Pollen–pistil interaction studies in mango (Mangifera indica L.) cultivars

The increasing intensity and frequency of extreme heat events threaten crop productivity globally. Certain phases of plant reproduction necessary for fertilization are highly sensitive to extreme heat, particularly during pollen development, germination, and tube elongation. However, few studies have assessed the effects of extreme heat on pollen performance in perennial crop plants. To understand how northern highbush blueberry pollen responds to high temperatures, we quantified pollen germination and pollen tube growth in vitro using four commercially relevant cultivars (Bluecrop, Elliott, Jersey, and Liberty) in climate-controlled chambers. We also tested recovery from high heat in ‘Bluecrop’ to determine whether pollen tubes can still germinate and grow after short bursts of extreme heat. We found the highest proportion of germinated pollen tetrads and the greatest pollen tube growth at 20 and 30 °C, and the lowest levels at 10 and 40 °C, with nearly complete inhibition at 40 °C. Exposure to between 30 and 40 °C revealed significant reduction in pollen germination and tube growth above 35 °C across all cultivars and assessment times. Exposure to 37.5 °C for only 4 hours resulted in substantial reductions in pollen germination and pollen tube growth, even after pollen was moved to optimal conditions of 25 °C. Extreme heat exposure, even for a short duration, significantly limits blueberry pollen germination and tube development. This is expected to have cascading effects on fruit set and crop yield. The nonreversibility of the effects on pollen highlights the need to prevent fields reaching damaging temperatures by developing crop monitoring and management strategies to protect crops during bloom.

Nitric oxide (NO) plays essential roles in many biotic and abiotic stresses in plant development procedures, including pollen tube growth. Here, effects of NO on cold stress inhibited pollen germination and tube growth in Camellia sinensis were investigated in vitro. The NO production, NO synthase (NOS)-like activity, cGMP content and proline (Pro) accumulation upon treatment with NO scavenger cPTIO, NOS inhibitor L-NNA, NO donor DEA NONOate, guanylate cyclase (GC) inhibitor ODQ or phosphodiesterase (PDE) inhibitor Viagra at 25°C (control) or 4°C were analyzed. Exposure to 4°C for 2 h reduced pollen germination and tube growth along with increase of NOS-like activity, NO production and cGMP content in pollen tubes. DEA NONOate treatment inhibited pollen germination and tube growth in a dose-dependent manner under control and reinforced the inhibition under cold stress, during which NO production and cGMP content promoted in pollen tubes. L-NNA and cPTIO markedly reduced the generation of NO induced by cold or NO donor along with partly reverse of cold- or NO donor-inhibited pollen germination and tube growth. Furthermore, ODQ reduced the cGMP content under cold stress and NO donor treatment in pollen tubes. Meanwhile, ODQ disrupted the reinforcement of NO donor on the inhibition of pollen germination and tube growth under cold condition. Additionally, Pro accumulation of pollen tubes was reduced by ODQ compared with that receiving NO donor under cold or control condition. Effects of cPTIO and L-NNA in improving cold-treated pollen germination and pollen tube growth could be lowered by Viagra. Moreover, the inhibitory effects of cPTIO and L-NNA on Pro accumulation were partly reversed by Viagra. These data suggest that NO production from NOS-like enzyme reaction decreased the cold-responsive pollen germination, inhibited tube growth and reduced Pro accumulation, partly via cGMP signaling pathway in C. sinensis.

