5.6 - hawksbeard Dittrichia D. graveolens D. graveolens stinkwort - - 3.7 Guizotia G. abyssinica G. abyssinica niger seed 6.3 - - Helianthus H. annuus H. annuus annual 21.9 11.1 7.4 sunflower Hypochaeris H. radicata H. radicata catsear 3.1 - - Lactuca Lactuca sp lettuce 6.3 - - Scorzoneroides S. autumnalis S. autumnalis autumn 6.3 - - hawkbit Senecio S. inaequidens S. inaequidens narrow- 3.1 - - leaved ragwort Sonchus - Sonchus sp sow thistles 3.1 - - Taraxacum - Taraxacum sp dandelions 21.9 11.1 - Tripleurospermum - Tripleurospermum mayweeds - - 5.6 sp Boraginales Boraginaceae Echium E. vulgare E. vulgare blueweed - 5.6 - Brassicales Brassicaceae Raphanus - Raphanus sp radishes - - 9.3 Sinapis S. alba S. alba white - 5.6 3.7 mustard Brassica - Brassica sp cole crops 28.1 50.0 68.5 B. juncea B. juncea brown - 22.2 33.3 mustard B. napus B. napus rapeseed 6.3 22.2 38.9 B. oleracea B. oleracea cabbage - - 13.0 B. rapa B. rapa bird rape 6.3 16.7 38.9 Cardamine C. hirsuta C. hirsuta hairy - 5.6 - bittercress Caryophyllales Amaranthaceae Chenopodium - Chenopodium sp goosefoots 12.5 16.7 5.6 CaryophyllaceaeC erastium - Cerastium sp mouse-ear 3.1 - - chickweeds Sagina S. apetala S. apetala annual 3.1 - - pearlwort Silene - Silene sp campions 3.1 - - S. latifolia S. latifolia white 3.1 - - campion S. vulgaris S. vulgaris bladder - - 1.9 campion Stellaria S. media S.media chickweed - - 7.4 Cucurbitales Cucurbitaceae Cucumis - Cucumis sp 40.6 - 25.9 Cucurbita - Cucurbita sp gourd 65.6 27.8 51.9 C. pepo C. pepo field 43.8 16.7 35.2 pumpkin Dipsacales Adoxaceae Sambucus S. nigra S. nigra elder 6.3 - - Ericales Balsaminaceae Impatiens - Impatiens sp snapweeds - - 5.6 I. parviflora I. parviflora small - - 5.6 balsam Fabales Fabaceae Glycine G. max G. max soya bean 6.3 11.1 - - 89 - Chapter 4 | Diet analysesx - Table 1 continued - Order Family Genus Species MOTU Common FOO % name WP TD SD Lotus - Lotus sp bird’s-foot 3.1 - - trefoils Pisum P. sativum P. sativum pea - - 20.4 Robinia - Robinia sp locusts - 11.1 - Trifolium T. pratense T. pratense red clover 3.1 - - T. repens T. repens white clover 6.3 - - Vicia - Vicia sp vetches - - 55.6 V. hirsuta V. hirsuta hairy vetch - - 38.9 V. lathyroides V. lathyroides spring vetch - - 11.1 V. sativa V. sativa common - - 31.5 vetch V. sepium V. sepium bush vetch - - 7.4 V. tetrasperma V. tetrasperma smooth - - 9.3 vetch Fagales Betulaceae Betula - Betula sp birches 9.4 11.1 - Carpinus - Carpinus sp hornbeams 6.3 - - Fagaceae Fagus - Fagus sp beeches 9.4 - 11.1 Juglandaceae Juglans J. regia J. regia walnut 3.1 - 1.9 Gentianales Rubiaceae Galium - Galium sp bedstraws 3.1 - - Lamiales Plantaginaceae Hippuris - Hippuris sp mare's tails - - 5.6 Plantago P. lanceolata P. lanceolata buckhorn 18.8 - 5.6 plantain Veronica V. chamaedrys V. chamaedrys cat’s eyes - - 3.7 Liliales Liliaceae Lilium - Lilium sp lilies 3.1 - - Malpighiales Euphorbiaceae Euphorbia E. helioscopia E. helioscopia sun spurge - 11.1 3.7 Mercurialis M. annua M. annua annual - - 5.6 mercury Linaceae Linum - Linum sp flax plants - 5.6 - Malvales Malvaceae Tilia - Tilia sp linden 9.4 - - T. platyphyllos T. platyphyllos large-leaved 6.3 - - linden Myrtales Lythraceae Lythrum L. salicaria L. salicaria purple - 5.6 - loosestrife Onagraceae Epilobium - Epilobium sp willowherbs - 5.6 1.9 Oenothera - Oenothera sp evening - - 5.6 primroses Pinales Pinaceae Picea - Picea sp spruces 9.4 - 22.2 Pinus - Pinus sp pines - 5.6 - P. sylvestris P. sylvestris European 9.4 5.6 9.3 red pine Poales Cyperaceae Carex - Carex sp sedges 31.3 16.7 22.2 Poaceae - - Poaceae grasses 96.9 94.4 88.9 Agrostis - Agrostis sp bentgrasses 12.5 - 1.9 Alopecurus A. myosuroides A. myosuroides black-grass - - 1.9 A. pratensis A. pratensis meadow 15.6 5.6 - foxtail Arrhenatherum - Arrhenatherum sp oatgrasses 12.5 11.1 - A. elatius A. elatius false oat- 12.5 11.1 - grass Avena - Avena sp oats 15.6 5.6 11.1 Dactylis D. glomerata D. glomerata cock’s-foot 6.3 - 11.1 Elymus - Elymus sp couch - - 3.7 grasses Festuca - Festuca sp fescues 9.38 - - Holcus - Holcus sp soft-grasses - 11.11 - Hordeum H. vulgare H. vulgare barley 21.9 33.3 27.8 Lolium - Lolium sp ryegrasses 15.6 - 3.7 L. perenne L. perenne perennial 15.6 - 3.7 ryegrass Molinia M. caerulea M. caerulea purple 28.1 - 14.8 moorgrass Panicum P. miliaceum P. miliaceum proso millet 21.9 50.0 18.5 Phalaris - Phalaris sp 9.4 - - - 90 - x - Table 1 continued - Order Family Genus Species MOTU Common FOO % name WP TD SD Poa - Poa sp meadow- 15.6 11.1 5.6 grasses P. trivialis P. trivialis rough 12.5 - - bluegrass Secale S. cereale S. cereale rye 9.4 16.7 18.5 Setaria Setaria sp bristle 6.3 11.1 5.6 grasses Trisetum T. flavescens T. flavescens yellow 3.1 - - oatgrass Triticum - Triticum sp wheat 68.8 66.7 77.8 T. aestivum T. aestivum common 25.0 16.7 35.2 wheat T. dicoccon T. dicoccon emmer 9.4 - - wheat T. spelta T. spelta dinkel wheat - - 11.1 Zea Z. mays Z. mays maize 34.4 16.7 16.7 Ranunculales Ranunculaceae Ranunculus - Ranunculus sp buttercups - 27.8 - Rosales Cannabaceae Cannabis C. sativa C. sativa hemp 25.0 27.8 27.8 Elaeagnaceae Hippophae H. rhamnoides H. rhamnoides sea- - - 1.9 buckthorn Rosaceae Amelanchier - Amelanchier sp shadbushes 6.3 - - Potentilla - Potentilla sp cinquefoils 9.4 - - Prunus - Prunus sp 15.6 - 3.7 P. avium P. avium bird cherry 3.1 - - Rosa - Rosa sp roses 3.1 11.1 - Rubus - Rubus sp 6.3 22.2 1.9 Urticaceae Urtica U. dioica U. dioica common 18.8 16.7 11.1 nettle Sapindales Sapindaceae Acer - Acer sp maples 18.8 - 9.3 A. platanoides A. platanoides norway 6.3 - - maple A. A. pseudoplatanus sycamore 6.3 - - pseudoplatanus Saxifragales Crassulaceae Sedum - Sedum sp stonecrops 3.1 - - Solanales Convolvulaceae Convolvulus C. arvensis C. arvensis field 3.1 - - bindweed Solanaceae Solanum S. lycopersicum S. lycopersicum tomato 6.3 - 14.8 Diet composition – plants Of all faecal samples successfully amplified with the plant primers (n = 104), at least one valid MOTU was found in every sample with an average of 9.3 ± 5.8 MOTUs per sample (Stock Dove = 10.0 ± 5.0, Woodpigeon = 9.8 ± 7.5, Turtle Dove = 6.4 ± 4.1). A total of 118 MOTUs were found, with 54.2% of MOTUs determined at species level, 44.9% at genus level and 0.8% at family level (Table 1). Most MOTUs were found in samples of Woodpigeons (79 MOTUs), followed by Stock Doves (67) and Turtle Doves (44). All MOTUs belonged to the class Spermatopsida, distributed among 23 orders and 34 families (Table 1; Fig. 1). Asteraceae, Brassicaceae, Cucurbitaceae, Fabaceae and Poaceae have been the most frequently represented plant families, occurring with a FOO of over 50% in at least one of the tested species (Fig. 1). These five families contained varying numbers of genera (17, 4, 2, 6 and 19, respectively), that occurred with varying FOO% (Fig. S2; Table S5). - 91 - Chapter 4 | Diet