蓝花丹结实率低的传粉生物学和繁育系统初探

为解释蓝花丹自然结实率低的原因,该研究从传粉生物学和繁育系统入手,采用TTC法与联苯胺-过氧化氢法分别测定花粉活力和柱头可授性的动态变化;用花粉-胚珠比(P/O比)、杂交指数(OCI)估算蓝花丹繁育系统类型并通过人工控制授粉试验验证。结果表明:(1)蓝花丹L型雌器官与S型雄器官、L型雄器官与L型雌器官成熟时间重叠区域较多,雌雄性器官在成熟时间上无显著差异;而S型雌器官与L型雄器官、S型雌器官与S型雄器官成熟时间重叠区域较少,但蓝花丹持续开花的模式缓解了雌雄性器官时间差异而引起的生殖隔离。(2)蓝花丹L型花P/O为502.00±52.30,S型花P/O为482.70±87.91,OCI为4,综合判断蓝花丹的繁育系统属于专性异交型且具有异型自交不亲和的特性。综上可初步解释蓝花丹自然结实率低的原因为内外因素共同作用的结果,其中雌雄性器官同熟时间短不是主要原因,而其本身较强的自交不亲和性可能是自然结实率低下的关键性内因。由于蓝花丹的专性异交繁育系统使其成功授粉需要传粉媒介,但因引种地与原产地环境存在显著差异,同时开花模式不集中导致的缺乏传粉者完成异花授粉或许是引起异交成功率降低的主要外因。因此,提高蓝花丹的结实率应从克服其自身自交不亲和性以及适当引入安全传媒昆虫入手。     

异型花柱植物喀什补血草的传粉者功能群与花粉转移效率

异型花柱是受遗传控制的花柱多态现象, 被达尔文认为是植物通过在传粉者体表不同部位滞落花粉以促进型间花粉准确传递的一种适应。该现象虽已受到广泛关注, 但在一些花型变异较大且不稳定的传粉系统中, 不同传粉者对各花型繁殖所产生的影响仍知之甚少。该研究以分布于新疆天山南坡的一个有同长花柱共存的异型花柱植物喀什补血草(Limonium kaschgaricum)种群为研究对象, 对其花型构成及频率、传粉者及花粉转移效率等进行了调查分析。结果表明: 1)种群中除了存在雌/雄蕊长度交互对应的长(L)/短(S)花柱型花外, 还有雌/雄蕊同长的花(H型), 且各花型花的花冠口直径、花冠筒长及花粉量等参数间无差异, 但花粉纹饰和柱头乳突细胞形态具二型性。其中, H型花的花粉和柱头形态与L型花(或S型花)的一致。2)花型内和自花授粉均不亲和; 型间授粉时, 花粉和柱头形态不同的花型间亲和, 反之不亲和。3)种群内存在长/短吻两类传粉昆虫。在以短吻传粉者为主的盛花初、中期, L和H型花柱头上的异型花粉数均显著高于S型花的, 且L和S型花高位性器官间的异型花粉传递效率高于低位性器官间的; 而在以长吻传粉者为主的盛花后期, L和S型花的柱头间异型花粉数无显著差异, 且高/低位性器官间具有相同的异型花粉转移效率; 与传粉者出现时期相对应的、在花期不同阶段开放花的结实率也明显不同。4)长/短吻昆虫具明显不同的传粉功能, 短吻昆虫只能对L和H型花进行有效传粉, 且访花频率和型间花粉转移效率较低, 为低效传粉者; 而长吻昆虫对各花型均能有效传粉, 具高的访花频率和型间花粉转移效率, 为高效传粉者。因为长吻昆虫的阶段性出现所形成的不稳定传粉系统, 使低效的短吻昆虫可能会成为种群中花型变异的驱动力, 并使S型花受到更大的选择压力。H型花克服了柱头缩入的弊端, 可能会成为不稳定传粉系统下的一个替代花型而持续存在。     

