Structure of floral nectaries and female-biased nectar production in protandrous species Geranium macrorrhizum and Geranium phaeum.

Nectar is a major floral reward offered to pollinators by plants. In dichogamous plant species, differences in nectar production across sexual phases often occur, but both the male- and female-phase flowers have to attract pollinators to achieve effective pollination. Nectar-producing structures, i.e. floral nectaries, are a key component of floral organisation and architecture, and the knowledge of their structure and function contributes to better understanding of the plant-pollinator interactions. In the present study, we investigated the morphology and structure of nectaries and the nectar production pattern in two protandrous species Geranium macrorrhizum and G. phaeum. The flowers of the studied species have been shown to exhibit varied availability of nectar for insect visitors. Their nectaries differ in the shape, size and thickness. The other differences include the localisation of the stomatal field, the size and number of nectarostomata, the presence of non-glandular and glandular trichomes, the presence of tannin idioblasts, the mode of secretion and the occurrence of plastids functioning probably as autophagosomes and autolysosomes, whose presence in nectary cells has been described for the first time. The flowers of the studied species started nectar secretion in the non-receptive phase before pollen presentation and nectar was produced throughout both sexual phases. The nectar production was gender biased towards the female phase in the nectar amount, nectar sugar concentration and total sugar secreted in the nectar. We postulate that the nectar production patterns in G. phaeum and G. macrorrhizum might have evolved as a response to pollinators' pressure.

Taxonomic significance of macro and micro-morphology of Geranium L. species Using Scanning Electron Microscopy.

Macro and micromorphology of different parts that is, sepals, petals, flowers, indumentum, and leaf shape of 22 Geranium L. taxa from Iran were studied. These taxa representing the eight sections of the genus as sect. Dissecta, sect. Geranium, and sect. Tuberosa (of subgen. Geranium); sect. Batrachioidea, sect. Divaricata, sect. Lucida, sect. Ruberta and sect. Trilopha (of subgen. Robertium), that were collected from different geographical habitats of Iran. The biometric study involved 27 quantitative and 33 qualitative characters which were studied by use of stereomicroscopy and scanning electron microscopy (SEM). Statistical analyses were done by use of PAST software. Principal Component Analysis defined the most variable characters. Results showed that the diagnostic features were; length, width, apex of sepals, petals, trichomes type in pedicles and sepals along with their density, the shape length and width of leaves. In general, the present study revealed that the species could be differentiated by macro and micro-morphological characters. Taxa of two subgenera were clearly separated based on selected characters. These characters were found useful for the taxonomic identification and species delimitation in almost all taxa studied at the sub generic level, although some species of section Geranium and section Dissecta are mixed and nested in both subgenea.

Species-level phylogeny, fruit evolution and diversification history of Geranium (Geraniaceae).

The cosmopolitan genus Geranium L. (Geraniaceae) consists of c. 350 species distributed in temperate habitats worldwide, with most of its diversity concentrated in the Mediterranean region. Unlike other genera in Geraniaceae, the species of Geranium present contrasting seed discharge syndromes, i.e. the 'Erodium-type' (ET), the 'carpel-projection type' (CP), the 'seed-ejection type' (SE), and the 'inoperative type' (IT), which have been used to delimit major groups within the genus. However, phylogenetic relationships within Geranium are unknown and so is the evolution of the different seed discharge mechanisms. Here, we used a calibrated multispecies coalescent approach to infer the species-level phylogeny and divergence times of the genus based on chloroplast (rbcL, trnL-trnF) and nuclear (ITS) DNA sequences. Our sampling represents most of the morphological variation described in the genus. We reconstruct the evolution of the seed discharge mechanism using ancestral state reconstruction (ASR) techniques on the multispecies coalescent tree, and assess the association between fruit type evolution and species diversification using stochastic birth-death and trait-dependent diversification models. Finally, we reconstruct the early biogeographic history of the genus using discrete and continuous biogeographic analyses of species distribution centroids, including fossil evidence and tip dates. Our results show that fruit type is homoplasious and that the classification based on fruit type in Geranium is artificial. The taxonomy and putative apomorphic characters for Geranium are discussed. ASR of the fruit characters suggests that ET may represent the ancestral state in Geranium and from which CP originated twice, IT presumably once, and SE twice. The independent appearance of the SE syndrome is in both cases associated with increases in diversification rates in the genus. The biogeographic analysis centers the origin and early 10Ma diversification of Geranium on the Mediterranean region. The evolution of seed discharge mechanism about 5Ma might have allowed the species of Geranium to increase in geographic range and to ultimately, diversify.

