HD-Zip proteins modify floral structures for self-pollination in tomato.

Cleistogamy is a type of self-pollination that relies on the formation of a stigma-enclosing floral structure. We identify three homeodomain-leucine zipper IV (HD-Zip IV) genes that coordinately promote the formation of interlocking trichomes at the anther margin to unite neighboring anthers, generating a closed anther cone and cleistogamy (flower morphology necessitating strict self-pollination). These HD-Zip IV genes also control style length by regulating the transition from cell division to endoreduplication. The expression of these HD-Zip IV genes and their downstream gene, Style 2.1, was sequentially modified to shape the cleistogamy morphology during tomato evolution and domestication. Our results provide insights into the molecular basis of cleistogamy in modern tomato and suggest targets for improving fruit set and preventing pollen contamination in genetically modified crops.

Measuring factors affecting honey bee (Hymenoptera: Apidae) attraction to soybeans using bioacoustics monitoring.

Soybean (Glycine max (L.) Merr.) is an important agricultural crop around the world, and previous studies suggest that honey bees (Apis mellifera Linnaeus) can be a component for optimizing soybean production through pollination. Determining when bees are present in soybean fields is critical for assessing pollination activity and identifying periods when bees are absent so that bee-toxic pesticides may be applied. There are currently several methods for detecting pollinator activity, but these existing methods have substantial limitations, including the bias of pan trappings against large bees and the limited duration of observation possible using manual techniques. This study aimed to develop a new method for detecting honey bees in soybean fields using bioacoustics monitoring. Microphones were placed in soybean fields to record the audible wingbeats of foraging bees. Foraging activity was identified using the wingbeat frequency of honey bees (234 ±â€…14 Hz) through a combination of algorithmic and manual approaches. A total of 243 honey bees were detected over 10 days of recording in 4 soybean fields. Bee activity was significantly greater in blooming fields than in non-blooming fields. Temperature had no significant effect on bee activity, but bee activity differed significantly between soybean varieties, suggesting that soybean attractiveness to honey bees is heavily dependent on varietal characteristics. Refinement of bioacoustics methods, particularly through the incorporation of machine learning, could provide a practical tool for measuring the activity of honey bees and other flying insects in soybeans as well as other crops and ecosystems.

Spillover effects from invasive Acacia alter the plant-pollinator networks and seed production of native plants.

Invasive flowering plants can disrupt plant-pollinator networks. This is well documented where invasives occur amongst native plants; however, the potential for 'spillover' effects of invasives that form stands in adjacent habitats are less well understood. Here we quantify the impact of two invasive Australian species, Acacia saligna and Acacia longifolia, on the plant-pollinator networks in fynbos habitats in South Africa. We compared networks from replicate 1 ha plots of native vegetation (n = 21) that were subjected to three treatments: (1) at least 400 m from flowering Acacia; (2) adjacent to flowering Acacia, or (3) adjacent to flowering Acacia where all Acacia flowers were manually removed. We found that native flowers adjacent to stands of flowering Acacia received significantly more insect visits, especially from beetles and Apis mellifera capensis, and that visitation was more generalized. We also recorded visitation to, and the seed set of, three native flowering species and found that two received more insect visits, but produced fewer seeds, when adjacent to flowering Acacia. Our research shows that 'spillover' effects of invasive Acacia can lead to significant changes in visitation and seed production of native co-flowering species in neighbouring habitats-a factor to be considered when managing invaded landscapes.

A novel O-methyltransferase Cp4MP-OMT catalyses the final step in the biosynthesis of the volatile 1,4-dimethoxybenzene in pumpkin (Cucurbita pepo) flowers.

BACKGROUND: Floral scents play a crucial role in attracting insect pollinators. Among the compounds attractive to pollinators is 1,4-dimethoxybenzene (1,4-DMB). It is a significant contributor to the scent profile of plants from various genera, including economically important Cucurbita species. Despite its importance, the biosynthetic pathway for the formation of 1,4-DMB was not elucidated so far. RESULTS: In this study we showed the catalysis of 1,4-DMB in the presence of 4-methoxyphenol (4-MP) by protein extract from Styrian oil pumpkin (Cucurbita pepo) flowers. Based on this finding, we identified a novel O-methyltransferase gene, Cp4MP-OMT, whose expression is highly upregulated in the volatile-producing tissue of pumpkin flowers when compared to vegetative tissues. OMT activity was verified by purified recombinant Cp4MP-OMT, illustrating its ability to catalyse the methylation of 4-MP to 1,4-DMB in the presence of cofactor SAM (S-(5'-adenosyl)-L-methionine). CONCLUSIONS: Cp4MP-OMT is a novel O-methyltransferase from C. pepo, responsible for the final step in the biosynthesis of the floral scent compound 1,4-DMB. Considering the significance of 1,4-DMB in attracting insects for pollination and in the further course fruit formation, enhanced understanding of its biosynthetic pathways holds great promise for both ecological insights and advancements in plant breeding initiatives.

