Phylotranscriptomics of Swertiinae (Gentianaceae) reveals that key floral traits are not phylogenetically correlated

Establishing how lineages with similar traits are phylogenetically related remains critical for understanding the origin of biodiversity on Earth. Floral traits in plants are widely used to explore phylogenetic relationships and to delineate taxonomic groups. The subtribe Swertiinae (Gentianaceae) comprises more than 350 species with high floral diversity ranging from rotate to tubular corollas and possessing diverse nectaries. Here we performed phylogenetic analysis of 60 species from all 15 genera of the subtribe Swertiinae sensu Ho and Liu, representing the range of floral diversity, using data from the nuclear and plastid genomes. Extensive topological conflicts were present between the nuclear and plastome trees. Three of the 15 genera represented by multiple species are polyphyletic in both trees. Key floral traits including corolla type, absence or presence of lobe scales, nectary type, nectary position, and stigma type are randomly distributed in the nuclear and plastome trees without phylogenetic correlation. We also revealed the likely ancient hybrid origin of one large clade comprising 10 genera with diverse floral traits. These results highlight the complex evolutionary history of this subtribe. The phylogenies constructed here provide a basic framework for further exploring the ecological and genetic mechanisms underlying both species diversification and floral diversity.     

A phylogenetic analysis of subtribe Pleurothallidinae (Orchidaceae)

A cladistic analysis of subtribe, Pleurothallidinae (Orchidaceae) is based on 45 anatomical/ morphological characters. The ingroup members comprise 24 genera; the large genus Pleurothallis consists of two subgenera and ten species complexes. Three taxa representing subtribes Laeliinae and Arpophyllinae are designated as outgroup. Eight most parsimonious trees were discovered using computer assisted software (length = 230; CI = 0.27). The hypothesis that subtribe Pleurothallidinae has undergone a unilinear reduction in the number of pollinia is not supported by this study. Although the eight-pollinia state as represented by Octomeria apparently is plesiomorphic, the two-pollinia and four-pollinia states arose early in the phylogeny of the subtribe. Both two-and four-pollinia states subsequently reappeared as parallelisms. The six-pollinia state exhibited in Brachionidium is autapomorphic. This cladistic analysis suggests that Pleurothallis is not a natural genus and, perhaps may be divided into several discrete genera.     

A phylogenetic analysis of subtribe Pleurothallidinae (Orchidaceae)

A cladistic analysis of subtribe, Pleurothallidinae (Orchidaceae) is based on 45 anatomical/ morphological characters. The ingroup members comprise 24 genera; the large genus Pkurothallis consists of two subgenera and ten species complexes. Three taxa representing subtribes Laeliinae and ArpophyUinae are designated as outgroup. Eight most parsimonious trees were discovered using computer assisted software (length = 230; CI = 0.27). The hypothesis that subtribe Pleurothallidinae has undergone a unilinear reduction in the number of pollinia is not supported by this study. Although the eight-pollinia state as represented by Octomeria apparently is plesiomorphic, the two-pollinia and four-pollinia states arose early in the phytogeny of the subtribe. Both two-and four-pollinia states subsequently reappeared as parallelisms. The six-pollinia state exhibited in Brachionidium is autapomorphic. This cladistic analysis suggests that Pkurothallis is not a natural genus and, perhaps may be divided into several discrete genera.     

