Seasonal changes in pollen limitation and femaleness along the snowmelt gradient in a distylous alpine herb,Primula modesta

Flowering phenology of alpine plants is strongly determined by the timing of snowmelt, and the conditions of pollination of widely distributed plants vary greatly during their flowering season. We examined the reproductive success of the distylous alpine herb, Primula modesta, along the snowmelt gradient under natural conditions, and compared it with the result of artificial pollination experiments. In addition, the compositions and visit frequencies of pollinators to the flower of P. modesta were examined during the flowering period. The pin and thrum plants of P. modesta growing at the same site have an equal ability to produce seeds if a sufficient amount of legitimate pollen grains are deposited on the stigma surface. However, under natural conditions, their seed‐set success was often (even if not always) restricted by pollen limitation, and the functional gender of the pin and thrum plants biased to the female and male, respectively, associated with their growing sites. These variations were not ascribed to resource limitation nor biased morph ratio but to the seasonal changes in pollination situations, a replacement of pollinator types from long‐ to short‐tongued pollinators resulted in unidirectional pollen transfer from long stamens (thrum plants) to long styles (pin plants). The functional gender specialization may enhance the evolution of dioecy from heterostyly, but the severe pollen limitation may cause the breakdown of heterostyly into homostyly. To consider the evolutionary pathway of heterostylous plants, an accumulation of the empirical data is required demonstrating how phenological synchrony between plants and pollinators is decided and to what degree this relationship is stable over years, along with estimates of selection and gene flow in individual plants.     

Temporal variations in the linkages between plants and flower visitors and the pollination success of Primula modesta along the snowmelt gradient

Although pollination networks between plants and flower visitors are diverse and flexible, seed production of many plant species is restricted by pollen limitation. Obligate outcrossers often suffer from low pollinator activity or severe interspecific competition for pollinator acquisition among co-flowering species. This study focused on seasonal changes in plant–flower visitor linkages in an alpine ecosystem and examined whether and how this seasonality affected the seed-set of Primula modesta, a self-incompatible distylous herb having long-tubed flowers. First, we recorded the linkages between plants and flower visitors along the snowmelt gradient. Then, pollination experiment was conducted to estimate the degree of pollen limitation over the course of flowering season of P. modesta. Flower visitors were classified by their tongue length based on the morphological matching with P. modesta flowers. As the season progressed, plant–visitor linkages became more diverse and generalized, and the visitation frequency to P. modesta flowers increased. In the later part of the season, however, the seed set of P. modesta was significantly reduced due to severe pollen limitation, presumably because of increased competition for long-tongued pollinators among co-flowering species. The present study revealed that pollinator availability for specialist species may be restricted even when plant–visitor linkages are diverse and generalized as a whole. In the case of P. modesta, morphological matching and competition for pollinators might be the main factors explaining this discrepancy.     

Parental effects and gender specialization in a tropical heterostylous shrub

Abstract.— Male sterility in hermaphroditic species may represent the first step in the evolution toward dioecy. However, gender specialization will not proceed unless the male-sterile individuals compensate for fitness lost through the male function with an increase in fitness through the female function. In the distylous shrub Erythroxylum havanense , thrum plants are partially male-sterile. Using data collected throughout eight years, we investigated whether thrum individuals have an increased performance as female parents, thereby compensating for their loss of male fitness. We found that thrum plants outperformed pins in the probabilities of seed maturation and germination and long-term growth of the seedlings. In turn, pollen from pin plants achieved greater pollen tube growth rates. Our results suggest that the superior performance of the progeny of thrum maternal plants is a consequence of better seed provisioning via effects of the maternal environment, cytotype or nuclear genes. Overall, our results suggest that E. havanense is evolving toward a dioecious state through a gynodioecious intermediate stage. This evolutionary pathway is characterized by an unusual pattern of gender dimorphism with thrums becoming females and pins becoming males. We propose that this pattern may be better explained by the interaction between male-sterility cytoplasmic genes and the heterostyly supergene.     

Characterization of microsatellite loci in Primula modesta Bisset et Moore (Primulaceae)

Eleven microsatellite loci were isolated from alpine plant Primula modesta. Enriched repeat libraries were constructed and screened by colony hybridization. Forty‐eight polymerase chain reaction (PCR) primer pairs were designed, of which 11 pairs producted successful amplification. A total of 31–35 adult individuals were genotyped for allelic diversity. The polymorphism ranged from three to 14 alleles, and the heterozygosity ranged from 0.161 to 0.828. No linkage disequilibrium was found between these 11 loci.     

