Morphological variation pattern of Aquilegia ecalcarata and its relatives

Aquilegia ecalcarata Maxim. is the only spurless species within the genus Aquilegia and comprises a monophyletic clade with A. yabeana Kitag., A. kansuensis Brühl, and A. rockii Munz. Our previous study on the genetic diversity of those four species revealed that the populations of A. ecalcarata can be divided into two groups, indicating possible genetic difference within A. ecalcarata. However, it is not clear whether the genetic difference is related to the morphological variation among species and groups of A. ecalcarata populations. To answer that question, the morphological variation patterns based on 22 floral and 19 vegetative traits from 42 populations, covering the entire distribution of A. ecalcarata and its relatives, were analyzed in the present study. The result showed that: (i) the differences among the four species were reflected in the floral rather than the vegetative traits; (ii) populations of A. yabeana and A. rockii fell into one cluster each, and each of the six clusters occupied its own distribution range; (iii) one of two A. ecalcarata clusters fell into a subgroup and shared common floral traits with A. rockii; (iv) the individuals of A. ecalcarata form. ecalcarata and form. semicalcarata were often mixed in the same population; and (v) the populations of A. kansuensis were split into two clusters, which differed obviously in floral traits. These results will provide an important morphological basis for the redefinition of species and lay a foundation for the further exploration of the “spurless” A. ecalcarata.     

The diversity of elaborate petals in Isopyreae (Ranunculaceae): a special focus on nectary structure

Elaborate petals are highly diverse in morphology, structure, and epidermal differentiation and play a key role in attracting pollinators. There have been few studies on the elaborate structure of petals in the tribe Isopyreae (Ranunculaceae). Seven genera in Isopyreae (Aquilegia, Semiaquilegia, Urophysa, Isopyrum, Paraquilegia, Dichocarpum, and Leptopyrum) have petals that vary in morphology, and two genera (Enemion and Thalictrum) have no petals. The petals of nine species belonged to 7 genera in the tribe were studied to reveal their nectary structure, epidermal micromorphology and ancestral traits. The petal nectaries of Isopyreae examined in this study were located at the tip of spurs (Aquilegia yabeana and A. rockii), or the bottom of shallow sacs (Semiaquilegia adoxoides, Urophysa henryi, Isopyrum manshuricum, and Paraquilegia microphylla), a cup-shaped structure (Dichocarpum fargesii) and a bilabiate structure (Leptopyrum fumarioides). The petal nectary of eight species in Isopyreae (except A. ecalcarata) was composed of secretory epidermis, nectary parenchyma, and vascular tissues, and some sieve tubes reached the secretory parenchyma cells. Among the eight species with nectaries examined in the present study, A. yabeana had the most developed nectaries, with 10–15 layers of secretory parenchyma cells. The epidermal cells of mature petals of the nine species were divided into 11 types. Among these 11 types, there were two types of secretory cells and two types of trichomes. Aquilegia yabeana and A. rockii had the highest number of cell types (eight types), and I. manshuricum and L. fumarioides had the lowest number of cell types (three types). Aquilegia ecalcarata had no secretory cells, and the papillose conical polygonal secretory cells of D. fargesii were different from those of the other seven species with nectaries. Trichomes were found only in Aquilegia, Semiaquilegia, Urophysa, and Paraquilegia. The ancestral mode of nectar presentation in Isopyreae was petals with hidden nectar (70.58%). The different modes of nectar presentation in petals may reflect adaptations to different pollinators in Isopyreae.

     

Early stages of speciation with gene flow in the Amazilia Hummingbird (Amazilis amazilia) subspecies complex of Western South America

Disentangling the factors underlying the diversification of geographically variable species with a wide geographical range is essential to understanding the initial stages and drivers of the speciation process. The Amazilia Hummingbird, Amazilis amazilia, is found along the Pacific coast from northern Ecuador down to the Nazca Valley of Peru, and is currently classified as six phenotypically differentiated subspecies. We aimed to resolve the evolutionary relationships of the six subspecies, to assess the geographical pattern and extent of evolutionary divergence, and to test for introgression using both a mtDNA marker and a genome‐by‐sequencing dataset from 86 individuals from across the species range. The consensus phylogenetic tree separated the six subspecies into three distinct clades, corresponding with the Ecuador lowlands (A. amazilia dumerilii), the Ecuador highlands (A. amazilia alticola and A. amazilia azuay), and the Peruvian coast (A. amazilia leucophoea, A. amazilia amazilia, and A. amazilia caeruleigularis). However, an unresolved mtDNA network suggests that the diversification of the subspecies was recent and rapid. We found evidence of gene flow among the subspecies A. amazilia dumerilii, A. amazilia alticola, and A. amazilia leucophoea, with strong genetic isolation of the subspecies A. amazilia azuay in the isolated Yunguilla Valley of Ecuador. Finally, environmental data from each subspecies’ capture locations were concordant with the three distinct clades. Overall, our results suggest that both expansions into new habitats and geographic isolation shaped the present‐day phylogeny and range of the A. amazilia subspecies, and that A. amazilia azuay may be genetically divergent enough to be considered a separate species.     

