Selfed Pistils Research Articles

Reproductive systems and low outbreeding barriers between Jacaranda cuspidifolia and J. mimosifolia (Jacarandeae, Bignoniaceae)

The genus Jacaranda shows notable karyotype stability and a prevailing self‐sterile breeding system with evidence of late‐acting self‐incompatibility in several species. However, some studies have indicated self‐compatibility in J. mimosifolia, a species cultivated worldwide in tropical and subtropical areas. Jacaranda cuspidifolia is a closely related species with natural distribution broadly overlapping with that of J. mimosifolia, and manual heterospecific pollination studies have indicated that these species are interfertile, but there is no report on the breeding system of the former. In this study, we used hand‐pollination experiments, pistil longevity, epi‐fluorescence and histological analysis of post‐pollination events to determine the breeding system of J. cuspidifolia. We also employed intra and interspecific crosses and seed germination tests to reevaluate the breeding system of J. mimosifolia and the inter‐fertility between the two species. Some fruits were initiated from self‐pollinated pistils in J. mimosifolia, but none of them reached maturity. On the other hand, complete absence of fruit development by self‐pollination was verified in J. cuspidifolia, while in situ pollen tube growth and histological analysis of post‐pollination events in selfed pistils revealed the characteristic ovule penetration, fertilization and endosperm initiation also observed in other bignoniaceous species with late‐acting self‐incompatibility. Low outbreeding barriers seem to operate between these species because reciprocal interspecific crosses and hybrid seed germination tests indicated they are bilaterally interfertile. However, fruit/seed production and seed germinability were significantly lower when pistils of J. cuspidifolia were pollinated with pollen of J. mimosifolia, compared with crosses in the opposite direction, which indicates a partial unilateral incompatibility. This result is discussed in the context of the possible occurrence of self‐compatibility in J. mimosifolia. The low level of incongruity operating between the two species also points to their recent evolutionary divergence.

Evidence for post-zygotic self-incompatibility in Handroanthus impetiginosus (Bignoniaceae).

Late-acting self-incompatibility (LSI) has been defined as a genetically controlled self-sterility mechanism that prevents seed set by selfing, despite normal pollen tube growth and ovule penetration in self-pollinated pistils. In species of the Bignoniaceae with LSI, such as Handroanthus impetiginosus, the selfed pistils are characterized by a marked delay in ovule penetration, fertilization, and endosperm initiation, followed by uniform pistil abscission. This highlights the contentious possibility of a post-zygotic self-incompatibility system. However, previous studies were unable to confirm fusion of the sperm and egg cell nuclei in selfed ovules. In the present study, the cytology of the embryo sac, double fertilization, and pistil longevity was investigated in H. impetiginosus using comparative nuclei microspectrofluorometry of DAPI-stained sections of self- vs. unpollinated pistils. Differences in both pistil longevity and ovary size between self- and unpollinated flowers at the time of pistil abscission were significant. Zygotes with double the DNA content in their nuclei relative to unfertilized egg cell nuclei were verified in selfed ovules from the first day after pollination onward, and G1 karyogamy appeared to have occurred. Our cytological analysis clearly indicates that ovules of self-pollinated pistils in H. impetiginosus are fertilized before pistil abscission but no embryogenesis initiation occurs, which strongly supports the idea of a post-zygotic self-incompatibility mechanism.

Sporophytic apomixis in polyembryonic Handroanthus serratifolius (Vahl) S.O. Grose (Bignoniaceae) characterizes the species as an agamic polyploid complex

