Forest top canopy bacterial communities are influenced by elevation and host tree traits.

BACKGROUND: The phyllosphere microbiome is crucial for plant health and ecosystem functioning. While host species play a determining role in shaping the phyllosphere microbiome, host trees of the same species that are subjected to different environmental conditions can still exhibit large degrees of variation in their microbiome diversity and composition. Whether these intra-specific variations in phyllosphere microbiome diversity and composition can be observed over the broader expanse of forest landscapes remains unclear. In this study, we aim to assess the variation in the top canopy phyllosphere bacterial communities between and within host tree species in the temperate European forests, focusing on Fagus sylvatica (European beech) and Picea abies (Norway spruce). RESULTS: We profiled the bacterial diversity, composition, driving factors, and discriminant taxa in the top canopy phyllosphere of 211 trees in two temperate forests, Veluwe National Parks, the Netherlands and Bavarian Forest National Park, Germany. We found the bacterial communities were primarily shaped by host species, and large variation existed within beech and spruce. While we showed that there was a core microbiome in all tree species examined, community composition varied with elevation, tree diameter at breast height, and leaf-specific traits (e.g., chlorophyll and P content). These driving factors of bacterial community composition also correlated with the relative abundance of specific bacterial families. CONCLUSIONS: While our results underscored the importance of host species, we demonstrated a substantial range of variation in phyllosphere bacterial diversity and composition within a host species. Drivers of these variations have implications at both the individual host tree level, where the bacterial communities differed based on tree traits, and at the broader forest landscape level, where drivers like certain highly plastic leaf traits can potentially link forest canopy bacterial community variations to forest ecosystem processes. We eventually showed close associations between forest canopy phyllosphere bacterial communities and host trees exist, and the consistent patterns emerging from these associations are critical for host plant functioning.

Opportunities to reduce pollination deficits and address production shortfalls in an important insect-pollinated crop.

Pollinators face multiple pressures and there is evidence of populations in decline. As demand for insect-pollinated crops increases, crop production is threatened by shortfalls in pollination services. Understanding the extent of current yield deficits due to pollination and identifying opportunities to protect or improve crop yield and quality through pollination management is therefore of international importance. To explore the extent of "pollination deficits," where maximum yield is not being achieved due to insufficient pollination, we used an extensive dataset on a globally important crop, apples. We quantified how these deficits vary between orchards and countries and we compared "pollinator dependence" across different apple varieties. We found evidence of pollination deficits and, in some cases, risks of overpollination were even apparent for which fruit quality could be reduced by too much pollination. In almost all regions studied we found some orchards performing significantly better than others in terms of avoiding a pollination deficit and crop yield shortfalls due to suboptimal pollination. This represents an opportunity to improve production through better pollinator and crop management. Our findings also demonstrated that pollinator dependence varies considerably between apple varieties in terms of fruit number and fruit quality. We propose that assessments of pollination service and deficits in crops can be used to quantify supply and demand for pollinators and help to target local management to address deficits although crop variety has a strong influence on the role of pollinators.

The aerobiome uncovered: Multi-marker metabarcoding reveals potential drivers of turn-over in the full microbial community in the air.

Air is a major conduit for the dispersal of organisms at the local and the global scale. Most research has focused on the dispersal of plants, vertebrates and human disease agents. However, the air represents a key dispersal medium also for bacteria, fungi and protists. Many of those represent potential pathogens of animals and plants and have until now gone largely unrecorded. Here we studied the turnover in composition of the entire aerobiome, the collective diversity of airborne microorganisms. For that we performed daily analyses of all prokaryotes and eukaryotes (including plants) using multi-marker high-throughput sequencing for a total of three weeks. We linked the resulting communities to local weather conditions, to assess determinants of aerobiome composition and distribution. We observed hundreds of microbial taxa, mostly belonging to spore-forming organisms including fungi, but also protists. Additionally, we detected many potential human- and plant-pathogens. Community composition fluctuated on a daily basis and was linked to concurrent weather conditions, particularly air pressure and temperature. Using network analyses, we identified taxonomically diverse groups of organisms with correlated temporal dynamics. In part, this was due to co-variation with environmental conditions, while we could also detect specific host-parasite interactions. This study provides the first full inventory of the aerobiome and identifies putative drivers of its dynamics in terms of taxon composition. This knowledge can help develop early warning systems against pathogens and improve our understanding of microbial dispersal.