Frequency-dependent pollinator discrimination acts against female plants in the gynodioecious Geranium maculatum.

BACKGROUND AND AIMS: Gynodioecy, the co-occurrence of female and hermaphroditic individuals, is thought to be an intermediate step between hermaphroditism and separate sexes, a major transition in flowering plants. Because retaining females in a population requires that they have increased seed fitness (to compensate for the lack of pollen fitness), factors that affect seed fitness are of great importance to the evolution of this mating system and have often been studied. However, factors negatively affecting female fitness are equally important and have been largely neglected. One such factor stems from female flowers being less attractive to insects than hermaphrodite flowers, thereby decreasing their relative fitness. METHODS: To test the severity and consequences of this type of pollinator discrimination in Geranium maculatum, experimental populations with the range of sex ratios observed in nature were created, ranging from 13 % to 42 % females. Pollinators were observed in order to measure the strength of discrimination, and pollen deposition and seed production of both sexes were measured to determine the fitness consequences of this discrimination. Additionally a comparison was made across the sex ratios to determine whether discrimination was frequency-dependent. KEY RESULTS: It was found that female flowers, on average, were visited at half of the rate of hermaphrodite flowers, which decreased their pollen receipt and seed production. Additionally, females were most discriminated against when rare, due to both changes in the pollinators' behaviour and a shift in pollinator composition. CONCLUSIONS: The results suggest that pollinator discrimination negatively affects females' relative fitness when they are rare. Thus, the initial spread of females in a population, the first step in the evolution of gynodioecy, may be made more difficult due to pollinator discrimination.

Herbivore pressure by weevils associated with flower color polymorphism in Geranium thunbergii (Geraniaceae).

Although floral herbivory has recently received increased attention as an important factor influencing plant reproduction, relatively little is known about how its frequency and intensity vary depending on traits of host plants. Here we report that herbivore pressure by a weevil, Zacladus geranii, is associated with a flower color polymorphism of Geranium thunbergii (Geraniaceae). Pink and white flower color morphs have been reported in G. thunbergii, and we found in a three-year field survey in multiple populations that, generally, adult weevils more preferentially visited white flowers than pink flowers. Consistently, we found more severe damage by weevil larvae in white flowers. Overall herbivore pressure for G. thunbergii varied strongly between populations, and the difference seems to be partly explained by the co-occurrence of a related plant species, Geranium yezoense, in a population, as weevils preferred it to both color morphs of G. thunbergii, thereby relaxing overall herbivore pressure for G. thunbergii. Nonetheless, despite such high variability, the preference of weevils for white morphs over pink morphs of G. thunbergii was found across multiple populations. We discuss possible mechanisms causing the association between flower color and herbivore preference as well as its evolutionary consequences.

Quantitative analysis of the thermal requirements for stepwise physical dormancy-break in seeds of the winter annual Geranium carolinianum (Geraniaceae).