欧报春二型花柱的繁育特性研究

二型花柱植物通常具有自交不亲和性,在不同植物中所表现的可育性不同。欧报春(Primula vulgaris)是典型的二型花柱植物,为了探究欧报春的繁育特性,通过温室栽培,对欧报春的长花柱和短花柱的花部特征、花粉和花柱形态、花粉活力、柱头可授性、花粉/胚珠比、杂交指数、杂交亲和性、花粉管观察进行研究。结果表明,(1)长花柱和短花柱的花冠直径、花冠筒长度、柱头高度、花药高度和花筒中部直径均表现出两型性;(2)花粉和柱头观察发现花粉极轴长、花粉赤道轴长、花柱直径、柱头乳突细胞和花粉数量均有差异;(3)长花柱的花粉/胚珠比为384.20,短花柱为369.70,属于兼性异交类型;长花柱和短花柱的花粉活力和柱头可授性能在较长时间内维持较高活力;(4)长花柱杂交指数值为5,短花柱为4,表明繁育系统类型为异交,部分自交亲和,需要传粉者;6种授粉组合均能结实,异型花间授粉的结实数明显高于同型自花授粉和异株同型授粉;短花柱为母本的异型花间授粉组合亲和性优于其他组合;欧报春存在自交不亲和性,长花柱的自交不亲和性低于短花柱。     

濒危植物蒙古扁桃花粉活力和柱头可授性研究

用过氧化氢酶法测定蒙古扁桃花粉的活力和寿命,用联苯胺-过氧化氢法测定柱头可授性。结果表明,蒙古扁桃花粉活力在散粉后第2天最高,3d后活力迅速下降,三种类型的植株的花粉活力下降速度存在差异,其活力可持续30d左右。短柱头植株柱头在开花后1d可分泌黏液,长柱头和中柱头花植株的柱头在开花后2d才分泌黏液,短柱头分泌黏液的持续时间较后者长。柱头可授性持续时间同种之间也存在差异,长柱头和中柱头植株可达6d,而短柱头花植株可持续10d左右。     

束花石斛的繁育生物学特性

为了探究束花石斛（Dendrobium chrysanthum）的繁育生物学特性,对其开花习性、花粉活力、柱头分泌特性、人工授粉结实率、自然结实种子育性和贮藏特性进行研究,结果表明：（1）束花石斛花芽萌动期为19 d,现蕾期为23 d,群体花期为28 d,单花寿命10~15 d;（2）开花1~3 d花粉活力保持在90%左右,开花至第9天时花粉活力仅为23.7%,初始3 d开花的花粉活力与其他时间取样的活力差异均达极显著水平（P<0.01）,花粉活力随开花时间延长呈由强到弱的趋势;（3）束花石斛的柱头腔从开花第2天开始有分泌物,第6 d时分泌物最多,柱头过氧化氢染色结果表明在开花5~6 d时柱头可授性最强,柱头可授性随开花时间延长呈由弱到强再到弱的趋势;（4）选取花朵开放4 d的花粉和柱头人工授粉,同株异花和同花授粉未见结实。野外结实果质量2.02~4.83 g,种子长0.628~0.845 mm,宽0.082~0.110 mm,长宽比为7.798,成胚率达97.35%,种子活力为90.80%,室温条件保存6个月的种子活力维持在73.20%。上述结果说明,束花石斛单花花期不长,开花初期花粉活力最高,柱头发育要滞后于花粉的发育;束花石斛不存在无融和生殖,人工同株自花授粉不育。但自然状况下能正常结实,种子发育良好,胚活力较高,干燥种子在自然条件下有一定的耐贮特性。本研究揭示了束花石斛的开花生物学特性和雌雄蕊异熟特点,其交配系统应为自交不亲和,为其野生种质资源的保育和人工精准授粉提供科学依据。     

刺五加花粉活力和柱头可授性的研究

用TTC法测定了刺五加花粉的活力和寿命,用联苯胺一过氧化氢法、开花后套袋法测定了其柱头可授期,用扫描电子显微镜观察了刺五加柱头的亚显微形态特征。结果表明,刺五加长花丝植株的花粉活力比中花丝植株的高;花粉活力及其下降快慢随生境不同而有所差异;通常情况下,花粉寿命约4天。刺五加长花丝植株的柱头一直不具有可授性;中花丝植株和短花丝植株的柱头,因所处生镜不同,在开花后5-7、6-8、或7-9天具可授性;生境越郁闭,柱头具可授性的时间越晚。具可授性的柱头有独特的形态特征,即：柱头外翻、5或4裂、湿性、乳突细胞呈猫耳朵状。     