Floral visitors of sesame (Sesamum indicum L.): Elucidating their nectar-robbing behaviour and impacts on the plant reproduction.

Nectar robbing is common in angiosperms, especially in long tubular flowers or flowers with spurs that keep nectar out of reach of visitors. However, the robbing behaviour of bees is less understood. Here, we studied the sesame visitors, their robbing behaviour, and the impacts of robbing on plant reproductive fitness. Diverse insect species (primarily members of Hymenoptera) visited sesame flowers. The most effective pollinators were Amegilla zonata, Apis cerana, Apis dorsata, Apis florea, Ceratina binghami, Halictus acrocephalus and Xylocopa amethystina. Almost all visitors with variable percentages revealed the nectar-robbing phenomenon. Robbing activity depended on a complex of multiple attributes, including the visitor's body size, the corolla tube length, the availability and accessibility of nectar, and the resource-collecting task allocation of bees. Robbing activity varied according to flower-visiting species, flowering period and daytime. Robbing was comparatively higher in the late flowering period at 10.00-14.00 h. In the case of robbing visits, flower handling time was lower, and the visitation rate remained higher than non-robbing visits. Robbing visits did not significantly affect fruit and seed sets of sesame. Therefore, we can interpret the nectar-robbing interactions on sesame as commensal, with pollinators benefitting without altering the plant's reproductive fitness.

Variable resource allocation pattern, biased sex-ratio, and extent of sexual dimorphism in subdioecious Hippophae rhamnoides.

Evolutionary maintenance of dioecy is a complex phenomenon and varies by species and underlying pathways. Also, different sexes may exhibit variable resource allocation (RA) patterns among the vegetative and reproductive functions. Such differences are reflected in the extent of sexual dimorphism. Though rarely pursued, investigation on plant species harbouring intermediate sexual phenotypes may reveal useful information on the strategy pertaining to sex-ratios and evolutionary pathways. We studied H. rhamnoides ssp. turkestanica, a subdioecious species with polygamomonoecious (PGM) plants, in western Himalaya. The species naturally inhabits a wide range of habitats ranging from river deltas to hill slopes. These attributes of the species are conducive to test the influence of abiotic factors on sexual dimorphism, and RA strategy among different sexes. The study demonstrates sexual dimorphism in vegetative and reproductive traits. The sexual dimorphism index, aligned the traits like height, number of branches, flower production, and dry-weight of flowers with males while others including fresh-weight of leaves, number of thorns, fruit production were significantly associated with females. The difference in RA pattern is more pronounced in reproductive traits of the male and female plants, while in the PGM plants the traits overlap. In general, habitat conditions did not influence either the extent of sexual dimorphism or RA pattern. However, it seems to influence secondary sex-ratio as females show their significant association with soil moisture. Our findings on sexual dimorphism and RA pattern supports attributes of wind-pollination in the species. The observed extent of sexual dimorphism in the species reiterates limited genomic differences among the sexes and the ongoing evolution of dioecy via monoecy in the species. The dynamics of RA in the species appears to be independent of resource availability in the habitats as the species grows in a resource-limited and extreme environment.

Advances and knowledge gaps on climate change impacts on honey bees and beekeeping: A systematic review.

