248
Relationships between flowering phenology and female reproductive success in the Japanese tree species Magnolia stellata Suzuki Setsuko, Ichiro Tamaki, Kiyoshi Ishida, and Nobuhiro Tomaru
Abstract: We have examined the earliness, duration, and amplitude of flowering genets in a Magnolia stellata (Sieb. et Zucc.) Maxim. population in relation to their size, environmental factors (temperature and light), and female reproductive success (ovule survival rate) over three consecutive years. Average flowering durations of individual flowers, genets, and the whole population in these 3 years were 10.2, 15.2, and 29.0 d, respectively. A bisexual phase (with both female and male phase flowers) in genets spanned 62.9% of the total flowering period, suggesting that geitonogamy can occur. The earliness, duration, and amplitude of flowering genets were all significantly correlated, indicating that genets flower early and long periods have high flowering amplitudes. The three parameters were also significantly correlated with the size of the genets (represented by the diameter at breast height of its thickest ramet) and relative photosynthetic photon flux density at the top of their crowns. Therefore, genets that are large and located in well-lit sites tend to have many flowers, and blossom both earlier and longer. Later-flowering genets have higher female reproductive success, probably because M. stellata is protogynous. Significantly positive correlation between flowering amplitude and female reproductive success suggests that large numbers of flowers increase the attractiveness of genets for pollinators, and this outweighs the negative effects of geitonogamy. Key words: geitonogamy, photoenvironment, plant size, protogyny, seed production, star magnolia. Re´sume´ : Les auteurs ont examine´ la pre´cocite´, la dure´e et l’ampleur des genets floraux, dans des populations de Magnolia stellata (Sieb. et Zucc.) Maxim., en relation avec leur dimension, des facteurs environnementaux (tempe´rature, lumie`re), et le succe`s reproductif femelle (taux de survie des ovules), au cours de trois anne´es conse´cutives. On cours de ces trois saisons, on a observe´ des dure´es de floraisons moyennes des fleurs individuelles, des genets et des populations dans leur ensemble, de 10,2, 15,2 et 29,0, respectivement. Chez les genets, une phase bisexue´e (phase de floraison a` la fois maˆle et femelle) s’e´tale sur 62.9 % de la dure´e totale de la pe´riode de floraison, sugge´rant la pre´sence possible de geitonogamie. Il y a une corre´lation significative entre la pre´cocite´, la dure´e et l’amplitude des genets floraux, ce qui indique que les genets fleurissent toˆt et les longues pe´riodes ont de grandes amplitudes florales. On observe e´galement une corre´lation significative entre les trois parame`tres et la dimension des genets (repre´sente´e par le diame`tre a` hauteur de poitrine de ses rame`tes les larges) et la densite´ relative du flux de protons photosynthe´tique au sommet de leurs houppiers. Conse´quemment, les genets de fortes dimensions se retrouvant dans des sites bien e´claire´s ont tendance a` posse´der plusieurs fleurs, avec des boutons plus haˆtifs et de plus longue dure´e. Les genets fleurissant plus tard montrent un meilleur succe`s reproductif femelle, probablement parce que le M. stellata est protogyne. La corre´lation positive entre l’amplitude de floraison et le succe`s reproductif femelle sugge`re que le grand nombre de fleurs augmente l’attirance des genets pour les pollinisateurs, et que ceci contrebalance les effets ne´gatifs de la geitonogamie. Mots-cle´s : geitonogamie, photoenvironnement, dimension de la plante, protogynie, production de semences, magnolia e´toile´. [Traduit par la Re´daction]
Introduction Phenology is defined as the characterization of the seasonal timing of life cycle events (Rathcke and Lacey 1985). Optimizing the timing of these events is important for the Received 18 June 2007. Published on the NRC Research Press Web site at botany.nrc.ca on 29 February 2008. S. Setsuko,1 I. Tamaki, and N. Tomaru. Laboratory of Forest Ecology and Physiology, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan. K. Ishida. Kansai Research Center, Forestry and Forest Products Research Institute, Momoyama, Fushimi, Kyoto 612-855, Japan. 1Corresponding
author (e-mail:
[email protected]).
