Consistent predictors of microbial community composition across spatial scales in grasslands reveal low context-dependency.

Environmental circumstances shaping soil microbial communities have been studied extensively. However, due to disparate study designs, it has been difficult to resolve whether a globally consistent set of predictors exists, or context-dependency prevails. Here, we used a network of 18 grassland sites (11 of those containing regional plant productivity gradients) to examine (i) if similar abiotic or biotic factors predict both large-scale (across sites) and regional-scale (within sites) patterns in bacterial and fungal community composition, and (ii) if microbial community composition differs consistently at two levels of regional plant productivity (low vs. high). Our results revealed that bacteria were associated with particular soil properties (such as base saturation) and both bacteria and fungi were associated with plant community composition across sites and within the majority of sites. Moreover, a discernible microbial community signal emerged, clearly distinguishing high and low-productivity soils across different grasslands independent of their location in the world. Hence, regional productivity differences may be typified by characteristic soil microbial communities across the grassland biome. These results could encourage future research aiming to predict the general effects of global changes on soil microbial community composition in grasslands and to discriminate fertile from infertile systems using generally applicable microbial indicators.

Analysis of 3-nitropropionic acid in Fabaceae plants by HPLC-MS/MS.

INTRODUCTION: 3-Nitropropionic acid (3-NPA) is a toxic compound that can accumulate in esterified form in the Fabaceae family. In the Lotae tribe, many species have been identified as 3-NPA producers (e.g., Securigera varia), while some of the genetically close Lotae plants were formerly reported as 3-NPA-free (e.g., Lotus corniculatus and Anthyllis vulneraria). These plants are used as forage and have a tradition in ethnomedicine, also, the extracts of A. vulneraria are used in cosmetics. OBJECTIVES: Our aim was to investigate the 3-NPA content of these selected Fabaceae species and to develop a validated quantitative method to evaluate 3-NPA concentrations in extracts of different herbal parts and cosmetic products. MATERIALS AND METHODS: A UHPLC-ESI-Orbitrap-MS/MS method was applied for detection and identification of 3-NPA derivatives in the form of glucose esters. For the quantitative analysis, an optimized sample processing method was developed. The free 3-NPA content was determined using HPLC-ESI-MS/MS. RESULTS: 3-NPA esters could be detected in all three species, but their quantity showed a high variation. S. varia contained 0.5-1.0 g/100 g of 3-NPA, while in L. corniculatus samples only trace quantities were detectable, below the LOQ (25 ng/ml). Most of the A. vulneraria samples showed similarly low concentrations, but one sample had 3-NPA levels comparable to S. varia. 3-NPA could not be detected in the tested cosmetics containing A. vulneraria extracts. CONCLUSIONS: Using highly sensitive analytical methods, new 3-NPA-containing species were identified. The developed validated quantitative method is suitable for the determination of 3-NPA concentrations in herbal samples.

Fine-scale spatial genetic structure across the species range reflects recent colonization of high elevation habitats in silver fir (Abies alba Mill.).

Variation in genetic diversity across species ranges has long been recognized as highly informative for assessing populations' resilience and adaptive potential. The spatial distribution of genetic diversity within populations, referred to as fine-scale spatial genetic structure (FSGS), also carries information about recent demographic changes, yet it has rarely been connected to range scale processes. We studied eight silver fir (Abies alba Mill.) population pairs (sites), growing at high and low elevations, representative of the main genetic lineages of the species. A total of 1,368 adult trees and 540 seedlings were genotyped using 137 and 116 single nucleotide polymorphisms (SNPs), respectively. Sites revealed a clear east-west isolation-by-distance pattern consistent with the post-glacial colonization history of the species. Genetic differentiation among sites (FCT = 0.148) was an order of magnitude greater than between elevations within sites (FSC = 0.031), nevertheless high elevation populations consistently exhibited a stronger FSGS. Structural equation modelling revealed that elevation and, to a lesser extent, post-glacial colonization history, but not climatic and habitat variables, were the best predictors of FSGS across populations. These results suggest that high elevation habitats have been colonized more recently across the species range. Additionally, paternity analysis revealed a high reproductive skew among adults and a stronger FSGS in seedlings than in adults, suggesting that FSGS may conserve the signature of demographic changes for several generations. Our results emphasize that spatial patterns of genetic diversity within populations provide information about demographic history complementary to non-spatial statistics, and could be used for genetic diversity monitoring, especially in forest trees.