青篱柴通过促进亲和花粉生长而提高传粉精确性

异型花柱植物的适应意义在于提高亲和花粉的传递, 从而促进异交, 但是有不少研究发现柱头上落置了大量不亲和花粉。目前, 很少有人研究柱头是否有利于亲和花粉的萌发和花粉管伸长, 以及通过去雄处理排除花内和植株内不亲和花粉干扰从而验证二型花柱植物是否可以促进异交。本研究以亚麻科青篱柴属(Tirpitzia)的二型花柱植物青篱柴(T. sinensis)为研究对象, 分居群调查了青篱柴长、短柱型的植株数量, 测量不同柱型的花粉与柱头大小以及观察其表面纹饰, 测定不同开花时间长、短柱型的柱头活性与花粉活力, 统计自然状态下柱头上落置的不同表型花粉比例, 进一步统计花内、株内去雄和自然对照下柱头上落置的不同表型花粉比例, 计算型间、型内和混合授粉花粉管长度和花粉萌发率, 人工授粉检测其是否型内异交不亲和与自交不亲和。结果表明, 自然居群中长、短柱型青篱柴植株数量没有显著性差异; 短柱型青篱柴花粉体积显著大于长柱型, 且二者花粉表面纹饰不同, 但不同柱型的柱头表面积与表面纹饰无差异; 青篱柴开花第一天的花粉活力显著高于开花第二天, 而柱头活性在开花前两天无显著性差异; 自然状态下柱头上落置的不亲和花粉比例显著高于亲和花粉; 在长柱型青篱柴中, 花内和株内去雄显著提高了柱头上亲和花粉的落置比例; 柱头上授型内花粉, 其花粉萌发率和花粉管的长度都显著低于授型间花粉; 授粉结果表明青篱柴为型内异交不亲和与自交不亲和。二型花柱植物通过促进亲和花粉的萌发和花粉管的伸长而有利于型间授粉, 从而显著提高其传粉精确性。     

南蛇藤花粉和柱头的生物学特性研究

以南蛇藤花粉和柱头为研究对象,用日立TM-1000扫描电镜(SEM)观察花粉和柱头形态;用TTC(2,3,5-triphenyl tetrazolium chloride)法测定不同贮藏温度对雄花花粉活力的影响;用固体培养基萌发法测定不同温度下雄花花粉的萌发率。结果表明:南蛇藤花粉粒呈长球形,赤道观细长椭圆形,极面观近三裂圆形,具三孔沟,网状雕纹。南蛇藤雌花、两性花的柱头形态及不同发育期的变化规律相似,而雄花柱头形态明显区别于雌花和两性花的。不同的贮藏温度对雄花花粉活力的影响差异明显,贮藏温度越高,花粉失活越快,说明低温利于花粉贮藏。南蛇藤雄花花粉萌发的最适温度约25℃。     

花柱和柱头的结构

雌蕊由柱头、花柱和子房３部分组成。柱头是接受花粉的表面,通常将柱头分为２种类型：干砂和湿柱头。干柱头在传粉时没有分泌物的形成,表达干燥并覆盖有一层蛋白质薄膜;湿柱头在传粉时是湿润的,表面被一层分必物所覆盖,有皮细胞人有腺细胞的特征。花柱也被分为２种类型：开放型和闭合型。开放型花柱即中空花柱,花柱中央为花柱道,花柱道周围分布有一层特殊的通道细胞;闭合型花柱即实心花柱,也有２种类型,一类其中央有引导     

蓝花丹二型花雌、雄蕊发育进程及内源激素对花柱生长的影响

为了解二型花雌、雄蕊发育进程及内源激素对长、短花柱生长发育的影响,以蓝花丹(Plumbago auriculata Lam.)为材料,观察分析了长花柱(L型)、短花柱(S型)花朵内雌、雄蕊的发育特征,并分别检测了L、S型花柱中的内源激素水平。结果显示:蓝花丹雌、雄蕊发育进程基本符合逻辑斯蒂变化曲线,并可划分为5个时期,即T1初始发育期、T2转折期(一)、T3快速发育期、T4转折期(二)、T5平稳发育期;在整个发育进程中,L型花朵中雌蕊的生长速率始终高于雄蕊;S型花朵中雌蕊的生长速率在T3期由快转慢,导致T3~T5期雌蕊的生长速率始终低于雄蕊,从而形成了雌蕊低于雄蕊的短花柱特征。这说明花柱的分化是在二型花雌、雄蕊快速发育的T3期开始出现,并逐渐形成花柱异长植物最显著的花部形态特征。IAA、IPA和GA含量均在T1~T3期增加、T4~T5期降低,且在L型花柱中的含量始终高于S型,而ABA含量的变化趋势与这3种生长促进类激素相反,说明在蓝花丹花柱发育过程中,IAA、IPA和GA可能参与调控花柱的伸长生长,而ABA主要在发育后期促使花柱成熟。     