Floral traits and isolation of three sympatric Aquilegia species in the Qinling Mountains, China

Floral isolation has been considered to be an important reproductive mechanism governing the species diversification in many genera. In a classic example Aquilegia, sympatric species from North America with diverse floral traits are generally associated with specialized pollinators that prohibit interspecific hybridization. It remains unclear whether species diversification in the genus from Eurasia is also maintained by floral isolation. We investigated floral phenology, floral characteristics and pollinators in three sympatric Aquilegia species (A. ecalcarata, A. incurvata and A. yabeana) in the Qinling Mountains, Shanxi Province, China from 2001 to 2005. The spurless A. ecalcarata flowers earlier than the other two species with nectar spurs but their floral phenology overlaps. Major pollinators of A. ecalcarata are syrphid flies while bumblebees are major for A. incurvata and A. yabeana. Therefore our observations confirm that mechanical isolation through differential pollinators could contribute reproductive isolation between spurless and spurred species, as demonstrated by studies from North America. Whether floral isolation plays a major role in the reproductive isolation between two spurred species (A. incurvata and A. yabeana), however, remains to be seen. Further studies are required to quantify the potential role of geographical isolation because they occupy different habitats.     

Georeferenced phylogenetic analysis of a global collection of wild and cultivated Citrullus species

The geographical origin of watermelon (Citrullus lanatus) remains debated. While a first hypothesis suggests the center of origin to be West Africa, where the endemic sister species C. mucosospermus thrives, a second hypothesis suggests northeastern Africa where the white‐fleshed Sudanese Kordophan melon is cultivated. In this study, we infer biogeographical and haplotype genealogy for C. lanatus, C. mucosospermus, C. amarus, and C. colocynthis using noncoding cpDNA sequences (trnT‐trnL and ndhF‐rpl32 regions) from a global collection of 135 accessions. In total, we identified 38 haplotypes in C. lanatus, C. mucosospermus, C. amarus, and C. colocynthis; of these, 21 were found in Africa and 17 appear endemic to the continent. The least diverse species was C. mucosospermus (5 haplotypes) and the most diverse was C. colocynthis (16 haplotypes). Some haplotypes of C. mucosospermus were nearly exclusive to West Africa, and C. lanatus and C. mucosospermus shared haplotypes that were distinct from those of both C. amarus and C. colocynthis. The results support previous findings that revealed C. mucosospermus to be the closest relative to C. lanatus (including subsp. cordophanus). West Africa, as a center of endemism of C. mucosospermus, is an area of interest in the search of the origin of C. lanatus. This calls for further historical and phylogeographical investigations and wider collection of samples in West and northeastern Africa.     

Floral isolation between Aquilegia formosa and Aquilegia pubescens

The acquisition of floral nectar spurs is correlated with increased species diversity across multiple clades. We tested whether variation in nectar spurs influences reproductive isolation and, thus, can potentially promote species diversity using two species of Aquilegia, Aquilegia formosa and Aquilegia pubescens, which form narrow hybrid zones. Floral visitors strongly discriminated between the two species both in natural populations and at mixed-species arrays of individual flowers. Bees and hummingbirds visited flowers of A. formosa at a much greater rate than flowers of A. pubescens. Hawkmoths, however, nearly exclusively visited flowers of A. pubescens. We found that altering the orientation of A. pubescens flowers from upright to pendent, like the flowers of A. formosa, reduced hawkmoth visitation by an order of magnitude. In contrast, shortening the length of the nectar spurs of A. pubescens flowers to a length similar to A. formosa flowers did not affect hawkmoth visitation. However, pollen removal was significantly reduced in flowers with shortened nectar spurs. These data indicate that floral traits promote floral isolation between these species and that specific floral traits affect floral isolation via ethological isolation while others affect floral isolation via mechanical isolation.     

