Succession comprises a sequence of threshold-induced community assembly processes towards multidiversity.

Research on successions and community assembly both address the same processes such as dispersal, species sorting, and biotic interactions but lack unifying concepts. Recent theoretical advances integrated both research lines proposing a sequence of stochastic and deterministic processes along successional gradients. Shifts in ecosystem states along successional gradients are predicted to occur abruptly once abiotic and biotic factors dominate over dispersal as main driver. Considering the multidiversity composed of five organismal groups including plants, animals, and microbes, our results imply that stochastic, likely dispersal-dominated, processes are replaced by rather deterministic processes such as environmental filtering and biotic interactions after around 60 years of succession in a glacier forefield. The niche-based character of later successional processes is further supported by a decline in multi-beta-diversity. Our results may update concepts of community assembly by considering multiple taxa, help to bridge the gap between research on successions and community assembly, and provide insights into the emergence of multidiverse and complex ecosystems.

Accuracy of mutual predictions of plant and microbial communities vary along a successional gradient in an alpine glacier forefield.

Receding glaciers create virtually uninhabited substrates waiting for initial colonization of bacteria, fungi and plants. These glacier forefields serve as an ideal ecosystem for studying transformations in community composition and diversity over time and the interactions between taxonomic groups in a dynamic landscape. In this study, we investigated the relationships between the composition and diversity of bacteria, fungi, and plant communities as well as environmental factors along a successional gradient. We used random forest analysis assessing how well the composition and diversity of taxonomic groups and environmental factors mutually predict each other. We did not identify a single best indicator for all taxonomic and environmental properties, but found specific predictors to be most accurate for each taxon and environmental factor. The accuracy of prediction varied considerably along the successional gradient, highlighting the dynamic environmental conditions along the successional gradient that may also affect biotic interactions across taxa. This was also reflected by the high accuracy of predictions of plot age by all taxa. Next to plot age, our results indicate a strong importance of pH and temperature in structuring microbial and plant community composition. In addition, taxonomic groups predicted the community composition of each other more accurately than environmental factors, which may either suggest that these groups similarly respond to other not measured environmental factors or that direct interactions between taxa shape the composition of their communities. In contrast, diversity of taxa was not well predicted, suggesting that community composition of one taxonomic group is not a strong driver of the diversity of another group. Our study provides insights into the successional development of multidiverse communities shaped by complex interactions between taxonomic groups and the environment.

Humans Share More Preferences for Floral Phenotypes With Pollinators Than With Pests.

Studies on the selection of floral traits usually consider pollinators and sometimes herbivores. However, humans also exert selection on floral traits of ornamental plants. We compared the preferences of bumblebees (Bombus terrestris), thrips (Frankliniella occidentalis), and humans for flowers of snapdragon. From a cross of two species, Antirrhinum majus and Antirrhinum linkianum, we selected four Recombinant Inbred Lines (RILs). We characterised scent emission from whole flowers and stamens, pollen content and viability, trichome density, floral shape, size and colour of floral parts. We tested the preferences of bumblebees, thrips, and humans for whole flowers, floral scent bouquets, stamen scent, and individual scent compounds. Humans and bumblebees showed preferences for parental species, whereas thrips preferred RILs. Colour and floral scent, in combination with other floral traits, seem relevant phenotypes for all organisms. Remarkably, visual traits override scent cues for bumblebees, although, scent is an important trait when bumblebees cannot see the flowers, and methyl benzoate was identified as a key attractant for them. The evolutionary trajectory of flowers is the result of multiple floral traits interacting with different organisms with different habits and modes of interaction.

Divergent assembly processes? A comparison of the plant and soil microbiome with plant communities in a glacier forefield.

Community assembly is a result of dispersal, abiotic and biotic characteristics of the habitat as well as stochasticity. A direct comparison between the assembly of microbial and 'macrobial' organisms is hampered by the sampling of these communities in different studies, at different sites or on different scales. In a glacier forefield in the Austrian Alps, we recorded the soil and plant microbiome (bacteria and fungi) and plants that occurred in the same landscape and in close proximity in the same plots. We tested five predictions deduced from assembly processes and revealed deviating patterns of assembly in these community types. In short, microbes appeared to be less dispersal limited than plants and soil microbes, and plants strongly responded to abiotic factors whereas the leaf microbiome was plant species specific and well buffered from environmental conditions. The observed differences in community assembly processes may be attributed to the organisms' dispersal abilities, the exposure of the habitats to airborne propagules and habitat characteristics. The finding that assembly is conditional to the characteristics of the organisms, the habitat and the spatial scale under consideration is thus central for our understanding about the establishment and the maintenance of biodiversity.

gcProfileMakeR: An R Package for Automatic Classification of Constitutive and Non-Constitutive Metabolites.