POLLINATION IN FLORAL SCENT MORPHS OF POLEMONIUM VISCOSUM: A MECHANISM FOR DISRUPTIVE SELECTION ON FLOWER SIZE

Plants of Polemonium viscosum have flowers that are either sweet or skunky in scent. The two morphs are preferentially pollinated by insects of strongly contrasting body size: bumblebee queens specialize on sweet flowers, flies on skunky ones. In this study 13 characters were examined in plant specimens from five populations to identify major components of intraspecific variation in flower and inflorescence morphology and test their correlation with floral scent. Factor analysis identified four major axes of morphological variation. The first explained 22% of the variance among specimens and correlated strongly with four flower size characters: sepal length, corolla tube length, corolla lobe width, and corolla lobe length. Floral scent morphs differed significantly in the multivariate representation of flower size defined by these characters. Sweet flowers had wider corolla lobes, longer corolla tubes, and longer sepals than skunky ones. Corolla lobe width accounted for the greatest amount of intermorph divergence. Divergence in flower size between morphs was maintained in mixed populations at four locations in alpine Colorado, with corollas of sweet flowers significantly broader or more flared than those of skunky flowers. Patterns of pollen receipt suggest that this difference is adaptive. In the sweet morph, pollination intensity and purity increased significantly with corolla flare. Conversely, in the skunky morph, corolla flare had little influence on pollination intensity and had a strong negative effect on purity. These findings suggest that selection for effective pollination should favor intraspecific divergence in flower size in Polemonium viscosum.     

Are flowers physiological sinks or faucets? Costs and correlates of water use by flowers of Polemonium viscosum

Water loss through inflorescences may place extreme demands on plant water status in arid environments. Here we examine how corolla size, a trait known to influence pollination success, affects the water cost of flowering in the alpine skypilot, Polemonium viscosum. In a potometry experiment, water uptake rates of inflorescences were monitored during bud expansion and anthesis. Corolla volume of fully expanded flowers predicted water uptake during bud expansion (R ²=0.61, P=0.0375) and corolla surface area predicted water uptake during anthesis (R ²=0.59, P=0.044). To probe mechanisms underlying the relationship between corolla size and water uptake, cell dimensions and densities were measured in several regions of fully expanded corollas. Corolla length was positively correlated with cell length in the middle of the corolla tube and cell diameter in the corolla lobe (Pearson's r from 0.26–0.33, n=86, P ≤ 0.05). Cell density was negatively correlated with cell dimensions in the upper corolla tube and lobe (Pearson's r from –0.39 to –0.42, P ≤ 0.0015). These findings suggest that more water may be required to maintain turgor in large corollas in part because their tissues have lower cell wall densities. The carbon cost of water use by flowers was assessed in krummholz and tundra habitats for P. viscosum flowering, respectively, during dry and wet portions of the growing season. For plants in full flower, average leaf water potentials were significantly more negative (P=0.0079) at mid-day in the krummholz (June) than in the tundra (July), but were similar before dawn (P=0.631). Photosynthetic rate at the time of flowering declined significantly with increasing corolla size in the krummholz (P=0.0376), but was unrelated to corolla size on the tundra (P>0.72). Plants losing water through large corollas may close leaf stomata to maintain turgor. If photosynthesis limits growth in this perennial species, then the water cost of producing large flowers should exacerbate the cost of reproduction under dry conditions. Such factors could select for flowers with smaller corollas in the krummholz, countering pollinator-mediated selection and helping maintain genetic variation in corolla size components of P. viscosum. Received: 5 May 1998 / Accepted: 2 October 1998     

The filament curtain: a structure important to systematics and pollination biology in the Acanthaceae

The filament curtain is a complex structure found in Acanthaceae inside the corolla. It usually divides the corolla into two compartments, enclosing the main nectar bulk behind two adjacent filament bands. Four independent floral characters are involved in the structure of the filament curtain: a fusion of the filaments, decurrent filaments along the corolla wall, a slanting border between the synstapetal and apostapetal corolla regions (i.e. stamen corolla tube and corolla tube s.s .), and geniculate lower, lateral corolla lobe traces at this border. The distribution of a filament curtain within Acanthaceae suggests an enlarged tribe Ruellieae s.l ., including Louteridieae and Trichanthereae, excluding subtribe Barleriinae, and raises a question mark as to the systematic position of the genera Calacanthus, Glossochilus, Lankesteria and Mhitfieldia . The evolutionary origin of the filament curtain is probably connected with its functions in pollination biology, which are proposed to be those of restricted nectar access, prevention of nectar evaporation, lever arm function facilitating dorsal pollen deposition, and stabilizing of posticous position of anthers and style. Four types of filament curtains are distinguished: phaulopsoid, corolla fold, reduced and strobilanthoid. Variation in the filament curtain structure appears to be related to different pollination syndromes of the flowers. Mapping of this variation on to an existing phylogeny shows an evolution from a phaulopsoid type diverging into the other three types, possibly a development responding to shifts of pollinators.     

