Paulo Silveira

Effect of pollination on floral longevity and costs of delaying fertilization in the out-crossing Polygala vayredae Costa (Polygalaceae).

BACKGROUND AND AIMS The effect of pollination on flower life span has been widely studied, but so far little attention has been paid to the reproductive consequences of delayed pollination in plants with long floral life spans. In the present study, Polygala vayredae was used to answer the following questions. (1) How does male and female success affect the floral longevity of individual flowers? (2) How does delaying fertilization affect the female fitness of this species? METHODS Floral longevity was studied after experimental pollinations involving male and/or female accomplishment, bagging and open pollination. The reproductive costs of a delay in the moment of fertilization were evaluated through fruit set, seed-ovule ratio and seed weight, after pollination of flowers that had been bagged for 2-18 d. KEY RESULTS Senescence of the flowers of P. vayredae was activated by pollen reception on the stigmatic papillae, while pollen removal had no effect on floral longevity. Nonetheless, a minimum longevity of 8 d was detected, even after successful pollination and pollen dissemination. This period may be involved with the enhancement of male accrual rates, as the female accomplishment is generally achieved after the first visit. Floral life span of open-pollinated flowers was variable and negatively correlated with pollinator visitation rates. Delayed pollination had a major impact on the reproductive success of the plant, with fruit set, seed-ovule ratio and seed weight being significantly diminished with the increase of flower age at the moment of fertilization. CONCLUSIONS A strong relationship between pollination and floral longevity was observed. Flowers revealed the ability to extend or reduce their longevity, within some limits, in response to the abundance of efficient pollinators (i.e. reproductive fulfilment rates). Furthermore, with scarce or unpredictable pollinators, a long floral life span could maintain the opportunity for fertilization but would also have reproductive costs on production of offspring. Reduced female fitness late in the flowers life could shift the cost-benefit balance towards a shorter life span, partially counteracting the selection for longer floral life span potentially mediated by scarce pollination services.

Flow cytometric and cytogenetic analyses of Iberian Peninsula Festuca spp.

Festuca L. has an important diversification centre in the Iberian Peninsula. We used chromosome counting, fluorescence (FISH) and genomic in situ hybridization (GISH), and DNA flow cytometry (FCM) to clarify the taxonomic position of several taxa, to search for phylogenetic relationships and to assess the extent and pattern of genome variation in fescues. The chromosome number of Festuca duriotagana var. barbata is determined for the first time and new ploidy level estimations are given for F. rothmaleri and F. summilusitana. In the latter species, besides the reported decaploid level, dodecaploidy was found in some populations, which points to the existence of an unrecognized taxon. Moreover, these differences were confirmed by FCM and a high positive correlation was found with the type of substrate where F. summilusitana was growing. For each section, a decrease of genome size with increase of polyploidy was observed. In general, in situ hybridization techniques failed to reveal phylogenetic relationships among the selected species. In FISH, a variation in the number of rDNA sites was observed in some species. GISH results indicate that F. henriquesii is not a progenitor of the studied polyploid species.

Consequences of nectar robbing for the fitness of a threatened plant species

The effect of nectar robbing on plant fitness is poorly understood and restricted to a few plant species. Furthermore, the available studies generally evaluate the effects of nectar robbing on female fitness, disregarding the male component. Here we measured the effects of the nectar-robbing bumblebees on male (measured as pollen analogue flow distance) and female (measured as seed production) reproductive success in the insect-dependent Polygala vayredae, a narrow endemic species from the pre-Pyrenees (Spain). Intense nectar robbing by bumblebees significantly reduced the nectar available to legitimate pollinators in the studied population, and this reduction affected both male and female fitness. Significant differences were observed in fluorescent dye dispersion between robbed and non-robbed flowers within the population. Fluorescent dyes from non-robbed flowers were dispersed to larger distances and over a larger number of flowers when compared with robbed ones. Moreover, significant differences were observed in both fruit set and seed ovule ratios between the two groups, with non-robbed flowers presenting higher reproductive outcomes. However, no effect on seed weight was detected among treatments. The data obtained suggest that in this species, nectar robbing has important indirect and negative effects on plant fecundity, through both male and female functions, due to a modification in the foraging behaviour of legitimate visitors.