Pollen germination, along with pollen tube growth, is an essential process for the reproduction of flowering plants. The germinating pollen with tip-growth characteristics provides an ideal model system for the study of cell growth and morphogenesis. As an essential step toward a detailed understanding of this important process, the objective of this study was to comprehensively analyze the transcriptome changes during pollen germination and pollen tube growth. Using Affymetrix Arabidopsis (Arabidopsis thaliana) ATH1 Genome Arrays, this study is, to our knowledge, the first to show the changes in the transcriptome from desiccated mature pollen grains to hydrated pollen grains and then to pollen tubes of Arabidopsis. The number of expressed genes, either for total expressed genes or for specifically expressed genes, increased significantly from desiccated mature pollen to hydrated pollen and again to growing pollen tubes, which is consistent with the finding that pollen germination and tube growth were significantly inhibited in vitro by a transcriptional inhibitor. The results of Gene Ontology analyses showed that expression of genes related to cell rescue, transcription, signal transduction, and cellular transport was significantly changed, especially for up-regulation, during pollen germination and tube growth. In particular, genes of the calmodulin/calmodulin-like protein, cation/hydrogen exchanger, and heat shock protein families showed the most significant changes during pollen germination and tube growth. These results demonstrate that the overall transcription of genes, both in the number of expressed genes and in the levels of transcription, was increased. Furthermore, the appearance of many novel transcripts during pollen germination as well as tube growth indicates that these newly expressed genes may function in this complex process.

SummaryThe effect of a constant (10, 15, 20 or 25°C) or a diurnal maximum/minimum (15/5, 20/10, 25/15 or 30/20°C) incubation temperature on in vitro pollen germination and pollen tube growth in the pistils of two poly-embryonic (‘Kensington’ and ‘Nam Dok Mai’), and two mono-embryonic (‘Irwin’ and ‘Sensation’) mango cultivars was studied. In in vitro experiments where pollen was incubated in a liquid germination medium for 24 h in darkness, little difference was found between pollen germination of mono- and poly-embryonic cultivars. Averaged over the four cultivars, 53.9% of pollen germinated at 10°C, this increased to 76.2% when the incubation temperature was increased to 15°C, thereafter up to 25°C the percentage germination remained stable but germination decreased slightly to 68.2% at 30°C. Similarly, there was no difference in percent germination between cultivars when pollen was incubated under diurnal temperature regimes. Mean pollen germination of all four cultivars was 52.3% at 15/5°C and pollen germination increased by 10% when the temperature was raised to 30/20°C. When self-pollinated flowers were incubated for 24 h on a semi-solid agar medium at 10°C, pollen tube growth of the four cultivars was retarded and no pollen tubes reached the ovaries. As the temperature was increased from 15 to 25°C, the mean number of pollen tubes in ovaries increased from 0.04 to 0.25. At 30°C, the mean number of pollen tubes that entered ovaries decreased to 0.04. After incubation under diurnal temperature regimes, the mean number of pollen tubes in ovaries of all four cultivars at 15/5°C was 0.23 and increased to 0.42 when the temperature increased to 30/20°C. At each incubation temperature, there were significant differences in pollen tube growth between cultivars, but there were no differences between the temperature response of pollen from mono- and poly-embryonic cultivars.

Although many studies have emphasized the importance of auxin in plant growth and development, the thorough understanding of its effect on pollen-pistil interactions is largely unknown. In this study, we investigated the role of free IAA in pollen-pistil interactions during pollen germination and tube growth in Nicotiana tabacum L. through using histo and subcellular immunolocalization with auxin monoclonal antibodies, quantification by HPLC and ELISA together with GUS staining in DR5::GUS-transformed plants. The results showed that free IAA in unpollinated styles was higher in the apical part and basal part than in the middle part, and it was more abundant in the transmitting tissue (TT). At the stage of pollen germination, IAA reached its highest content in the stigma and was mainly distributed in TT. After the pollen tubes entered the styles, the signal increased in the part where pollen tubes would enter and then rapidly declined in the part where pollen tubes had penetrated. Subcellular localization confirmed the presence of IAA in TT cells of stigmas and styles. Accordingly, a schematic diagram summarizes the changing pattern of free IAA level during flowering, pollination and pollen tube growth. Furthermore, we presented evidence that low concentration of exogenous IAA could, to a certain extent, facilitate in vitro pollen tube growth. These results suggest that IAA may be directly or indirectly involved in the pollen-pistil interactions. Additionally, some improvements of the IAA immunolocalization technique were made.