Phylogeny of Barnadesioideae (Asteraceae) inferred from DNA sequence data and morphology

Subfamily Barnadesioideae (Asteraceae) consists of nine genera and 91 species endemic to South America. They include annual and perennial herbs, arching shrubs and trees up to 30 m tall. Presumed sister to all other Asteraceae, its intergeneric relationships are key to understanding the early evolution of the family. Results of the only molecular study on the subfamily conflict with relationships inferred from morphology. We investigate inter- and intrageneric relationships in Barnadesioideae with novel DNA sequence data and morphological characters using parsimony, likelihood and Bayesian inference. All results verify Barnadesioideae as monophyletic and sister to the rest of the family. A basal split within the subfamily is recognized, with Chuquiraga, Doniophyton and Duseniella in one clade, and Arnaldoa, Barnadesia, Dasyphyllum, Fulcaldea, Huarpea and possibly Schlechtendalia in another. The largest genus, Dasyphyllum, is revealed as biphyletic with the two clades separating along subgeneric and geographic lines. Schlechtendalia, suggested as the earliest diverging lineage of the subfamily by morphological studies and parsimony analyses, is found in a more derived position under model-based inference methods. Competing phylogenetic hypotheses, both previous and present, are evaluated using likelihood-based tests. Evolutionary trends within Barnadesioideae are inferred: hummingbird pollination has developed convergently at least three times. An early vicariance in the subfamily’s distribution is revealed. X = 9 is supported as the ancestral base chromosome number for both Barnadesioideae and the family as a whole.     

Morphological phylogenetics of theAsteraceae-Asteroideae,with notes on character evolution

The relationships within theAsteraceae-Asteroideae are scrutinized by means of cladistic analyses of morphological and chemical data. New data are presented, and novel interpretations of features previously used in cladistic analyses are made. Examples are insertion of anther filaments in the corolla, and morphology of disc floret styles, respectively. The results include a polyphyletic or paraphyleticInuleae s. l., and a paraphyleticHeliantheae s.l. in which theEupatorieae constantly form a subclade. Comparisons are made with previous cladistic analyses of morphological data, and those acquired from molecular data. Speculations about the evolution of certain organs are presented.     

INTERACTING ROLES OF GIBBERELLIN AND ETHYLENE IN COROLLA EXPANSION OF IPOMOEA NIL (CONVOLVULACEAE)

The relatively rapid and extensive corolla expansion in Ipomoea nil (L.) Roth cv. ‘Scarlett O'Hara’ appears to be initiated by a shift in the balance between a growth promoter (gibberellin, GA) and a growth inhibitor (ethylene) two days before anthesis. The effects of applied growth regulators in vitro, were measured as a change in area of segments from whole young corollas 16–17 mm long. Applied gibberellic acid (GA₃) strongly promoted growth, while a GA action inhibitor (ancymidol) reduced growth. Inhibitors of GA biosynthesis (AMO-1618, paclobutrazol, tetcyclasis, and chlorocholine chloride) had little effect, implying that the GAs were not being synthesized within the corolla segments, at least at this stage of corolla growth. Both the level of endogenous GAs and response of the segments to sucrose increased as the corolla size also increased from 15 to 20 mm in length. Applied 1-aminocyclopropane-l-carboxylic acid (ACC), an ethylene precursor, inhibited the response of corollas to applied GA₃. Applications of ethylene biosynthesis inhibitors (AVG and cobalt ions) promoted growth of the corolla segments. Rapid ethylene production by corollas 15–17 mm long (48 hr before flower opening) appeared to inhibit the growth that may have been induced by low levels of endogenous GAs. Older corollas (longer than 18 mm) had very low levels of ethylene production and much higher levels of GA-like substances; these changes apparently allow the corollas to begin to expand in vivo and to strongly respond to sucrose applications alone in vitro. Acid-induced growth does not seem to be an important component of corolla expansion; applied fusicoccin (a proton efflux promoter) and sodium orthovanadate (a proton efflux inhibitor) had no significant effect on growth of the 16–17 mm long corollas. Buffers (Good's and phosphate) over a wide pH range had no effect on corolla expansion. Taken together, our results indicate that the regulation of corolla expansion depends (at least in part) on compensatory shifts in the levels of two plant growth regulators [PGRs], ethylene and GA.     