How does secondary pollen presentation affect the fitness of Polygala vayredae (Polygalaceae)

Secondary pollen presentation is the relocation and presentation of pollen in floral structures (termed pollen presenters) other than the anthers. These pollen presenters are often found close to the stigma and have been hypothesized to increase the accuracy of pollen transfer, although no experimental studies have been done. We examined the function of the pollen presenter and its efficiency in pollen dispersal, female fitness, and the degree of interference created by self-pollen in the shrublet Polygala vayredae, an insect-pollinated species with secondary pollen presentation. Herkogamy, a mechanism generally involved in the reduction of self-interference, was also evaluated. Significant pollen was lost (49% of total pollen) during the secondary relocation in the pollen presenter. However, pollen was exported from the pollen presenter, and subsequent pollen losses were similar to those in species with primary pollen presentation. Despite the presence of a self-incompatibility system, the numbers of developed pollen tubes as well as fruit and seed production were significantly reduced by the self-pollen interference created at the stigmatic papillae level. The extent of herkogamy correlated positively with female fitness. The secondary pollen presentation mechanism may in fact be an accurate system for pollen transport, but it may also have its costs. Further comparative studies involving species with primary and secondary pollen presentation are needed to fully understand the advantages and disadvantages of secondary pollen presentation.

So many visitors and so few pollinators: variation in insect frequency and effectiveness governs the reproductive success of an endemic milkwort

AbstractPlant–pollinator interactions are one of the most important and variable mutualisms having major implications for plant fitness. The present study evaluates the interactions between an endemic milkwort, Polygala vayredae, and its floral visitors by studying the temporal variability, foraging behaviour and effectiveness of floral visitors in three populations during three consecutive years. The flowers were visited by a diverse array of insects, totalling 24 different species. However, only four species were effective pollinators, depositing pollen on stigmas after one visit, while the remaining species behaved as nectar robbers, secondary nectar robbers or nectar thieves and were completely ineffective for pollination. Among the effective pollinators, two groups with distinct foraging behaviours were observed: the nectar collecting long-tongued bees Bombus pascuorum and Anthophora sp. and the pollen collectors Eucera longicornis and Halictus sp. No significant differences were observed among pollinators in their efficiency in pollen deposition on stigmas, but significant differences were observed in the foraging behaviour between nectarivorous and pollen collectors. Variation in the abundance and assemblage of floral visitors was observed at the temporal scale and among populations, with the effective pollinators being generally scarce. Consequently, the reproductive outcome in this species was low and significantly variable among populations and years. The results highlight the importance of studying floral visitor effectiveness when determining pollinator assemblages.

Dispersal mechanisms of the narrow endemic Polygala vayredae: dispersal syndromes and spatio-temporal variations in ant dispersal assemblages

This study assesses the dispersal mechanisms of the narrow endemic Polygala vayredae, analysing the functioning of its dispersal syndromes (anemochory and myrmecochory), the spatio-temporal variability of the disperser assemblage, foraging behaviour and dispersal ability, and the role of the elaiosome in ant attraction and seed germination. The dispersion of diaspores begins when either (1) capsules or seeds fall beneath the mother plant (barochory) or (2) the seeds are directly collected in the suspended capsules by ants (myrmecochory). As capsules frequently open and expose/disseminate seeds before leaving the mother plant, the adaptation for anemochory appears to be reduced and rarely functional, possibly with only occasional events of long-distance dispersal (e.g. under extreme weather conditions). P. vayredae is essentially myrmecochorous and a diverse array of ant species are involved in seed manipulation, with the elaiosome playing a major role in ant attraction. From the plant’s perspective for dispersal, the majority of ant species had a positive interaction with the seeds, but negative and potential neutral interactions were also observed. Overall, dispersal distances were limited and were mainly determined by ant body size. The frequency of interactions and the ant assemblage varied significantly both spatially and temporally, and these factors may have an effect on directing or disrupting the selection of plant traits. Low seed predation and similar germination rates of intact seeds compared with seeds without elaiosome indicate that seed predator avoidance and seed germination improvement after ant manipulation are not among the selective advantages of myrmecochory operating at present. Dispersal mechanisms that enhance seed dispersal within the population and only occasionally lead to long-distance dispersal events, along with the rarity and patchiness of suitable habitats, may be the main factors explaining the actual density and narrow distribution of this species.

Pistil anatomy and pollen tube development in Polygala vayredae Costa (Polygalaceae)

Low seed ovule ratios have been observed in natural populations of Polygala vayredae Costa, a narrowly endemic species from the oriental pre-Pyrenees. To evaluate physical and nutritional constraints and pollen tube attrition in this endemic species, stigma and style anatomy, as well as pollen tube development along the pistil were investigated using light and fluorescence microscopy. The structural morphology of the stigmatic region was also examined with scanning electron microscopy. Pollen grains that reached the stigmatic papillae came into contact with a lipid-rich exudate and germinated easily. Although a large number of pollen grains reach the stigmatic papillae, few pollen tubes were able to grow into the style towards the ovary. The style was hollow, with the stylar channel beginning a few cells below the stigmatic papillae. Initially, the stylar channel area was small compared to other levels of the style, and was surrounded by lipid-rich, highly metabolic active cells. Furthermore, lipid-rich mucilage was detected inside the stylar channel. At subsequent style levels towards the ovary, no major reserves were detected histochemically. The reduced intercellular spaces below the stigmatic papillae and the reduced area of the stylar channel at its commencement are suggested to physically constrain the number of pollen tubes that can develop. In subsequent levels of the style, the stylar channel could physically support a larger number of pollen tubes, but the lack of nutritional reserves cannot be disregarded as a cause of pollen tube attrition. Finally, the number of pollen tubes entering the ovary was greater than the number of ovules, suggesting that interactions occurring at this level play a major role in the final reproductive outcome in this species.