Pollen flow in a population of Primula vulgaris Huds.

A British population of Primula vulgaris was visited four times between 1971 and 1976 for study of the nature and functioning of distyly. The stainability and production of pollen from pins exceeded those of thrums. About 20% of pin pollen and 48% of thrum pollen was removed from anthers by insect visitors. Pins and thrums were present in a 1:1 ratio in 1971 but pins outnumbered thrums in 1976. Thrums produced more seeds per flower than pins though the number of ovules produced by each was similar. Two methods for collecting stigmas from open flowers and analyzing them for pollen loans produced somewhat different results. For pin stigmas, the pollen load consisted of 2–23% thrum pollen; for thrum stigmas, the pollen load consisted of 0–71% pin pollen with most stigmas having less than 50% pin pollen. In general, intermorph pollen flow is less than would be expected if pollen flow were random. It is probable that most intramorph pollen on stigmas is a result of self- or geitonogamous pollination. The extensive literature concerning the natural pollinators of the primrose is reviewed. Although Darwin's hypothesis concerning the functional significance of distyly in promoting intermorph pollination was never quantified, the pollen flow patterns observed in P. vulgaris are unexpectedly deviant and are similar to those patterns observed in several unrelated heterostylous species in other plant families.     

Female self-incompatibility type in heterostylous Primula is determined by the brassinosteroid-inactivating cytochrome P450 CYP734A50

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Skewed morph ratios lead to lower genetic diversity of the heterostylous Primula veris in fragmented grasslands

• Populations of heterostylous plant species are ideally composed of equal frequencies of two (distylous) or three (tristylous) morphologically different floral morphs. Intra-morph incompatibility helps to avoid inbreeding and to maintain genetic diversity, supporting plant fitness and long-term viability. Habitat fragmentation can lead to skewed morph ratios and thereby reduce the abundance of compatible mates. This, in turn, can result in a loss of genetic diversity. We tested whether the genetic diversity of heterostylous plants is affected by morph ratio bias using populations of the distylous grassland plant Primula veris in recently fragmented grasslands.
• We recorded morph frequencies and population sizes in 30 study populations of P. veris on two Estonian islands characterised by different degrees of habitat fragmentation. Examining variation of thousands of single nucleotide polymorphisms (SNPs) and heterostyly-specific genetic markers, we quantified overall and morph-specific genetic diversity and differentiation in these populations.
• Morph frequencies deviated more in smaller populations. Skewed morph ratios had a negative effect on the genetic diversity of P. veris in more fragmented grasslands. In the populations of better-connected grassland systems, genetic differentiation among S-morphs was higher than among L-morphs.
• Our study shows that deviations from morph balance are stronger in small populations and have a negative impact on the genetic diversity of the distylous plant P. veris. Together with the direct negative effects of habitat loss and decreased population size on the genetic diversity of plants, morph ratio bias may intensify the process of genetic erosion, thus exacerbating the local extinction of heterostylous species.

     

Factors limiting seed production of Taxus brevifolia (Taxaceae) in Western Oregon

Seed production of Pacific yew (Taxus brevifolia), an understory conifer, was studied at four sites in western Oregon over 2 yr. The effects of pollen supplementation, overstory canopy, and predator exclusion on ovule attrition were examined. Supplemental hand-pollination of ten trees at two sites resulted in significantly increased rates of ovule development and a doubling of seed efficiency (ratio of seeds to ovules). However, seed efficiency still averaged <15% on branches receiving supplemental pollen, so pollination was not a primary factor limiting seed production. The number of developing ovules was positively associated with overstory openness, but seed production was not. Seed efficiency was negatively associated with overstory openness. Branches bagged to exclude vertebrate seed predators had higher seed production than unbagged branches at three of four sites for 2 yr. In contrast to unbagged branches, seed production on bagged branches was positively associated with overstory openness, as was the effectiveness of bagging. Therefore, both vertebrate predation and overstory were important in limiting seed production, and these factors interacted. Factors limiting seed production varied in importance among the four sites and between years, illustrating the importance of examining multiple limiting factors over several sites and years.     