Botany 86: 248–258 (2008)
survival, growth, and reproduction of many organisms, but especially for those such as established plants that are unable to move to more favorable habitats in adverse conditions. Flowering phenology is one of the most important life-history traits that affect the reproductive success of plants, for several reasons. For instance, plants flowering asynchronously may fail to produce seeds, since no pollen from potential mates may be available when their flowers are receptive. In addition, flowering phenology is often correlated with both abiotic factors, such as temperature (Smithberg and Weiser 1968; Vasek and Sauer 1971), moisture (Opler et al. 1976; Reich and Borchert 1982), and light conditions (Osunkoya 1999), as well as biotic factors, such as the availability of pollinators (Robertson 1895; Schemske
doi:10.1139/B07-135
#
2008 NRC Canada
Setsuko et al.
et al. 1978; Chan and Appanah 1980). The size of the plants can also affect their flowering and fruiting phenology (Ollerton and Lack 1998; McIntosh 2002) because large plants with ample resources are not only able to produce more flowers and fruits than smaller plants, but they can flower over longer periods of the year (Schmitt 1983). Thus, these (and other) environmental variables and traits of plants have both quantitative and phenological effects on their flower production and, hence, reproductive success. However, these environmental variables and plant size may be correlated (e.g., the light condition is likely to be correlated with temperature, and plant size may be correlated with the light conditions, because plants in well-lit sites may have higher growth rates than those in shaded sites), and often have confounding effects on the relationships between flowering phenology and reproductive success. These confounding effects may lead to misinterpretation of the results of phenological investigations, as pointed out by Mauricio and Mojonnier (1997). Therefore, it is important to clarify the relationship between these variables, and then examine the relationships between flowering phenology and reproductive success. This can provide valuable information regarding the biotic and abiotic conditions required for the successful production and maturation of fruits and seeds. The species in the genus Magnolia are basal angiosperms that retain many primitive morphological characters (Takhtajan 1980; Thien et al. 2000). The floral biology of this genus is also considered to be primitive: the flowers generally do not secrete nectar (Thien 1974; Yasukawa et al. 1992) and they are protogynous; a common trait in hermaphrodite flowers of primitive woody angiosperms (Godley and Smith 1981; Bernhardt and Thien 1987). In addition, their pollination depends primarily on beetles (Thien 1974; Bernhardt and Thien 1987; Kikuzawa and Mizui 1990; Ishida 1996), which are generally considered to be less effective pollinators than bees (Ramsey 1988). Accordingly, they have several adaptive features, including protogyny and the opening of tepals during the day [observed in eight North American Magnolia species (Thien 1974) and the Japanese species Magnolia obovata Thunb. (Kikuzawa and Mizui 1990)] that are putatively highly specialized characteristics that encourage exclusive pollination by beetles (Thien 1974). In addition, certain Japanese and Mexican Magnolia species (such as M. obovata and Magnolia tamaulipana A. Va´zquez, respectively) are known to emit copious floral scent (Azuma et al. 1997), which is recognized as an important mechanism whereby basal angiosperms attract pollinators (Thien et al. 1975; Azuma et al. 1997; Thien et al. 2000). Although there are several published studies on flowering phenology of Magnolia species, these studies make no reference to reproductive success (e.g., Kikuzawa and Mizui 1990; Ishida 1996; Dieringer et al. 1999). Star magnolia, Magnolia stellata (Sieb. et Zucc.) Maxim. (Magnoliaceae), is a threatened tree species endemic to the Tokai region of central Japan (Environment Agency of Japan 2000). This species has several biologically interesting floral and flowering characteristics. Flowers of M. stellata appear before leaf emergence in early spring, when the pollinator availability is low. In common with other members of the genus, the species is protogynous and produces no nectar-like secretions during the flowering season, suggest-
249
ing that the female phase flowers of M. stellata have no rewards for pollinators, although the male phase flowers release pollen, which can provide rewards for pollinators. Unlike some congenerics (see above) it emits extremely small amounts of floral scent (Azuma et al. 1997), also suggesting very small attraction for pollinators. In addition, flowering times of individual M. stellata flowers are not completely synchronized within a tree, and thus male phase flowers and female phase flowers coexist at the same time (see Results). Clearly, the flowers produced by the species have a number of characteristics that are regarded as being generally unfavorable, so it is of interest to identify flowering patterns that may increase their reproductive success to increase our fundamental understanding of links between phenological and other flowering parameters and reproductive success. We investigated flowering phenology at individual flower, genet, and population levels, and the flowering amplitude of genets, in a population of M. stellata over three consecutive years. The data acquired were then used to evaluate the relationships linking the flowering phenology and amplitude of genets to both their size and environmental factors (temperature and light). Finally, we examined the relationships between female reproductive success and flowering phenology, amplitude, size, and light environment.
