Pollen viability, stigma receptivity and their effect on fruit set of passionfruit at different flower developmental stages

Abstract:【Objective】The study examined the pollen viability and stigma receptivity at different developmental stages of passionfruit flowers and their effects on the fruit setting and fruit traits, so as to find out the effective pollination period (EPP).【Methods】In this study, the purple passionfruit of Guibai No.1 was used as the material, and the grafted seedings were planted in the experimental orchard of institute of Horticulture of Guangxi Academy of Agricultural Sciences (108.24° E, 22.85° N) in September 2020 by the root-restriction method, with a spacing of 3 m × 2 m. The experiment was conducted from April 15, 2022 to July 20, 2022. For determination of pollen viability, the flowering time, pistil and stamen development of Guibai No.1 were observed starting from the day before the experiment began. The number of the flower bud samples was 100 from 20 plants each with 5. The sample collection was carried out at 5 h (-5 h), 3 h (-3 h) and 1 h (-1 h) before flowering, or 0 (0 h), 1 h (1 h), 3 h (3 h),5 h (5 h), 7 h (7 h), 9 h (9 h) and 20 h (20 h) after flowering. Pollen germination rate and pollen tube length at different times were determined with pollen germination in vitro in liquid medium. The stigma receptivity was determined by benzidine-H2O2 method, and sampling methods were the same as those for the determination of pollen viability. The stigmas of flowers at different times were placed in a petri dish containing benzidine-H2O2 solution (1% benzidine∶3% hydrogen peroxide∶water = 4∶11∶22) and completely submerged in the solution. After 5 min, the stigmas were observed under a stereoscopic microscope OLYMPUS SZX10, and a Canon 7500 camera was used to take pictures and record. The degree of stigma receptivity was determined by the number of bubbles generated around the stigma. Stronger receptivity results in generation of more of bubbles around the stigma. The receptivity was classified into different levels. In grade 1 marked as +, there were a few bubbles and the stigma was weak, and the score was assigned 1; in grade 2 marked as ++, there were many bubbles and the stigma receptivity was medium with a score of 2; in grade 3 marked at +++, there were a large number of bubbles and the stigma was strong with a score of 3. For observation of pollen and stigma by scanning electron microscope, anthers and stigmas at different times were placed in different EP tubes and fixed in 2.5% glutaraldehyde fixation solution for 2 h, and then stored under 4 ℃. The fixed samples were rinsed with 0.1 mol · L^-1 phosphate buffer PB (pH 7.4) for 3 times, 15 min each time, dehydrated successively in 30%, 50%, 70%, 80%, 90%, and 95% ethanol solutions, and then in 100% ethanol twice, 15 min each time, and finally dried in a QuorumK850 critical point dryer. A Yulon Times LJ-16 ion sputtering instrument was used for gold coating for 30s, and a Carl Zeiss EVOLS10 scanning electron microscope was used for sample observation. The representative fields of view were selected to observe the equator, polar axis, pollen germination groove, germination hole, and other characters of the pollen. The analysis software was Image-Pro Plus 6.0. The equatorial axis length and polar axis length of 15 pollen grains were measured randomly, and the average was counted. For determination fruit setting and fruit characteristics at different times, pollens were collected gently, and then evenly smeared onto the stigmas with a soft brush. 20 flowers were pollinated at each sampling time, and three days were repeated at the same time point as three repetitions. All the tested flowers were emasculated the day before the experiment and pollinated with pollens of other flowers at corresponding time points. 10 days after pollination, fruit setting was recorded, and the fruit setting rate (% ) was calculated at (number of fruits/total pollinated flowers) ×100. When the fruit matured, the single fruit weight, fruit horizontal diameter, fruit vertical diameter, flesh recovery, juice yield, seed number of single fruit as a repetion，and other characters were recorded.【Results】The pollen of passionfruit displayed some viability at 5 h before flowering, and then pollen viability increased continuously, reaching a peak at 3 h after flowering, and then dropped to only 10.63% at 9 h and to 0% at 20 h. The stigma receptivity was significantly correlated with pollen viability, with a correlation coefficient of 0.711, which indicates that the flower of passionfruit is of monochogamy. Scanning electron microscopy showed that there was no obvious change in the appearance of pollen at different times before and after flowering. The stigma shrank chiefly on the surface of mastoid cells at 7 h after flowering, and the stigma shrank obviously at 20 h after flowering, which indicates that the receptivity of stigma can be judged by observing the appearance of the mastoid cells, but pollen viability cannot be judged by its appearance. The fruit setting rate was the best when pollination was carried out 3 h before flowering, but slightly worse at flowering and 1 h after flowering. After that, the fruit setting rate decreased continuously with the extension of time, and the fruit setting rate was 0% at 9 h after flowering. Pollen viability and stigma receptivity were significantly correlated with fruit setting rate, but not with single fruit weight, fruit transverse diameter, fruit longitudinal diameter and flesh re-covery, which indicates that pollen viability and stigma receptivity only affected fruit set but not fruit development.【Conclusion】Flowers of passionfruit are characterized by monochogamy, the viability of the pollen and stigma is short and quickly loses with time after flowering. The effective pollination period (EPP) is about 10 h from 3 h before flowering to 7 h after flowering.