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  1. Zamia integrifolia L.f. (Cycadales), a threatened cycad native to Florida, depends on 2 native beetle species for pollination: Rhopalotria slossoni (Chevrolat; Coleoptera: Belidae) and Pharaxanotha floridana (Casey; Coleoptera: Erotylidae). Both insects are brood-site pollina- tion mutualists, known to live and feed within the pollen (male) cone. However, for pollination to occur, beetles must also visit ovulate (fe- male) cones, which have been assumed to offer no benefits to them as food or nurseries. We tested the potential for beetle pollinator use of ovulate cones by performing no-choice behavior and feeding trials for adults of both beetle species on both ovulate cones and pollen cones of Z. integrifolia. Rhopalotria slossoni beetles showed greater survival on ovulate cone tissues despite showing no significant difference in to- tal tissue mass consumed between cone sexes. Conversely, P. floridana consumed more tissue mass from ovulate cone scales yet showed no difference in survivorship on ovulate vs. pollen cone scales. Although neither beetle species is found in large numbers on ovulate cones in the field, our laboratory study suggests that both species could po- tentially benefit from feeding on ovulate cone tissues, questioning the standing hypothesis that Z. integrifolia pollination occurs by deceit. 
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    Free, publicly-accessible full text available September 1, 2024
  2. Abstract Background

    The evening primrose family (Onagraceae) includes 664 species (803 taxa) with a center of diversity in the Americas, especially western North America. Ongoing research in Onagraceae includes exploring striking variation in floral morphology, scent composition, and breeding system, as well as the role of these traits in driving diversity among plants and their interacting pollinators and herbivores. However, these efforts are limited by the lack of a comprehensive, well-resolved phylogeny. Previous phylogenetic studies based on a few loci strongly support the monophyly of the family and the sister relationship of the two largest tribes but fail to resolve several key relationships.

    Results

    We used a target enrichment approach to reconstruct the phylogeny of Onagraceae using 303 highly conserved, low-copy nuclear loci. We present a phylogeny for Onagraceae with 169 individuals representing 152 taxa sampled across the family, including extensive sampling within the largest tribe, Onagreae. Deep splits within the family are strongly supported, whereas relationships among closely related genera and species are characterized by extensive conflict among individual gene trees.

    Conclusions

    This phylogenetic resource will augment current research projects focused throughout the family in genomics, ecology, coevolutionary dynamics, biogeography, and the evolution of characters driving diversification in the family.

     
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  3. Studies of pollination biology often focus on visual and olfactory aspects of attraction, with few studies ad- dressing behavioral responses and morphological adaptation to primary metabolic attributes. As part of an in-depth study of obligate nursery pollination of cycads, we find that Rhopalotria furfuracea weevils show a strong physiological response and behavioral orientation to the cone humidity of the host plant Zamia furfur- acea in an equally sensitive manner to their responses to Z. furfuracea-produced cone volatiles. Our results demonstrate that weevils can perceive fine-scale differences in relative humidity (RH) and that individuals exhibit a strong behavioral preference for higher RH in binary choice assays. Host plant Z. furfuracea pro- duces a localized cloud of higher than ambient humidity around both pollen and ovulate cones, and R. furfuracea weevils preferentially land at the zone of maximum humidity on ovulate cones, i.e., the cracks between rows of megasporophylls that provide access to the ovules. Moreover, R. furfuracea weevils exhibit striking antennal morphological traits associated with RH perception, suggesting the importance of humidity sensing in the evolution of this insect lineage. Results from this study suggest that humidity functions in a signal-like fashion in this highly specialized pollination system and help to characterize a key pollination- mediating trait in an ancient plant lineage. 
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    Free, publicly-accessible full text available May 1, 2024
  4. Abstract Premise

    Floral scent is a complex trait that mediates many plant–insect interactions, but our understanding of how floral scent variation evolves, either independently or in concert with other traits, remains limited. Assessing variation in floral scent at multiple levels of biological organization and comparing patterns of variation in scent to variation in other floral traits can contribute to our understanding of how scent variation evolves in nature.

    Methods

    We used a greenhouse common garden experiment to investigate variation in floral scent at three scales—within plants, among plants, and among populations—and to determine whether scent, alone or in combination with morphology and rewards, contributes to population differentiation inOenothera cespitosasubsp.marginata. Its range spans most of the biomes in the western United States, such that variation in both the abiotic and biotic environment could contribute to trait variation.

    Results

    Multiple analytical approaches demonstrated substantial variation among and within populations in compound‐specific and total floral scent measures. Overall, populations were differentiated in morphology and reward traits and in scent. Across populations, coupled patterns of variation in linalool, leucine‐derived compounds, and hypanthium length are consistent with a long‐tongued moth pollination syndrome.