The Western honey bee Apis mellifera is a managed species that provides diverse hive products and contributing to wild plant pollination, as well as being a critical component of crop pollination systems worldwide. High mortality rates have been reported in different continents attributed to different factors, including pesticides, pests, diseases, and lack of floral resources. Furthermore, climate change has been identified as a potential driver negatively impacting pollinators, but it is still unclear how it could affect honey bee populations. In this context, we carried out a systematic review to synthesize the effects of climate change on honey bees and beekeeping activities. A total of 90 articles were identified, providing insight into potential impacts (negative, neutral, and positive) on honey bees and beekeeping. Interest in climate change's impact on honey bees has increased in the last decade, with studies mainly focusing on honey bee individuals, using empirical and experimental approaches, and performed at short-spatial (<10 km) and temporal (<5 years) scales. Moreover, environmental analyses were mainly based on short-term data (weather) and concentrated on only a few countries. Environmental variables such as temperature, precipitation, and wind were widely studied and had generalized negative effects on different biological and ecological aspects of honey bees. Food reserves, plant-pollinator networks, mortality, gene expression, and metabolism were negatively impacted. Knowledge gaps included a lack of studies at the apiary and beekeeper level, a limited number of predictive and perception studies, poor representation of large-spatial and mid-term scales, a lack of climate analysis, and a poor understanding of the potential impacts of pests and diseases. Finally, climate change's impacts on global beekeeping are still an emergent issue. This is mainly due to their diverse effects on honey bees and the potential necessity of implementing adaptation measures to sustain this activity under complex environmental scenarios.

La abeja occidental Apis mellifera es una especie manejada que proporciona diversos productos de la colmena y servicios de polinización, los cuales son cruciales para plantas silvestres y cultivos en todo el mundo. En distintos continentes se han registrado altas tasas de mortalidad, las cuales son atribuidas a diversos factores, como el uso de pesticidas, plagas, enfermedades y falta de recursos florales. Además, el cambio climático ha sido identificado como un potencial factor que afecta negativamente a los polinizadores, pero aún no está claro cómo podría afectar a las poblaciones de abejas melíferas. En este contexto, realizamos una revisión sistemática de la literatura disponible para sintetizar los efectos del cambio climático en las abejas melíferas y las actividades apícolas. En total, se identificaron 90 artículos que proporcionaron información sobre los posibles efectos (negativos, neutros y positivos) en las abejas melíferas y la apicultura. El interés por el impacto del cambio climático en las abejas melíferas ha aumentado en la última década, con estudios centrados principalmente en individuos de abejas melíferas, utilizando enfoques empíricos y experimentales y realizados a escalas espaciales (<10 km) y temporales (<5 años) cortas. Además, los análisis ambientales fueron basaron principalmente en datos a corto plazo (meteorológicos) y se concentraron sólo en algunos países. Variables ambientales como la temperatura, las precipitaciones y el viento fueron ampliamente estudiadas y tuvieron efectos negativos generalizados sobre distintos aspectos biológicos y ecológicos de las abejas melíferas. Además, las reservas alimenticias, las interacciones planta-polinizador, la mortalidad, la expresión génica y el metabolismo se vieron afectados negativamente. Entre los vacios de conocimiento cabe mencionar la falta de estudios a nivel de colmenar y apicultor, la escasez de estudios de predicción y percepción, la escasa representación de las grandes escalas espaciales y a mediano plazo, el déficit de análisis climáticos y la escasa comprensión de los impactos potenciales de plagas y enfermedades. Por último, las repercusiones del cambio climático en la apicultura mundial siguen siendo un tema emergente, que debe estudiarse en los distintos países. Esto se debe principalmente a sus diversos efectos sobre las abejas melíferas y a la necesidad potencial de aplicar medidas de adaptación para mantener esta actividad crucial en escenarios medioambientales complejos.

Heat and desiccation tolerances predict bee abundance under climate change.

Climate change could pose an urgent threat to pollinators, with critical ecological and economic consequences. However, for most insect pollinator species, we lack the long-term data and mechanistic evidence that are necessary to identify climate-driven declines and predict future trends. Here we document 16 years of abundance patterns for a hyper-diverse bee assemblage1 in a warming and drying region2, link bee declines with experimentally determined heat and desiccation tolerances, and use climate sensitivity models to project bee communities into the future. Aridity strongly predicted bee abundance for 71% of 665 bee populations (species × ecosystem combinations). Bee taxa that best tolerated heat and desiccation increased the most over time. Models forecasted declines for 46% of species and predicted more homogeneous communities dominated by drought-tolerant taxa, even while total bee abundance may remain unchanged. Such community reordering could reduce pollination services, because diverse bee assemblages typically maximize pollination for plant communities3. Larger-bodied bees also dominated under intermediate to high aridity, identifying body size as a valuable trait for understanding how climate-driven shifts in bee communities influence pollination4. We provide evidence that climate change directly threatens bee diversity, indicating that bee conservation efforts should account for the stress of aridity on bee physiology.

Providing rigor in bee colony strength auditing methods.

