Automated extraction of seed morphological traits from images

The description of biological objects, such as seeds, mainly relies on manual measurements of few characteristics, and on visual classification of structures, both of which can be subjective, error prone and time-consuming. Image analysis tools offer means to address these shortcomings, but we currently lack a method capable of automatically handling seeds from different taxa with varying morphological attributes and obtaining interpretable results. Here, we provide a simple image acquisition and processing protocol and introduce Traitor, an open-source software available as a command-line interface (CLI), which automates the extraction of seed morphological traits from images. The workflow for trait extraction consists of scanning seeds against a high-contrast background, correcting image colours, and analysing images with the software. Traitor is capable of processing hundreds of images of varied taxa simultaneously with just three commands, and without a need for training, manual fine-tuning or thresholding. The software automatically detects each object in the image and extracts size measurements, traditional morphometric descriptors widely used by scientists and practitioners, standardised shape coordinates, and colorimetric measurements. The method was tested on a dataset comprising of 91,667 images of seeds from 1228 taxa. Traitor's extracted average length and width values closely matched the average manual measurements obtained from the same collection (concordance correlation coefficient of 0.98). Further, we used a large image dataset to demonstrate how Traitor's output can be used to obtain representative seed colours for taxa, determine the phylogenetic signal of seed colour, and build objective classification categories for shape with high levels of visual interpretability. Our approach increases productivity and allows for large-scale analyses that would otherwise be unfeasible. Traitor enables the acquisition of data that are readily comparable across different taxa, opening new avenues to explore functional relevance of morphological traits and to advance on new tools for seed identification

Automated extraction of seed morphological traits from images

Abstract The description of biological objects, such as seeds, mainly relies on manual measurements of few characteristics, and on visual classification of structures, both of which can be subjective, error prone and time‐consuming. Image analysis tools offer means to address these shortcomings, but we currently lack a method capable of automatically handling seeds from different taxa with varying morphological attributes and obtaining interpretable results. Here, we provide a simple image acquisition and processing protocol and introduce Traitor, an open‐source software available as a command‐line interface (CLI), which automates the extraction of seed morphological traits from images. The workflow for trait extraction consists of scanning seeds against a high‐contrast background, correcting image colours, and analysing images with the software. Traitor is capable of processing hundreds of images of varied taxa simultaneously with just three commands, and without a need for training, manual fine‐tuning or thresholding. The software automatically detects each object in the image and extracts size measurements, traditional morphometric descriptors widely used by scientists and practitioners, standardised shape coordinates, and colorimetric measurements. The method was tested on a dataset comprising of 91,667 images of seeds from 1228 taxa. Traitor's extracted average length and width values closely matched the average manual measurements obtained from the same collection (concordance correlation coefficient of 0.98). Further, we used a large image dataset to demonstrate how Traitor's output can be used to obtain representative seed colours for taxa, determine the phylogenetic signal of seed colour, and build objective classification categories for shape with high levels of visual interpretability. Our approach increases productivity and allows for large‐scale analyses that would otherwise be unfeasible. Traitor enables the acquisition of data that are readily comparable across different taxa, opening new avenues to explore functional relevance of morphological traits and to advance on new tools for seed identification

Using activated charcoal to remove substances interfering with the colorimetric assay of inorganic phosphate in plant extracts

Aim Organic substances in leaves of several southwest Australian native species interfere with sensitive colorimetric assays and prevent quantification of inorganic phosphate concentration ([Pi]). We aimed to develop a reproducible routine procedure for treating leaf extracts with activated charcoal (AC) to remove interfering substances, allowing the determination of [Pi] by the malachite green spectrophotometric assay. Methods Leaf extracts of native plants from southwest Australia in 1% (v/v) acetic acid were treated with 10 mg mL−1 acid-washed AC for removal of interfering substances. Standard solutions (0 to 18 μM Pi) with and without AC treatment were compared to quantify Pi loss. A spiking and recovery test was performed to validate the AC treatment. Results Leaf extracts treated with AC exhibited distinguishable absorbance peaks for the malachite green-orthophosphate complex between 630 and 650 nm, as opposed to untreated samples. The Pi-adsorption by AC represented a relatively larger fraction of [Pi] in solutions at 0–4 μM Pi range and stabilised at higher [Pi] when maximum adsorption capacity of AC reached at 11.7 μg Pi g−1AC. The Pi recovery after AC treatment in spiked samples ranged between 100 and 111%. Conclusion The AC treatment successfully removed interfering substances from samples but caused Pi loss. Thus, quantification of [Pi] in AC-treated extracts requires sample [Pi] ≥ 6 μM Pi and the use of AC-treated standards. The error of the AC treatment was minor compared with environmental variability of leaf [Pi]. The AC treatment was a reproducible time- and cost-effective method to remove interfering substances from leaf extracts

Overcoming challenges on using native seeds for restoration of megadiverse resource-poor environments: a reply to Madsen et al.