Molecular phylogenetics of the subtribeGentianinae (Gentianaceae) inferred from the sequences of internal transcribed spacers (ITS) of nuclear ribosomal DNA

The internal transcribed spacer (ITS) regions of 18S–25S nuclear ribosomal DNA from representatives of 23 species of the subtribeGentianinae and one outgroup species (Centaurium capitatum) were analyzed by polymerase chain reaction amplification and direct DNA sequencing. Within the taxa analyzed, the length of the ITS1 region varied from 221 to 233 bp, ITS2 from 226 to 234 bp. Of the aligned sequences of 497 positions, 151 sites involved gaps or nucleotide ambiguity, 133 were invariable and 213 showed divergence. In pairwise comparisons among the taxa of the subtribeGentianinae and the outgroup, sequence divergence ranged from 1.3% to 34.1% in ITS1, from 0 to 28.1% in ITS2 and from 0.6% to 27.5% in combined ITS1 and ITS2. Phylogenetic trees generated from ITS sequences were highly resolutive and principally concordant with morphological classifications for the major phylogenetic divisions in the subtribe. An ancient divergence leading to two evolutionary lines was suggested in the subtribe by both DNA sequence and morphological data. One line encompasses the generaGentiana, Crawfurdia andTripterospermum, morphologically characterized by their glands on the base of ovary and their plicate corolla, while the other line involves all other members of the subcribe surveyed, characterized by their epipetalous glands and simple corolla without plicae.Megacodon, with glands on the base of ovary but without plicae on its corolla, was revealed to be more related to the latter group than to the former.Comastoma, Gentianella andGentianopsis were shown to be well-defined monophyletic genera.Pterygocalyx showed much closer affinity toGentianopsis than to any other genus. Some conflictions were detected in the genusSwertia.     

Congruence between comparative morphology and molecular phylogenies: Evolution of the male genital skeletal/muscular system in the subtribe Polyommatina (Lepidoptera,Lycaenidae)

The skeleton and musculature of male genitalia were studied in species of a model butterfly group (subtribe Polyommatina, Lycaenidae). In total, we analyzed 45 species of the tribe Polyommatini most of which were previously used in the molecular phylogenetic study (Talavera et al., 2013). The studied morphological characters were mapped on the molecular trees, which allowed us to reveal trends of morphological changes and to estimate the age of their origin. As a result, chronology of evolution of skeleton and musculature traits was established. It was shown that periods of slow morphological evolution alternated in the subtribe Polyommatina with those of a high rate of origin of new traits. For example, topography of the intravalvar muscles has not changed for 26 MY preserving their initial fan-shaped attachment. The evolution of intravalvar muscles started 10 MYA, proceeded slowly during the first 5 MY, and then accelerated during the last 5 MY resulting in the extensive splitting of the musculature in most monophyletic lineages. Mapping the morphological characters on the phylogeny demonstrated that the rates of skeleton and muscle evolution within the skeleton/musculature apparatus were different. In most cases the intravalvar musculature evolved much faster than the skeleton. The cladistic interpretation of states of morphological traits was found to be consistent with phylogenetic reconstructions based on analysis of multiple molecular markers. Moreover, morphological synapomorphies were found for the lineages Alpherakya + Glabroculus and Aricia + (Alpherakya + Glabroculus), which had low statistical support in molecular phylogenetic analysis. Additionally, in some cases molecular studies helped to reveal trends in the evolution of morphological traits. For example, the unpaired uncus and the compact juxta are not plesiomorphic for Cupidina as previously thought; instead, they were shown to have evolved secondarily within this subtribe.     