BACKGROUND AND AIMS: Physical dormancy (PY)-break in some annual plant species is a two-step process controlled by two different temperature and/or moisture regimes. The thermal time model has been used to quantify PY-break in several species of Fabaceae, but not to describe stepwise PY-break. The primary aims of this study were to quantify the thermal requirement for sensitivity induction by developing a thermal time model and to propose a mechanism for stepwise PY-breaking in the winter annual Geranium carolinianum. METHODS: Seeds of G. carolinianum were stored under dry conditions at different constant and alternating temperatures to induce sensitivity (step I). Sensitivity induction was analysed based on the thermal time approach using the Gompertz function. The effect of temperature on step II was studied by incubating sensitive seeds at low temperatures. Scanning electron microscopy, penetrometer techniques, and different humidity levels and temperatures were used to explain the mechanism of stepwise PY-break. KEY RESULTS: The base temperature (Tb) for sensitivity induction was 17·2 °C and constant for all seed fractions of the population. Thermal time for sensitivity induction during step I in the PY-breaking process agreed with the three-parameter Gompertz model. Step II (PY-break) did not agree with the thermal time concept. Q10 values for the rate of sensitivity induction and PY-break were between 2·0 and 3·5 and between 0·02 and 0·1, respectively. The force required to separate the water gap palisade layer from the sub-palisade layer was significantly reduced after sensitivity induction. CONCLUSIONS: Step I and step II in PY-breaking of G. carolinianum are controlled by chemical and physical processes, respectively. This study indicates the feasibility of applying the developed thermal time model to predict or manipulate sensitivity induction in seeds with two-step PY-breaking processes. The model is the first and most detailed one yet developed for sensitivity induction in PY-break.

A method for quantifying rotational symmetry.

Here, a new approach for quantifying rotational symmetry based on vector analysis was described and compared with information obtained from a geometric morphometric analysis and a technique based on distance alone. A new method was developed that generates a polygon from the length and angle data of a structure and then quantifies the minimum change necessary to convert that polygon into a regular polygon. This technique yielded an asymmetry score (s) that can range from 0 (perfect symmetry) to 1 (complete asymmetry). Using digital images of Geranium robertianum flowers, this new method was compared with a technique based on lengths alone and with established geometric morphometric methods used to quantify shape variation. Asymmetry scores (s) more clearly described variation in symmetry and were more consistent with a visual assessment of the images than either comparative technique. This procedure is the first to quantify the asymmetry of radial structures accurately, uses easily obtainable measures to calculate the asymmetry score and allows comparisons among individuals and species, even when the comparisons involve structures with different patterns of symmetry. This technique enables the rigorous analysis of polysymmetric structures and provides a foundation for a better understanding of symmetry in nature.

Root, stem and leaf anatomy of Geranium palustre Torner Cent. (Geraniaceae).

An anatomical study on the root, stem and leaf of Geranium palustre Torner Cent. has been performed in order to continue our previous researches on this species. Technique of inclusion into paraffin and optical microscope have been used. Root, stem and leaf transverse sections were studied. They contain calcium oxalate crystals as inclusions. The root presents elements of secondary structure (suber, secondary cortex, secondary xylem and phloem). The vascular bundles in the stem can be classified as internal or external and differ in the presence or absence of an own sclerenchyma sheet depending on their age. The vascular bundles of stem are collateral. Leaf structure is bi-facial. Stem and leaf surface have no glandular hairs.

Sex allocation of females and hermaphrodites in the gynodioecious Geranium sylvaticum.

Seed production and patterns of sex allocation were studied in female and hermaphroditic plants in two gynodioecious populations of Geranium sylvaticum (Geraniaceae). Females produced more flower buds and seeds than hermaphrodites in one of the two study populations. The other female traits measured (pistil biomass, seed number per fruit, individual seed mass) did not differ between the gender morphs. The relative seed fitness of hermaphrodites differed between the study populations, with hermaphrodites gaining less of their fitness through female function in the population with a high frequency of females. However, the amount and size of pollen produced by hermaphrodites did not differ between populations. The number of flower buds was positively correlated with seed production in females, whereas in hermaphrodites a positive correlation between number of buds and seed production was found in only one of the two study populations. These results suggest that fitness gain through female function is labile in hermaphrodites of this species, and is probably affected by environmental factors such as the sex ratio of the population.