Brassica rapa Domestication: Untangling Wild and Feral Forms and Convergence of Crop Morphotypes

The study of domestication contributes to our knowledge of evolution and crop genetic resources. Human selection has shaped wild Brassica rapa into diverse turnip, leafy, and oilseed crops. Despite its worldwide economic importance and potential as a model for understanding diversification under domestication, insights into the number of domestication events and initial crop(s) domesticated in B. rapa have been limited due to a lack of clarity about the wild or feral status of conspecific noncrop relatives. To address this gap and reconstruct the domestication history of B. rapa, we analyzed 68,468 genotyping-by-sequencing-derived single nucleotide polymorphisms for 416 samples in the largest diversity panel of domesticated and weedy B. rapa to date. To further understand the center of origin, we modeled the potential range of wild B. rapa during the mid-Holocene. Our analyses of genetic diversity across B. rapa morphotypes suggest that noncrop samples from the Caucasus, Siberia, and Italy may be truly wild, whereas those occurring in the Americas and much of Europe are feral. Clustering, tree-based analyses, and parameterized demographic inference further indicate that turnips were likely the first crop type domesticated, from which leafy types in East Asia and Europe were selected from distinct lineages. These findings clarify the domestication history and nature of wild crop genetic resources for B. rapa, which provides the first step toward investigating cases of possible parallel selection, the domestication and feralization syndrome, and novel germplasm for Brassica crop improvement.     

Phylogeography of sugar kelp: Northern ice‐age refugia in the Gulf of Alaska

Many Northeast (NE) Pacific fishes and invertebrates survived Pleistocene glaciations in northern refugia, but the extent that kelps survived in northern areas is uncertain. Here, we test the hypothesis that populations of sugar kelp (Saccharina latissima) persisted in the Gulf of Alaska during ice‐age maxima when the western margin of the Cordilleran ice sheet covered coastal areas around the NE Pacific Ocean. We estimated genetic diversities within and phylogeographical relationships among 14 populations along 2,800 km in the NE Pacific and Bering Sea with partial sequences of mitochondrial DNA 5′‐cytochrome oxidase subunit I (COI, bp = 624, n = 543), chloroplast DNA ribulose‐1,5‐bisphosphate carboxylase large subunit‐3′ (rbcL, bp = 735, n = 514), and 11 microsatellite loci. Concatenated sequences of rbcL and COI showed moderate levels of within‐population genetic diversity (mean h = 0.200) but substantial differences among populations (Φ_ST = 0.834, p < .0001). Microsatellites showed moderate levels of heterozygosity within populations (mean H _E = 0.391). Kelps in the same organellar lineage tended to cluster together, regardless of geographic origins, as indicated in a principal coordinate analysis (PCoA) of microsatellite genotypes. The PCoA also showed evidence of nuclear hybridizations between co‐occurring organellar lineages. Individual admixture plots with population clusters of K = 2, 6, and 9 showed increasing complexity with considerable historical admixture between some clusters. A time‐calibrated phylogeny placed divergences between rbcL‐COI lineages at 1.4 million years at most. The time frames of mutation in the rbcL‐COI lineages and microsatellite population clusters differed among locations. The existence of ancient lineages in the Gulf of Alaska, moderate levels of genetic diversity, and the absence of departures from neutrality are consistent with northern refugia during multiple Croll‐Milankovitch climate cycles in the Pleistocene Epoch.     

Plastid genomes reveal evolutionary shifts in elevational range and flowering time of Osmanthus (Oleaceae)

Species of Osmanthus are economically important ornamental trees, yet information regarding their plastid genomes (plastomes) have rarely been reported, thus hindering taxonomic and evolutionary studies of this small but enigmatic genus. Here, we performed comparative genomics and evolutionary analyses on plastomes of 16 of the 28 currently accepted species, with 11 plastomes newly sequenced. Phylogenetic studies identified four main lineages within the genus that are here designated the: “Caucasian Osmanthus” (corresponding to O. decorus), “Siphosmanthus” (corresponding to O. sect. Siphosmanthus), “O. serrulatus + O. yunnanensis,” and “Core Osmanthus: (corresponding to O. sect. Osmanthus + O. sect. Linocieroides). Molecular clock analysis suggested that Osmanthus split from its sister clade c. 15.83 Ma. The estimated crown ages of the lineages were the following: genus Osmanthus at 12.66 Ma; “Siphosmanthus” clade at 5.85 Ma; “O. serrulatus + O. yunnanensis” at 4.89 Ma; and “Core Osmanthus: clade at 6.2 Ma. Ancestral state reconstructions and trait mapping showed that ancestors of Osmanthus were spring flowering and originated at lower elevations. Phylogenetic principal component analysis clearly distinguished spring‐flowering species from autumn‐flowering species, suggesting that flowering time differentiation is related to the difference in ecological niches. Nucleotide substitution rates of 80 common genes showed slow evolutionary pace and low nucleotide variations, all genes being subjected to purifying selection.     