Wild insect diversity increases inter-annual stability in global crop pollinator communities.

While an increasing number of studies indicate that the range, diversity and abundance of many wild pollinators has declined, the global area of pollinator-dependent crops has significantly increased over the last few decades. Crop pollination studies to date have mainly focused on either identifying different guilds pollinating various crops, or on factors driving spatial changes and turnover observed in these communities. The mechanisms driving temporal stability for ecosystem functioning and services, however, remain poorly understood. Our study quantifies temporal variability observed in crop pollinators in 21 different crops across multiple years at a global scale. Using data from 43 studies from six continents, we show that (i) higher pollinator diversity confers greater inter-annual stability in pollinator communities, (ii) temporal variation observed in pollinator abundance is primarily driven by the three-most dominant species, and (iii) crops in tropical regions demonstrate higher inter-annual variability in pollinator species richness than crops in temperate regions. We highlight the importance of recognizing wild pollinator diversity in agricultural landscapes to stabilize pollinator persistence across years to protect both biodiversity and crop pollination services. Short-term agricultural management practices aimed at dominant species for stabilizing pollination services need to be considered alongside longer term conservation goals focussed on maintaining and facilitating biodiversity to confer ecological stability.

Protist taxonomic and functional diversity in soil, freshwater and marine ecosystems.

Protists dominate eukaryotic diversity and play key functional roles in all ecosystems, particularly by catalyzing carbon and nutrient cycling. To date, however, a comparative analysis of their taxonomic and functional diversity that compares the major ecosystems on Earth (soil, freshwater and marine systems) is missing. Here, we present a comparison of protist diversity based on standardized high throughput 18S rRNA gene sequencing of soil, freshwater and marine environmental DNA. Soil and freshwater protist communities were more similar to each other than to marine protist communities, with virtually no overlap of Operational Taxonomic Units (OTUs) between terrestrial and marine habitats. Soil protists showed higher γ diversity than aquatic samples. Differences in taxonomic composition of the communities led to changes in a functional diversity among ecosystems, as expressed in relative abundance of consumers, phototrophs and parasites. Phototrophs (eukaryotic algae) dominated freshwater systems (49% of the sequences) and consumers soil and marine ecosystems (59% and 48%, respectively). The individual functional groups were composed of ecosystem- specific taxonomic groups. Parasites were equally common in all ecosystems, yet, terrestrial systems hosted more OTUs assigned to parasites of macro-organisms while aquatic systems contained mostly microbial parasitoids. Together, we show biogeographic patterns of protist diversity across major ecosystems on Earth, preparing the way for more focused studies that will help understanding the multiple roles of protists in the biosphere.

DNA of centuries-old timber can reveal its origin.

Oak wood was highly appreciated and widely used for construction in past centuries. As population sizes expanded in some regions of Europe, local forests were depleted of high-quality timber. Therefore, regions of soaring economies were importing timber initially from the European market and eventually from other continents. Origin of archaeological or historical timber is usually identified by means of dendroprovenancing, i.e. statistical matching of tree-ring-width (TRW) series of timber of unknown origin with TRW reference datasets. However, this method has pitfalls and limitations and therefore alternative techniques are needed. Here, we used three different DNA analysis methods to investigate the potential of using ancient (a)DNA, extracted from oak timber derived from historical buildings and shipwrecks from a variety of countries. All the material had also been analysed dendrochronologically, so its dating and provenance is demonstrated. We included heartwood samples in this analysis, for which DNA extraction is especially challenging as it contains chemicals that inhibit DNA amplification. We succeeded in amplifying DNA for at least one marker from 56% of samples (including heartwood samples), yielding crucial information that allowed us to identify the potential source area of centuries old timber buildings in Latvia and Denmark and of 750-year-old shipwreck material from Germany. Our results prove the strong potential of DNA analyses for identifying timber origin to the regional scale, but by combining these with the dendrochronological results, we can control the exactitude of the aDNA approach and demonstrate a more nuanced examination of the timber sources for these historic structures.

A global synthesis reveals biodiversity-mediated benefits for crop production.