Floral morph composition and pollen limitation in the seed set of <Emphasis Type="Italic">Nymphoides indica</Emphasis> populations

We investigated the floral morph composition and seed set of the populations in a heterostylous aquatic plant, Nymphoides indica O. Kuntze (Menyanthaceae). The bias of floral morph ratio in the populations was negatively correlated with the seed set. In the populations with biased floral morph ratio, the pollen on the stigma of the common morphs were dominated by pollen of the same morph, resulting in reduced seed set. The addition of pollen of the opposite morph on the stigmas of the common morph resulted in an increase of fruit and seed set rates. The intermorph pollination rate and the fruit and seed set rates of Nymphoides indica were highest when two floral morphs were close in the populations. Various insect species including Lepidoptera, Diptera and Hymenoptera visited the flowers. However, geitonogamous pollination was common because of the short flying distance of the pollinators. These results indicate that the seed set of Nymphoides indica is limited by the shortage of compatible pollens and that the success of intermorph pollination is influenced by spatial distribution of the two floral morphs. As Nymphoides indica is endangered, conservation of the populations with high fecundity is proposed by us.     

ZmRPN1 confers quantitative variation in pollen number and boosts hybrid seed production in maize

The number of pollen grains is a critical determinant of reproductive success in seed plants and varies among species and individuals. However, in contrast with many mutant-screening studies relevant to anther and pollen development, the natural genetic basis for variations in pollen number remains largely unexplored. To address this issue, we carried out a genome-wide association study in maize, ultimately revealing that a large presence/absence variation in the promoter region of ZmRPN1 alters its expression level and thereby contributes to pollen number variation. Molecular analyses showed that ZmRPN1 interacts with ZmMSP1, which is known as a germline cell number regulator, and facilitates ZmMSP1 localization to the plasma membrane. Importantly, ZmRPN1 dysfunction resulted in a substantial increase in pollen number, consequently boosting seed production by increasing female–male planting ratio. Together, our findings uncover a key gene controlling pollen number, and therefore, modulation of ZmRPN1 expression could be efficiently used to develop elite pollinators for modern hybrid maize breeding.     

Species Biology of Primula sieboldii for the Conservation of its Lowland-habitat Population: I. Inter-clonal Variations in the Flowering Phenology, Pollen Load and Female Fertility Components

Abstract Intra-population variations of several characteristics related to fertility were assessed for a nature-reserve population of Primula sieboldii in the flood plain of the Arakawa River. Although there were large intra- and inter-clonal variations of fruit and seed sets as well as other female fertility components in the 13 clones examined, female fertility was generally low, except in a homomorph clone. Pollinator availability was assessed by continuous monitoring of insect visits with 8 mm film. No insect visits were recorded during the monitoring of total 68 flowers over 16 hours in the nature-reserve population. In contrast, a high pollinator visitation rate was recorded in a reference population of P. sieboldii on the flank of Mt. Yatugatake. Thus, the low rates of seed production in the nature reserve was due to a shortage of compatible pollens as a result of the limitation of pollinator availability. Stigmas of flowers in various clones occasionally were loaded with pollen grains whose sizes were not significantly different from those of anther pollens of the same flowers. However, pollen germination on stigmas was observed only in the flowers of a homo-morph clone whose flowers have pin-type stigmas and anthers with compatible thrum-type pollens at the same height. Exceptionally high fertility of the homo-morph clone would be associated with its inbreeding habit.     

Factors affecting fruit and seed production in Dactylorhixa maculata (Orchidaceae)

It has been argued that some of resources needed for fruit and seed production in terrestrial orchids originate from storage in underground biomass. Resources for female reproductive traits may also originate from current photosynthesis. Orchid mycorrhiza may also influence fruit and seed production. The extent to which current photosynthetic activity and nutrient uptake via mycorrhizal fungi affect fruit and seed production has only rarely been studied experimentally. This paper examines the importance of photosynthesis and mycorrhiza for fruit and seed production, leaf area and plant status in the next season for Dactylorhiza maculata (L.) Soó in two populations. Plants were hand-pollinated in two populations to assure successful fertilization. Photosynthesis in the main leaves and in the green parts of inflorescence was prevented and/or mycorrhizal function inhibited by applying thiabendazole to plants. None of the treatments affected relative capsule production, but all treatments resulted in significant decreases in capsule dry weight and the number of seeds produced. The fact that seed production remained high in spite of the experimental treatments indicates that resources stored in the underground corms were used in fruit development and seed production. Inhibition of photosynthesis decreased leaf area in the year following treatments, while the other treatments had no effect on subsequent growth.     