Pollen morphology of Chamaebuxus (DC.) Schb., Chodatia Paiva and Rhinotropis (Blake) Paiva (Polygala L., Polygalaceae)

Abstract Polygala L. is a large and highly diverse genus with complex taxonomy, but pollen morphological information for this taxon is scarce. In the present study, pollen characters have been used to assess the taxonomic delimitation and phylogenetic relationships of three newly established subgenera of Polygala: Chamaebuxus, Chodatia and Rhinotropis (sensu Paiva). The pollen morphology of 22 species has been examined using light microscopy and scanning electron microscopy of acetolysed material. The pollen of 15 of the species is examined for the first time. The pollen grains are isopolar, radially symmetrical, tectate and, typically, polyzonocolporate with numerous colpi running parallel to the polar axis, and an endocingulum around the equator. Two pollen types can be distinguished: Type I, which includes species belonging to Rhinotropis, and Type II, which includes species from Chamaebuxus and Chodatia. The two pollen types are described and the pollen of the three studied subgenera is illustrated. Despite the low infrageneric morphological diversity observed within the genus Polygala, quantitative characters of pollen grains support the current classification of the subgenera Chamaebuxus, Chodatia and Rhinotropis, and reveal a closer relationship between the first two taxa. Pollen characters are shown to be a useful and informative tool for assessing taxonomic position and phylogenetic relationships within Polygalaceae, especially at higher taxonomic levels.

Bacteria from nodules of wild legume species: Phylogenetic diversity, plant growth promotion abilities and osmotolerance

The demand for food with high nutritional value that can sustain the growth of human population while safeguarding sustainability deserves urgent attention. A possible strategy is the inoculation of crops with plant growth promoting (PGP) bacteria. Plants are naturally colonized by bacteria that can exert beneficial effects on growth and stress tolerance. N2 fixation by rhizobia in the root nodules of legumes is a well-known PGP effect. These bacteria can be used as inoculants to boost legumes productivity and can be especially interesting if they are able to survive to abiotic stresses, such as drought. Herein we report the phylogenetic diversity of bacteria colonizing the root nodules of several wild legume species, from four geographic locations in Portugal with different bioclimates. Interestingly, the vast majority of strains belonged to Flavobacterium, Pseudomonas and other genera apart from rhizobia. PGP abilities other than N2 fixation (production of indol acetic acid, siderophores and volatile organic compounds) and osmotolerance were screened. Location and host plant species did not influence PGP abilities and osmotolerance. Taken together, results evidenced that bacterial strains from wild legumes displaying PGP abilities and osmotolerance can be regarded as good candidates for inoculants of a broad range of hosts, including non-legumes.

Contribution to the knowledge of the pollen morphology in the tribe Orobancheae Lam. & DC. (Orobanchaceae)

Abstract The pollen grains of 53 species and 57 taxa of two genera (Cistanche, Orobanche s. l.) of tribe Orobancheae were studied with light microscopy (LM). Thirty-five of these species were also studied with scanning electron microscopy (SEM). Two pollen types (Orobanche and Phelipanche) and six pollen groups (I–VI) were described. A principal component analysis was performed to help clarify some taxonomic problems at generic, specific and infraspecific levels. Some transition forms exist between the two pollen types. We consider that these data support the elevation of Trionychon to the rank of subgenus (Phelipanche). The pollen morphology of the subgenus Boulardia and of the genus Cistanche is quite similar to that of subgenera Orobanche and Phelipanche, respectively. Our results support the position of Boulardia as the sister taxon to Orobanche s. s. The contribution of pollen morphology to the resolution of several taxonomic problems related to specific and infra-specific taxa was discussed. Within the Orobancheae, the tricolpate, isopolar, microreticulate-scabrate pollen grains are plesiomorphic and the apolar, granulate, inaperturate pollen grains are apomorphic. Some taxa (e.g. O. cernua, O. cumana, O. lavandulacea and O. mutelii) present pollen heteromorphism, which could be related to the existence of geographical clines. The polymorphic (inaperturate, colpate), scabrate-perforate, tectate-columellate pollen grains present in some species (O. cernua, O. cumana) possibly represent an intermediate evolutionary type. Considering the separation of specific and infraspecific taxa in the Orobancheae, LM is apparently more useful than SEM, although information from SEM is clearly relevant as a complement for LM.