Human land use threatens global biodiversity and compromises multiple ecosystem functions critical to food production. Whether crop yield-related ecosystem services can be maintained by a few dominant species or rely on high richness remains unclear. Using a global database from 89 studies (with 1475 locations), we partition the relative importance of species richness, abundance, and dominance for pollination; biological pest control; and final yields in the context of ongoing land-use change. Pollinator and enemy richness directly supported ecosystem services in addition to and independent of abundance and dominance. Up to 50% of the negative effects of landscape simplification on ecosystem services was due to richness losses of service-providing organisms, with negative consequences for crop yields. Maintaining the biodiversity of ecosystem service providers is therefore vital to sustain the flow of key agroecosystem benefits to society.

Combined effects of agrochemicals and ecosystem services on crop yield across Europe.

Simultaneously enhancing ecosystem services provided by biodiversity below and above ground is recommended to reduce dependence on chemical pesticides and mineral fertilisers in agriculture. However, consequences for crop yield have been poorly evaluated. Above ground, increased landscape complexity is assumed to enhance biological pest control, whereas below ground, soil organic carbon is a proxy for several yield-supporting services. In a field experiment replicated in 114 fields across Europe, we found that fertilisation had the strongest positive effect on yield, but hindered simultaneous harnessing of below- and above-ground ecosystem services. We furthermore show that enhancing natural enemies and pest control through increasing landscape complexity can prove disappointing in fields with low soil services or in intensively cropped regions. Thus, understanding ecological interdependences between land use, ecosystem services and yield is necessary to promote more environmentally friendly farming by identifying situations where ecosystem services are maximised and agrochemical inputs can be reduced.

Testing projected wild bee distributions in agricultural habitats: predictive power depends on species traits and habitat type.

Species distribution models (SDM) are increasingly used to understand the factors that regulate variation in biodiversity patterns and to help plan conservation strategies. However, these models are rarely validated with independently collected data and it is unclear whether SDM performance is maintained across distinct habitats and for species with different functional traits. Highly mobile species, such as bees, can be particularly challenging to model. Here, we use independent sets of occurrence data collected systematically in several agricultural habitats to test how the predictive performance of SDMs for wild bee species depends on species traits, habitat type, and sampling technique. We used a species distribution modeling approach parametrized for the Netherlands, with presence records from 1990 to 2010 for 193 Dutch wild bees. For each species, we built a Maxent model based on 13 climate and landscape variables. We tested the predictive performance of the SDMs with independent datasets collected from orchards and arable fields across the Netherlands from 2010 to 2013, using transect surveys or pan traps. Model predictive performance depended on species traits and habitat type. Occurrence of bee species specialized in habitat and diet was better predicted than generalist bees. Predictions of habitat suitability were also more precise for habitats that are temporally more stable (orchards) than for habitats that suffer regular alterations (arable), particularly for small, solitary bees. As a conservation tool, SDMs are best suited to modeling rarer, specialist species than more generalist and will work best in long-term stable habitats. The variability of complex, short-term habitats is difficult to capture in such models and historical land use generally has low thematic resolution. To improve SDMs' usefulness, models require explanatory variables and collection data that include detailed landscape characteristics, for example, variability of crops and flower availability. Additionally, testing SDMs with field surveys should involve multiple collection techniques.

Fern spore longevity in saline water: can sea bottom sediments maintain a viable spore bank?

Freshwater and marine sediments often harbor reservoirs of plant diaspores, from which germination and establishment may occur whenever the sediment falls dry. Therewith, they form valuable records of historical inter- and intraspecific diversity, and are increasingly exploited to facilitate diversity establishment in new or restored nature areas. Yet, while ferns may constitute a considerable part of a vegetation's diversity and sediments are known to contain fern spores, little is known about their longevity, which may suffer from inundation and--in sea bottoms--salt stress. We tested the potential of ferns to establish from a sea or lake bottom, using experimental studies on spore survival and gametophyte formation, as well as a spore bank analysis on sediments from a former Dutch inland sea. Our experimental results revealed clear differences among species. For Asplenium scolopendrium and Gymnocarpium dryopteris, spore germination was not affected by inundated storage alone, but decreased with rising salt concentrations. In contrast, for Asplenium trichomanes subsp. quadrivalens germination decreased following inundation, but not in response to salt. Germination rates decreased with time of storage in saline water. Smaller and less viable gametophytes were produced when saline storage lasted for a year. Effects on germination and gametophyte development clearly differed among genotypes of A. scolopendrium. Spore bank analyses detected no viable spores in marine sediment layers. Only two very small gametophytes (identified as Thelypteris palustris via DNA barcoding) emerged from freshwater sediments. Both died before maturation. We conclude that marine, and likely even freshwater sediments, will generally be of little value for long-term storage of fern diversity. The development of any fern vegetation on a former sea floor will depend heavily on the deposition of spores onto the drained land by natural or artificial means of dispersal.