International audienceMadsen et al. (2016) reviewed several major limiting factors to establishment of seedlings in nonforest ecosystems (NFE), and proposed seed enhancement technologies to overcome these restoration barriers. However, biodiverse nutrient-poor NFE present additional hurdles that preclude landscape-scale seed-based restoration and were not mentioned in their review. Here, we discuss issues related to native seed availability and provenance, and shortfalls in knowledge on seed quality testing and dormancy release that severely hamper restoration of degraded nutrient-impoverished NFE. We present alternatives for overcoming these challenges and highlight the need for investments to find more practical and cost-effective options for broad-scale restoration

Ontogenetic shifts in plant ecological strategies

International audience1. Ontogenetic shifts in plants are ubiquitous, but their ecological and evolutionary significance remain largely unknown. Juveniles have few opportunities to accumulate sufficient biomass to withstand damage, whereas adults allocate resources to reproduction. Thus, age-specific environmental filters are expected to drive selection towards specific resource-investment strategies for each developmental stage. 2. We tested whether species exhibited strategies that favour faster rates of biomass accumulation during early developmental stages to cope with vulnerability, shifting towards habitat-dependent strategies to optimise resource acquisition and use later on. We also hypothesised that juveniles exhibit greater intraspecific variability of strategies than adults, as a result of continuous filtering of traits throughout the plant's development towards adulthood. 3. We measured key leaf traits-leaf area (LA), leaf dry matter content (LDMC) and specific leaf area (SLA)-and calculated scores for competitor (C), stress-tolerator (S), ruderal (R) strategies of the CSR system for juveniles of 54 species and conspecific adults of 27 species naturally occurring either in unproductive (grassland) or productive (forest) habitats. 4. Juveniles exhibited higher SLA and LA and lower LDMC, and thus a more R-strategy in both habitats when compared with adults, but were not necessarily "R" in absolute terms. We also found smaller intraspecific variation for the R-strategy in adults compared with juveniles in both habitats, whereas the variation of the other strategies was habitat dependent. 5. Ontogenetic shifts in ecological strategies appear to have been selected as a response to ontogeny-dependent filters. Thus, strategies that favour less costly leaves and faster growth rates (relatively R-selected) in juveniles shift towards C and/or S strategies in adulthood, depending on habitat productivity. Nevertheless, habitat-dependent specialisation seems to be a major driver of ecological strategy selection in juveniles. Our study reveals ontogeny-dependent strategies, offering a new approach to integrate plant development and functional specialisation

Intraspecific variation in fruit–frugivore interactions: effects of fruiting neighborhood and consequences for seed dispersal

We thank GW Fernandes for allowing research in his property and the use of his facilities for the study. We also thank PHV da Silva and LB Fantecelle for fieldwork assistance. The comments made by three anonymous reviewers significantly improved the manuscript. TJ Guerra, RLC Dayrell, AJ Arruda and AL Teixido received scholarships from CAPES, JVS Messeder received a scholarship from FAPEMIG and FAO Silveira receives research productivity grant from CNPq. Financial support was provided by CAPES, CNPq and FAPEMIG (APQ02317-14).The extent of specialization/generalization continuum in fruit–frugivore interactions at the individual level remains poorly explored. Here, we investigated the interactions between the Neotropical treelet Miconia irwinii (Melastomataceae) and its avian seed dispersers in Brazilian campo rupestre. We built an individual-based network to derive plant degree of interaction specialization regarding disperser species. Then, we explored how intraspecific variation in interaction niche breadth relates to fruit availability on individual plants in varying densities of fruiting conspecific neighbors, and how these factors affect the quantity of viable seeds dispersed. We predicted broader interaction niche breadths for individuals with larger fruit crops in denser fruiting neighborhoods. The downscaled network included nine bird species and 15 plants, which varied nearly five-fold in their degree of interaction specialization. We found positive effects of crop size on visitation and fruit removal rates, but not on degree of interaction specialization. Conversely, we found that an increase in the density of conspecific fruiting neighbors both increased visitation rate and reduced plant degree of interaction specialization. We suggest that tracking fruit-rich patches by avian frugivore species is the main driver of density-dependent intraspecific variation in plants’ interaction niche breadth. Our study shed some light on the overlooked fitness consequences of intraspecific variation in interaction niches by showing that individuals along the specialization/generalization continuum may have their seed dispersed with similar effectiveness. Our study exemplifies how individual-based networks linking plants to frugivore species that differ in their seed dispersal effectiveness can advance our understanding of intraspecific variation in the outcomes of fruit–frugivore interactions.Depto. de Biodiversidad, Ecología y EvoluciónFac. de Ciencias BiológicasTRUEpu