The primary method used to audit honey bee (Apis mellifera Linnaeus, 1758 [Hymenoptera: Apidae]) colony strength for almond pollination services, Nasr et al.'s (1990) frame-top cluster count method, is a subjective visual audit that relies on an auditor's spot assessment and may lack rigor and repeatability. We created novel, open-source software for the analysis of frame-top cluster count photographic assessments to improve methodological rigor and repeatability. We evaluated 2 existing visual audit methods, created 3 novel audit method variations, and determined between-method conversion factors using linear modeling. The software has potential applications in apiological research, apiarist and orchardist colony auditing, as well as training future generations of apiarists in auditing techniques. The software enhances the rigor and repeatability of Nasr et al.'s (1990) frame-top cluster count population assessment. In this article, we introduce the novel open-source software and between-method regression equations and review the tested visual assessment methods and their application.

A new orchid species expands Darwin's predicted pollination guild in Madagascar.

The world-renowned pollination system of the long-spurred orchid Angraecum sesquipedale Thouars and the long-tongued hawkmoth Xanthopan praedicta (Rothschild & Jordan, 1903), from Madagascar, is the best-known example of the predictive power of evolutionary ecology1,2, yet its actual degree of specialisation remains poorly described due to the incompleteness of the pollination record of X. praedicta. Here, we describe another species from Madagascar, an angraecoid orchid distantly related to the genus Angraecum Bory, that has evolved these extreme adaptations to a single pollinator after a pollinator shift. It bears the longest spur of any flowering plant, relative to flower diameter, reaching 33 cm. The discovery of a species with such an exceptionally long spur is a rare event, the most recent dating to 19653. This novelty is described here as Solenangis impraedicta (Figure 1A-F) and discussed in a phylogenetic framework. Its conservation status is assessed as Endangered.

Pollen chemical and mechanical defences restrict host-plant use by bees.

Plants produce an array of chemical and mechanical defences that provide protection against many herbivores and pathogens. Putatively defensive compounds and structures can even occur in floral rewards: for example, the pollen of some plant taxa contains toxic compounds or possesses conspicuous spines. Yet little is known about whether pollen defences restrict host-plant use by bees. In other words, do bees, like other insect herbivores, tolerate the defences of their specific host plants while being harmed by non-host defences? To answer this question, we compared the effects of a chemical defence from Lupinus (Fabaceae) pollen and a putative mechanical defence (pollen spines) from Asteraceae pollen on larval survival of nine bee species in the tribe Osmiini (Megachilidae) varying in their pollen-host use. We found that both types of pollen defences reduce larval survival rate in some bee species. These detrimental effects were, however, mediated by host-plant associations, with bees being more tolerant of the pollen defences of their hosts, relative to the defences of plant taxa exploited by other species. This pattern strongly suggests that bees are adapted to the pollen defences of their hosts, and that host-plant use by bees is constrained by their ability to tolerate such defences.

Multitrophic Higher-Order Interactions Modulate Species Persistence.

AbstractEcologists increasingly recognize that interactions between two species can be affected by the density of a third species. How these higher-order interactions (HOIs) affect species persistence remains poorly understood. To explore the effect of HOIs stemming from multiple trophic layers on a plant community composition, we experimentally built a mesocosm with three plants and three pollinator species arranged in a fully nested and modified network structure. We estimated pairwise interactions among plants and between plants and pollinators, as well as HOIs initiated by a plant or a pollinator affecting plant species pairs. Using a structuralist approach, we evaluated the consequences of the statistically supported HOIs on the persistence probability of each of the three competing plant species and their combinations. HOIs substantially redistribute the strength and sign of pairwise interactions between plant species, promoting the opportunities for multispecies communities to persist compared with a non-HOI scenario. However, the physical elimination of a plant-pollinator link in the modified network structure promotes changes in per capita pairwise interactions and HOIs, resulting in a single-species community. Our study provides empirical evidence of the joint importance of HOIs and network structure in determining species persistence within diverse communities.

Warmer autumns and winters could reduce honey bee overwintering survival with potential risks for pollination services.