Variation in ploidy level and phenology can result in large and unexpected differences in demography and climatic sensitivity between closely related ferns.

PREMISE OF THE STUDY: Current environmental changes may affect the dynamics and viability of plant populations. This environmental sensitivity may differ between species of different ploidy level because polyploidization can influence life history traits. We compared the demography and climatic sensitivity of two closely related ferns: the tetraploid Polystichum aculeatum and one of its diploid parents, Polystichum setiferum. METHODS: Matrix models were used to assess the effects of life history variation on population dynamics under varying winter conditions. We analyzed the contributions of all key aspects of the fern life cycle to population growth. Our study is the first to also include the gametophyte generation. KEY RESULTS: Projected population growth rate (λ) was much higher for the tetraploid P. aculeatum (1.516) than for P. setiferum (1.071) under normal winter conditions. During a year with harsh winter conditions, population growth of P. aculeatum was strongly reduced. This finding contradicts our expectation that the winter-hardy fronds of this species would allow high survival of harsh winters. Differences in λ between species and between years with different winter conditions were mostly caused by variation in gametophyte-related recruitment rates, a finding that shows the importance of including gametophytes in fern demographic studies. CONCLUSIONS: Our results indicate that populations of closely related ferns can show large differences in population performance, mainly related to recruitment rates and frond phenology, and that these differences may depend greatly on climatic conditions. Our findings provide a first indication that (allo)polyploidization in ferns can have a significant effect on population dynamics.

Inter-and intraspecific variation in fern mating systems after long-distance colonization: the importance of selfing.

BACKGROUND: Previous studies on the reproductive biology of ferns showed that mating strategies vary among species, and that polyploid species often show higher capacity for self-fertilization than diploid species. However, the amount of intraspecific variation in mating strategy and selfing capacity has only been assessed for a few species. Yet, such variation may have important consequences during colonization, as the establishment of any selfing genotypes may be favoured after long-distance dispersal (an idea known as Baker's law). RESULTS: We examined intra-and interspecific variation in potential for self-fertilization among four rare fern species, of which two were diploids and two were tetraploids: Asplenium scolopendrium (2n), Asplenium trichomanes subsp. quadrivalens (4n), Polystichum setiferum (2n) and Polystichum aculeatum (4n). Sporophyte production was tested at different levels of inbreeding, by culturing gametophytes in isolation, as well as in paired cultures with a genetically different gametophyte. We tested gametophytes derived from various genetically different sporophytes from populations in a recently planted forest colonized through long-distance dispersal (Kuinderbos, the Netherlands), as well as from older, less disjunct populations.Sporophyte production in isolation was high for Kuinderbos genotypes of all four species. Selfing capacity did not differ significantly between diploids and polyploids, nor between species in general. Rather selfing capacity differed between genotypes within species. Intraspecific variation in mating system was found in all four species. In two species one genotype from the Kuinderbos showed enhanced sporophyte production in paired cultures. For the other species, including a renowned out crosser, selfing capacity was consistently high. CONCLUSIONS: Our results for four different species suggest that intraspecific variation in mating system may be common, at least among temperate calcicole ferns, and that genotypes with high selfing capacity may be present among polyploid as well as diploid ferns. The surprisingly high selfing capacity of all genotypes obtained from the Kuinderbos populations might be due to the isolated position of these populations. These populations may have established through single-spore colonization, which is only possible for genotypes capable of self-fertilization. Our results therewith support the idea that selection for selfing genotypes may occur during long-distance colonization, even in normally outcrossing, diploid ferns.

Isolation of polymorphic microsatellite markers and tests of cross-amplification in four widespread European calcicole ferns.