Evolutionary Drivers of Diversification and Distribution of a Southern Temperate Stream Fish Assemblage: Testing the Role of Historical Isolation and Spatial Range Expansion

This study used phylogenetic analyses of mitochondrial cytochrome b sequences to investigate genetic diversity within three broadly co-distributed freshwater fish genera (Galaxias, Pseudobarbus and Sandelia) to shed some light on the processes that promoted lineage diversification and shaped geographical distribution patterns. A total of 205 sequences of Galaxias, 177 sequences of Pseudobarbus and 98 sequences of Sandelia from 146 localities across nine river systems in the south-western Cape Floristic Region (South Africa) were used. The data were analysed using phylogenetic and haplotype network methods and divergence times for the clades retrieved were estimated using *BEAST. Nine extremely divergent (3.5–25.3%) lineages were found within Galaxias. Similarly, deep phylogeographic divergence was evident within Pseudobarbus, with four markedly distinct (3.8–10.0%) phylogroups identified. Sandelia had two deeply divergent (5.5–5.9%) lineages, but seven minor lineages with strong geographical congruence were also identified. The Miocene-Pliocene major sea-level transgression and the resultant isolation of populations in upland refugia appear to have driven widespread allopatric divergence within the three genera. Subsequent coalescence of rivers during the Pleistocene major sea-level regression as well as intermittent drainage connections during wet periods are proposed to have facilitated range expansion of lineages that currently occur across isolated river systems. The high degree of genetic differentiation recovered from the present and previous studies suggest that freshwater fish diversity within the south-western CFR may be vastly underestimated, and taxonomic revisions are required.     

Characterization of Salix nigra floral insect community and activity of three native Andrena bees

Salix nigra (black willow) is a widespread tree that hosts many species of polylectic hymenopterans and oligolectic bees of the genus Andrena. The early flowering of S. nigra makes it an important nutritive resource for arthropods emerging from hibernation. However, since S. nigra is dioecious, not all insect visits will lead to successful pollination. Using both visual observation and pan‐trapping, we characterized the community of arthropods that visited S. nigra flowers and assessed differences among male and female trees as well as the chemical and visual drivers that influenced community composition across 3 years. We found that male trees consistently supported higher diversity of insects than female trees and only three insect species, all Andrena spp., consistently visited both sexes. Additionally, Andrena nigrae, which was the only insect that occurred more on female than male flowers, correlated strongly to volatile cues. This suggests that cross‐pollinators cue into specific aspects of floral scent, but diversity of floral visitors is driven strongly by visual cues of yellow male pollen. Through time, the floral activity of two Andrena species remained stable, but A. nigrae visited less in 2017 when flowers bloomed earlier than other years. When native bee emergence does not synchronize with bloom, activity appears to be diminished which could threaten species that subsist on a single host. Despite the community diversity of S. nigra flowers, its productivity depends on a small fraction of species that are not threatened by competition, but rather rapidly changing conditions that lead to host‐insect asynchrony.     

Cellulase immobilized magnetic nanoparticles for green energy production from Allamanda schottii L: Sustainability research in waste recycling