Functional dioecy in <Emphasis Type="Italic">Morinda parvifolia</Emphasis> (Rubiaceae), a species with stigma-height dimorphism

The evolution of dioecy from heterostyly has been well documented, but detailed studies on this transitional process are rare. Here we report the occurrence of cryptic dioecy in a perennial liana species with stigma-height dimorphism, Morinda parvifolia Bartl. ex DC. (Rubiaceae). Floral morphology, ancillary characters and cross compatibility of long-styled (L-morph) and short-styled (S-morph) were examined. L-morph and S-morph display obvious pistil dimorphisms, with the stigma of S-morph lacking papillae cells. Both floral morphs show similar pollen morphology, although pollen viability is higher in S-morph than in L-morph. S-morph flowers produce viable pollen grains but much reduced stigma and set no fruits, functioning as males; L-morphs, although with viable pollen grains and receptive stigmas, exhibit strong self- and intramorph incompatibility, with self- and intramorph pollen tubes arrested in the stigma lobes and the upper part of style, respectively, resulting in L-morphs functioning only as females. The species thus has physiological androdioecy but functional dioecy. This might be the first case showing the possibility that androdioecy could be a mid-stage in the pathway of dioecy evolving from stigma-height dimorphism.     

茜草科二型花柱植物滇丁香传粉精确性研究

异型花柱是一种受遗传因素控制的花型多态性现象,其适应意义在于提高不同花型间传粉精确性,从而促进异交。为检验二型花柱植物滇丁香（Luculia pinceana）是如何促进异交并确定有效传粉者的类型,实验统计自然种群中长、短柱型的植株数量,测量花部特征,统计自然种群植株柱头上的型间花粉落置的数量和比例,比较型内、型间花粉的萌发率以及花粉管长度,比较不同人工授粉处理结实情况,观察传粉昆虫类型并测量虫体特征。结果表明（1）滇丁香自然居群中长、短柱型植株的数量没有显著性差异。长柱型的筒深、筒直径、开口直径、雄蕊长和柱头长均显著小于短柱型,而长柱型的叶片长和宽、雌蕊长、花药体积、花粉粒体积、柱头厚均显著大于短柱型。（2）自然种群植株柱头上型间花粉所占比例显著低于型内花粉的比例。（3）型间授粉的花粉萌发率以及花粉管长度均大于型内授粉和混合授粉。（4）型间授粉的结实率显著高于型内授粉的结实率,型内授粉具有一定的结实。（5）夜行性长喙条背天蛾（Cechenena lineosa）是滇丁香的有效传粉者,其吻长和滇丁香的花筒深相适应。本研究表明二型花柱植物滇丁香主要是通过与其花部特征相匹配的长喙天蛾实现有效的传粉,通过促进型间花粉的萌发和花粉管的伸长而有利于型间授粉结实,提高传粉精确性。     

Pollinator shifts along an elevation gradient mediate different response in self-pollination in heterostylous Primula nivalis

Pollinator shift and its influence on floral traits have been well documented to explain the diversity of angiosperms, but such effects are still less known at the intraspecific level, especially the responses of different morphs of distylous plants. We hypothesized that the pollen transfer efficiency would decrease if the pollinator shifted from a long-tongued to short-tongued insect across populations, and plants would evolve towards selfing in response to the stressed pollination environment. Given the gender specialization between flower morphs, the long styled (L-morph) plants would increase female reproduction. Our study showed that the short-tongued Bombus tianshanicus, the most-visit pollinator in high elevation populations of distylous Primula nivalis, was less efficient in pollen transfer than long-tongued Bombylius major. The plants evolved to promote selfing through reducing the anther–stigma separation and increasing intramorph self-compatibility. The hand pollination experiment showed that after intramorph selfing, the fruit set and seed set increased with increasing elevation. Moreover, anther and stigma were closer in the L-morph than in the short styled (S-morph) plants, and the L-morphs showed higher pollen transfer efficiency than the S-morphs. Along with increased self-compatibility, the fruit set and seed set of L-morph plants were significantly higher than those of S-morph plants. We described the pollinator shifts along an elevation gradient in a distylous plant and the response of plants by promoting selfing, which confirmed our hypothesis and supported the pollinator-shift model. Our study also highlighted the different response in self-compatibility between flower morphs.     