SNP marker development in Pinus sylvestris L. in stress-responsive genes characterized from Pinus cembra L. transcriptomes.

Massively parallel sequencing of cDNA is an efficient route for generating sequence collections that represent expressed genes under different environmental control. The analysis of their sequence helps in developing molecular markers, such as SNPs, which represent a useful tool in detecting adaptive signals in populations. In this study novel PCR markers, based on stress responsive genes, were designed from the transcriptome of the haploxylon Swiss stone pine (Pinus cembra L.) and tested for SNPs in the diploxylon Scots pine (Pinus sylvestris L.). 84 primers were tested on P. sylvestris DNA samples originating from three different types of habitat. After sequencing and BLAST search of the amplified products, parts of 19 different candidate genes were analysed by considering the polymorphic sites, insertions/deletions as well as synonymous and non-synonymous SNPs. In a total of 3735 sites no indels, eight synonymous and 11 non-synonymous SNPs were found. By providing de novo molecular markers developed in P. cembra and tested for transferability in Scots pine, our results give support for the use of de novo markers targeting conserved regions across different pines. The SNPs detected may have important applications in further studies of adaptive genetic variation, providing tools to study relevant genes important in the long-term adaptation of pine species.

Living on the edge: morphological, karyological and genetic diversity studies of the Hungarian Plantago maxima populations and established ex situ collection.

BACKGROUND: The analysis of genetic diversity of protected plant species can greatly support conservation efforts. Plantago maxima Juss. ex Jacq. is a perennial species distributed along the Eurasian steppe. The westernmost range edge of the species' distribution is located in the Pannonian basin, in Hungary where it is represented by a few, fragmented and highly endangered populations. We studied population diversity of all Hungarian range edge, natural populations, and one established ex situ population. One population from the centre of distribution (Kazakhstan) was implemented in the cpDNA haplotype study to compare the peripheral vs. central populations. We performed morphometric trait-based analysis, chromosome studies (morphometric analyses and FISH) and genetic diversity evaluations using inter simple sequence repeats (ISSR) and cpDNA trnL-trnF to evaluate differences between the in situ and ex situ populations as well as central vs. peripheral populations. RESULTS: Our results showed no obvious morphological differences among the in situ and ex situ populations in the period between 2018 and 2020. One ex situ subpopulation develops flowers three years in a row from 2019, which is a favourable indicator of the introduction success. Hungarian populations are exclusively diploids (2n = 2x = 12). The karyogram consists of 5 metacentric and 1 acrocentric chromosome pair. Plantago maxima has one 35S and two 5S rDNA loci, located on the acrocentric chromosome pair. Eight variable ISSR primers yielded 100 fragments, of which 74.6% were polymorphic (mean He = 0.220). A high level of genetic variation within population was observed (92%) while the genetic differentiation among the populations was only 8%. STRUCTURE analysis revealed that the largest Kunpeszér population separated from the rest of the Hungarian populations, indicating a high rate of admixture among the other ones. Based on the trnL-trnF sequence analysis the Hungarian populations represent a single haplotype, which can indicate a reduced diversity due to isolation and recent population decline. By contrast, Kazakh population represents a distinct haplotype compared to the Hungarian samples. CONCLUSIONS: The present study draws the attention to the high conservation value of the Plantago maxima populations from the westernmost range edge of the species' distribution.

PADAPT 1.0 - the Pannonian Dataset of Plant Traits.

The existing plant trait databases' applicability is limited for studies dealing with the flora and vegetation of the eastern and central part of Europe and for large-scale comparisons across regions, mostly because their geographical data coverage is limited and they incorporate records from several different sources, often from regions with markedly different climatic conditions. These problems motivated the compilation of a regional dataset for the flora of the Pannonian region (Eastern Central Europe). PADAPT, the Pannonian Dataset of Plant Traits relies on regional data sources and collates data on 54 traits and attributes of the plant species of the Pannonian region. The current version covers approximately 90% of the species of the region and consists of 126,337 records on 2745 taxa. By including species of the eastern part of Europe not covered by other databases, PADAPT can facilitate studying the flora and vegetation of the eastern part of the continent. Although data coverage is far from complete, PADAPT meets the longstanding need for a regional database of the Pannonian flora.

Imprints of selection in peripheral and ecologically marginal central-eastern European Scots pine populations.