analysesx Figure 1 Diet composition of Common Woodpigeons Columba palumbus (WP), European Turtle Doves Streptopelia turtur (TD) and Stock Doves C. oenas (SD). Summary of plant families found in faecal samples represented as frequency of occurrence (FOO). ‘*’ indicates a significant difference (p < 0.05) in the occurrence (presence/absence data) of respective families between two species (Table A4). Diet differences among species of Columbiformes The frequency of most plant families (77.1%) did not differ significantly between the columbiform species. However, significant differences were present in eight of the families (Fig. 1; Table S4). Overall, the community analyses showed that the diet composition differed between the columbiform species at plant family level (Fig. 2) as indicated by permutation tests (Permutation test for differences: F93,2 = 7.1, p < 0.001). However, the difference in species explained only 13.3% of the overall variation (R² = 0.133; Fig. 2). Likewise, the result at genus level (Fig. S3) pointed out differences between the species’ diet composition (Permutation test for differences: F101,2 = 4.8, p < 0.001), though this difference also explained a rather small proportion (8.7%) of the overall variation (R² = 0.087). According to Pianka's measure of overlap at family level Woodpigeons and Stock Doves showed the highest dietary overlap (Ojk = 0.768), followed by Stock Doves and Turtle Doves (Ojk = 0.636). Turtle Doves and Woodpigeons had the least plant families in common (Ojk = 0.551). Also at genus level, Woodpigeons and Stock Doves showed the highest dietary overlap (Ojk = 0.605). The overlap between Turtle Doves and Woodpigeons (Ojk = 0.504) as well as Turtle Doves and Stock Doves (Ojk = 0.504) was equal. Most MOTUs were assigned to the category ‘natural’ (55.1%), followed by ‘cultivated’ (15.3%) and ‘tree’ (14.4%). The remaining MOTUs were categorised as ‘brassica’ (5.9%) or ‘fed’ (5.1%). Some MOTUs (4.2%) could not be clearly assigned (‘NA’) to one of the categories (Table S3). In all three species, most MOTUs were assigned to the category ‘natural’ (Woodpigeon = 52.9%, Turtle Dove = 48.7%, Stock Dove = 46.8%), followed by ‘cultivated’ for Turtle Doves (17.9%) and Stock Doves (19.4%) and ‘tree’ for Woodpigeons (17.6%; Fig. 3). - 92 - x None of the proportion of categories varied significantly between the columbiform species (pairwise t-test all χ2 < 2.6, df = 1, p > 0.110). Figure 2 Differences in the diet composition at plant family level in three columbiform species (Common Woodpigeon Columba palumbus (WP); European Turtle Dove Streptopelia turtur (TD); Stock Dove C. oenas (SD)), using Non-metric Multidimensional Scaling (NMDS, function metaMDS in the R package ‘VEGAN’). Depicted are (A) the distribution of the plant families (the word ending ‘- ceae’ was removed to avoid overlapping) and (B) the distribution of samples and 95% confidence ellipses. - 93 - Chapter 4 | Diet analysesx Figure 3 Proportion of dietary component categories of three columbiform species (Common Woodpigeon Columba palumbus (WP), European Turtle Dove Streptopelia turtur (TD), Stock Dove C. oenas (SD)). The categories reflect the likely source of the dietary item (MOTU, Table S3). Proportion is given as percent [%] based on the presence data of MOTUs per species. ‘NA’ indicates that the MOTU could not be assigned clearly to a category. Discussion The main aim of the present study was to analyse the diet composition of Common Woodpigeons, European Turtle Doves and Stock Doves, using NGS technology and metabarcoding, and to compare the diet composition found between the species and to the results of previous studies. Diet reconstruction based on NGS technology and comparison with previous studies Animal constituents The DNA data from faecal samples of the columbiform species show a diverse range of taxa, dominated mainly by plant constituents, while animal prey was present very rarely. In line with our study, most other studies found no or only little numbers of animal material. In a review, the proportion of invertebrate components in the diet of all three species of Columbiformes was below 5% (Holland et al., 2006). Murton et al. (1964) reported the intake of cocoons of earthworms for Stock Doves. We found earthworm DNA in the faeces of one Stock Dove individual. However, with the applied method we cannot determine the development stage, e.g. cocoon, larvae or imago, of consumed prey. Woodpigeons sampled in Algeria and Spain were exclusively herbivorous (Jimenez et al., 1994; Kaouachi et al., 2021). In other studies, animal constituents were observed with small volumes and low frequency in Woodpigeon diet (Ó hUallachain & Dunne, 2013; Gutiérrez- Galán et al., 2017; Negrier et al., 2021). - 94 - x Animal prey was present in only 2.7% of Turtle Doves in Spain or completely absent in other years (Gutiérrez-Galán & Alonso, 2016). Another study on Turtle Dove diet from Spain (Jimenez et al., 1992), finding three gastropods (Helicella sp.) in 64 samples, states the animal fraction in the diet as insignificant compared to seeds. Animal material, particularly calcareous shells, might be mainly consumed to cover the need for calcium (Möckel, 1988; Glutz von Blotzheim & Bauer, 1987), but shells would likely not result in detectable DNA in the faeces. Moreover, the birds might also meet their mineral requirements through the consumption of small clods of earth (Glutz von Blotzheim & Bauer, 1987; Downs et al., 2019) and together, this might explain the absence of animal DNA in the individuals examined here. A few insect species were detected in the faecal samples (Table S3). Insects and Crustacea have occasionally been found in previous studies, e.g. Cecidomyiidae larvae or Coleoptera in Stock Doves (Möckel, 1988), Coccoidea, larvae and cocoons of Lepidoptera in Woodpigeons (Murton et al., 1964; Glutz von Blotzheim & Bauer; 1987, Ó hUallachain & Dunne, 2013). However, these animals were likely not consumed on purpose but taken ‘accidentally’ while eating plant components or during plumage care. Plant constituents Results obtained in the diet of Columbiformes showed a wide diversity in consumed plants. The applied molecular DNA metabarcoding approach detected a larger number of plant families than former analyses based on visual or observational identification of food items (Table S6-8). For Stock Doves, we detected 22 plant families (Table S6). Previously, seeds and fruits of plants from the families Poaceae, Fabaceae, Brassicaceae, Polygonaceae and Caryophyllaceae were described as the most important food items for Stock Doves (Glutz von Blotzheim & Bauer, 1987), whereby especially seeds of wild and cultivated vetches Vicia sp. (Fabaceae) comprise a major part (Murton et al., 1965; Möckel, 1988). In line with this, four of the five aforementioned plant