Neopolyploidy has been associated with gametophytic apomixis and breakdown of gametophytic self-incompatibility. Nevertheless, Bignoniaceae presents agamic polyploid complexes with neopolyploidy associated to sporophytic apomixis. Apomictic populations are commonly polyploid, polyembryonic and self-fertile, while diploids are mostly late-acting self-incompatible (LSI) and monoembryonic. Contrastingly, Handroanthus serratifolius shows hexaploid monoembryonic and polyembryonic populations, although breeding system has been studied only for monoembryonic individuals, which are LSI. Our aim here was to investigate breeding system and early embryology in polyembryonic individuals of H. serratifolius to define if they form an agamic polyploid complex. Experimental pollinations and histological analyzes of ovules and young seeds were carried out. Megasporogenesis and megagametogenesis occurred as in other sexual species of Bignoniaceae. The polyembryonic individuals were self-fertile and double fertilization was observed both after self and cross-pollinated pistils. Adventitious embryos originated from the hypostasis and integument of the ovule, indicating sporophytic apomixis. Adventitious embryo precursor cells occurred in all pistils, including unpollinated ones. But unpollinated pistils aborted possibly due to absence of endosperm, and pollination was required for fruit-set (pseudogamy). It is possible that the self-fertility in polyembryonic individuals ensues as the initial endosperm of self-fertilized ovules supply early adventitious embryos development, and these embryos would later prevent the abortion of selfed pistils. The sporophytic apomixis in polyembryonic populations and the occurrence of sexual monoembryonic populations of H. serratifolius allows us to consider the species part of an agamic polyploid complex. But in contrast with other Handroanthus agamic complexes, both apomictic and sexual LSI plants were hexaploid.

Ovarian self-incompatibility inNarcissus papyraceus(Amaryllidaceae) is the result of a pre-zygotic response

Ovarian self-incompatibility, including pre- and post-zygotic reactions, is a complex mechanism for which we still lack many details relating to its function and significance. The joint presence of ovarian self-incompatibility with style polymorphism is a rare combination that is found in the genus Narcissus. Usually, style polymorphic species have heteromorphic (diallelic and linked to style length locus) incompatibility, which prevents fertilization between individuals of the same morph, thereby helping to maintain equal proportions of floral morphs in populations. However, when present, self-incompatibility in Narcissus is not linked to style polymorphism and cross-fertilization within each morph is possible. Hence, self-incompatibility in Narcissus is of particular interest when attempting to unravel the nature of the rejection reaction and aiming to assess possible cryptic differences in the fertilization process in intra- and inter-morph crosses, which might ultimately explain the wide variation of morph-ratio in the field. We examined the breeding system of Narcissus papyraceus, a style-dimorphic species that has biased morph ratios in most of its populations. We studied pollen-tube growth in the pistil and ovule fate after experimentally controlled hand pollinations. The growth of pollen tubes in self- and intra- and inter-morph crosses was similar up to the point of micropyle penetration in both morphs but, subsequently, a pre-zygotic failure appeared to affect male and female gametophytes in selfed pistils. A high proportion of ovules from self-pollinated flowers showed signs of collapse and self-pollen tubes were blocked or behaved abnormally before entering the embryo sac. Self-incompatibility was stronger in the long-styled morph than in the short-styled morph. We did not find any conclusive sign of differential functioning between intra- and inter-morph cross-pollinations in any morph. These results enable us to rule out the possible effects of pollen–pistil interactions in N. papyraceus as a cause of morph-ratio biases and confirm the exceptional nature of the self-incompatibility mechanism in this polymorphic species.

Contrasting breeding systems in six species of Tillandsia L. (Bromeliaceae) from woody areas of Santa Fe Province: Argentina

The reproductive biology of six native Tillandsia species that co-occurred in woody areas of Santa Fe Province (Argentina) was studied for floral biology, breeding systems and reproductive success. Flower longevity, floral phases, hand self- and cross-pollinations or natural pollinations and fluorescence microscopy observations of post-pollination events were analysed. Also pollen/ovule (P/O) ratio and pre-emergent reproductive success (PERS) were compared between species. In T. aëranthos, T. ixioides and T. meridionalis, self-pollinated flowers mostly failed to form fruits, and fluorescence microscopy studies showed that in selfed pistils pollen tube growth was arrested in the style. It is likely that these species have homomorphic, gametophytic self-incompatibility. In contrast, T. capillaris, T. tricholepis and T. recurvata regularly formed fruits following autonomous self-pollination. Floral traits, the P/O ratios and PERS indices correlate with the breeding systems found in these two groups of species: the three selfing species have inconspicuous closed flowers, and they have low P/O ratios and high PERS values. In contrast, the three self-incompatible (SI) species have conspicuous flowers with relatively high P/O and low PERS values. We described here three new cases of SI in Tillandsia species subgenera Anoplophytum and confirm the autogamous and cleistogamous status in three species of Tillandsia subgenera Diaphoranthema.