Knowledge of the molecular mechanisms underlying the stress response in plants is essential to understand evolutionary processes that result in long-term persistence of populations. Populations inhabiting marginal ecological conditions at the distribution range periphery may have preserved imprints of natural selection that have shaped functional genetic variation of the species. Our aim was to evaluate the extent of selection processes in the extremely fragmented, peripheral and isolated populations of Scots pine in central-eastern Europe. Autochthonous populations of the Carpathian Mts. and the Pannonian Basin were sampled and drought stress-related candidate genes were re-sequenced. Neutrality tests and outlier detection approaches were applied to infer the effect and direction of selection. Populations retained high genetic diversity by preserving a high number of alleles and haplotypes, many of them being population specific. Neutrality tests and outlier detection highlighted nucleotide positions that are under divergent selection and may be involved in local adaptation. The detected genetic pattern confirms that natural selection has played an important role in shaping modern-day genetic variation in marginal Scots pine populations, allowing for the long-term persistence of populations. Selection detected at functional regions possibly acts to maintain diversity and counteract the effect of genetic erosion.

Current range characteristics of Swiss stone pine (Pinus cembra L.) along the Carpathians revealed by chloroplast SSR markers.

We investigated the diversity pattern of nine Swiss stone pine (Pinus cembra L.) populations along the Carpathian range including the High Tatras, by using six chloroplast DNA microsatellites (cpSSR). Our aim was to detect genetically distinct regions by clustering of populations, and to tackle possible historical colonization routes. Our analysis referred to an investigated geographical range with the two most distant populations situated at about 500 air km. We found that the most diverse populations are situated at the two edges of the investigated part, in the Retezat Mts. (South Carpathians) and the High Tatras, and diversity decreases towards the populations of the Eastern Carpathians. Hierarchical clustering and NMDS revealed that the populations of the South Carpathians with the Tatras form a distinct cluster, significantly separated from those of the Eastern Carpathians. Moreover, based on the most variable chloroplast microsatellites, the four populations of the two range edges are not significantly different. Our results, supported also by palynological and late glacial macrofossil evidences, indicate refugial territories within the Retezat Mts. that conserved rich haplotype composition. From this refugial territory Pinus cembra might have colonized the Eastern Carpathians, and this was accompanied by a gradual decrease in population diversity. Populations of the High Tatras might have had the same role in the colonizing events of the Carpathians, as positive correlation was detected among populations lying from each other at a distance of 280 km, the maximum distance between neighbouring populations.

Speciation by triparental hybridization in genus Sorbus (Rosaceae).

Hybridization associated with polyploidization and apomixis is a frequent mechanism of speciation. Sorbus is a genus with ongoing hybridization resulting in a polyploid complex with different parental lineage. Triparens is the smallest hybridogenous subgenus of Sorbus so far known to comprise only two taxa, S. intermedia and S. × liljeforsii that combine the genomes of three taxa (S. aria agg., S. aucuparia and S. torminalis). To elucidate the origins of S. dacica, S. paxiana and S. tauricola, three new trigenomic candidates formerly believed to be of biparental origin with either S. aria agg. × S. aucuparia or S. aria agg. × S. torminalis lineage we combined data from HPLC and chloroplast DNA analysing additional 33 related taxa as well. We concluded that the 'torminalis-type' flavonoid profile and the 'aucuparia-type' plastid indicate the participation of both S. torminalis and S. aucuparia resulting in the formation of S. dacica, S. paxiana and S. tauricola. Sorbus aria agg. as the third ancestor and as a necessary link to meet genes of S. torminalis and S. aucuparia in one genome is obvious from morphological features (densely tomentose undersides of leaves). The tetraploid cytotypes and obligate pseudogamy of S. dacica and S. paxiana were determined by flow cytometry and are published here for the first time. The most probable evolutionary scenario for Triparens species is: 1. a diploid sexual S. aucuparia as pollen acceptor hybridized with a tetraploid apomictic taxon from the S. aria agg. producing a triploid apomictic taxon with 'aucuparia-type' plastid inherited maternally; 2. during a second crossing event this subgenus Soraria hybrid as maternal progenitor hybridized with the sexual diploid S. torminalis (providing gene(s) of apigenin O-glucuronide synthesis) forming a tetraploid Triparens hybrid with 'aucuparia-type' plastid and 'torminalis-type' flavonoids.

