Pollen structure and germination of Crocus thomasii Ten. (Iridaceae)

Abstract

Mature pollen of C. thomasii Ten. (Iridaceae) has been studied from a morphological and physiological point of view and compared to that of C. sativus L., C. thomasii pollen is roundish, with a 83 μm diameter and a 14% of anomalous grains. Percentage of the positive alcoholdehydrogenase (ADH +) pollen grains is about 90% whereas the in vitro germinated rate reaches 55%. Exine is homogeneously thickened (2.5 μm) with randomly distributed thinner zones and 5 μm thick intine. Cytoplasm of vegetative and generative cells is very rich of small and large smooth vesicles and vesicles coated by ribosomes. The generative cell shows a thin ondulated pectocellulosic wall. Its nucleus is intensely fluorescent after treatment with the 4,6-diamidino-2-phenylindole (DAPI) fluorochrome. Although many ultrastructural aspects of the C. thomasii pollen are common in C. sativus. L., C. thomasii pollen is smaller, more regularly structured and germinates in vivo and in vitro in higher percentage than that of C. sativus.     

Crocus ilvensis sp. nov. (sect. Crocus,Iridaceae), endemic to Elba Island (Tuscan Archipelago,Italy)

A new species of Crocus sect. Crocus is described as an endemic of Elba Island (Tuscan Archipelago, Italy): C. ilvensis Peruzzi & Carta sp. nov. (2n=8). The new species was wrongly referred to C. corsicus Vanucchi (2n=18), C. etruscus Parl. (2n=8) or C. vernus Hill. subsp. vernus (2n=8, 16) by previous authors. Its karyotype structure, asymmetry and chromosome dimensions show affinity with C. etruscus, and no clear relations with other related taxa. From a morphological point of view, C. ilvensis appears intermediate between C. etruscus and C. vernus subsp. vernus, showing however a peculiar combination of character‐states. The new species is completely allopatric with other Crocus sect. Crocus taxa and its possible role in the origin of tetraploid races of C. vernus is discussed.     

Ultrastructure of chromoplasts and other plastids in Crocus sativus L. (Iridaceae)

Saffron (Crocus sativus L. Iridaceae) chromoplasts and other plastids were studied by electron microscope to determine their structure, origin and pigment localization. Plastids from pistils of floral buds and flowers at anthesis, dried and decoloured stigmas, and green and senescent leaves were examined. Results indicated that mature saffron chromoplasts occur in the red parts of stigmas and have a reticulo-tubular structure. They contain a reticulum of tubules and plastoglobules. Tubules formed dilated vesicles mainly while plastoglobules appeared numerous and scattered on the whole chromoplast. Chromoplasts appeared in red stigma of very young floral buds. They originated from amyloplasts, the only plastids occurring in the colourless basal portion of style, as well as in the parenchyma of ovary and corm. Transition forms of plastid as amylo-chromoplast, occur in the yellow parts of stigma and style. Senescent leaves did not show plastids with structure similar to the chromoplast of red stigma. Red pigmented and scented stigmas might cooperate in saffron reproduction by attracting pollinator.     

Ultrastructure and Germination Percentage of Crocus biflorus Miller subsp. biflorus (Iridaceae) Pollen

Pollen of Crocus biflorus Miller subsp. biflorus from natural habitats of Tusculum (Frascati, near Rome, Italy) has been studied in order to compare its structure and physiology to pollen of other Crocus species belonging to the Crocus sativus group. Mature pollen grains are rounded, 60 μm in diameter, in-aperturate (but with surface incisions where exine is lacking). DAPI staining reveals a spindle-shaped generative nucleus which is intensely fluorescent, and vegetative nucleus which is less fluorescent, and is elongated with numerous lobes. At anthesis the pollen is bicellular, but about 2% of tricellular grains occur among the pollen grains released from the anthers as well as on both naturally or handpollinated stigmas. Pollen germination is low in vitro, but higher in vivo. The pollen tubes are of normal shape. An electron-dense surface coat is sometimes visible on the exine, which in many cases, is detached from the exine. The vegetative cytoplasm is very rich in glycolipid bodies surrounded by endoplasmic reticulum. The generative cell has a lobed cell wall and is surrounded by the vegetative nucleus.     