families were present in Stock Doves sampled in our study (FOO%: Poaceae = 88.9%, Fabaceae = 64.8%, Brassicaceae = 70.4%, Caryophyllaceae = 9.3%, Fig. 1). The important proportion of vetches for Stock Dove diet is also supported by our data. Vicia DNA could be traced in 55.6% of all Stock Dove samples with the hairy vetch Vicia hirsuta being the most frequent (Table 1), whereas Vicia DNA was not found in Woodpigeon or Turtle Dove samples. Nine plant families have not been mentioned as being part of Stock Dove diet in previous studies (Table S6). While seven of these families had a FOO lower than 15%, Pinaceae (27.8%) and Cucurbitaceae (55.6%) were present in more individuals. Overall, our results support the previous assumptions that especially Poaceae, Brassicaceae and Fabaceae, particularly Vicia sp., are characteristic for Stock Dove diet. Generally, the diet of Stock Doves seems less intensely studied compared to Woodpigeons and Turtle Doves. - 95 - Chapter 4 | Diet analysesx The diet of Woodpigeons has been studied more extensively, likely because they were appraised as a pest of growing crops (Ückermann, 1985). The main food of Woodpigeons, considered a granivorous-frugivorous species, consists of different kinds of seeds and green plant parts. Acorns, beechnuts, maple and other tree seeds are consumed. Cereals, different berries, e.g. elder or ivy and drupes are also eaten, as well as buds, young leaves and young shoots of deciduous trees and conifers. In some areas and times of the year, leaves of rape, cabbage and clover can constitute the main part of the food (Ückermann, 1985; Gutiérrez- Galán et al., 2017). Woodpigeons are regarded as generalist/opportunistic feeders, feeding on various food items and switching to alternative species when preferred ones are unavailable, leading to a pronounced seasonal variation (Gutiérrez-Galán, et al. 2017; Kaouachi et al., 2021). This generalist feeding is also reflected in our results, as Woodpigeon samples contained the highest number of plant families (n = 25; Fig. 1). However, Woodpigeons were the only species of which we also had samples from the non-breeding season and this may lead to the result of a more variable diet due to the seasonal variation in available and preferred food items. Most of the plant families detected in our samples were already described as part of Woodpigeon diet. Only Crassulaceae, Juglandaceae, Liliaceae, Malvaceae and Sapindaceae were not mentioned in previous studies (Table S7). Some MOTUs found for Woodpigeons in our study might differ from previous studies, as many studies concentrated on sampling in rural and agricultural areas, whereas most of our Woodpigeon samples originated from (sub)urban habitats. Once a typical and exclusive woodland species, Woodpigeons colonised cities of Western and Central Europe since the early 19th century (Fey et al., 2015). Urban areas typically contain novel food items, such as non-native species, intentionally (e.g. bird feeders) or unintentionally provisioned food (e.g. garbage in landfills). Therefore, many wildlife species shift their diets to use these ‘novel’ food resources (Murray et al., 2018). Some examples for the food items that were most likely found in urban areas solely are the MOTUs Amelanchier sp. (ornamental shrub), Sedum sp. (ornamental garden plant; roof covering in green roofs) and Lilium sp.(ornamental plant). Other MOTUs likely originate from food provided in bird feeders (Table S3, category ‘fed’): Relatively frequently found in Woodpigeon faecal samples were e.g. sunflower Helianthus annuus, niger seed Guizotia abyssinica and proso millet Panicum miliaceum (Table 1). Provided seeds in urban areas, e.g. wheat, maize or millet, have probably also contributed to the high FOO% of Poaceae (96.9%) in Woodpigeons, but it also is known that individuals from (sub-)urban areas move out to agricultural areas to feed upon farmland there (Slater et al., 2001). Overall, the diet of the Woodpigeon fits into the known pattern with some peculiarities in the diet of individuals from urban areas. - 96 - x Turtle Doves are considered obligate granivorous birds (Fisher et al., 2018). Glutz von Blotzheim and Bauer (1987) name seeds of Polygonaceae, Papaveraceae, Brassicaceae, Asteraceae, Poaceae, Pinaceae, Faboideae and Chenopodium sp. to constitute the main diet on breeding grounds. While Poaceae (FOO = 94.4%), Brassicaceae (61.1%), Asteraceae (27.8%), Faboideae (16.7%), Chenopodium sp. (16.7%) and Pinaceae (5.6%) occurred in our Turtle Dove samples, Polygonaceae and Papaveraceae were not detected. The feeding ecology of Turtle Doves changed significantly from non-cultivated, natural arable plants, primarily weed seeds, to mainly cultivated plants, such as rape and wheat, from the 1960s to the late 1990s in the UK (Browne & Aebischer, 2003). Also, seeds provided at bird feeders were recently found in Turtle Dove diet (Dunn et al., 2018). Fumitory Fumaria sp. historically formed the mainstay of Turtle Dove diet. Individuals sampled in the UK also commonly ate other natural plants, e.g. S. media, scarlet pimpernel Anagallis arvensis, cock’s-foot Dactylis glomerata, Poa sp., Geraniaceae and Amaranthaceae (Murton et al., 1964; Fisher et al., 2018; Dunn et al., 2018). After the 1990s cultivated seeds, mainly Triticum sp. and rape Brassica napus, were the main food items in the UK (Browne & Aebischer, 2003). Our results also reflect the dietary shift from wild plants to cultivated ones On the one hand MOTUs categorized as ‘natural’, except for Ranunculus sp. (27.8%) and Rubus sp. (22.2%), occurred with FOO lower than 20%, while cultivated ones reached higher FOO (Triticum sp.= 66.7%; Brassica sp. = 50.0%, including B. napus with 22.2%). On the other hand, we did not find some historically important food items, particularly Fumaria sp. and S. media, even though they grow in Germany and the Netherlands (Sparrius et al., 2014; Metzing et al., 2018). Similar to our results, these natural plant species, classified as important in Turtle Dove diet, in particular in the UK, were also absent in other regions (Romania and Slovakia: Glutz von Blotzheim & Bauer, 1987; Russia: Murton et al., 1965; Spain: Gutiérrez- Galán & Alonso, 2016). The comparison with previous studies shows that only the plant family Poaceae was present in Turtle Dove diet in all the represented European countries (Table S8). To our knowledge, the families Betulaceae, including the MOTU Betula sp., Cyperaceae (MOTU: Carex sp.) and Lythraceae (MOTU: Lythrum salicaria) were so far not mentioned as part of Turtle Dove diet (Table S8). The observed regional dietary differences may be due to climatic and biogeographical differences as well as variation in habitat, e.g. agricultural landscape vs forest, and occurrence and availability of certain plant species (Gutiérrez-Galán & Alonso, 2016; Mansouri et al., 2019). Dietary composition differences between species The degree of dietary overlap between the studied columbiform species pairs was slightly lower than observed by Dunn et al. (2018) with Pianka’s measure, ranging from 0.7 to 0.9 compared to 0.5 to 0.8 in our study. Dietary overlap between the species suggests that some resources are shared and the species might compete for food. However, it has been suggested that the related columbiform species select different feeding sites, occupy different ecological - 97 - Chapter 4 | Diet analysesx niches or utilise superabundant supplies if taking the same food items, indicating rather little or no competition between them (Murton et al., 1964; Jimenez et al., 1994). For some plant families and genera, we found significant differences in their occurrence for the tested species (Table S4, S5). In addition, the permutation tests indicated significant variance in diet composition among the species (Fig. 2, S3). However, both at plant family and genus level, the differences among species explained only a rather small proportion of the overall variation (13.3 and 8.7%, respectively). This implies a rather pronounced variability within species, which is also supported by the rather strongly varying number of MOTUs detected per sample (1 to 33). Furthermore, with the use of DNA metabarcoding, we cannot distinguish which part of the plant was eaten and the different species may feed on different parts of the same plant species, e.g. Woodpigeons eat the young leaves of Brassicaceae, whereas Turtle Doves feed on Brassica seeds. This can result in the degree of dietary overlap being overestimated. Another limitation of the method is that only presence/absence data of food items are obtained and thus, quantitative assessment of the proportion of consumed plants is not possible. Even though most MOTUs were assigned to the category ‘natural’ for all three species (Fig. 3), it cannot be assumed that these proportionally form the main part of the diet. Based on previous studies, seeds and plant material of ‘cultivated’ species are expected to constitute the main fraction of the diet nowadays. In Woodpigeons sampled in Spain, 97.6% in volume corresponded to cultivated plants (Jimenez et al., 1994). Wheat and rape seeds averaged 61% of the seeds eaten by Turtle Doves in the UK (Browne & Aebischer, 2003). In Stock Doves, wheat and barley made up 80-90% of the diet in April (Möckel, 1988). Application of results for conservation measures While the population trends of Woodpigeons and Stock Doves are moderately increasing, Turtle Doves are declining across their entire European breeding range (-80% since the 1980s, PECBMS, 2021). The population decline occurred concurrently with decreases in the abundance of many noncultivated, natural arable plants and along with a decrease in reproductive output (Calladine et al., 1997; Browne & Aebischer, 2004; Dunn et al., 2018). Changes in farming practice and agricultural intensification have caused a serious decrease in weed seed abundance in European farmland areas and the availability of seeds from natural arable plants has declined. Therefore, it has been suggested that the dietary change is associated with a reduction in food availability during important periods of the breeding season, when seeding natural arable plants have become scarce, and this may constitute a stressor for Turtle Dove populations (Browne & Aebischer, 2004; Dunn et al., 2015, 2018; Gutiérrez- Galán & Alonso, 2016). For instance, the condition of Turtle Dove nestlings fed with cultivated seeds was poorer than that of those fed with wild seeds (Dunn et al., 2018) and the availability of wild plant seeds is considered one of the main breeding habitat requirements (Dunn & Morris, 2012). - 98 - x Thus, the development of an extensive, seed-provisioning option is considered vital for the conservation of Turtle Doves and options to enhance food availability should favour the provision of wild seeds rather than cultivated seeds (Dunn et al., 2015). However, most existing options, e.g. agri‑environment schemes (AES) or agri-environmental policies (AEP), are suboptimal in providing accessible food for Turtle Doves (Dunn et al., 2015). A tailored sown mix, based on plant species known to be present in Turtle Dove diet historically, has been devised by an RSPB/Natural England project aiming to provide optimal foraging conditions. However, even though the sown plots provided more seeds compared to control plots, sown plots developed a too dense vegetation structure to attract foraging Turtle Doves. Therefore, modifications for the tailored sown mix were recommended (Dunn et al., 2015). Updated and improved knowledge of the seeds included in Turtle Dove diet will help to plan and carry out tailored management options as well as optimise feeding during rehabilitation and possible ex situ conservation projects, in particular as data on the Turtle Dove diet is fragmentary and limited (Mansouri et al., 2019). Our results and their comparison with previous studies highlight the presence of regional differences in Turtle Dove diet composition and that some plant species, (historically) considered important food items in some regions, might not be the major part of Turtle Dove diet in other regions despite their existence in the studied areas. Further studies should focus on identifying regional dietary differences as they might play an important role in planning tailored seed mixes. It is probably advisable to plan the composition of seed mixtures according to locally preferred and known wild plant species in order to achieve the best possible acceptance of the managed foraging and feeding areas by the Turtle Doves. - 99 - Chapter 4 | Diet analysesx Statements Ethical approval All applicable institutional and/or national guidelines for the care and use of animals were followed. Capture and handling were carried out under licenses of the regional council Hesse (license number TVA-51/2017), the state office for occupational safety, consumer protection and health, Brandenburg (license number 2347-11-2018) and the Nederlandse Ringcentrale (project AVD 801002015342). Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Funding Financial support was received from the Naturschutz Bund Deutschland e.V. (BirdLife Germany) and the Hessische Gesellschaft für Ornithologie und Naturschutz (HGON) for fieldwork in Germany. JFM work was partly funded by the Hessen State Ministry for Higher Education, Research and the Arts, Germany, as part of the LOEWE priority project Nature 4.0 – Sensing Biodiversity. Acknowledgements Thanks are due to the many landowners who allowed access to their land for bird catching, helpers in the field (amongst others: Hagen Deutschmann, Lennart Wegner, Jennifer Greiner, Leslie Koch and Thiemo Karwinkel) and Klaus Klehm for providing Stock Dove samples. Marc Kümmel, Sven Griep and Alexander Goesmann of the Bioinformatics group of the Justus- Liebig University Giessen developed the Galaxy tool. We thank the staff of the Clinic for Birds, Reptiles, Amphibians and Fish of JLU Giessen for the sampling of admitted patient birds and the association to support avian medicine Giessen (Verein zur Foerderung der Vogelmedizin in Giessen e.V.) for support. - 100 - x References Afgan, E., Baker, D., Batut, B., et al. (2018). The Galaxy platform for accessible, reproducible and collaborative biomedical analyses: 2018 update. Nucleic Acids Res, 46, W537-W544. https://doi.org/10.1093/nar/gky379. 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Pompanon, F., Deagle, B.E., Symondson, W.O.C., Brown, D.S., Jarman, S.N., Taberlet, P. (2012). Who is eating what: diet assessment using next generation sequencing. Mol Ecol, 21, 1931-1950. https://doi.org/10.1111/j.1365-294X.2011.05403.x. R Core Team (2021) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. https://www.R-project.org/. Slater, P. (2001). Breeding ecology of a suburban population of Woodpigeons Columba palumbus in northwest England. Bird Study, 48, 361-366. Sparrius, L.B., Odé, B., Beringen, R. (2014). Basisrapport voor de Rode Lijst Vaatplanten 2012. FLORON-rapport 57. FLORON, Nijmegen. Ückermann, E. (1985). Die Ringeltaube (Columba palumbus). Zur Ökologie und Biologie flugbetriebsgefährdender Vogelarten. Vogel und Luftverkehr, 5, 45-54. Valentini, A., Pompanon, F., Taberlet, P. (2008). DNA barcoding for ecologists. Trends Ecol Evol, 24, 110-117. https://doi.org/10.1016/j.tree.2008.09.011. Wood, J.E. (1954). Food habits of furbearers of the upland post oak region in Texas. J Mammal, 35, 406-415. Zhang, J. (2016). R package ‘spaa’ SPecies Association Analysis. https://github.com/helixcn/spaa - 103 - Chapter 4 | Diet analysesx Information on the ‘Supplementary Material’ can be found in the appendix - 104 - x - 105 - CHAPTER 5 PREVALENCE AND GENETIC DIVERSITY OF AVIAN HAEMOSPORIDIAN PARASITES IN WILD BIRD SPECIES OF THE ORDER COLUMBIFORMES Yvonne R. Schumm, Dimitris Bakaloudis, Christos Barboutis, Jacopo G. Cecere, Cyril Eraud, Dominik Fischer, Jens Hering, Klaus Hillerich, Hervé Lormée, Viktoria Mader, Juan F. Masello, Benjamin Metzger, Gregorio Rocha, Fernando Spina, Petra Quillfeldt Parasitology Research (2021) 120: 1405 - 1420. DOI: 10.1007/s00436-021-07053-7 - 106 - - 107 - Chapter 5 | Avian haemosporidian parasitesx - 108 - - 109 - Chapter 5 | Avian haemosporidian parasitesx - 110 - - 111 - Chapter 5 | Avian haemosporidian parasitesx - 112 - - 113 - Chapter 5 | Avian haemosporidian parasitesx - 114 - - 115 - Chapter 5 | Avian haemosporidian parasitesx - 116 - - 117 - Chapter 5 | Avian haemosporidian parasitesx - 118 - - 119 - Chapter 5 | Avian haemosporidian parasitesx - 120 - - 121 - Chapter 5 | Avian haemosporidian parasitesx - 122 - APPENDIX SUPPLEMENTARY MATERIAL The following supplementary material is saved on a CD (attached to this thesis). The files are stored in folders sorted and named according to the chapters: CHAPTER 1 | Electronic supplementary material  Supplementary Figure 1 Assignment to likely wintering origin (moulting areas of winter- grown primary feathers) of European turtle doves Streptopelia turtur (N = 181) sampled in seven different European countries (labeled and gray shaded) predicted from a multivariate normal probability distribution function based on tenth primary feather (P10) δ2H and δ13C isotope assignments of individual birds. Assignment probabilities of individuals (0 to 1) were summed according to the maximum value obtained in a pixel during the assignment process for the single countries (shown in red: a: Spain, b: France, c: Germany, d: Italy, e: Malta, f: Bulgaria, and g: Greece) representing the percent of individuals potentially originating from a cell in the isoscape. The assignment is restricted to a hitherto described turtle dove wintering range (red outline)  Supplementary Figure 2 Assignments to likely wintering origin (moulting areas of winter- grown primary feathers) of European turtle doves Streptopelia turtur predicted from a normal probability distribution function based on tenth primary feather (P10) δ2H isotope assignments of individual birds. (a): All individuals (n = 181), sampled in seven different European countries (labeled and shaded gray). Separated for (b): the western flyway (n = 121) and (c): the central/eastern flyway (n = 55). Assignment probabilities of individuals (0 to 1) were summed according to the maximum value obtained in a pixel during the assignment process for the respective sample set representing the percent of individuals potentially originating from a cell in the isoscape. The assignments are restricted to a hitherto described turtle dove wintering range (outline in red) CHAPTER 2 | Electronic supplementary material  Electronic supplementary material File 1, including: o Supplementary Table 1 Percentage [%] of land cover classes occurring in the 95% Epanechnikov kernels of breeding habitats of satellite-tracked European turtle dove individuals o Supplementary Table 2 Percentage [%] of land cover classes occurring in the 95% Epanechnikov kernels of wintering habitats of satellite-tracked European turtle dove Streptopelia turtur individuals - 123 - Appendixx o Supplementary Table 3 Results of the principal component analysis (PCA) conducted for seven habitat variables obtained through the Environmental Data Automated Track Annotation System (Env-DATA) on Movebank (movebank.org) for positions of satellite-tracked European turtle doves Streptopelia turtur o Supplementary Table 4 Studies on habitat selection patterns and habitat requirements of European turtle doves Streptopelia turtur o Supplementary Figure 1 Individual satellite tracks of European turtle doves Streptopelia turtur during migration between European breeding (red circles) and African wintering grounds (blue circles). Tracks are given for the individuals (a-e) which reached the wintering grounds and for individuals (f-n) which did not reach the sub-Saharan wintering grounds. Black circles correspond to stopover sites and ‘ǂ’ indicates stopover moult sites. Catching and tagging position is displayed as yellow cross and a black cross indicates the end of data transmission. Autumn migration is shown as solid black line and spring migration as dashed black line. Partial tracks are displayed in grey. Duration at the different sites is given in dd.mm (ST = Start of data transmission; ET = End of data transmission). Background colours indicate the terrain and grey lines indicate national borders (Background map: Stamen terrain (map tiles by Stamen Design: http://maps.stamen.com; data by OpenStreetMap: www.openstreetmap.org)) o