STUDIES ON THE FLORAL ANATOMY OF THE CUNONIACEAE

Twenty-two genera representing sixty-two species of Cunoniaceae and Davidsonia were examined with respect to floral anatomy. Sepals are vascularized by three traces with the lateral traces of adjacent sepals united. Pancheria is unique for the family with species in which the sepals are vascularized by a single, undivided bundle. Petals, when present, and stamens, are uniformly one-trace structures. A general tendency exists within the family for the principal floral bundles to unite in various ways, with fusions evident between calyx, corolla, and androecial vascular supplies. Carpel number ranges from two to five and the gynoecium is generally surrounded by a prominent disc. Gynoecia of Ceratopetalum and Pullea are “half-inferior.” The number of ovules per carpel locule ranges from one to numerous. Ventral carpel sutures range from open to completely sealed at the level of placentation. Carpels of the apocarpous genus Spiraeanthemum (incl. Acsmithia) are vascularized by a dorsal bundle and either three or four bundles constituting the ovular and wing vasculation in the ventral position, a condition unlike other members of the family. Ovules are supplied by the median ventral bundle. More advanced bicarpellate gynoecia within the family are predominately vascularized by a dorsal and two ventral bundles although a variable number of additional lateral wall traces may be present. A major trend exists toward fusion of the ventral bundles of adjacent carpels in the ovary of both bicarpellate and multicarpellate plants. At the base of the styles the fused ventral strands separate and extend along with the dorsal carpellary bundles into styles of adjacent carpels. In Pullea the ventral bundles terminate within the ovules. The united ventral carpellary bundles in Aphanopetalum, Gillbeea, and Aistopetalum lie in the plane of the septa separating adjacent carpels. Ovules are vascularized by traces originating from the vascular cylinder at the base of the gynoecium or by traces branching from the ventral bundles. Ovular traces in each carpel are united, or remain as discrete bundles, prior to entering the placenta. Tannin and druses are common throughout all floral parts. Although floral anatomy generally supports the position of Cunoniaceae near Saxifragaceae and Davidsoniaceae, the evolutionary relationship of the Cunoniaceae to the Dilleniaceae is uncertain.     

COMPARATIVE ANATOMY OF THE PETIOLE AND INFRAGENERIC RELATIONSHIPS IN JATROPHA (EUPHORBIACEAE)

Anatomical features of the petiole in several species of Jatropha L. (Euphorbiaceae) are presented as evidence in support of infrageneric relationships. A trilacunar 3-trace nodal pattern is typical for the genus. The vascular supply to the stipules is derived from the branching of the two peripheral leaf traces. The number of vascular bundles range from 11 through 9, 7, 5 and 3, and occur in a ring, as free traces, a medullated cylinder, or as U-shaped free traces. The reduction from nine to three bundles is correlated with the gross morphological features while 11, which occurs only in the section Peltatae (Pax) Dehgan & Webster, presents an increase. Reduction in the number of petiolar traces follows the evolutionary advancement of various taxa. This reduction in traces corresponds with south-north distribution of the species and consequential adaptation to colder and more arid climates in Central America and Africa. Smaller leaves, fewer primary veins and fewer vascular traces have resulted as a response to reduced need for water. Presence of dorsal (super-numerary) bundles which supply the petiolar glands in subgenus Jatropha (= Adenoropium Pax) is considered significant, since African taxa of the section (subsection Pubescentes Pax) have retained these bundles despite the loss of petiolar glands. The latter glands are prominent in the South American and Indian species. Sectional lines in the genus can, therefore, be drawn generally on the basis of numerical constancy and relative uniformity in the arrangement of petiolar traces. The continuity of vascular bundles from the stem into the petiole and variations of bundle arrangements are depicted in three-dimensional drawings.     