Handling by avian frugivores affects diaspore secondary removal.

The balance between the costs and benefits of fleshy fruit production depends on the feeding behavior of their seed dispersers, which might effectively disperse seeds to farther areas or drop beneath parent plants some diaspores they handle during frugivory bouts. Nevertheless, the consequences of variation in fruit handling by primary seed dispersers on the secondary removal of diaspores remains poorly understood. We conducted a field study to determine how variation in fruit handling by avian frugivores affects short-term secondary removal of Miconia irwinii (Melastomataceae) diaspores by the ground-dwelling fauna in campo rupestre vegetation, southeastern Brazil. We conducted factorial experiments manipulating: (1) different outcomes of primary fruit/seed removal by birds, (2) distances of diaspore deposition from conspecifics, and (3) the access of ants and vertebrates to diaspores. We showed that secondary removal of diaspores was highly variable at the population scale, with an overall low removal rate by the ground-dwelling fauna (13% seeds, 19% fruits). However, we found that gut-passed seeds embedded in bird feces were less removed than seeds expelled from fruits. Gut-passed seeds were more likely to be removed by ant species acting as secondary dispersers, whereas pulp-free seeds dropped by birds were likely to interact with potential seed predators, including ants and rodents. We found no clear effect of dispersal from parent plant vicinity on seed removal, but fruit removal was significantly higher near parent plants. Partially defleshed fruits were more removed than intact fruits. The removal of fruits by ant and vertebrate rescuers, including lizards and birds, might reduce the costs of interactions with less effective dispersers that drop partially defleshed fruits under parent plants. Our study highlights that variation in fruit handling by primary avian seed dispersers mediate subsequent interactions among discarded diaspores and ground-dwelling animals, potentially affecting final seed fates. Moreover, we argue that escape-related benefits of dispersal can be contingent on how primary dispersers handle and discard seeds

Topsoil disturbance reshapes diaspore interactions with ground‐foraging animals in a megadiverse grassland

International audienceQuestions: Anthropogenic disturbances are known to be followed by extremely poor recovery in edaphic grasslands. However, the role of interactions with diaspore predators and secondary dispersers, which compose the dispersal filter and modulate plant community recovery, has been overlooked. We performed field experiments to investigate how soil disturbances affect diaspore interactions with the ground‐foraging fauna to better understand how disturbance influences regeneration potential.Location: Campo rupestre vegetation, megadiverse edaphic grasslands, southeastern Brazil.Methods: We used diaspores from five native species to compare removal rates between disturbed (top soil removal) and preserved sites; we also controlled invertebrate and vertebrate access to determine their role. In addition, we assessed differences in the diaspore removal effectiveness (DRE) and the structure of animal–diaspore interactions through network based‐approach.Results: For three species, the average diaspore removal rates was relatively high (between 30% and 70%). Invertebrates were the most common removal agents in both disturbed and preserved sites. Interactions with foraging fauna and removal rates were 20% smaller in disturbed sites. Considering all diaspore removal events in disturbed sites, 24% resulted in the displacement from disturbed to preserved sites, but no diaspore was transported from preserved to disturbed ones. The animal–diaspore network was exclusively composed by ant–diaspore interactions and was more diverse and robust in preserved sites compared with disturbed ones. Seed predator ants (Pheidole and Dorymyrmex) were more common in disturbed sites. Furthermore, significant differences were found in the DRE between ant species and site types, suggesting specificity in the provision of dispersal services.Conclusions: Topsoil removal affected removal proportions, DRE and ant–diaspore interaction network structure. The lack of diaspore dispersal towards disturbed sites indicates that soil removal affects secondary seed removal dynamics, impeding recovery potential. Disturbance negatively affected diaspore fate by reshaping interactions with ground‐foraging secondary seed dispersers and predators, constraining the development of seed bank and thus impacting upon vegetation dynamics and resilience