Supplementary Figure 2 Breeding and post-breeding sites of European turtle dove #161050. Presented are the three years we received data during the entire breeding period (2017 = green, 2018 = orange, 2019 = blue). Shown are the filtered Argos location fixes as points and calculated Epanechnikov kernels (95% and 50% Kernel Utilization Distributions ‘KUD’ as dashed and solid lines, respectively). Duration at the different sites is given in dd.mm. Background map: Stamen terrain (map tiles by Stamen Design: http://maps.stamen.com; data by OpenStreetMap: www.openstreetmap.org) o Supplementary Figure 3 Overview chart of migration pattern of 13 satellite tracked European turtle doves Streptopelia turtur. Shown are the countries used for stopovers during autumn and spring migration and wintering for turtle doves with different breeding areas (colors indicate the country or federal state the breeding site was located). Country codes are given as two-letter code (ISO-3166-1 ALPHA- 2). ‘X’ represents that no stopover was done. Digits in brackets or at the lines indicate the number of individuals using the site. ‘Western’, ‘Central’ and ‘Eastern’ indicate the used flyway. ‘*’ as we have data from individual #161050 for consecutive years every of the years is depicted as ‘single’ count. If the data transmission has ended, the line was not continued from the last site - 124 - o Supplementary Figure 4 Habitat niches of satellite-tracked European turtle doves Streptopelia turtur (n = 5) for wintering (blue) and breeding (red) grounds. Obtained from Argos-positions and kernel densities of principal component scores of environmental parameters (n = 9) obtained through the Environmental Data Automated Track Annotation System (Env-DATA) on Movebank. a: individual #161046; b: #161048; c: #161050; d: #181091; e: #181092. Environmental parameters determining respective PC1 and PC2 as well as their eigenvalues can be found in detail in Supplementary Table 3  Electronic supplementary material File 2, excel file including: o Keys for 'TrackingData' sheets and 'EnvData' sheet o Raw Argos tracking data CHAPTER 3 | Electronic supplementary material  Electronic supplementary material File 1, including: o Supplementary Table 1 Circumstances of recovery of ringed Common Woodpigeons Columba palumbus. All individuals here were recovered during wintering timea in the different countries. o Supplementary Table 2 Home range (95% KUD, Epanechnikov kernel) and core area (50% KUD, Epanechnikov kernel) size of Common Woodpigeons Columba palumbus equipped with GPS-GSM transmitters. KUDs are given for individuals in Lisbon, Portugal (L; n = 10), Giessen, Germany (G; n = 19) and wintering sites of migrating individuals per month throughout the annual cycle [Mean ± SE]. Given in the brackets for 50% and 95% KUD is the number of months and individuals (no. months / no. individuals). Given in brackets in the column ‘Movements’ are the number of individuals visiting the farmland outside the city and the total number of individuals (no. visiting farmland / total no. of individuals). o Supplementary Table 3 Average proportions [%] of land cover categories in monthly home ranges calculated on the basis of GPS locations of tagged Common Woodpigeons Columba palumbus. Numbers are given for individuals from two regions and with different migrations strategies (L: Lisbon, Portugal; G: Giessen, Germany, residents and individuals during the non-wintering season; MD: Individuals using another distinct site during the wintering season than during the breeding season, but migratory movements occurred within Germany; MF: Woodpigeons migrating to France). Land cover categories were named according CORINE Land Cover (CLC) nomenclature. - 125 - Appendixx o Supplementary Figure 1 Overview of validated ring recoveries of Common Woodpigeon Columba palumbus with breeding and wintering grounds within Germany. Coordinates where ringed Woodpigeons were spotted or caught during breeding time (pink circles) and during wintering time (blue squares) are connected by a straight dashed line. In case of identical coordinates during breeding and wintering time, the location is displayed by a yellow star symbol. Background colours indicate the terrain, black lines national borders and grey lines borders of the federal states in Germany (Background map: Stamen Design http://maps.stamen.com; data by OpenStreetMap: www.openstreetmap.org)) o Supplementary Figure 2 Annual cycle of Common Woodpigeons Columba palumbus equipped with Argos-transmitters during the non-breeding season in Portugal and France. The map gives the spatial organisation with spring migration (solid line) and autumn migration (dashed line) between the breeding sites (circles) in Germany and the nonbreeding sites (squares). The star symbol indicates that breeding and non-breeding time were spent at the same location. The triangles indicate stopover sites and crosses that the last Argos-position was transmitted outside the breeding or non-breeding site. The inset shows the temporal organisation with percentages of time for period spent at the breeding site (dark grey), the non-breeding site (light grey) and on migration (striped = spring migration; dotted = autumn migration) and average arrival and departure for each respective period. Background colours indicate the terrain and grey lines indicate national borders (Background map: Stamen terrain (map tiles by Stamen Design: http://maps.stamen.com; data by OpenStreetMap: www.openstreetmap.org)) o Supplementary Figure 3 Alteration of longitudes of Argos locations of tracked Common Woodpigeon Columba palumbus within the annual cycle. The average longitude per decade is presented. Grey shaded time periods correspond to spring and autumn migration periods (for all individuals combined) - 126 - o Supplementary Figure 4 Diagram representing the migratory systems based on satellite tracking data of the studied Common Woodpigeon Columba palumbus populations: Resident population of Portugal, Lisbon (n = 12 winter periods from 10 individuals equipped with GPS transmitters) and partial migratory population of Germany, Hesse (GPS: n = 30 winter periods from 19 individuals; Argos: n = 7 winter periods from 5 individuals only data of the second year was included). Annual seasons are separated by month (given as numbers 01-12) and displayed by colour (yellow = summer, orange = autumn, blue = winter, green = spring). During the months (November – March) of the non-breeding season migrants and residents of partial migratory population winter in different habitats, whereas individuals of Lisbon share the same habitat the entire year. Height of boxes is proportional to the sample sizes of the two breeding populations and proportion of wintering strategies o Supplementary Figure 5 Annual cycle of migrating Common