This study presents ethanol''s fabrication by fermenting the golden trumpet flower (Allamanda schottii L) with the yeast strain Saccharomyces cerevisiae. The changes in different parameters during fermentation were studied and optimized while producing the ethanol and the end product was subjected to emission test study by blending petrol and ethanol. The Allamanda floral substrate contains 65% polysaccharides. The strain S. cerevisiae was obtained in the form of baker’s yeast from a domestic shop. For 100 ml of slurry, the highest bioethanol yield recorded was about 18.75 ml via optimization of different culture conditions, including a 1:8 ratio for slurry preparation, maintained under 35 ⁰C, 5.5 pH, 72 h. old inoculum with a quantity of 3.75 g 100 ml⁻¹, fermented for120 h. The highest yield of bioethanol was acquired under the addition of urea. This technique & design is capable of industrial-scale fabrication of bioethanol by using A. schottii floral substrates. This research was conducted to fabricate ethanol by fermentation (A. schottii L) floral substrate with S. cerevisiae. The optimum physiochemical parameters required to obtain the highest yield of bioethanol from A. schottii flower by fermentation was studied. The immobilization strategy with a cheap agricultural substrate and magnetic nanoparticles were also studied. The engine performance and emission studies were done with different blends of petrol and bio-ethanol.     

Evolution of RXLR-Class Effectors in the Oomycete Plant Pathogen Phytophthora ramorum

Phytophthora plant pathogens contain many hundreds of effectors potentially involved in infection of host plants. Comparative genomic analyses have shown that these effectors evolve rapidly and have been subject to recent expansions. We examined the recent sequence evolution of RXLR-class effector gene families in the sudden oak death pathogen, P. ramorum. We found that P. ramorum RXLR effectors have taken multiple evolutionary paths, including loss or gain of repeated domains, recombination or gene conversion among paralogs, and selection on point mutations. Sequencing of homologs from two subfamilies in P. ramorum’s closest known relatives revealed repeated gene duplication and divergence since speciation with P. lateralis. One family showed strong signatures of recombination while the other family has evolved primarily by point mutation. Comparison of a small number of the hundreds of RXLR-class effectors across three clonal lineages of P. ramorum shows striking divergence in alleles among lineages, suggesting the potential for functional differences between lineages. Our results suggest future avenues for examination of rapidly evolving effectors in P. ramorum, including investigation of the functional and coevolutionary significance of the patterns of sequence evolution that we observed.     

Genomic divergence in sympatry indicates strong reproductive barriers and cryptic species within Eucalyptus salubris

Genetic studies are increasingly detecting cryptic taxa that likely represent a significant component of global biodiversity. However, cryptic taxa are often criticized because they are typically detected serendipitously and may not receive the follow‐up study required to verify their geographic or evolutionary limits. Here, we follow‐up a study of Eucalyptus salubris that unexpectedly detected two divergent lineages but was not sampled sufficiently to make clear interpretations. We undertook comprehensive sampling for an independent genomic analysis (3,605 SNPs) to investigate whether the two purported lineages remain discrete genetic entities or if they intergrade throughout the species’ range. We also assessed morphological and ecological traits, and sequenced chloroplast DNA. SNP results showed strong genome‐wide divergence (F _ST = 0.252) between two discrete lineages: one dominated the north and one the southern regions of the species’ range. Within lineages, gene flow was high, with low differentiation (mean F _ST = 0.056) spanning hundreds of kilometers. In the central region, the lineages were interspersed but maintained their genomic distinctiveness: an indirect demonstration of reproductive isolation. Populations of the southern lineage exhibited significantly lower specific leaf area and occurred on soils with lower phosphorus relative to the northern lineage. Finally, two major chloroplast haplotypes were associated with each lineage but were shared between lineages in the central distribution. Together, these results suggest that these lineages have non‐contemporary origins and that ecotypic adaptive processes strengthened their divergence more recently. We conclude that these lineages warrant taxonomic recognition as separate species and provide fascinating insight into eucalypt speciation.     

Allopatric and Sympatric Drivers of Speciation in Alviniconcha Hydrothermal Vent Snails

Despite significant advances in our understanding of speciation in the marine environment, the mechanisms underlying evolutionary diversification in deep-sea habitats remain poorly investigated. Here, we used multigene molecular clocks and population genetic inferences to examine processes that led to the emergence of the six extant lineages of Alviniconcha snails, a key taxon inhabiting deep-sea hydrothermal vents in the Indo-Pacific Ocean. We show that both allopatric divergence through historical vicariance and ecological isolation due to niche segregation contributed to speciation in this genus. The split between the two major Alviniconcha clades (separating A. boucheti and A. marisindica from A. kojimai, A. hessleri, and A. strummeri) probably resulted from tectonic processes leading to geographic separation, whereas the splits between co-occurring species might have been influenced by ecological factors, such as the availability of specific chemosynthetic symbionts. Phylogenetic origin of the sixth species, Alviniconcha adamantis, remains uncertain, although its sister position to other extant Alviniconcha lineages indicates a possible ancestral relationship. This study lays a foundation for future genomic studies aimed at deciphering the roles of local adaptation, reproductive biology, and host–symbiont compatibility in speciation of these vent-restricted snails.     