The avoidance of self-interference in the endemic daffodil Narcissus cyclamineus (Amaryllidaceae)

Hermaphrodite flowers usually possess floral traits to avoid the negative effects derived from inbreeding depression and/or self-interference between pollen export and reception, both acting as the main selective pressures on those floral traits. The avoidance of self-interference is widely accepted as the primary force promoting the separation between sexes within the flowers in time (dichogamy) and/or space (herkogamy) for self-incompatible species, which are already protected from the negative effects of inbreeding depression by the incompatibility system. Different degrees of incompatibility, herkogamy, and dichogamy have been reported for the genus Narcissus. However, the only mechanism for the separation of sexes reported up to date for Narcissus cyclamineus is herkogamy, while the presence of dichogamy and the type of incompatibility in this species remain uncertain. In this study, we analyze the patterns of sexual reproduction in N. cyclamineus to ascertain whether there is any selective pressure favouring sexual segregation or its maintenance and their mechanisms. N. cyclamineus is self-incompatible and dichogamy can be rejected for this species. Even though the species is self-incompatible, when cross-pollination is preceded by self-pollination the number of ovules available for legitimate crosses is diminished (ovule discounting). Pollinators are scarce during the flowering period, resulting in pollen limitation. It is suggested that both the scarcity of pollinators and ovule discounting may be acting synergically to promote herkogamy or its maintenance in this species.     

Chromosome numbers of Turkish Crocus (Liliiflorae,Iridaceae) and their geographical distribution

The genus Crocus is known for its widely varying chromosome numbers (from 2n = 6 to 2n = 70) with varying numbers occurring even within species, as it is the case for Crocus biflorus Miller (2n = 8, 10, 12, 14, 16, 18, 20, 22, 24). After we found morphological diverse C. biflorus populations in Turkey doubts arose about their rank of being subspecies of the Italian C. biflorus (2n = 8). Here we publish the chromosome numbers for 76 populations of C. biflorus sensu lato distributed all over Turkey. The chromosome numbers ranged from 2n = 8 to 2n = 36, with the higher numbers occurring in the mountain ranges of the Anatolian Diagonal and east of it, while lower numbers were found only southwest of these mountains. Closely related taxa with similar distribution mostly differ in their chromosome numbers. This led us to assume that chromosomal changes influence speciation processes in the genus. Therefore, chromosome numbers may represent an important character for the establishment of a new taxonomic treatment of the Crocus species, especially within section Nudiscapus. (© 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Self-incompatibility inCrocus vernus subsp.vernus (Iridaceae)

Outcross, self- and mixed pollinations were performed inCrocus vernus subsp.vernus, a species with bicellular pollen, dry stigmas and hollow style. No differences were noted among the above pollinations concerning the germination of pollen and the growth of pollen tubes until the top of ovary. Within 45 min after pollinations 62% of pollen grains germinated. Pollen tubes penetrated the papilla cuticle extending along the papilla wall; on entry into stigmatic lobes they continued growth in the stylar secretion to ovarian locules. Here, however, self-pollen tubes failed to reach or to enter the ovule micropyle; while pollen tubes from either outcross- or mixed pollinations grew until fertilizing ovules. These observations gave evidence of a self-incompatibility system inCrocus, which appeared to be neutralized by mentor effect. The ovary as site of incompatibility response is discussed.     

Pollen-pistil interactions in Hermodactylus tuberosus Mill. (Iridaceae)

ABSTRACT

Hermodactylus tuberosus Mill. (Iridaceae) is a species with a very short style and a tricarpellar unilocular ovary. For ornamental purposes it is usually reproduced by means of tubers, although both vegetative and seed reproduction occur in nature. However, its capsules do not contain many seeds in comparison to the high number of ovules inside the ovary. To understand this conflicting process a study on H. tuberosus pollen-pistil interaction was carried out. Pollen viability was evaluated by different techniques. Germination rate was tested in vitro and in vivo after stigma, intrastylar and intraovarian pollinations. A high percentage of the pollen grains were found to be viable resulting in a high percentage of in vitro germination and a low rate of germination on the stigma. Indeed, many pollen tubes developed after intrastylar and intraovarian pollinations. Several anomalies characterized pollen germination and pollen tube growth in vitro and in vivo. Observations from field trials indicated that capsules were only produced from open or hand, cross- pollinated flowers, confirming that the plant is self- incompatible and not apomictic. In addition, all the mature capsules contained only a few mature seeds suggesting the existence of pre- and postzygotic seed set inhibition.     