Long-term survival and successful conservation? Low genetic diversity but no evidence for reduced reproductive success at the north-westernmost range edge of Poa badensis (Poaceae) in Central Europe.

Many steppe species reach their (north)westernmost distribution limit in western Central Europe. This also applies to Poa badensis, a rare steppe plant of calcareous rock/sand vegetation. To explore potential differences in reproductive success and genetic composition of peripheral populations, we analysed the absolute (north)westernmost occurrences in Western Germany and populations at the western margin (Eastern Austria) and the centre (Central Hungary) of the Pannonicum, representing a part of the continuous range. Specifically, we discuss the genetic and reproductive constitution of the (north)westernmost exclave and draw conclusions on the species' biogeographical and conservation history in this region. Therefore, we used two independent molecular marker systems (AFLPs, cpDNA sequences) and a set of performance parameters. Overall, lowest regional genetic diversity was found in Western Germany, which is mainly a result of the specific history of two populations. However, this low genetic diversity was not accompanied by reduced reproductive success. The Eastern Austrian populations showed reduced genetic diversity and predominantly reduced performance, interpreted as a consequence of small population sizes. Central Hungarian populations showed the overall highest genetic diversity and comparatively high performance values. We observed high admixture and haplotype sharing between Austrian and Hungarian populations, indicating gene flow among these regions. In contrast, we interpreted the increased population differentiation within, and the clear distinctiveness of the German exclave as a long-term isolation of these (north)westernmost occurrences. Our results, overall, prove the good constitution of these populations and, together with their particular biogeographical history, highlight their conservation value.

Identification, Structural and Functional Characterization of Dormancy Regulator Genes in Apricot (Prunus armeniaca L.).

In the present study, we identified and characterized the apricot (Prunus armeniaca L.) homologs of three dormancy-related genes, namely the ParCBF1 (C-repeat binding factor), ParDAM5 (dormancy-associated MADS-BOX) and ParDAM6 genes. All highly conserved structural motifs and the 3D model of the DNA-binding domain indicate an unimpaired DNA-binding ability of ParCBF1. A phylogenetic analysis showed that ParCBF1 was most likely homologous to Prunus mume and Prunus dulcis CBF1. ParDAM5 also contained all characteristic domains of the type II (MIKCC) subfamily of MADS-box transcription factors. The homology modeling of protein domains and a phylogenetic analysis of ParDAM5 suggest its functional integrity. The amino acid positions or small motifs that are diagnostic characteristics of DAM5 and DAM6 were determined. For ParDAM6, only a small part of the cDNA was sequenced, which was sufficient for the quantification of gene expression. The expression of ParCBF1 showed close association with decreasing ambient temperatures in autumn and winter. The expression levels of ParDAM5 and ParDAM6 changed according to CBF1 expression rates and the fulfillment of cultivar chilling requirements (CR). The concomitant decrease of gene expression with endodormancy release is consistent with a role of ParDAM5 and ParDAM6 genes in dormancy induction and maintenance. Cultivars with higher CR and delayed flowering time showed higher expression levels of ParDAM5 and ParDAM6 toward the end of endodormancy. Differences in the timing of anther developmental stages between early- and late-flowering cultivars and two dormant seasons confirmed the genetically and environmentally controlled mechanisms of dormancy release in apricot generative buds. These results support that the newly identified apricot gene homologs have a crucial role in dormancy-associated physiological mechanisms.

A Reference Genome Sequence for the European Silver Fir (Abies alba Mill.): A Community-Generated Genomic Resource.

Silver fir (Abies alba Mill.) is a keystone conifer of European montane forest ecosystems that has experienced large fluctuations in population size during during the Quaternary and, more recently, due to land-use change. To forecast the species' future distribution and survival, it is important to investigate the genetic basis of adaptation to environmental change, notably to extreme events. For this purpose, we here provide a first draft genome assembly and annotation of the silver fir genome, established through a community-based initiative. DNA obtained from haploid megagametophyte and diploid needle tissue was used to construct and sequence Illumina paired-end and mate-pair libraries, respectively, to high depth. The assembled A. alba genome sequence accounted for over 37 million scaffolds corresponding to 18.16 Gb, with a scaffold N50 of 14,051 bp. Despite the fragmented nature of the assembly, a total of 50,757 full-length genes were functionally annotated in the nuclear genome. The chloroplast genome was also assembled into a single scaffold (120,908 bp) that shows a high collinearity with both the A. koreana and A. sibirica complete chloroplast genomes. This first genome assembly of silver fir is an important genomic resource that is now publicly available in support of a new generation of research. By genome-enabling this important conifer, this resource will open the gate for new research and more precise genetic monitoring of European silver fir forests.