    Conclusions

    The considerable variation in floral scent detected within populations suggests that, similar to other floral traits, variation in floral scent may have a heritable genetic component. Differences in patterns of population differentiation in floral scent and in morphology and rewards indicate that these traits may be shaped by different selective pressures.

     
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  5. Abstract Previous studies have considered floral humidity to be an inadvertent consequence of nectar evaporation, which could be exploited as a cue by nectar-seeking pollinators. By contrast, our interdisciplinary study of a night-blooming flower, Datura wrightii , and its hawkmoth pollinator, Manduca sexta , reveals that floral relative humidity acts as a mutually beneficial signal in this system. The distinction between cue- and signal-based functions is illustrated by three experimental findings. First, floral humidity gradients in Datura are nearly ten-fold greater than those reported for other species, and result from active (stomatal conductance) rather than passive (nectar evaporation) processes. These humidity gradients are sustained in the face of wind and are reconstituted within seconds of moth visitation, implying substantial physiological costs to these desert plants. Second, the water balance costs in Datura are compensated through increased visitation by Manduca moths, with concomitant increases in pollen export. We show that moths are innately attracted to humid flowers, even when floral humidity and nectar rewards are experimentally decoupled. Moreover, moths can track minute changes in humidity via antennal hygrosensory sensilla but fail to do so when these sensilla are experimentally occluded. Third, their preference for humid flowers benefits hawkmoths by reducing the energetic costs of flower handling during nectar foraging. Taken together, these findings suggest that floral humidity may function as a signal mediating the final stages of floral choice by hawkmoths, complementing the attractive functions of visual and olfactory signals beyond the floral threshold in this nocturnal plant-pollinator system. 
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  6. Research on floral volatiles has grown substantially in the last 20 years, which has generated insights into their diversity and prevalence. These studies have paved the way for new research that explores the evolutionary origins and ecological consequences of different types of variation in floral scent, including community-level, functional, and environmentally induced variation. However, to address these types of questions, novel approaches are needed that can handle large sample sizes, provide quality control measures, and make volatile research more transparent and accessible, particularly for scientists without prior experience in this field. Drawing upon a literature review and our own experiences, we present a set of best practices for next-generation research in floral scent. We outline methods for data collection (experimental designs, methods for conducting field collections, analytical chemistry, compound identification) and data analysis (statistical analysis, database integration) that will facilitate the generation and interpretation of quality data. For the intermediate step of data processing, we created the R package bouquet , which provides a data analysis pipeline. The package contains functions that enable users to convert chromatographic peak integrations to a filtered data table that can be used in subsequent statistical analyses. This package includes default settings for filtering out non-floral compounds, including background contamination, based on our best-practice guidelines, but functions and workflows can be easily customized as necessary. Next-generation research into the ecology and evolution of floral scent has the potential to generate broadly relevant insights into how complex traits evolve, their genomic architecture, and their consequences for ecological interactions. In order to fulfill this potential, the methodology of floral scent studies needs to become more transparent and reproducible. By outlining best practices throughout the lifecycle of a project, from experimental design to statistical analysis, and providing an R package that standardizes the data processing pipeline, we provide a resource for new and seasoned researchers in this field and in adjacent fields, where high-throughput and multi-dimensional datasets are common. 
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  7. Evison, Sophie (Ed.)
    1. Precise pollen placement on floral visitors can improve pollen transfer, but in many plant species, pollen is deposited onto the flexible proboscises of long-tongued insects. These proboscises are curled and uncurled between floral visits, potentially causing pollen to be lost or displaced. Rates of pollen movement and loss resulting from proboscis curling, and hence the potential quality of long-tongued insects as pollinators, are unknown. 2. Here, pollen loss and movement on the proboscises of Manduca sexta (Sphingidae) hawkmoths was experimentally measured. It was predicted that (i) proboscis curling causes pollen loss; (ii) pollen that is not lost is displaced from its deposition site; and (iii) repeated curls result in more displacement. Pollen from Datura wrightii, an important nectar plant for M. sexta, was placed distal to the knee bend on M. sexta proboscises, and the number and location of grains was recorded after proboscis curls. 3. Consistent with the hypotheses, proboscis curling caused significant pollen loss. (i) A single curl resulted in the loss of almost 75% of the pollen from the placement site; after repeated curling, 98% of grains were lost from this site. (ii) A single curl was also sufficient to displace pollen across all surfaces of the proboscis, but (iii) further curling did not affect its distribution across surfaces. 4. Together, these results suggest that precise pollen placement on the proboscises of hawkmoths would be unlikely to increase pollen transfer success. Strategies by which flowering plants might mitigate the effects of pollen loss from visitors with flexible pollen-pickup structures are discussed. 
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