Embryo origin and development in Crocus sativus L. (Iridaceae)

Abstract

Fruit and seed set were obtained in the triploid Crocus sativus (saffron) either by hand crossing with pollen of the diploid C. cartwrightianus or by free pollination. The morphology of the capsules and seeds was similar in both Crocus species. The embryo is comparatively small, whereas the endosperm is formed of numerous layers of cells with hemicellulose-thickened walls and a cytoplasm containing few lipid and protein reserves. Seed germination gave rise to a white prophyll from which a green leaf emerged. At the base of this leaf, a small corm without tunics developed, which, after leaf desiccation, became dormant. These corms sown separately in pots were again capable of growth and emitted a single long green leaf. Vegetative development of new corms was similar in the diploid and hybrid saffron plants. In sites of past saffron cultivation the beds were found to contain degenerated corms and corms at different development stages. The absence of fruit and seed set in emasculated and self- and out-pollinated plants suggests that saffron does not form apomictic embryos.     

The effectiveness of some mechanisms of reproductive isolation in Arum maculatum and A. italicum (Araceae)

Mechanisms of reproductive isolation were studied in two species of Arum , A. italicum and A. maculatum , growing in England and the south of France. The study focused on three potential mechanisms for reproductive isolation: the effectiveness of dichogamy as a barrier to autogamy; the ability of self and outcrossed pollen to germinate on stigmas at different stages of anthesis and to effect pollination; and postzygotic barriers to selfing. Dichogamy was found to provide a very effective barrier to within-inflorescence selfing in these species, as no seeds were produced by spontaneous self-pollination (i.e. autogamy) in any population of either species. However, the study found that geitonogamy (cross-pollination between inflorescences of a same individual or clone) was possible, as genotypes frequently produced several inflorescences and stigmas were found to be receptive to pollen from before anthesis until their contraction at the end of the female phase of flowering. Hand pollination with self pollen from clone inflorescences produced as numerous and heavy seeds as outcross pollination. In addition, the germination and growth of geitonogamously produced seed was similar to that of outcrossed seed, suggesting that this potential postzygotic barrier is absent or weak. These findings suggest that geitonogamous seed production may be unrestricted by significant pre- or postzygotic barriers. The possible advantages of geitonogamy in Arum are discussed.  © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society , 2006, 150 , 323–328.     

藏红花细胞悬浮培养动力学研究

基于已经建立的藏红花细胞悬浮培养体系,研究了其悬浮培养动力学.结果表明,悬浮培养时细胞的生长周期约为20 d,在20 d时生物量达到最大,为12.3 g DW/L,但是藏红花素的合成周期大约为28 d,在28 d时藏红花素的含量和产量达到最大,分别为95.8 mg/g和0.92 g/L.藏红花素的积累与细胞的生长之间的关系为半生长偶联型,为其生物反应器放大培养提供了参数等理论依据.     

Reversible inhibition of the pollen germination and the stigma penetration in Crocus vernus ssp. vernus (Iridaceae) following fumigations with NO2, CO, and O3 Gases

We assessed the pollen hydration, the pollen germination, and the stigma papilla penetration of CROCUS VERNUS subsp. VERNUS (Iridaceae) after 2 h fumigations with O (3), NO (2), and CO gases within humidified (90 - 100 % RH) box experiments. When the pollen and the pistil were separately fumigated, the pollen retained the capacity to emit a tube which penetrated papilla, and the stigma papillae retained the receptivity; when the pistils were first pollinated and then fumigated, the capacity of pollen to hydrate was not affected, but the germination was significantly reduced. The vulnerability to gases became evident at 0.3 ppm O (3), 0.2 ppm NO (2), and 0.5 ppm CO. The inhibition curves as a function of the gas concentrations were of an exponential type, and they saturated at 2 ppm NO (2), 25 ppm CO, and 0.5 ppm O (3), with germination percentages of 17 %, 27 %, and 60 %, respectively. Both the pollen germination and the papilla penetration were fully restored by prolonging for 60 - 90 min the incubation at 90 - 100 % RH, after the cessation of fumigations. The vulnerability of the pollen-papilla system is discussed.     