Shortened anther-stigma distance reduces compatible pollination in two distylous Primula species

Aims Distyly has been regarded as an adaptation to improve compatible pollination between two floral morphs with reciprocal herkogamy. The hypothesis that the different positions of anthers and stigmas within flowers as well as their reciprocal position between morphs, reduce the probability of self pollination raised by Darwin has been rarely tested. In this study, we measured stigmatic pollen loads in response to reduced reciprocal herkogamy in two Primula species.Methods To see whether reciprocal herkogamy can increase compatible and/or reduce incompatible pollen deposition, thus promoting compatible pollination, we shortened the distance between anthers and stigmas within the flowers by changing the position of the corolla tube, to which the anthers were fused, i.e. reduced herkogamy in natural populations of Primula secundiflora and P. poissonii and quantified stigmatic pollen loads in the field over 2 years.Important findings In both species, stigmatic pollen loads were significantly higher in the long-styled (L-morph) than in the short-styled morph (S-morph) in both control and manipulated flowers, but percentage of compatible pollen in S-morph were higher. Flowers manipulated to halve the anther–stigma distance showed a similar pattern for 2 years: total pollen grain counts on stigmas did not differ significantly but compatible pollen grains in L- and S-morphs were significantly decreased in both species. The percentage of compatible pollen loads was decreased by 68.7% in P. secundiflora and 65.3% in P. poissonii in L-morphs, while it decreased by 30.6% and 2.9% in S-morphs, respectively. Our manipulation of the relative position of anthers and stigmas in the two distylous species indicated that a lower degree of herkogamy reduced compatible but incompatible pollen transfer was likely to increase. The higher proportion of compatible pollen in the S-morph than in the L-morph in the two Primula species could be attributed to the accessibility of two-level sexual organs, floral orientations and pollinator behaviors. This is a first attempt to manipulate intraflower herkogamy for understanding adaptation of heterostyly, shedding insights into how the reciprocal herkogamy promotes compatible pollination.     

Floral morphology and breeding system in Polygonum hastato-sagittatum Mak. (Polygonaceae)

Distyly has been clearly confirmed in Polygonum hastato-sagittatum Mak, which is dimorphic for the intrinsic features of style height, anther height and a suite of ancillary pollen and stigma polymorphisms. The stigma width of long styled flowers is larger than that of short ones, and L-morph (long) stigmas have longer and less number of papillae than the S-morphs (short). Flowers produced significantly smaller pollen grains, but in greater numbers, in L-morph than in S-morph. In P. hastato-sagittatum, style-morph ratios were associated with population size: larger populations always exhibited 1:1 morph ratios, or were slightly L-biased, whereas smaller populations were often strongly L-biased, and only occasionally S-biased. Populations containing only the L-morph were occasionally found, but populations fixed for the S-morph were not observed. The biased style-morph ratios maybe result from morph-specific differences in assortative mating or artificial disturbance. The study supports the theory that evolution of heterostyly reduces the conflict that can occur in sexually monomorphic animal-pollinated species, achieving efficient cross-pollination but simultaneously avoiding self-interference between female and male sexual organs.     