Honey bees and other pollinators are critical for food production and nutritional security but face multiple survival challenges. The effect of climate change on honey bee colony losses is only recently being explored. While correlations between higher winter temperatures and greater colony losses have been noted, the impacts of warmer autumn and winter temperatures on colony population dynamics and age structure as an underlying cause of reduced colony survival have not been examined. Focusing on the Pacific Northwest US, our objectives were to (a) quantify the effect of warmer autumns and winters on honey bee foraging activity, the age structure of the overwintering cluster, and spring colony losses, and (b) evaluate indoor cold storage as a management strategy to mitigate the negative impacts of climate change. We perform simulations using the VARROAPOP population dynamics model driven by future climate projections to address these objectives. Results indicate that expanding geographic areas will have warmer autumns and winters extending honey bee flight times. Our simulations support the hypothesis that late-season flight alters the overwintering colony age structure, skews the population towards older bees, and leads to greater risks of colony failure in the spring. Management intervention by moving colonies to cold storage facilities for overwintering has the potential to reduce honey bee colony losses. However, critical gaps remain in how to optimize winter management strategies to improve the survival of overwintering colonies in different locations and conditions. It is imperative that we bridge the gaps to sustain honey bees and the beekeeping industry and ensure food and nutritional security.

Flower opening dynamics, pollen-ovule ratio, stigma receptivity and stigmatic pollen germination (in-vivo) in Chaenomeles speciosa (Sweet) Nakai.

Although Chaenomeles is widely used in horticulture, traditional Chinese medicine and landscape greening, insufficient research has hindered its breeding and seed selection. This study investigated the floral phenology, floral organ characteristics, palynology, and breeding systems of Chaenomeles speciosa (Sweet) Nakai. The floral characteristics of C. speciosa were observed both visually and stereoscopically. The microstructures of the flower organs were observed using scanning electron microscopy. Pollen stainability was determined using triphenyl tetrazolium chloride staining. Stigma receptivity was determined using the benzidine-H2O2 method and the post-artificial pollination pollen germination method. The breeding system was assessed based on the outcrossing index and pollen-ovule ratio. The flowers of C. speciosa were bisexual with a flowering period from March to April. The flowering periods of single flowers ranged from 8 to 19 d, and those of single plants lasted 18-20 d. The anthers were cylindrical, with the base attached to the filament, and were split longitudinally to release pollen. The flower had five styles, with a connate base. The ovaries had five carpels and five compartments. The inverted ovules were arranged in two rows on the placental axis. The stigma of C. speciosa was dry and had many papillary protrusions. In the early flowering stage (1-2 d of flowering), the pollen exhibited high stainability (up to 84.24%), but all stainability was lost at 7 d of flowering. Storage at - 20 °C effectively delayed pollen inactivation. The stigma receptivity of C. speciosa lasted for approximately 7 days, and the breeding system was classified as outcrossing with partial self-compatibility.

Distribution of exchangeable Ca2+ during the process of Larix decidua Mill. pollination and germination.

The involvement of Ca2+ ions in angiosperms sexual processes is well established, while in gymnosperms, such knowledge remains limited and is still a topic of discussion. In this study, we focused on Larix decidua, using Alizarin-red S staining and the pyroantimonate method to examine the tissue and subcellular distribution of free and loosely bound Ca2+ ions at different stages of the male gametophyte's development and its interaction with the ovule. Our findings show that in larch, both the germination of pollen grains and the growth of pollen tubes occur in an environment rich in Ca2+. These ions play a crucial role in the adhesion of the pollen grain to the stigmatic tip and its subsequent movement to the micropylar canal. There is a significant presence of free and loosely bound Ca2+ ions in both the fluid of the micropylar canal and the extracellular matrix of the nucellus. As the pollen tube extends through the nucellus, we observed a notable accumulation of Ca2+ ions just above the entry to the mature archegonium, a region likely crucial for the male gametophyte's directional growth. Meanwhile, the localized presence of free and loosely bound Ca2+ ions within the egg cell cytoplasm may inhibit the pollen tubes growth and rupture, playing an important role in fertilization.

Hybrid Mimulus flowers attract a new pollinator.

Hybridization is common in flowering plants and is believed to be an important force driving adaptation and speciation. The flowers of hybrids often exhibit new trait combinations, which, theoretically, could attract new species of pollinators. In this study, we found that the hybrids between a hummingbird-pollinated species Mimulus cardinalis and a self-pollinated species Mimulus parishii attract bumblebees (Bombus impatiens), a pollinator not attracted to either of the progenitor species. This novel attraction is explained by new combinations of floral traits in hybrids, including, most importantly, petal color, in addition to nectar concentration and corolla size. To understand how petal color variation is perceived by bumblebees, we performed reflectance spectroscopy and multispectral imaging to model the flower appearance in bee vision. This analysis showed that color variation would impact the ease of detection. We also found that YUP, the genetic locus responsible for a large portion of floral color variation and previously shown to be important in bee interactions with other Mimulus species, also played an important role in this novel attraction. These results together suggest that the attraction of new pollinators to hybrid plants could be an underexplored avenue for pollinator shift and speciation.