Woodpigeons Columba palumbus equipped in Hesse (DE) with GPS-GSM transmitters. The map gives the spatial organisation with spring migration (solid line) and autumn migration (dashed line) between the tagging and breeding sites in Germany (star symbol: C = Caldern, G = Giessen) and the non-breeding, i.e. wintering sites (squares). Black dashed lines depict movements between wintering sites. The triangles indicate stopover sites and the cross that the last GPS position was transmitted on the wintering site. The inset shows the temporal organisation with percentages of time for period spent at the breeding site (dark grey), the non- breeding site (light grey) and on migration (striped = spring migration; dotted = autumn migration) and average arrival and departure for each respective period. Background colours indicate the terrain and black lines indicate borders (Background map: Stamen terrain (map tiles by Stamen Design: http://maps.stamen.com; data by OpenStreetMap: www.openstreetmap.org)) o Supplementary Figure 6 Winter movements of Common Woodpigeons Columba palumbus equipped in Hesse (DE) with GPS-GSM transmitters. Individuals that wintered within Germany in proximity to their breeding ground in Giessen are shown, as well as the movements of individual #190759 between Giessen (G) and Herborn (H). The map shows spring movements (solid line) and autumn movements (dashed line) between the breeding sites (star symbol) and the non- breeding, i.e. wintering sites (squares). Black dashed lines depict movements between wintering sites and the cross that the last GPS position was transmitted on the wintering site. Background colours indicate CLC land cover categories (Corine Land Cover CLC 2018 v.2020_20u1 raster land cover data; Copernicus Land Monitoring Service) - 127 - Appendixx o Supplementary Figure 7 Core area size (50% KUD) of Common Woodpigeons Columba palumbus. Shown are Woodpigeons from two regions (Lisbon, Portugal and Giessen, Germany) and with different migrations strategies (Giessen (DE): residents and individuals during the non-wintering season; Migration (DE): Individuals using another distinct site during the wintering season than during the breeding season, but migratory movements occurred within Germany; Migration (FR): Woodpigeons migrating to France). Boxplots denote the median value, interquartile range (25–75th percentiles) and range of core area size. The star symbol represents the mean value. Outliers are plotted as individual points o Supplementary Figure 8 Home range size (95% KUD) of Common Woodpigeons Columba palumbus. Shown are Woodpigeons from two regions (Lisbon, Portugal and Giessen, Germany) and with different migrations strategies (Giessen (DE): residents and individuals during the non-wintering season; Migration (DE): Individuals using another distinct site during the wintering season than during the breeding season, but migratory movements occurred within Germany; Migration (FR): Woodpigeons migrating to France). Boxplots denote the median value, interquartile range (25–75th percentiles) and range of home range size. The star symbol represents the mean value. Outliers are plotted as individual points CHAPTER 4 | Electronic supplementary material - Electronic supplementary material File 1, including: o Supplementary material A1 Initial tests for amplicon PCR o Supplementary material A2 Amplicon and index PCR setups and cycling conditions o Supplementary material A3 GALAXY workflow o Supplementary Table 1 Collected faecal samples from three species of the order Columbiformes (TD = European Turtle Dove Streptopelia turtur, SD = Stock Dove Columba oenas, WP = Common Woodpigeon C. palumbus) at different sampling sites in Germany and the Netherlands from the years 2013 to 2020 and results of amplicon PCR amplifications of plant and metazoan DNA o Supplementary Table 2 Seed mixes used at baited sites in Germany and Netherlands to attract columbiform species o Supplementary Table 3 Best blast results for each of the 118 detected valid MOTUs using the UniPlant primer pair (Plant) and mICOIintF/dgHCO-2198 (Metazoa) corresponding accession number, the identity with the blast reference sequence, the sequence length and the bitscore. If determination to species level was not clearly determinable, MOTUs were assigned to the lowest shared taxonomic level. Order of the plant MOTUs equates to their order in Table 1 - 128 - o Supplementary Table 4 Differences in the occurrence (presence/absence data) of plant families determined in the diet of three columbiform species (WP = Common Woodpigeon Columba palumbus, TD = European Turtle Dove Streptopelia turtur, SD = Stock Dove C. oenas). Differences among species were tested pairwise with chi² tests. Statistically significant results (p < 0.05) and associated families are marked bold o Supplementary Table 5 Differences in the occurrence (presence/absence data) of plant genera determined in the diet of three columbiform species (WP = Common Woodpigeon Columba palumbus, TD = European Turtle Dove Streptopelia turtur, SD = Stock Dove C. oenas). Genera of the five most frequently represented families are shown. Differences among species were tested pairwise with chi² tests. Statistically significant results (p < 0.05) and associated genera are marked bold o Supplementary Table 6 Stock Dove Columba oenas diet composition: Compilation of our results and results from previous studies. Given are the plant families and animal prey taxa found in the diet of Stock Doves based on different methods. The plant or animal item is marked with “x” if it was found in the respective study o Supplementary Table 7 Common Woodpigeon Columba palumbus diet composition: Compilation of our results and results from previous studies. Given are the plant families and animal prey taxa found in the diet of Woodpigeons based on different methods. The plant or animal item is marked with “x” if it was found in the respective study o Supplementary Table 8 European Turtle Dove Streptopelia turtur diet composition: Compilation of our results and results from previous studies. Given are the plant families and animal prey taxa found in the diet of Turtle Doves based on different methods. The plant or animal item is marked with “x” if it was found in the respective study o Supplementary Figure 1 Sampling locations of faecal samples of three species from the order of Columbiformes (Common Woodpigeon Columba palumbus, European Turtle Dove Streptopelia turtur, Stock Dove C. oenas) in Germany and the Netherlands. Black triangles represent temporarily baited sites and white triangles sites without bait. 1: Helgoland, 2: Wilhelmshaven, 3: Zak van Zuid-Beveland, 4: Lieberoser Heide, 5: Lausitz, 6: Caldern, 7: Zeulenroda, 8: Giessen, 9: Hungen- Villingen, 10: Cleeberg, 11: Eichkopf, 12: Weilbacher Kiesgruben, 13: Groß- Umstadt. The