The reproductive biology of Cybistax antisyphilitica (Bignoniaceae), a characteristic tree of the South American savannah-like “Cerrado” vegetation

Bignoniaceous woody species are very important ecological components of neotropical forests, but the reproductive biology of many species, such as Cybistax antisyphilitica, remains virtually unknown. Most species of Bignoniaceae are characteristically self-sterile, despite typically exhibiting normal pollen tube growth throughout the style, combined with slow rates of ovule penetration, fertilisation and endosperm initiation in selfed pistils. Uniform abortion occurs within a few days of anthesis, indicating the occurrence of late-acting self-incompatibility (LSI). However, breeding system studies have been performed in fewer than 7% of species, and other types of breeding systems (e.g., self-compatibility and apomixis) have been reported in this family. In the present study, the reproductive biology of C. antisyphilitica was investigated by field observation of flower visitors and floral events. Moreover, reproductive biology of this species was examined through experimental pollinations, analyses of pollen tube growth and ovule penetration using fluorescence microscopy, verification of pistil longevity, and a histological analysis of unpollinated vs. self-pollinated pistils. Finally, morphological aspects, quantities and germination were investigated in seeds that resulted from different pollination treatments. Natural pollination was effected by large- and medium-sized bees, and their visiting behaviour favours a high proportion of geitonogamy and no pollen limitation. Self-pollinated flowers produced no fruits, and all of the characteristic post-pollination events cited above were verified, witnessing the occurrence of LSI with post-zygotic rejection of selfed pistils in C. antisyphilitica. Although some indications of extended pistil longevity were found in selfed pistils, this feature seemed to be affected by unidentified environmental factors. The seeds were always monoembryonic and with high viability. A larger variation in the number of viable seeds was found in fruits derived from natural pollination. A low fruit set was observed after both natural and cross-pollination, and most crossed fruits underwent abortion at several points during the juvenile phase, even when protected against herbivory. The formation of surplus flowers/juvenile fruits and the apparently wasteful selfing mechanism control implied in LSI are discussed in the context of the perennial life style of tropical woody plant species.

Sistema reprodutivo do Ipê-Branco: Tabebuia roseo-alba (Ridley) Sandwith (Bignoniaceae)

Estudos sobre sistemas reprodutivos têm indicado o predomínio da autoincompatibilidade de ação tardia (AIT) em Bignoniaceae, embora poucas espécies tenham sido investigadas e ocorram outros tipos de sistemas reprodutivos na família. O presente estudo objetivou determinar o sistema reprodutivo de T. roseo-alba através de experimentos de polinizações controladas, análise histológica dos eventos posteriores à polinização, verificação do desenvolvimento in situ dos tubos polínicos e testes de germinação de sementes. Apesar de os tubos polínicos penetrarem e fecundarem a maioria dos óvulos em pistilos autopolinizados, o aborto de 100% dos mesmos foi verificado e, embora sua abscisão tenha ocorrido entre o quarto e o sexto dia após o início da antese, observou-se um ligeiro crescimento dos óvulos e do ovário precedendo a abscisão, porém inferior ao crescimento nos pistilos submetidos à polinização cruzada. A endospermogênese inicial e a formação do tubo proembriônico também foram mais lentas nos pistilos autopolinizados. A longevidade dos pistilos autopolinizados foi maior que a de pistilos não polinizados, e a taxa de germinação de sementes foi de 93%, sendo todas as sementes monoembriônicas. Os resultados demonstram que T. roseo-alba é espécie auto-estéril, destituída de poliembrionia e que apresenta AIT pós-zigótica.