Pollination is an essential prerequisite for the production of many fruit and seed crops, including apple. In apple, successful fertilization requires pollen transfer to the stigma, pollen germination, and successful pollen tube growth resulting in fruit set. Precise selection of the most effective pollinizers for commercial orchards is not possible however, until these processes are more fully understood. The present study was undertaken to compare pollinizers in terms of pollen tube growth and fruit set. On trees of ‘Honeycrisp’, ‘Fuji’, and ‘Gala’ from which bees were excluded, flowers were hand-pollinated using pollen collected from crabapple (‘Ralph Shay’ or Malus floribunda ), ‘Delicious’ and ‘Golden Delicious’. Flowers were harvested at one, two, three, and four days after pollination (DAP). Pollen source had a significant influence on pollen germination on the stigmatic surface, number of pollen tubes penetrating the stigma, distance of pollen tube growth down the style, and pollen tubes reaching the base of the style. In ‘Honeycrisp’ and ‘Gala’, ‘Golden Delicious’ pollen grew the fastest, followed by ‘Delicious’ and crabapple. Neither ‘Ralph Shay’ nor Malus floribunda were effective pollinizers for ‘Honeycrisp’ and resulted in low fruit set suggesting incompatibility may be involved. However, both these crabapples were effective pollinizers for ‘Fuji’ and ‘Gala’. These results indicate that pollen source can have a tremendous impact on pollen tube growth and fruit set. The physiological basis for these effects is not clear, but implications for pollinizer selection are obvious.

SummaryThe timing of fungicide and antibiotic applications in fruit crops often overlaps flowering and pollination. Numerous studies report detrimental effects of chemical applications on pollination, fruit set and yield. However, field results can be quite variable even when similar compounds are tested on the same crop. Problematic is that field variables unrelated to the effects of pesticides on reproductive activities can mask results. In the current study, we specifically investigated the effects of selected pesticide sprays applied to stigmas, on subsequent pollen germination and tube growth. Using two model systems, apple and almond, sprays were applied to detached emasculated flowers under constant laboratory conditions using an electronically controlled robotic apparatus which simulated a field sprayer. flowers were pollinated at specific times after spraying, and pollen tube numbers and growth assessed using fluorescence microscopy. The pesticides evaluated were for apple: captan, myclobutanil and streptomycin; and for almond: azoxystrobin, myclobutanil, iprodione and cyprodinil. In apple, captan significantly reduced pollen germination, by 20% compared with water controls when pollination was conducted 18 h after treatment. The number of tubes that reached the basal part of the style within 20 h was not affected. In contrast, myclobutanil and streptomycin had no significant effect on pollen tube growth. With almond, none of the sprays significantly affected pollen germination or tube growth.

The inhibitory effects of benomyl on pollen tube growth have received little attention, particularly at the microscopic and immunohistochemical levels. Pollen germination and tube growth in the presence of benomyl were evaluated in Tradescantia virginiana to investigate the effects of this fungicide on pollen germination rate, tube growth and morphology, and microtubule (Mt) organization. Benomyl was incorporated in germination media at 0,480, 600 or 720 mg 1−1. Inhibition of pollen germination, cytoplasmic streaming, and tube elongation were associated with benomyl treatments. Benomyl also induced abnormal pollen tube morphology and Mt organization. Compared to controls, Mts in the treated tubes were characteristically fewer in number, fragmented, sinuous and increasingly disorganized. At the two highest benomyl concentrations, Mts were considerably fewer or absent in apical/subapical regions of the pollen tubes. This work verifies that benomyl incorporated into germination media at concentrations lower than recommended field rates inhibit pollen germination and tube growth, and that the effects are associated with alterations of Mt organization.