PREMISE OF THE STUDY: Studies on the biogeography and population genetics of the widespread European rock ferns Asplenium scolopendrium, A. trichomanessubsp. quadrivalens, Polystichum setiferum, and P. aculeatumwould potentially yield interesting new insights into the colonization capacities of ferns. Markers with sufficient resolution for detailed genetic studies are, however, not yet available. METHODS AND RESULTS: Using genome screening with intersimple sequence repeat (ISSR) primers, a total of 16 different microsatellite markers were isolated and characterized for the four species. Some of these markers could be exchanged within each congeneric pair. CONCLUSIONS: The developed primer sets will be very useful for analyses of the biogeography and population genetics of some widespread calcicole ferns. The observed cross-amplification rates suggest a high potential for application on additional species from the same genera.

Use of rbcL and trnL-F as a two-locus DNA barcode for identification of NW-European ferns: an ecological perspective.

Although consensus has now been reached on a general two-locus DNA barcode for land plants, the selected combination of markers (rbcL + matK) is not applicable for ferns at the moment. Yet especially for ferns, DNA barcoding is potentially of great value since fern gametophytes--while playing an essential role in fern colonization and reproduction--generally lack the morphological complexity for morphology-based identification and have therefore been underappreciated in ecological studies. We evaluated the potential of a combination of rbcL with a noncoding plastid marker, trnL-F, to obtain DNA-identifications for fern species. A regional approach was adopted, by creating a reference database of trusted rbcL and trnL-F sequences for the wild-occurring homosporous ferns of NW-Europe. A combination of parsimony analyses and distance-based analyses was performed to evaluate the discriminatory power of the two-region barcode. DNA was successfully extracted from 86 tiny fern gametophytes and was used as a test case for the performance of DNA-based identification. Primer universality proved high for both markers. Based on the combined rbcL + trnL-F dataset, all genera as well as all species with non-equal chloroplast genomes formed their own well supported monophyletic clade, indicating a high discriminatory power. Interspecific distances were larger than intraspecific distances for all tested taxa. Identification tests on gametophytes showed a comparable result. All test samples could be identified to genus level, species identification was well possible unless they belonged to a pair of Dryopteris species with completely identical chloroplast genomes. Our results suggest a high potential of the combined use of rbcL and trnL-F as a two-locus cpDNA barcode for identification of fern species. A regional approach may be preferred for ecological tests. We here offer such a ready-to-use barcoding approach for ferns, which opens the way for answering a whole range of questions previously unaddressed in fern gametophyte ecology.

Mixed mating system in the fern Asplenium scolopendrium: implications for colonization potential.

BACKGROUND AND AIMS: Human-mediated environmental change is increasing selection pressure for the capacity in plants to colonize new areas. Habitat fragmentation combined with climate change, in general, forces species to colonize areas over longer distances. Mating systems and genetic load are important determinants of the establishment and long-term survival of new populations. Here, the mating system of Asplenium scolopendrium, a diploid homosporous fern species, is examined in relation to colonization processes. METHODS: A common environment experiment was conducted with 13 pairs of sporophytes, each from a different site. Together they constitute at least nine distinct genotypes, representing an estimated approx. 95 % of the non-private intraspecific genetic variation in Europe. Sporophyte production was recorded for gametophytes derived from each parent sporophyte. Gametophytes were grown in vitro in three different ways: (I) in isolation, (II) with a gametophyte from a different sporophyte within the same site or (III) with a partner from a different site. KEY RESULTS: Sporophyte production was highest in among-site crosses (III), intermediate in within-site crosses (II) and was lowest in isolated gametophytes (I), strongly indicating inbreeding depression. However, intragametophytic selfing was observed in most of the genotypes tested (eight out of nine). CONCLUSIONS: The results imply a mixed mating system in A. scolopendrium, with outcrossing when possible and occasional selfing when needed. Occasional intragametophytic selfing facilitates the successful colonization of new sites from a single spore. The resulting sporophyte, which will be completely homozygous, will shed large amounts of spores over time. Each year this creates a bed of gametophytes in the vicinity of the parent. Any unrelated spore which arrives is then selectively favoured to reproduce and contribute its genes to the new population. Thus, while selfing facilitates initial colonization success, inbreeding depression promotes genetically diverse populations through outcrossing. The results provide further evidence against the overly simple dichotomous distinction of fern species as either selfing or outcrossing.