Materials and methods Study species and study site Magnolia stellata (synonym, Magnolia tomentosa Thunb.; Ueda 1986; Nooteboom 1994) is a deciduous, clone-forming (via sprouting and layering) tree of the Magnoliaceae. The species occurs in nutrient-poor wetlands at elevations of 40–700 m a.s.l. in the Tokai region of central Japan (Japan Association for Shidekobushi Conservation 1996). The trees blossom in early spring, and form protogynous, insect-pollinated flowers that are 7–10 cm in diameter, with 12–18 white to slightly pink tepals (Fig. 1A). The main flower visitors are small beetles belonging to Coleoptera (S. Setsuko, unpublished data, 2003). The species is self-compatible and no evidence exists of a common self-incompatibility system (Hirayama et al. 2007). The fruits are aggregated, with at most two red seeds per follicle, and the seeds are considered to be dispersed by birds (Callaway 1994). This study was conducted within the Yato River watershed (9.3 ha) in the Kaisho Forest, near Nagoya city, Aichi Prefecture, Japan (35811’ 25@N, 137806’ 55@E). This site supports a secondary forest that is mainly composed of Pinus densiflora Sieb. et Zucc., Ilex pedunculosa Miq., Quercus serrata Murray, and Clethra barbinervis Sieb. et Zucc. (Tamaki et al. 2005). The potential natural vegetation of this area is evergreen broad-leaved forest, but this forest is now mainly dominated by deciduous trees because humans utilized the trees for fuel wood over a long period of time in the past. However, since the use of this wood for fuel stopped, the abundance of the evergreen broad-leaved trees has been increasing via secondary succession (Prefecture of Aichi 1998). Field observations of flowering phenology In a previous study, we measured the spatial coordinates #
2008 NRC Canada
250
Botany Vol. 86, 2008
Fig. 1. Flowering phases of an individual flower of Magnolia stellata. (A) An individual flower, (B–D) close-ups illustrating the female phase, transitionary phase, and male phase of an individual flower, respectively.
and sizes of all ramets of M. stellata within the watershed population, and identified 175 genets by examining the connections between ramets aboveground in conjunction with microsatellite analysis (Setsuko et al. 2004). To clarify the flowering phenology of M. stellata, flowering events were observed at two levels: genets and individual flowers. To monitor the flowering phenology at the genet level, the flowering status (flowering or nonflowering) of all genets within the population was recorded every 3 d during each of the 2003, 2004, and 2005 flowering seasons. The numbers of open flowers in flowering genets were also counted at the same time. To monitor the flowering phenology at the individual flower level, five genets that had sufficient number of flowers for us to be able to observe the individual flowers and that also could be reached using a ladder, were selected. The individual flowers were tagged both in 2003 and 2004 to observe the conditions of their stigmas and anthers, and to determine the sexual phase of the flowers. The flowering pattern of individual flowers of M. stellata is as follows. The flower buds gradually swell until the hirsute bracts split
and are shed, following this, tepals appear. At this time, the stigmas are curved and are believed to be receptive, while the stamens are tightly appressed against the androphore, and we defined this floral stage as ‘‘the female phase’’ (Fig. 1B). Following this phase, the stigmas straighten and become oriented in a relatively parallel direction to the gynoecium, while the stamens diverge and start to release pollen. We defined this floral stage as ‘‘the male phase’’ (Fig. 1D). However, in considerable proportions of flowers the stigmas had straightened while the stamens remained appressed to the axis for a variable period between the female and male phases. In such cases, the phase of the flower could not be classified as either female or male (Fig. 1C), and was denoted transitionary. Finally, most of the stamens and tepals detach from the axis and flowering ceases. In both flowering seasons, all flowers of four genets (designated Y83, Y137, Y145, and Y150) were tagged, but only the flowers on one ramet in the remaining genet (Y56, which bore about a third of all flowers of the genet) were tagged, since this genet produced too many flowers to observe them all. #
2008 NRC Canada
Setsuko et al.
251
Fig. 2. Flowering rates of M. stellata genets every 3 d (lower panels) and daily mean temperature during the flowering season (upper panels) in each of the three study years. Open circles and filled squares indicate the flowering rates and the cumulative flowering rates of genets on the census dates, respectively. Filled circles indicate the flowering rates of genets that were observed to have flowered for the first time on the census dates.