Metabolomes comprise constitutive and non-constitutive metabolites produced due to physiological, genetic or environmental effects. However, finding constitutive metabolites and non-constitutive metabolites in large datasets is technically challenging. We developed gcProfileMakeR, an R package using standard Excel output files from an Agilent Chemstation GC-MS for automatic data analysis using CAS numbers. gcProfileMakeR has two filters for data preprocessing removing contaminants and low-quality peaks. The first function NormalizeWithinFiles, samples assigning retention times to CAS. The second function NormalizeBetweenFiles, reaches a consensus between files where compounds in close retention times are grouped together. The third function getGroups, establishes what is considered as Constitutive Profile, Non-constitutive by Frequency i.e., not present in all samples and Non-constitutive by Quality. Results can be plotted with the plotGroup function. We used it to analyse floral scent emissions in four snapdragon genotypes. These included a wild type, Deficiens nicotianoides and compacta affecting floral identity and RNAi:AmLHY targeting a circadian clock gene. We identified differences in scent constitutive and non-constitutive profiles as well as in timing of emission. gcProfileMakeR is a very useful tool to define constitutive and non-constitutive scent profiles. It also allows to analyse genotypes and circadian datasets to identify differing metabolites.

The Effect of Post-harvest Conditions in Narcissus sp. Cut Flowers Scent Profile.

Narcissus flowers are used as cut flowers and to obtain high quality essential oils for the perfume industry. As a winter crop in the Mediterranean area, it flowers at temperatures ranging between 10 and 15°C during the day and 3-10°C during the night. Here we tested the impact of different light and temperature conditions on scent quality during post-harvest. These two types of thermoperiod and photoperiod. We also used constant darkness and constant temperatures. We found that under conditions of 12:12 Light Dark and 15-5°C, Narcissus emitted monoterpenes and phenylpropanoids. Increasing the temperature to 20°-10°C in a 12:12 LD cycle caused the loss of cinnamyl acetate and emission of indole. Under constant dark, there was a loss of scent complexity. Constant temperatures of 20°C caused a decrease of scent complexity that was more dramatic at 5°C, when the total number of compounds emitted decreased from thirteen to six. Distance analysis confirmed that 20°C constant temperature causes the most divergent scent profile. We found a set of four volatiles, benzyl acetate, eucalyptol, linalool, and ocimene that display a robust production under differing environmental conditions, while others were consistently dependent on light or thermoperiod. Scent emission changed significantly during the day and between different light and temperature treatments. Under a light:dark cycle and 15-5°C the maximum was detected during the light phase but this peak shifted toward night under 20-10°C. Moreover, under constant darkness the peak occurred at midnight and under constant temperature, at the end of night. Using Machine Learning we found that indole was the volatile with a highest ranking of discrimination followed by D-limonene. Our results indicate that light and temperature regimes play a critical role in scent quality. The richest scent profile is obtained by keeping flowers at 15°-5°C thermoperiod and a 12:12 Light Dark photoperiod.

A comparison of semi-quantitative methods suitable for establishing volatile profiles.

BACKGROUND: Full scent profiles emitted by living tissues can be screened by using total ion chromatograms generated in full scan mode and gas chromatography-mass spectrometry technique using Headspace Sorptive Extraction. This allows the identification of specific compounds and their absolute quantification or relative abundance. Quantifications ideally should be based on calibration curves using standards for each compound. However, the unpredictable composition of Volatile Organic Compounds (VOCs) and lack of standards make this approach difficult. Researchers studying scent profiles therefore concentrate on identifying specific scent footprints i.e. relative abundance rather than absolute quantities. We compared several semi-quantitative methods: external calibration curves generated in the sampling system and by liquid addition of standards to stir bars, total integrated peak area per fresh weight (FW), normalized peak area per FW, semi-quantification based on internal standard abundance, semi-quantification based on the nearest n-alkane and percentage of emission. Furthermore, we explored the usage of nearest components and single calibrators for semi-quantifications. RESULTS: Any of the semi-quantification methods based on a standard produced similar or even identical results compared to quantification by a true-standard for a compound, except for the method based on standard addition. Each method beholds advantages and disadvantages regarding level of accuracy, experimental variability, acceptance and retrieved quantities. CONCLUSIONS: Our data shows that, except for the method of standard addition to the biological sample, the rest of the semi-quantification methods studied give highly similar statistical results. Any of the methodologies presented here can therefore be considered as valid for scent profiling. Regarding relative proportions of VOCs, the generation of calibration curves for each compound analysed is not necessary.

Diel pattern of circadian clock and storage protein gene expression in leaves and during seed filling in cowpea (Vigna unguiculata).