新疆植被物候时空变化特征

基于MODIS-NDVI数据,提取新疆2001—2016年典型植被物候期,分析新疆不同生态分区的山地-绿洲系统植被物候期的时空演变趋势和空间分异特征,并结合同期气象数据,探讨植被物候与气候变化的响应关系。结论为:(1)新疆植被物候具有明显的纬向分布和垂直地带性分布特征,海拔在物候的地域分异中扮演着重要作用。新疆植被生长季开始时间(Start of season,SOS)集中于3月中旬至5月上旬,生长季结束时间(End of season,EOS)集中于10月中旬至12月下旬。(2)与全球大背景下典型植被物候特征变化趋势相反,新疆植被SOS呈推迟趋势,推迟幅度为1.9d/10a;EOS呈提前趋势,提前幅度为3.66d/10a;生长季长度(Length of season,LEN)呈缩短趋势,缩短幅度为5.6d/10a。除东疆地区外,全疆及不同分区均呈现出绿洲及平原SOS较早,山地区域较迟;全疆及不同分区均呈现出山地EOS结束较早,绿洲结束较迟;除东疆地区外,全疆及不同分区的LEN均为绿洲及平原区域山地,同样显示出垂直地带性分布的特征。(3)通过冗余分析(Redundancy analysis,RDA)解释了物候特征与气象因子关系的绝大部分信息,生长季开始时间受春季气温、前一年冬季降水量和日照时数的显著影响。夏季和秋季降水量是新疆植被生长季结束时间的重要影响因素,在总体上受气温和日照时数的影响较小。     

Reproductive ecology in the endemic Primula apennina Widmer (Primulaceae)

Abstract

Disassortative mating in distylous self-incompatible species should result in the equilibrium of morph types in natural populations. Deviation from isoplethy may affect pollen transfer, and in isolated populations it could lead to Allee effect and genetic drift. Pollen limitation has been found to occur in several distylous species, for which mating opportunities are actually reduced to half population. In this study, we investigated the reproductive features and pollination ecology of the narrow endemic Primula apennina. We recorded equilibrium of morph frequencies in the studied population, reflecting the comparable fecundity found in the two morphs. Long-styled flowers produce more pollen grains of smaller size than short-styled ones: we hypothesize that in thrum flowers, pollen is more easily removed by the insect pollinator Macroglossum stellatarum, resulting in equal pollen amounts carried to both short styles and long styles. This lower pollen transfer efficiency from long-styled to short-styled flowers is also reflected in legitimate pollen–ovule ratio values. Despite results show no evidence of imminent threats to population persistence at study site, the strict dependence on one or very few pollinator species, and ecological traits, may increase extinction risks in the long-term period.     

Male-biased sex allocation in late-blooming flowers driven by resource limitation in the clonal perennial Aconitum kusnezoffii (Ranunculaceae)

Flowering phenology and clonal growth are known to affect resource and pollen availability, and therefore select for adaptive or constrained sex allocation strategies to some degree. However, the consequences of temporal sex allocation patterns for reproductive fitness across the flower, inflorescence, and genet levels have rarely been examined. Moreover, experimental tests of the underlying regulatory mechanisms are scarce. We examined the association of flowering phenology and inflorescence position with temporal sex allocation and reproductive success in the protandrous perennial clonal herb, Aconitum kusnezoffii, over four consecutive growing seasons by examining more than 39 000 flowers. We also conducted controlled experiments to test the effects of resource and pollen limitation on the female reproductive success of lateral inflorescences. We found that some male functions were positively correlated with flowering phenology, whereas female reproductive success was negatively correlated with flowering phenology and inflorescence position. Lateral inflorescences invested more in male function than terminal inflorescences and therefore yielded fewer and smaller seeds. Resource limitation may serve as the key mechanism underlying this differentiated pattern. Decreased female reproductive success was consistently observed at the flower and inflorescence levels as flowering occurred later in the growth season. Late-blooming lateral inflorescences specialized in the male function, and their female reproductive success was constrained by early-blooming terminal inflorescences. This might be the first attempt to systematically demonstrate sex allocation strategy differentiation in a protandrous plant species at the inflorescence level. In addition, our study provides empirical evidence of dichogamy selecting for specialized sex allocation strategies among inflorescences.