In 2003, 2004, and 2005, 11 genets, (including the 5 mentioned above that were selected for the flowering phenology observations at the individual flower level in 2003 and 2004) were selected, and the fruit set, seed set per fruit, and ovule survival rate were estimated at the genet level. To do this the numbers of floral buds present before flowering were counted, then all the mature fruits were sampled in the summer. Finally, the numbers of carpels and filled seeds in the sampled fruits were counted. Environmental surveys An automatic thermometer (TR-51A, T & D Corporation, Matsumoto, Nagano, Japan) was installed in a shaded position, 1 m aboveground, at the study site to record temperatures once every hour throughout the flowering periods in each of the years 2003, 2004, and 2005. In June and July 2004, the photosynthetic photon flux density (PPFD) was measured both at the top of the crowns of 79 randomly sampled flowering genets and, simultaneously, at an open site using an LI-1400 datalogger (LI-COR). The relative photosynthetic photon flux density (rPPFD) at the top of each selected genet’s crown was then calculated as follows: rPPFD (%) = (PPFD at the top of genet’s crown) / (PPFD at the open site) 100. Data analyses To describe the flowering phenology of each genet, we used the two parameters (i) earliness of flowering (the date when flowering started) and (ii) flowering duration. In addition, we used the maximum number of open flowers of each genet on any day during the flowering period each year as a measure of its flowering amplitude (hereinafter simply referred to as flowering amplitude). As a measure of the size of each genet, the diameter at breast height of its thickest ramet (hereinafter DBHt) was used. The number of ovules in the sampled fruits was estimated by multiplying the number of carpels by two, because each carpel always contains two ovules. The percentages of fruit set and seed set per fruit were calculated for each genet in each year from the equations½ðnumber of fruits = number of flowersÞ 100, and ½ðnumber of filled seeds = number of ovulesÞ 100, respectively. In addition, as a measure of female reproductive success, the percentage of each genet’s ovules that developed into filled seeds was calculated, using the following equation. ovule survival rate = (fruit set) (seed set per fruit) 100 To investigate how the flowering phenology, amplitude, light condition, and size of the genets affect the female reproductive success, we used generalized linear mixed models (GLMMs) in which year was treated as a random effect.
In the analysis, fixed effect explanatory variables were initially the flowering earliness, duration, and amplitude of the focal genet, the flowering amplitude of neighboring genets (i.e., the sum of the maximum numbers of open flowers of each neighboring genet, defined as those within a certain ra#
2008 NRC Canada
252
Botany Vol. 86, 2008
Fig. 3. Flowering phenology of M. stellata genets in each of the 3 years. Each horizontal line represents the flowering duration of one of the genets, arranged in order of earliness and duration of flowering; a dot indicates the date when the number of open flowers on the genet per flowering day was maximal; and arrowhead brackets enclose the period during which the number of open flowers on the genet per flowering day was greater than half of the maximum number of open flowers for that genet on any day during the flowering period. The numbers following ‘‘Y’’ indicate the ID numbers of the genets for which the flowering phenology of individual flowers was also observed.
dius of the focal genet, on any day during the flowering period each year), and the rPPFD and DBHt of the focal genet. However, a multicollinearity problem that might have detrimental effects on the estimated regression parameters occurred between the flowering earliness and duration. Therefore, we omitted the flowering duration from the analysis. We then regressed the flowering earliness and amplitude of the focal genet, the flowering amplitude of neighboring genets, and rPPFD and DBHt of the focal genet against the focal genets’ female reproductive success parameter (ovule survival rates), using GLMMs with a binomial error distribution and logit-link function. Parameters of the model were estimated by the maximum likelihood method using R version 2.4.0 (The R Development Core Team
2003). To determine the most appropriate radius for neighboring genet, radii with 5 m increments in the range 5– 25 m were tested in the GLMMs, and the Akaike information criterion (AIC) was calculated for each of the models obtained. The AIC value was lowest when the radius was 5 m, so we set the radius to this distance for all further analyses.
Results Flowering phenology and amplitude of genets Magnolia stellata genets began to flower in late March or early April (Fig. 2). Of the 175 genets examined: 77, 76, and 78, (mean 77.0) flowered in the years 2003, 2004, and #
2008 NRC Canada
Setsuko et al.
253 Table 1. Spearman’s coefficients of rank correlation between earliness, duration, and amplitude for Magnonlia stellata genets flowering in the three study years.
Flowering duration
Flowering amplitude
Year 2003 2004 2005 2003 2004 2005
Flowering earliness –0.929 P