After nearly a decade of development, the scale of blueberry ( Vaccinium sp.) cultivation has increased, particularly in south China; however, this region is becoming increasingly challenged by temperature changes during the flowering phenophase. Understanding the effects of temperature on pollen germination and pollen tube growth in blueberry is thus important. Using the rabbiteye blueberry ( V. ashei ) ‘Brightwell’, different temperature treatments were carried out during open pollination and cross-pollination with the pollen from rabbiteye blueberry ‘Gardenblue’ in field, greenhouse, and controlled temperature experiments over two consecutive years. The differences in pollen germination, pollen tube dynamics, and ovule viability following different treatments were analyzed, and the critical temperatures were calculated using quadratic and modified bilinear equations to quantify the developmental responses to temperature. The results showed that the fruit set of the artificially pollinated plants inside the greenhouse was significantly higher than that outside the greenhouse. Furthermore, pollen germination and pollen tube growth gradually accelerated under the appropriate high-temperature range, resulting in reduced pollen tube travel time to the ovule. However, the percentage of the style traversed by the pollen tube did not increase at temperatures greater than 30 °C, and a high-temperature range could accelerate ovule degeneration. Therefore, impairment of pollen tube growth in the upper half of the style following pollen germination and ovule degeneration constituted important factors leading to reduced fruit setting under short periods of high temperature during the flowering phenophase in rabbiteye blueberry. This work advances our understanding of the effect of temperature on pollen germination, pollen tube growth, ovule longevity, and fruit setting in rabbiteye blueberry, and provides a foundation for continued cultivation and breeding enhancement. The findings propose that the tolerance of rabbiteye blueberry to a certain high-temperature range in the flowering phenophase should inform breeding strategies for temperature resistance and that temperature range is also an important indicator of suitable environments for cultivation to mitigate potential temperature stress.

The short-term effects of rare earth elements on pollen germination and tube growth were tested. Concentrations of 2.5 approximately 20 micro m lanthanum(La3+) or cerium (Ce3+)increased pollen germination and pollen tube growth, whereas concentrations higher than 40 micro m La3+ and Ce3+ inhibited this process. The most effective concentration of La3+ needed for promotion shifted from 10 to 40 micro m, depending on the Ca2+ concentration in the medium. Calmodulin (CaM) antagonist W7-agarose and anti-CaM antibody depressed La3+-promoted pollen germination and tube growth in a dose-dependent manner. La3+-CaM complexes (La3+-CaM) increased pollen germination and tube growth more than CaM or La3+ alone. Pertussis toxin (PTX) inhibited La3+-promoted pollen germination and tube growth. Cholera toxin (CTX) partially recovered the inhibition of the above La3+-promoted process by the anti-CaM antibody. Concentrations of 10-7 approximately 10-9 m La3+-CaM increased GTPase activity inside plasma membrane vesicles of the pollen tube, but apo-CaM or La3+ alone had no positive effects. The results suggest that apoplastic CaM may be involved in the promotion effects of lower concentrations of La3+ on pollen germination and tube growth, and the heterotrimeric G-protein on the plasma membrane may transduce La3+-activated CaM signalling. The present studies provide an apoplastic mechanism for short-term effects of rare earth elements at lower concentrations in the pollen system.

Pollen germination and pollen tube growth was quantified among various native Iranian wild almonds (P. dulcis (Mill.) D. A. Webb, P. eleaegnifolia Mill., P. orientalis Mill., P. lycioides Spach, P. reuteri Bioss. et Bushe, P. arabica Olivier, P. glauca Browick and P. scoparia Spach in order to identify differences in the tolerance of pollen to temperature variations. Pollen germination and pollen tube growth were observed after incubation in darkness in a germination medium for 24 h at 10–50°C at 5°C intervals. Maximum pollen germination of the wild almond species and specify that 60% was obtained for P. orientalis pollen and 98% for P. scoparia. Pollen tube length ranged from 860 μm was obtained in P. lycioides and 1490 μm in P. scoparia. A modified bilinear model best described the response to temperature of pollen germination and pollen tube length. Almond species variation was found for cardinal temperatures (Tmin, Topt and Tmax) of pollen germination percentage and pollen tube growth. Mean cardinal temperatures averaged over eight almond species were 14.7, 24.2, and 43.7°C for maximum percentage pollen germination and 14.48, 25.3, and 44.4 °C for maximum pollen tube length. The principal component analysis (PCA) identified maximum percentage pollen germination and pollen tube length of the species, and Tmax for the two processes as the most important pollen parameters in describing a species tolerance to high temperature. PCA also classified Prunus L. spp. into four groups according to the tolerance of pollen to temperature variations. The Tmin and Topt for pollen germination and tube growth, rate of pollen tube growth were less predictive in discriminating species for high temperature tolerance.