Floral biology and late-acting self-incompatibility in Jacaranda racemosa (Bignoniaceae)

Breeding-system studies have been conducted with 38 of the approximately800 species of Bignoniaceae, and self-incompatibility was found in 31 of these. In species for which the site of self-incompatibility barrier was studied, self-pollinated flowers consistently failed to develop into fruits, even though pollen tubes grew down to the ovary and penetrated most of the ovules. In this study, we have investigated the floral biology and the breeding system in Jacaranda racemosa Chamisso, with hand-pollination experiments and the histology of post-pollination events. Flower anthesis lasted 1–3 days, and although the frequency of flower visitation was extremely low, natural pollination seemed to be effected mainly by medium-sized bees. Because the conspicuous staminodium favours eventual pollination by small bees, a possible role of the staminodium in the increase of potential pollinators is suggested. Hand-pollinations indicated that J. racemosa is a self-sterile species. Histological analysis of post-pollination events indicated the occurrence of a kind of late-acting self-incompatibility in which the processes of ovule penetration, fertilisation and endosperm initiation were slower in selfed than in crossed pistils. Until the time of self-pollinated pistil abscission, no signs of endosperm malfunction or proembryo development were observed in selfed pistils. Therefore, inbreeding depression is an unlikely explanation for self-sterility in J. racemosa.

Late‐Acting Self‐Incompatibility and Other Breeding Systems inTabebuia(Bignoniaceae)

Although breeding system investigations were previously performed in only nine of the 100 Tabebuia species, indications of self‐incompatibility have been found in all of them, and the four species studied for the site of incompatibility reaction showed some kind of late‐acting self‐incompatibility. Polyembryony has been found in T. chrysotricha and T. ochracea, with adventitious origin of the extra embryos being shown in the latter. We investigated the breeding system in five species of Tabebuia by hand‐pollination experiments, fluorescence microscopy study of in situ pollen tube growth, and histological analysis of postpollination events. Although both T. chrysotricha and T. heptaphylla developed fruits by self‐pollination, polyembryony was verified only in the former, which indicates that self‐fertility in Tabebuia is not necessarily associated with apomixis. The remaining species were 100% self‐sterile. Although some penetrated ovules in crossed pistils of T. vellosoi were found at the 48‐h interval, none of the ovules in selfed pistil was penetrated at the same interval, with penetration of many ovules occurring 72 h postpollination. In T. impetiginosa, ovule penetration efficiency 48 h after pollination was higher in cross‐ than in self‐pollinated pistils, and the majority of the ovules in selfed pistils were penetrated and fertilized in 72 h. The incidences of ovule fertilization and endosperm initiation were significantly slower in selfed compared with crossed pistils, and a clear developmental slowdown of the endosperm occurred in selfed pistils before abscission, although no other signs of developmental malfunctions were detected. In all of the self‐sterile species, abortion of selfed pistils occurred in a small period after pollination, and no swelling of the ovary was observed in T. umbellata. All of these results agree with the occurrence of late‐acting self‐incompatibility in T. impetiginosa, T. umbellata, and T. vellosoi, and no evidence was found that self‐sterility in these species is due to inbreeding depression.

Does Late-acting Self-incompatibility (LSI) Show Family Clustering? Two More Species of Bignoniaceae with LSI:Dolichandra cynanchoides and Tabebuia nodosa

By means of hand pollination experiments and fluorescence microscopy the breeding systems and pollen tube growth were studied in Dolichandra cynanchoides and Tabebuia nodosa, two bignoniaceous species of the Chaco woodland of NE Argentina. Both species abscised all selfed pistils within 4–8 d although self pollen tubes grew successfully to the ovary and penetrated many ovules. Both species therefore show ‘late-acting self-incompatibility’ (LSI) or ‘ovarian sterility’ (OS) type control of selfing. LSI-OS has been reported in 17 species of the Bignoniaceae, and is also clustered in families such as the Bombacaceae-Sterculiaceae, Fabaceae and Myrtaceae. The implications of such family clustering are discussed, with regard to current hypotheses for the action of LSI-OS phenomena.