Intercontinental migration pattern and genetic differentiation of arctic-alpine Rhodiola rosea L.: A chloroplast DNA survey.

Our study describes genetic lineages and historical biogeography of Rhodiola rosea a widely distributed arctic-alpine perennial species of the Northern Hemisphere based on sequence analysis of six chloroplast regions. Specimens of 44 localities from the Northern Hemisphere have been sequenced and compared with those available in the GenBank. Our results support the migration of the species into Europe via the Central Asian highland corridor, reaching the European Alpine System (EAS) and also the western European edge, the British Isles. The EAS proved to be an important center of genetic diversity, especially the region of the Eastern Alps and the Dolomites where signs of glacial refugia was observed. Apart from those of the EAS, a common lineage was detected along the Atlantic coast from the British Isles toward Scandinavia as well as Iceland and the eastern parts of North America. Accordingly, the British Isles represent a main link between the northern Atlantic and southern EAS lineages.

No obvious genetic erosion, but evident relict status at the westernmost range edge of the Pontic-Pannonian steppe plant Linum flavum L. (Linaceae) in Central Europe.

We investigate patterns of genetic variation along an east-west transect of Central European populations of Linum flavum and interpret the Quaternary history of its peripheral populations, especially those at the westernmost isolated range edge, discussing their migrations and possible relict status. We defined our peripheral transect across three study regions from Central Hungary, eastern Austria to southwestern Germany. Using AFLP fingerprinting and cpDNA sequence variation (rpL16 intron, atpI-H), we analyzed 267 and 95 individuals, respectively, representing each study region by four populations. Hierarchical AMOVA (AFLPs) indicated significant variation among study regions (12% of total variance) and moderate differentiation between populations (10%). Population differentiation was high at the westernmost range edge (11.5%, Germany), but also in the east (13.4%, Hungary), compared to the Austrian study region (8.6%). Correspondingly, AFLP diversity was highest in the center of the study transect in eastern Austria. CpDNA haplotypes support a pattern of regional structuring with the strongest separation of the westernmost range edge, and some haplotype sharing among Austrian and Hungarian individuals. Equilibrating nucleotide versus haplotype diversity patterns, the highly diverse populations at the Pannonian range edge (Austria) indicate long-term persistence, while Central Pannonian populations are obviously effected by recent bottlenecks. Intermediate nucleotide, but high haplotype diversity within the westernmost exclave (Swabian Alb), is indicative of a founder bottleneck during its pre-LGM or early postglacial migration history, followed by sufficient time to accumulate cpDNA variation. The not obviously reduced genetic diversity and distinctiveness of L. flavum at the westernmost range edge suggest a long-term persistence (relict status) of populations in this region, where the species has survived probably even the Würm glaciation in extra-Mediterranean refugia. This genetic relict variation represents an important part of the overall genetic diversity found in the western periphery of this steppe plant and highlights the high conservation priority of respective gene pools.

Response to Comment on "Worldwide evidence of a unimodal relationship between productivity and plant species richness".

Tredennick et al. criticize one of our statistical analyses and emphasize the low explanatory power of models relating productivity to diversity. These criticisms do not detract from our key findings, including evidence consistent with the unimodal constraint relationship predicted by the humped-back model and evidence of scale sensitivities in the form and strength of the relationship.

Plant ecology. Worldwide evidence of a unimodal relationship between productivity and plant species richness.

The search for predictions of species diversity across environmental gradients has challenged ecologists for decades. The humped-back model (HBM) suggests that plant diversity peaks at intermediate productivity; at low productivity few species can tolerate the environmental stresses, and at high productivity a few highly competitive species dominate. Over time the HBM has become increasingly controversial, and recent studies claim to have refuted it. Here, by using data from coordinated surveys conducted throughout grasslands worldwide and comprising a wide range of site productivities, we provide evidence in support of the HBM pattern at both global and regional extents. The relationships described here provide a foundation for further research into the local, landscape, and historical factors that maintain biodiversity.