cross marks the location of the clinic for birds (‘Vetmed’, WP). For exact sample numbers per site see Table A1 - 129 - Appendixx o Supplementary Figure 2 Diet composition of Common Woodpigeons Columba palumbus (WP), European Turtle Doves Streptopelia turtur (TD) and Stock Doves C. oenas (SD). Genera of the five most frequently represented plant families (A: Asteraceae, B: Brassicaceae, C: Cucurbitaceae, D: Fabaceae and E: Poaceae) found in faecal samples represented as the frequency of occurrence (FOO%) per species. ‘*’ indicates significant difference (p < 0.05) in the occurrence (presence/absence data) of respective genera between two species (Table A5) o Supplementary Figure 3 Differences in the diet composition at plant genus level in three columbiform species (Common Woodpigeon Columba palumbus (WP); European Turtle Dove Streptopelia turtur (TD); Stock Dove C. oenas (SD)), using Non-metric Multidimensional Scaling (NMDS, function metaMDS in the R package ‘VEGAN’). Depicted are (A) the distribution of the plant genera (the first four or five letters of the genera are given) and (B) the distribution of samples and 95% confidence ellipses CHAPTER 5 | Electronic supplementary material  Supplementary Figure 1 Map of sampling locations for blood samples of columbiform birds  Supplementary Figure 2 Giemsa-stained blood smear of an adult, male European turtle dove (Streptopelia turtur) sampled 2019 on Antikythira Island, Greece  Supplementary Table 1 Lineage names and associated GenBank accession numbers for avian haemosporidian lineages (n = 109) used for phylogenetic tree construction based on a Bayesian analysis - 130 - x CURRICULUM VITAE - Removed from this version - - 131 - Appendixx LIST OF PUBLICATIONS Peer-reviewed paper Quillfeldt P, Schumm YR, Marek C, Mader V, Fischer D, Marx M (2018) Prevalence and genotyping of Trichomonas infections in wild birds in central Germany. PLoS ONE 13: e0200798. DOI: 10.1371/journal.pone.0200798 Schumm YR, Wecker C, Marek C, Wassmuth M, Bentele A, Willems H, Reiner G, Quillfeldt P (2019) Blood parasites in Passeriformes in central Germany: prevalence and lineage diversity of Haemosporida (Haemoproteus, Plasmodium and Leucocytozoon) in six common songbirds. PeerJ 6: e6259. DOI: 10.7717/peerj.6259 Schumm YR, Bakaloudis D, Barboutis C, Cecere JG, Eraud C, Fischer D, Hering J, Hillerich K, Lormée H, Mader V, Masello JF, Metzger B, Rocha G, Spina F, Quillfeldt P (2021) Prevalence and genetic diversity of avian haemosporidian parasites in wild bird species of the order Columbiformes. Parasitology Research 120: 1405-1420. DOI: 10.1007/s00436-021-07053-7 Castaño-Vázquez F, Schumm YR, Bentele A, Quillfeldt P, Merino S (2021) Experimental manipulation of cavity temperature produces differential effects on parasite abundances in blue tit nests at two different latitudes. International Journal for Parasitology: Parasites and Wildlife 14: 287-297. DOI: 10.1016/j.ijppaw.2021.03.010 Fecchio A, Clark NJ, Bell JA, Skeen HR, Lutz HL, De La Torre GM, Vaughan JA, Tkach VV, Schunck F, Ferreira FC, Braga EM, Lugarini C, Wamiti W, Dispoto JH, Galen SC, Kirchgatter K, Sagario MC, Cueto VR, González-Acuña D, Inumaru M, Sato Y, Schumm YR, Quillfeldt P, Pellegrino I, Dharmarajan G, Gupta P, Robin VV, Ciloglu A, Yildirim A, Huang X, Chapa-Vargas L, Álvarez-Mendizábal P, Santiago-Alarcon D, Drovetski SV, Hellgren O, Voelker G, Ricklefs RE, Hackett SJ, Collins MD, Weckstein JD, Wells K (2021) Global drivers of avian haemosporidian infections vary across zoogeographical regions. Global Ecology and Biogeography 30: 2393-2406. DOI: 10.1111/geb.13390 Schumm YR, Metzger B, Neuling E, Austad M, Galea N, Barbara N, Quillfeldt P (2021) Year‑round spatial distribution and migration phenology of a rapidly declining trans‑Saharan migrant - evidence of winter movements and breeding site fidelity in European turtle doves. Behavioral Ecology and Sociobiology 75: 152. DOI: 10.1007/s00265-021-03082-5 Marx M, Schumm YR, Kardynal KJ, Hobson KA; Rocha G, Zehtindjiev P, Bakaloudis D, Metzger B, Cecere JG, Spina F, Cianchetti-Benedetti M, Frahnert S, Voigt CC, Lormée H, Eraud C, Quillfeldt P (2022) Feather stable isotopes (δ2Hf and δ13Cf) identify the Sub- Saharan wintering grounds of turtle doves from Europe. European Journal of Wildlife Research 68: 21. DOI: 10.1007/s10344-022-01567-w - 132 - Conference contributions Masello JF, Rösner S, Schumm Y, Ehmig M, Lindner K, Quillfeldt P (2019) Movement ecology, energy landscapes, and the microhabitat choice of Common Woodpigeons Columba palumbus. 152. Annual Conference, Deutschen Ornithologen Gesellschaft, Marburg [Poster] Rösner S, Lindner K, Ehmig M, Strehmann F, Schumm YR, Quillfeldt P, Farwig N, Masello JF (2019) Competition, stress and parasite prevalence: A bird community approach in an interior forest ecosystem. 152. Annual Conference, Deutschen Ornithologen Gesellschaft, Marburg [Poster] Schumm Y, Metzger B, Barbara N, Neuling E, Lachmann L, Quillfeldt P (2019) Ist die Turteltaube ein Gewohnheitstier? Ergebnisse satellitentelemetrischer Untersuchungen an europäischen Turteltauben. 152. Annual Conference, Deutschen Ornithologen Gesellschaft, Marburg [Talk] Schumm Y (2020) Wildtauben in Hessen: Vergleich ökologischer Aspekte von Turtel-, Ringel- und Hohltaube. Fachsymposium 2020 – Vielfalt der Natur, NABU Landesverband Hessen, Wetzlar [Talk] Strehmann F, Lindner K, Becker M, Schumm YR, Quillfeldt P; Masello JF, Farwig N, Lindner K, Becker M, Schumm YR, Quillfeldt P, Masello JF, Farwig N, Schabo D, Rösner S (2021) Prevalence of blood parasites in a temperate forest bird community. 50th Annual Meeting of the Ecological Society of Germany, Austria and Switzerland. Online Conference [Electronic Poster] Schumm YR, Masello JF, Vreugdenhil-Rowlands J, Fischer D, Hillerich K, Quillfeldt P (2022) Who ate what? Diet composition of Common Woodpigeons, European Turtle Doves and Stock Doves determined by next-generation sequencing of plant and metazoan DNA in faecal samples. 13th European Ornithologists’ Union Congress. Online Conference [Electronic Poster] - 133 - Appendixx ACKNOWLEDGEMENTS - Removed from this version - - 134 - ILLUSTRATION CREDITS Credits for included illustrations from scientific publications are given in the respective figure captions. Laura Prause generously created the Woodpigeon illustration included on the thesis title page. Aleksandra Czylok generously created the Turtle Dove included in Figure 2 and the Woodpigeon with nestlings added to the last page. Photo of the Turtle Dove with Argos transmitter used as the title page for chapter 2 was taken by Mélibée Morel and the photo of the Woodpigeon with GPS-GSM/GPRS transmitter used for chapter 3 by Benjamin Metzger. Thanks to all artists and photographers for allowing me to use their works within my thesis. - 135 - Appendixx - 136 -