Hawthorn is one of the most important medicinal plants with more nutrients, medical, ornamental and sanitary properties and used as a main dry tolerant rootstock recently. Commercial part of the hawthorn is its fruits and the most important factors involved in fruit set are pollination, pollen tube growth and fertilization. This study was carried out to investigate pollen germination and tube growth of some East-Azerbaijan, Iran indigenous hawthorn genotypes in differentin-vitro media for receiving the suitable medium for pollen germination and identifying favourable genotypes for using in the future breeding and orchard establishment programs. Experiment accomplished based on completely randomized design (CRD) in four repeats. Pollens gathered from anthers and maintained at refrigerator until usage. In-vitro medium has different concentrations of sucrose (5, 10, 15 and 20%), boric acid (0.005, 0.01 and 0.02%) with (1.2%) agar. Pollens were cultured in different media and maintained about 24 h in 22ËšC and then, their growth was stopped with raising cholorophorm. Finally, pollen germination percentage and tube growth were measured under light-microscope in 7 randomized selected squares. Results showed that the best in-vitro medium for hawthorn pollen germination and tube growth were composed of 15% sucrose, 0.005 - 0.01% boric acid and 1.2% agar and significant differences observed between various media in the studied pollen traits.   Key words: Hawthorn, Crataegus, pollen germination, pollen tube growth, in vitro medium.

The Calcineurin B-Like Ca2+ Sensors CBL1 and CBL9 Function in Pollen Germination and Pollen Tube Growth in Arabidopsis

The process of pollen germination and tube growth in the pistil involves a series of cell-cell interactions, some facilitating fertilization while others prohibiting pollen tube access to the female gametophyte, either because of incompatibility or as a result of mechanisms to avert polyspermy and to ensure reproductive success. Understanding pollen tube growth and guidance to the female gametophyte has long been a pursuit among plant biologists, and observations indicate that diverse strategies may be adopted by different plant species. Recent studies in Arabidopsis, maize, and Torenia fournieri suggest that low molecular weight secretory molecules probably play major roles in the short-range attraction of pollen tubes to the female gametophyte. The process of pollen tube growth in the pistil occurs beneath several cell layers so much of the information that conveys the intimate partnership between penetrating pollen tubes and the female tissues has come from fixed samples and observations of in vitro pollen tube growth responses to female factors. A unique glimpse of the in vivo pollen germination and tube growth process is provided here by intra-vital two-photon excitation (TPE) microscopy of pollinated Arabidopsis pistils that remained on intact plants. Further discoveries of critical factors of male or female origins and how they control the pollen tube growth and fertilization process will broaden our understanding of the common themes and diverse strategies that plants have evolved to ensure reproductive success. The advancement of imaging technology to monitor pollination and fertilization and the development of probes to monitor various aspects of the pollen tube growth process, including pollen intracellular dynamics, will allow us to superimpose details obtained from studying pollen tube growth in culture conditions to interpret and understand the in vivo events.

Hand pollination experiments were conducted with seedling trees of cashew to determine the period of pistil receptivity to pollen, the timing of pollen tube growth, and compatibility between genotypes. The results were assessed by fluorescence microscopy of the pollen tubes, and by yield measurements. The pistil supported maximum pollen germination and tube growth when pollinated within 3 h of anthesis, and penetration of ovules by pollen tubes was significantly reduced following pollination later than 3 h after anthesis. Pollen tube growth in the style was rapid, with pollen tubes reaching the base of the style by 3 h after pollination, but penetration of the ovules was delayed until between six and 24 h. Specific combining ability was detected amongst cashew genotypes as measured by pollen tube growth, and both specific combining ability and general combining ability as measured by final fruit set. No difference in pollen germination and tube growth between self and cross pollination was measured in any...