Bee-pollination promotes rapid divergent evolution in plants growing in different soils.

Divergent evolution leads to variation among populations and thus promotes diversification. In plants, adaptation to different soils, pollinator guilds, and herbivores is thought to be a key ecological driver of adaptive divergence, but few studies have investigated this process experimentally. Here we use experimental evolution with fast cycling Brassica rapa plants to study the impact of soil, pollination, herbivory, and their interactions on divergent evolution in various traits during eight generations of selection. We found significant evolutionary changes in plant phenotypes caused by all three factors and their interactions. In the richer soil type, plants showed higher evolutionary rates, especially with bumblebee-pollination, which led to the evolution of increased attractiveness of plants to bumblebees. Plants that had experienced aphid-herbivory showed lower attractiveness. We found the strongest evolutionary divergence when plants evolved in different soils with bee-pollination rather than hand-pollination, irrespective of herbivory. This "soil-pollinator effect" impacted divergence in diverse suites of traits, for example leaf size, flowering time, flower petal length, some floral volatiles and leaf glucosinolates. We conclude that the interaction between soil and biotic pollination may be an important cause for divergent evolution of plants growing on different soil types, even without a shift in pollinator guilds.

Cross-pollination in seed-blended refuge and selection for Vip3A resistance in a lepidopteran pest as detected by genomic monitoring.

The evolution of pest resistance to management tools reduces productivity and results in economic losses in agricultural systems. To slow its emergence and spread, monitoring and prevention practices are implemented in resistance management programs. Recent work suggests that genomic approaches can identify signs of emerging resistance to aid in resistance management. Here, we empirically examined the sensitivity of genomic monitoring for resistance management in transgenic Bt crops, a globally important agricultural innovation. Whole genome resequencing of wild North American Helicoverpa zea collected from non-expressing refuge and plants expressing Cry1Ab confirmed that resistance-associated signatures of selection were detectable after a single generation of exposure. Upon demonstrating its sensitivity, we applied genomic monitoring to wild H. zea that survived Vip3A exposure resulting from cross-pollination of refuge plants in seed-blended plots. Refuge seed interplanted with transgenic seed exposed H. zea to sublethal doses of Vip3A protein in corn ears and was associated with allele frequency divergence across the genome. Some of the greatest allele frequency divergence occurred in genomic regions adjacent to a previously described candidate gene for Vip3A resistance. Our work highlights the power of genomic monitoring to sensitively detect heritable changes associated with field exposure to Bt toxins and suggests that seed-blended refuge will likely hasten the evolution of resistance to Vip3A in lepidopteran pests.

Investigating the longstanding mystery of stigma length differences between morphs of distylous plants.

Longer stigmas in short-styled morphs of distylous plants have been considered an adaptive characteristic that increases intermorph pollen deposition. The greater pollen deposition in short-styled stigmas may be a by-product of their longer length, making deposition effectiveness comparison between morphs unfeasible. Thus, investigating which morph has the relatively most efficient stigma (i.e., pollen deposition per unit length) can boost our understanding of the adaptive significance of longer stigmas. Here, we compared pollen deposition between morphs relative to stigma length and assessed whether short-styled stigmas are more, less, or equally effective in receiving pollen grains per unit length. We reviewed the literature to characterize the extent of sigma length differences between morphs using the two most speciose distylous genera as model systems: Palicourea and Psychotria (Rubiaceae). Then, we conducted a between-morph comparison of raw and relative pollen depositions in a Palicourea rigida population. We confirmed that short-styled stigmas were longer than long-styled stigmas in both Palicourea (194.31% longer) and Psychotria (52.59% longer) flowers. Furthermore, in the focal Pal. rigida, although short-styled stigmas were 268.04% longer and received 97.04% more pollen grains than long-styled stigmas, the relative pollen deposition had a reverse pattern, with short-styled stigmas being two-times less efficient in receiving pollen. Our results indicate that the longer length of the short-styled stigmas may have a compensatory effect, increasing the chances of pollen grains reaching the stigma and probably maintaining disassortative intermorph pollination in distylous plants.