Inflorescence and floral ontogeny in Crocus sativus L. (Iridaceae)

Inflorescence and floral ontogeny of the perennial, herbaceous crop Crocus sativus L. were studied using epi-illumination light microscopy. After production of leaves with helical arrangement a determinate inflorescence forms which becomes completely transformed into a single terminal flower. In some cases, bifurcation of the inflorescence meristem yields two or three floral meristems. The order of floral organs initiation is outer tepals – stamens – inner tepals – carpels. Stamens and outer tepals are produced from the lateral bifurcation of three common stamen-tepal primordia. Within each whorl, organs start developing unidirectionally from the adaxial side, except for the stamens which begin to grow from the abaxial side. Specialized features during organ development include interprimordial growth between tepals forming a perianth tube, fusion at the base of stamen filaments, and formation of an inferior ovary with unfused styles.     

Pollination of Oncocyclus irises (Iris: Iridaceae) by night-sheltering male bees

Irises in the section Oncocyclus (Siems.) Baker ( IRIS: Iridaceae) grow throughout the Middle East and have large and dark-coloured flowers but no nectar reward available to flower visitors. Consequently, no reward-collecting pollinators have been observed visiting the flowers during daytime. The only visitors are solitary male bees ( Eucera spp.: Apidae) that enter the flowers at dusk and stay there overnight. Here we describe the mating system of Oncocyclus irises, and the role of night-sheltering male bees in their pollination system. Pollen viability in I. haynei on Mt. Gilboa was very high (>90%) throughout all floral life stages. Stigmas were receptive in buds and in open flowers, but not in older ones. Self-pollination yielded no fruits in three species, confirming complete self-incompatibility in Oncocyclus irises. On average, 1.9 flowers were visited by each male bee before it settled for the night in the last one. Moreover, Iris pollen was present on the dorsal side of 38.8% of males caught sheltering in flower models mounted near an I. atrofusca population, indicating that pollen is transferred between flowers by night-sheltering solitary male bees. We have surveyed 13 flowering populations of six Oncocyclus species for the presence of night-sheltering male bees as well as for fruit set. We found a positive correlation, indicating that sexual reproduction in Oncocyclus irises is dependent on night-sheltering solitary male bees. Based on their complete self-incompatibility, the absence of nectar-collecting visitors during the day, and the transfer of pollen grains by the night-sheltering solitary male bees, we conclude that fertilization of Oncocyclus irises is totally dependent on pollination by night-sheltering solitary male bees.     

The reproductive biology of the Mediterranean endemic Cyclamen balearicum Willk. (Primulaceae)

Knowledge of the reproductive biology of endemic plants improves our understanding of how mating system may be related to patterns of species abundance and provides a basis for the development of rational conservation programmes. In this paper we present natural population data on the floral biology and reproductive ecology of the endemic Mediterranean species Cyclamen balearicum Willk. This is a long-lived, diploid perennial herb which occurs in southern France in five fragmented and isolated regions and on the Balearic Islands of Mallorca, Menorca, Ibiza, Cabrera and Draponera. Our observations indicate a particularly scarce pollinator activity (rare syrphid visits) and dispersal by ants over small distances. A controlled pollination experiment in a natural population showed that in southern France C. balearicum is fully self-compatible and that selfing is autonomous and probably delayed (i.e. following opportunities for outcrossing). The proximity of stigmas and anthers will favour autonomous selfing. The high pollen/ovule ratio indicates nevertheless that C. balearicum has a mixed mating system. Patterns of variation in stigma-anther separation and pollen production per flower suggest that not only has the current mating system of the species evolved from an outcrossing ancestor but that due to the fragmentation and isolation of populations greater levels of selfing have evolved in southern France (and to an intermediate degree on Ibiza and Menorca). On the island of Mallorca where larger continuous belts of C. balearicum habitat still exist the species has floral traits indicating a more outcrossed mating system. To our knowledge this is the first paper to document such trends in floral traits in the endemic component of the Mediterranean flora.     