Reproductive Biology ofDalbergia miscolobiumBenth. (Leguminosae-Papilionoideae) in SE Brazil: The Effects of Pistillate Sorting on Fruit-set

The breeding system ofDalbergia miscolobiumBenth. was investigated by means of controlled pollinations and studies of post-pollination events in pistils. This species does not have homomorphic self-incompatibility, but crossed flowers always have higher fruit set than selfs, despite the fact that embryo development may be initiated in selfed pistils. It is suggested that deleterious recessive alleles, which lead to abortion of most selfed seeds, together with some maternal resource limitation, result in low fruiting success.

Occurrence and Physiological Breakdown of Self-incompatibility in Easter Cactus

The existence of self-incompatibility (SI) was demonstrated in `Britton' and `Rose' Easter cactus (Rhipsalidopsis). In a full diallel cross among five clones, 18 out of 20 outcrosses resulted in 68% to 100% fruit set, whereas reciprocal crosses between two of the clones and all five self-pollinations failed to set fruit. Pollen tube growth was greatly inhibited in styles of selfed pistils, but there was no evidence of pollen tube inhibition in compatibly crossed pistils. Easter cactus exhibited characteristics typically found in sporophytic SI systems (trinucleate pollen, papillate stigmas, and scant stigmatic exudate) together with those associated with gametophytic SI systems (stylar inhibition of pollen tube growth and absence of reciprocal differences in outcrosses). Additional experiments were performed to determine the effects of bud pollinations, growth regulators (BA, GA3, and NAAm), and high temperatures (0- to 48-h exposure at 40C) on the SI response. Heat treatments were more effective than either bud pollinations or growth regulators in overcoming SI, and yielded an average of 7.2 viable seeds per treated flower when plants were incubated for 12 h at 40C and selfed immediately after incubation. Isozyme analysis of the S0 parent and putative S1 progeny confirmed that selfing had occurred following heat treatments. Using S1 progeny in breeding programs may extend the flower color range and lead to a greater diversity in other plant characteristics than presently exists in cultivated germplasm. Chemical names used: N-(phenylmethyl)-1H-purin-6-amine [benzyladenine (BA)], gibberellic acid (GA3), and α-naphthaleneacetamide (NAAm).

ニホンナシの和合及び不和合受粉後の花粉管の生長に伴うタンパク質の変動

The present study was performed to get a clue to a solution for the obfuscating mechanism with reference to self-incompatibility in Pyrus serotina.1. The growth of pollen tubes showed the slow rate from approximately 36 hours after pollination and stopped completely in 48 hours at the portion near the ovary in a selfed style, but the tubes in a crossed one continued their growth and fertilization occurred in about 96 hours after pollination.2. In the selfed style, the tips of pollen tubes began to swell in 36 hours after pollination and callose deposited abnormally on the surface of the tubes. However, in a crossed style, sharp tips of the tubes and no abnormality of callose deposition were observed at above time.3. Soluble proteins and proteins dissociated by Tween 20 in selfed pistils exhibited unchanged level till 48 hours after pollination and increased thereafter gradually. On the other hand, these in crossed pistils decreased leisurely up to 24 hours and displayed steep increase after that time.4. In the fraction of proteins dissociated by Tween 20, both electrophoretic patterns of proteins from selfed and crossed pistils exhibited little difference till 72 hours after pollination. However, in the fraction of soluble proteins, band 7 peak from the selfed pistils increased the height in 24 hours after pollination and then decreased. The band 7 from the crossed pistils displayed reverse patterns compared with that from the selfed pistils, that is to say, the band 7 decreased in 24 hours after pollination and then continued to increase gradually. Band 7 from the non-pollinated pistils showed intermediate patterns between above bands from selfed and crossed pistils.