Morph-ratio variation, population size and female reproductive success in distylous Pulmonaria officinalis (Boraginaceae)

Theory predicts that morph ratios in heterostylous populations are governed by negative frequency-dependent selection typically resulting in equal morph ratios at equilibrium. Previous work on the distylous perennial herb Pulmonaria officinalis, however, showed asymmetric mating between floral morphs and a weak self-incompatibility system, with the long-styled morph (L-morph) producing significantly higher seed set following intramorph crosses and even selfing than the short-styled morph (S-morph), two aspects thought to affect female fecundity and morph-ratio variation. Here, we evaluated morph ratios and population size of all known P. officinalis populations in the northern part of Belgium. Morph ratios deviated significantly from 1:1 (range 0.09-1 L-morph frequency, mean = 0.58). Relative fecundity of the S-morph (i.e. mean seed set of the S-morph/mean seed set of the L-morph) was on average 0.73, was positively related to the frequency of the L-morph, and reached 1 (similar levels of female fecundity) at an average L-morph frequency of 0.66 in the population. As some small populations had the S-morph in majority, our results suggest that local morph ratios are influenced both by the relative fecundity of L- and S-morph individuals and by stochastic processes in small populations.     

An experimental evaluation of self-interference in Narcissus assoanus: functional and evolutionary implications

Floral traits that reduce self-pollination in hermaphroditic plants have usually been interpreted as mechanisms that limit the genetic consequences of self-fertilization. However, the avoidance of sexual conflict between female and male function (self-interference) may also represent an important selection pressure for the evolution of floral traits, particularly in self-incompatible species. Here, we use experimental manipulations to investigate self-interference in Narcissus assoanus, a self-incompatible species with a stigma-height dimorphism in which the degree of spatial separation between sex organs (herkogamy) differs strikingly between the long- and short-styled morphs (hereafter L- and S-morphs). We predicted that weak herkogamy in the L-morph would cause greater self-pollination and hence self-interference. Experimental self-pollination reduced seed set when it occurred prior to, or simultaneously with, cross-pollination in the L-morph, but only if it occurred prior to cross-pollination in the S-morph. In the field, autonomous self-pollination was greater in the L-morph than the S-morph, but we found no evidence that self-interference reduced maternal or paternal fitness in either morph. One-day-old flowers of the L-morph have reduced stigma receptivity and hence exhibit protandry, whereas stigma receptivity and anther dehiscence are concurrent in the S-morph. This suggests that the two style morphs have alternative strategies for reducing self-interference: dichogamy in the L-morph and herkogamy in the S-morph. These results provide insight into the mechanisms that reduce sexual conflict in hermaphrodite plants and are of significance for understanding the evolution and maintenance of sexual polymorphisms.     

Cryptic dioecy in Mussaenda pubescens (Rubiaceae): a species with stigma-height dimorphism

Background and Aims

Evolutionary transitions from heterostyly to dioecy have been proposed in several angiosperm families, particularly in Rubiaceae. These transitions involve the spread of male and female sterility mutations resulting in modifications to the gender of ancestral hermaphrodites. Despite sustained interest in the gender strategies of plants, the structural and developmental bases for transitions in sexual systems are poorly understood.

Methods

Here, floral morphology, patterns of fertility, pollen-tube growth and floral development are investigated in two populations of the scandent shrub Mussaenda pubescens (Rubiaceae), native to southern China, by means of experimental and open-pollinations, light microscopy, fluorescence microscopy and scanning electron microscopy combined with paraffin sectioning.

Key Results

Mussaenda pubescens has perfect (hermaphroditic) flowers and populations with two style-length morphs but only weak differentiation in anther position (stigma-height dimorphism). Experimental pollinations demonstrated that despite morphological hermaphroditism, the species is functionally dioecious. The long-styled (L) morph possesses sterile pollen and functions as a female, whereas the short-styled (S) morph is female sterile and functions as a male. Self- and intra-morph pollinations of the S-morph were consistent with those expected from dimorphic incompatibility. The two populations investigated were both S-morph (male) biased. Investigations of early stages of floral development indicated patterns typical of hermaphroditic flowers, with no significant differences in organ growth between the floral morphs. Meiosis of microspore mother cells was of the simultaneous type with tetrads isobilateral in shape. The tapetal cells in anther walls of the L-morph became vacuolized during meiosis I, ahead of the uninucleate microspore stage in the S-morph. In the L-morph, the microspore nucleus degenerated at the tetrad stage resulting in male sterility. Microsporogenesis and male gametophyte development was normal in the S-morph. Failure in the formation of megaspore mother cells and/or the development of megagametophytes resulted in female sterility in the S-morph, compared with normal megasporogenesis in the L-morph.

Conclusions

In M. pubescens, cryptic dioecy has evolved from stigma-height dimorphism as a result of morph-specific sterility mutations.