艳丽耳草的二型花柱及异型自交不亲和系统

异型花柱是一种受遗传因素控制的花型多态性现象,包括二型花柱和三型花柱两种类型.本文以茜草科艳丽耳草(Hedyotis pulcherrima)为实验材料,通过对其野外居群的花型、花部形态及花粉特征等观察,发现艳丽耳草野外居群同时存在长花柱型花和短花柱型花,长/短花柱型花的数量比例为1∶1.两型花具有精确的交互式雌雄异位特征,并且该特征与花冠长度相关性显著.长/短花柱型花的柱头裂片长度、花粉大小及淀粉含量等具有二型性.花粉体外培养时花粉萌发率及花粉管生长速率无显著性差异.人工授粉后,艳丽耳草长/短花柱型花型间异交花粉管生长形态正常,授粉24 h后花粉管均已进入子房.而长/短花柱型花在自交及型内异交下均表现为不亲和,花粉管生长停止于柱头,花粉管顶端累积胼胝质并膨大.艳丽耳草没有无融合生殖现象,型间人工辅助异交授粉结实率为100％,显著高于自然结实率.本研究结果表明,艳丽耳草是典型的二型花柱植物,并具有异型自交不亲和系统.     

Patterns of style polymorphism in five species of the South African genus Nivenia (Iridaceae)

Background and Aims

Heterostylous plants have been characterized by the presence of two or three discrete morphs that differ in their sex organ position within populations. This polymorphism is widely distributed among the angiosperms, but detailed studies are limited to few taxonomic groups. Although a small representation, evolutionary meaningful variations of the heterostylous syndrome have been reported when precise measurements of the sexual whorls were taken. A thorough exploration of groups where heterostyly has been reported should offer new opportunities to further testing the evolutionary hypotheses explaining heterostyly. Here, the traits defining heterostyly were explored in half of the species in Nivenia, the only genus of Iridiaceae where heterostyly has been reported.

Methods

Detailed morphometric analysis of the flower sexual whorls and some traits considered as ancillary are supplied to determine for each population (a) the kind of stylar polymorphism, (b) the morph ratio and (c) the degree of reciprocity between sexual whorls. Also the rates of assortative (within morph) versus disassortative (between morphs) pollen transfer were estimated by analysing pollen loads on stigmas. The association between floral phenotypic integration and the reciprocity between sexual whorls was estimated; both characteristics have been quoted as dependent on the accuracy of the fit between pollinators and flowers and therefore related to the efficiency of pollen transfer.

Key Results

Different types of polymorphism, differing in their degree of reciprocity, were found in Nivenia. Effective disassortative mating appears to be common, since (a) all dimorphic populations show equal morph-ratios (isoplethy), and (b) the pollen placed on the stigmas of each morph is likely to be coming from the other (complementary) morph. The most reciprocal populations of the heterostylous species have also the highest values of phenotypical integration.

Conclusions

Stigma height dimorphism, as opposed to distyly, is proven for the first time in Nivenia. The presence of different types of polymorphism within the genus is consistent with hypotheses of the evolution of heterostyly. The role of the pollinators as the leading force of the transition seems to be apparent, since floral integration is related to reciprocity.     

Isozyme diversity in Iris Cristata and the threatened glacial endemic I. Lacustris (Iridaceae)

Iris cristata and I. lacustris differ markedly in geographic distribution, glacial history of current ranges, and ecology. We hypothesized that I. cristata, a widespread species of unglaciated regions of eastern North America, would exhibit genetic diversity typical of other widespread plant species, whereas the threatened I. lacustris, which occupies glaciated habitats on Great Lakes shorelines, would display little genetic variation. Iris lacustris lacked detectable polymorphisms in 18 isozyme loci, although we found evidence of possible incomplete gene silencing in four additional loci in some populations. In contrast, I. cristata was polymorphic at 73% of 15 loci examined, with an average of three alleles per locus. Genetic diversity (H_e) was 0.231. All species-level and population-level estimates of genetic diversity were higher than averages for plants having comparable life history traits. Nearly 98% of the total genetic diversity in I. cristata was apportioned within populations, and heterozygosity and fixation estimates suggest a high level of outcrossing in this species (t = 1.265). The long-lived perennial habit and high outcrossing rate in stable populations are proposed as factors contributing to high genetic diversity in I. cristata. The data are consistent with an hypothesis of a recent origin of I. lacustris from a very limited I. cristata gene pool exacerbated by repeated bottlenecks and founder effects as I. lacustris populations were displaced by lake-level changes over the past 11 000 yr.