MULTIPLE PATERNITY AND SELF-FERTILIZATION IN RELATION TO FLORAL AGE IN MIMULUS GUTTATUS (SCROPHULARIACEAE)

To gain some understanding of the mechanisms responsible for the intermediate levels of both multiple paternity and selfing observed previously in the common monkeyflower, Mimulus guttatus, we performed a field experiment to assess the role of 1) multiple paternity through sequential pollinator visits and 2) selfing through corolla abscission. In M. guttatus, flowers remain open for several days; then, in the process of corolla abscission anthers drag past the stigma. We predicted that multiple visitation by pollinators over this period should increase the degree of multiple paternity of sibling arrays, and that corolla dragging may be the primary cause of the observed selfing. In three northern California populations corollas were removed after being open for 1 day (preventing further mating), and the type of matings resulting was compared to matings from adjacent flowers with undisturbed corollas. On average, seed set was twice as great in flowers with unmanipulated corollas, indicating that about half of all matings in the lifespan of a M. guttatus flower occur after 1 day. Pollen supplementation revealed that pollen was limiting in one population. Electrophoretic assay of progeny showed multiple paternity was greater in unmanipulated flowers in the two populations that were not pollen limited, suggesting that sequential pollination may be an important source of multiple paternity. However, no difference was detected in the selfing rate between the two treatments, suggesting that corolla dragging was not a source of selfing even under pollen-limited conditions.     

ANATOMY OF THE GYNOECIUM IN TWO SPECIES OF BAKERIDESIA (MALVACEAE)

Structure of the gynoecium is described in two species of Bakeridesia, subgenus Bakeridesia (Malvaceae, tribe Malveae). The dorsal wall of each carpel bears a winglike projection with a marginal pair of pubescent, bluntly dentate wings. The projection arises as a single, solid ridge of tissue after the ovules are initiated and after the ventral carpellary margins are fused with the receptacle. Two multiseriate layers of fiber-sclereids line each locule and continue into the winglike projection where they are separated by parenchyma. Gynoecial vascularization is described in detail. The richly vascularized carpels are supplied by five traces: a median dorsal trace, which bifurcates into two dorsal bundles; two lateral traces; and two ventral traces. Adjacent ventral traces, lateral traces, and septal bundles are fused—i.e., they are held in common by neighboring carpels. The presence of lateral carpellary traces may be a primitive character in the tribe Malveae.     

A systematic histological study of palm fruits. V. Subtribe Archontophoeniciae (Arecaceae)

Pericarp histology in the Archontophoenicinae provides little to characterize the subtribe as a whole, revealing instead two separate trends with parallels in other subtribes of the Areceae. The data support a close relationship among the three genera occurring in New Caledonia:Chambeyronia, Actinokentia, andKentiopsis, in which there is a complex endocarp consisting of short, oblique fibrous bundles embedded in a thick mantle of brachysclereids, and a loose endocarp of heavily fibrous, flattened vascular bundles adjacent to a relatively thin locular epidermis. The data also support a close relationship between the two genera of the New Zealand/Tasman Sea region:Hedyscepe andRhopalostylis, in which the pericarp is more or less fibrous throughout, with purely fibrous bundles in the outer pericarp and heavily fibrous vascular bundles in the inner pericarp. These results confirm relationships revealed by other morphological data.Archontophoenix appears to be most like the New Caledonian genera in its pericarp structure, with a similar mantle of short fibrous bundles embedded in a a mantle of brachysclereids in the outer pericarp, although it differs significantly in other aspects of morphology and anatomy.     