BACKGROUND: Cowpea (Vigna unguiculata) is an important source of protein supply for animal and human nutrition. The major storage globulins VICILIN and LEGUMIN (LEG) are synthesized from several genes including LEGA, LEGB, LEGJ and CVC (CONVICILIN). The current hypothesis is that the plant circadian core clock genes are conserved in a wide array of species and that primary metabolism is to a large extent controlled by the plant circadian clock. Our aim was to investigate a possible link between gene expression of storage proteins and the circadian clock. RESULTS: We identified cowpea orthologues of the core clock genes VunLHY, VunTOC1, VunGI and VunELF3, the protein storage genes VunLEG, VunLEGJ, and VunCVC as well as nine candidate reference genes used in RT-PCR. ELONGATION FACTOR 1-A (ELF1A) resulted the most suitable reference gene. The clock genes VunELF3, VunGI, VunTOC1 and VunLHY showed a rhythmic expression profile in leaves with a typical evening/night and morning/midday phased expression. The diel patterns were not completely robust and only VungGI and VungELF3 retained a rhythmic pattern under free running conditions of darkness. Under field conditions, rhythmicity and phasing apparently faded during early pod and seed development and was regained in ripening pods for VunTOC1 and VunLHY. Mature seeds showed a rhythmic expression of VunGI resembling leaf tissue under controlled growth chamber conditions. Comparing time windows during developmental stages we found that VunCVC and VunLEG were significantly down regulated during the night in mature pods as compared to intermediate ripe pods, while changes in seeds were non-significant due to high variance. The rhythmic expression under field conditions was lost under growth chamber conditions. CONCLUSIONS: The core clock gene network is conserved in cowpea leaves showing a robust diel expression pattern except VunELF3 under growth chamber conditions. There appears to be a clock transcriptional reprogramming in pods and seeds compared to leaves. Storage protein deposition may be circadian regulated under field conditions but the strong environmental signals are not met under artificial growth conditions. Diel expression pattern in field conditions may result in better usage of energy for protein storage.

Genetic Analysis of Natural Variation in Antirrhinum Scent Profiles Identifies BENZOIC ACID CARBOXYMETHYL TRANSFERASE As the Major Locus Controlling Methyl Benzoate Synthesis.

The Antirrhinum genus has a considerable complexity in the scent profiles produced by different species. We have analyzed the genetic differences between A. majus and A. linkianum, two species divergent in the emission of methyl benzoate, methyl cinnamate, acetophenone, and ocimene. The genetic analysis showed that all compounds segregated in a Mendelian fashion attributable to one or two loci with simple or epistatic interactions. Several lines lacked methyl benzoate, a major Volatile Organic Compound emitted by A. majus but missing in A. linkianum. Using a candidate gene approach, we found that the BENZOIC ACID CARBOXYMETHYL TRANSFERASE from A. linkianum appeared to be a null allele as we could not detect mRNA expression. The coding region did not show significant differences that could explain the loss of expression. The intron-exon boundaries was also conserved indicating that there is no alternative splicing in A. linkianum as compared to A. majus. However, it showed multiple polymorphisms in the 5' promoter region including two insertions, one harboring an IDLE MITE transposon with additional sequences with high homology to the PLENA locus and a second one with somewhat lower homology to the regulatory region of the VENOSA locus. It also had a 778 bp deletion as compared to the A. majus BAMT promoter region. Our results show that the differences in scent emission between A. majus and A. linkianum may be traced back to single genes involved in discrete biosynthetic reactions such as benzoic acid methylation. Thus, natural variation of this complex trait maybe the result of combinations of wild type, and loss of function alleles in different genes involved in discrete VOCs biosynthesis. Furthermore, the presence of active transposable elements in the genus may account for rapid evolution and instability, raising the possibility of adaptation to local pollinators.

Phenotypic Space and Variation of Floral Scent Profiles during Late Flower Development in Antirrhinum.

The genus Antirrhinum comprises about 28 species with a center of origin in the Iberian Peninsula. They show an important diversity of growing niches. We have performed a comprehensive analysis of scent profiles in eight wild species, Antirrhinum linkianum, A. tortuosum, A. cirrigherum, A. latifolium, A. meonanthum, A. braun-blanquetii, A. barrelieri, and A. graniticum. We used also two laboratory inbred lines A. majus, 165E and Sippe50. We identified 63 volatile organic compounds (VOCs) belonging to phenylpropanoids, benzenoids, mono- and sesquiterpenes, nitrogen-containing compounds, and aliphatic alcohols previously described in plants. Twenty-four VOCs were produced at levels higher than 2% of total VOC emission, while other VOCs were emitted in trace amounts. The absolute scent emission varied during flower maturation and species. The lowest emitting was A. meonanthum while A. tortuosum had the largest emissions. Species were clustered according to their scent profiles and the resulting dendrogram matched the current species phylogeny. However, two accessions, A. majus Sippe 50 and A. braun-blanquetii, showed development-specific changes in their VOC composition, suggesting a precise control and fine tuning of scent profiles. Cluster analysis of the different scent components failed to identify a specific synthesis pathway, indicating a key role of scent profiles as blends. There is considerable degree of chemodiversity in scent profiles in Antirrhinum. The specific developmental stage plays an important role in scent quantitative emissions. The relative robustness of the bouquets could be an adaptation to local pollinators.