The association of carbohydrate changes in the shoot tip of cauliflower with flowering

Changes in levels of sugars and starch in the shoot tip of cauliflower, Brassica oleracea L. var. botrytis D. C. cv. Main Crop were studied during periods of growth which were inductive or non-inductive to flowering. Flowering was induced by growing plants for 2 weeks under 16 hr of light at 5°. During this period of floral induction there was a significant increase in sugar and starch content compared to that in vegetative plants grown at 20 to 26°. Sugar and starch content did not increase and flowering was prevented when light and CO₂ were excluded during growth at 5°. A 3-day dark period at 20° or a high temperature treatment at 33° with light following growth at 5° reduced the carbohydrate level and prevented flowering.     

Genetic diversity and structure of Rhododendron meddianum,a plant species with extremely small populations

Rhododendron meddianum is a critically endangered species with important ornamental value and is also a plant species with extremely small populations. In this study, we used double digest restriction-site-associated DNA sequencing (ddRAD) technology to assess the genetic diversity, genetic structure and demographic history of the three extant populations of R. meddianum. Analysis of SNPs indicated that R. meddianum populations have a high genetic diversity (π = 0.0772 ± 0.0024, H_E = 0.0742 ± 0.002). Both F_ST values (0.1582–0.2388) and AMOVA showed a moderate genetic differentiation among the R. meddianum populations. Meanwhile, STRUCTURE, PCoA and NJ trees indicated that the R. meddianum samples were clustered into three distinct genetic groups. Using the stairway plot, we found that R. meddianum underwent a population bottleneck about 70,000 years ago. Furthermore, demographic models of R. meddianum and its relative, Rhododendron cyanocarpum, revealed that these species diverged about 3.05 (2.21–5.03) million years ago. This divergence may have been caused by environmental changes that occurred after the late Pliocene, e.g., the Asian winter monsoon intensified, leading to a drier climate. Based on these findings, we recommend that R. meddianum be conserved through in situ, ex situ approaches and that its seeds be collected for germplasm.     

Phylogeography of the Japanese greater horseshoe bat Rhinolophus nippon (Mammalia: Chiroptera) in Northeast Asia: New insight into the monophyly of the Japanese populations

The Japanese greater horseshoe bat (Rhinolophus nippon) is distributed widely in East Asia. Within the species, R. nippon in Northeast Asia is regarded as the lineage that diverged most recently. However, the monophyly of the Japanese populations is unclear due to insufficient data about phylogenetic relationship of the western Japanese populations. To test the monophyly of the Japanese populations of R. nippon, we sampled R. nippon from western Japan and performed a phylogeographic analysis based on mitochondrial DNA cytochrome b and the D‐loop. The Northeast Asian lineage consisted of three main clades in eastern Japan (clade I), western Japan (clade II), and the continent as well as the Kumamoto population in westernmost Japan (clade III). The results of this study do not support the monophyly of the Japanese population. The findings suggest the “reverse colonization” of R. nippon from the Japanese Archipelago to the Eurasian continent, and provide important insight into the role of the island system in creation and supply of diversity to the continent.     

Floral phenology of an Andean bellflower and pollination by buff‐tailed sicklebill hummingbird

The Andean bellflowers comprise an explosive radiation correlated with shifts to specialized pollination. One diverse clade has evolved with extremely curved floral tubes and is predicted to be pollinated exclusively by one of two parapatric species of sicklebill hummingbirds (Eutoxeres). In this study, we focused on the floral biology of Centropogon granulosus, a bellflower thought to be specialized for pollination by Eutoxeres condamini, in a montane cloud forest site in southeastern Peru. Using camera traps and a pollination exclusion experiment, we documented E. condamini as the sole pollinator of C. granulosus. Visitation by E. condamini was necessary for fruit development. Flowering rates were unequivocally linear and conformed to the “steady‐state” phenological type. Over the course of >1800 h of monitoring, we recorded 12 E. condamini visits totaling 42 s, indicating traplining behavior. As predicted by its curved flowers, C. granulosus is exclusively pollinated by buff‐tailed sicklebill within our study area. We present evidence for the congruence of phenology and visitation as a driver of specialization in this highly diverse clade of Andean bellflowers.