Stamens and gibberellin in the regulation of corolla pigmentation and growth in Petunia hybrida

Removal of stamens, or even of only the anthers, at an early stage of corolla development, before the start of main anthocyanin production, inhibited both growth and pigmentation of attached corollas of Petunia. When only one or two stamens were removed from one side, the inhibition was restricted to the corolla side adjacent to the detached stamens. Application of gibberellic acid (GA₃) substituted for the stamens in its effect on both growth and pigmentation. In detached corollas, isolated at the early-green stage and grown in vitro in sucrose medium, GA₃ promoted growth and was essential for anthocyanin synthesis. A marked enhancement of anthocyanin production was observed 48 h before the increase in corolla growth rate. Corollas detached at later stages were able to continue their growth and pigmentation in sucrose without GA₃. When Paclobutrazol (-[(4-chlorophenyl)-ethyl]-(1,1-dimethylethyl)-H-1,2,4-triazol-1-ethanol), an inhibitor of gibberellin biosynthesis, was added to the growth medium of in-vitro-grown corollas, pigmentation was inhibited but there was no effect on corolla growth. Low levels of GA₃ counteracted the Paclobutrazol effect on pigmentation but did not affect growth. The above results indicate that the effect of GA₃ (and probably that of the stamens) on corolla growth is independent of its effect on pigmentation. Gibberellic acid and paclobutrazol had no effect on [¹⁴C]sucrose uptake by in-vitro-grown corollas. The activity of phenylalanine ammonialyase was correlated with the effect of stamens and GA₃ on pigmentation in corollas grown in vivo and in vitro.Abbreviations GA gibberellin - GA₃ gibberellic acid - PAC Paclobutrazol - PAL phenylalanine ammonia-lyase     

MECHANISM OF SELECTION FAVORING A WIDE TUBULAR COROLLA IN CAMPANULA PUNCTATA

A previous study of Campanula punctata pollinated by Bombus diversus showed greater male reproductive success in plants with wider corollas when male reproductive success was measured by paternity analysis. Pollen removal alone was not a good measure of male reproductive success. The aim of the current study is to elucidate the mechanism underlying the selection on corolla width in C. punctata. Multiple regression analysis of male reproductive success in experimental populations revealed that the advantage of a wide corolla to the male is independent of the corolla width of pollen recipients, indicating that the position of pollen deposited on Bombus may not be an important factor for selection. However, the number of pollen grains that fell on the petal or off the flower and were lost at the first visit of Bombus (pollen fall) decreased with increasing corolla width, so that most pollen grains removed from a wide flower were deposited onto the pollinator. In a narrow flower, a greater proportion of the pollen removed was wasted through pollen fall. Pollen removal does not reflect male reproductive success, at least in part because of the loss of pollen in some flowers through pollen fall. The positive relationship between wider corollas and reduced loss of pollen through pollen fall is likely to lead to greater male reproductive success of wider flowers under repeated visits of pollinators. Decreased pollen fall, probably due to more gentle contact between pollen and bees in flowers with wider corollas, may be one of the mechanisms underlying the selection on corolla width of C. punctata.     

Photosynthetic activities in the petunia corolla

Pink Petuniahybrida (cv Hit Parade Rosa) corollas were found to contain photosynthetically active chloroplasts. The corolla chloroplasts were similar to those of green leaves in size and structure. The chlorophyll (Chl) content of Petunia corollas increased during early stages of flower development, reaching a maximum just before anthesis. Chloroplasts isolated from corollas at this stage, carried out photosystem I-dependent electron transport at rates which were two-thirds of those measured in chloroplasts from green leaves, but full chain electron transport at only one-quarter of the rate carried out by chloroplasts from green leaves. Both the light saturated rate and the quantum yield for electron transport were lower in corolla chloroplasts, which also required lower intensities for light saturation. Reduced efficiency of photosystem II photoreactions in the corolla was also indicated by the ratio between variable and constant components of Chl fluorescence, which was lower in corollas compared to green leaves. The induction time of Chl fluorescence was at least three times shorter in corollas compared to green leaves, indicating a smaller number of functional photosystem II centers (per Chl) in the corolla. It is suggested that corolla chloroplasts of Petunia might have a role in flower developmental processes.