Factors affecting quantity of pollen dispersal of spray cut chrysanthemum (Chrysanthemum morifolium)

Abstract

Background: Spray cut chrysanthemum is a vital flower with high ornamental value and popularity in the world. However, the excessive quantity of pollen dispersal of most spray cut chrysanthemum is an adverse factor during its flowering stage, and can significantly reduce its ornamental value and quickly shorten its vase life. More seriously, excessive pollen grains in the air are usually harmful to people, especially for those with pollen allergies. Therefore, in order to obtain some valuable information for developing spray cut chrysanthemum with less-dispersed or non-dispersed pollen in the future breeding programs, we here investigated the factors affecting quantity of pollen dispersal of spray cut chrysanthemum with four cultivars, i.e. 'Qx-097', 'Noa', 'Qx-115', and 'Kingfisher', that have different quantity of pollen dispersal.

Results: 'Qx-097' with high quantity of pollen dispersal has 819 pollen grains per anther, 196.4 disk florets per inflorescence and over 800,000 pollen grains per inflorescence. The corresponding data for 'Noa' with low quantity of pollen dispersal are 406, 175.4 and over 350,000, respectively; and 219, 144.2 and nearly 160,000 for 'Qx-115' without pollen dispersal, respectively. 'Kingfisher' without pollen dispersal has 202.8 disk florets per inflorescence, but its anther has no pollen grains. In addition, 'Qx-097' has a very high degree of anther cracking that nearly causes a complete dispersal of pollen grains from its anthers. 'Noa' has a moderate degree of anther cracking, and pollen grains in its anthers are not completely dispersed. However, the anthers of 'Qx-115' and 'Kingfisher' do not crack at all. Furthermore, microsporogenesis and pollen development are normal in 'Qx-097', whereas many microspores or pollen degenerate in 'Noa', most of them abort in 'Qx-115', and all of them degrade in 'Kingfisher'.

Conclusions: These results suggest that quantity of pollen dispersal in spray cut chrysanthemum are mainly determined by pollen quantity per anther, and capacity of pollen dispersal. Abnormality during microsporogenesis and pollen development significantly affects pollen quantity per anther. Capacity of pollen dispersal is closely related to the degree of anther dehiscence. The entire degeneration of microspore or pollen, or the complete failure of anther dehiscence can cause the complete failure of pollen dispersal.

Figure 1

Flower morphology of four spray cut chrysanthemum cultivars. ‘Qx-097’ (A-D), ‘Noa’ (E-H), ‘Qx-115’ (I-L), and ‘Kingfisher’ (M-P). (A, B, E, F, I, J, M, N) Inflorescence morphology. (C, D, G, H, K, L, O, P) Disk floret. AN: Anther; MP: Mass of Pollen Grains; OV: Ovule; PE: Petal; PI: Pistil.

Figure 2

Microsporogensis of ‘Qx-097’ by TEM and paraffin section method. (A) Differentiation of archesporial cells. (B) Microsporocyte formation stage, showing differentiation of microspore mother cell and 3 anther walls (secondary parietal cell layer, endothecium and epidermis from the inner to the outer layer). (C) Microsporocyte formation stage, showing microspore mother cell tightly abutted and four complete anther walls. (D) Microspore mother cell at prophase I of meiosis, showing microspore mother cell in oval shape, discohesive cells and prominent nucleoli. (E) Microspores at first meiotic metaphase, chromosomes aligned on the metaphase plate. (F) Tetrad and condensed cytoplasm of tapetal layer. (G) Microspore mother cell in pachytene, showing mitochondria, plastids and numerous vacuoles. (H) The stage of tetrad, showing mitochondria, plastids, endoplasmic reticulum, enlargement of vacuoles, callose wall and cell wall which begin to develop. (I) Anther wall at microsporocyte meiosis stage, cytoplasm of endothecium layer, epidermal layer and tapetal layer cells become vacuolation, cytoplasm of tapetal layer cells which has large nuclei and obvious nucleoli condensed, middle layer degrades. AC: Archesporial cells; Ch: chromatin; CW: Callose wall; Dy: Dyad; En: Endothecium; Ep: Epidermis; ER: Endoplasmic reticulum; Mi: Mitochondrion; ML: Middle layer; Ms: Microspore mother cell; Nu: Nucleus; Pl: Plastid; PW: Pollen wall; SeL: Secondary parietal cell layer; Ta: Tapetum; Te: Tetrad; Va: Vacuole.

Figure 3

Pollen development of ‘Qx-097’ by TEM. (A) Early microspore, microspore cells have uniform and dense cytoplasm, abundant endoplasmic reticulum, mitochondria, plastids, some growing vacuoles and a nucleus which is in center of cells. (B) Anther wall at the early microspore stage, vacuolation of cytoplasm of tapetal layer with discohesive cells increase, middle layer is in further degradation. (C-D) Middle microspore, showing inner wall which begin to develop, spiked protuberances appear on the outside of the walls, recognizably germ pores, enlargement of vacuoles, off-centre nuclei and degraded cytoplasm and the number of density of ribosome, mitochondria and plastids decrease. (E) At middle microspore stage, tapetal cells contained abundant cell organelles and dense cytoplasm. (F) Late microspore stage, microspore cell wall, especially inner wall thickens, large central vacuole forms and squeeze the cytoplasm and nuclei to against the wall. (G) Late microspore stage, tapetal layer cells are hill-shaped and inner tangential walls are almost degraded completely, middle layer almost disappear and only has a few residue. (H) Early bicellular pollen stage, there is develop inner wall, exine formation by deposition of primexine and then sporopollenine, a large central vegetative cell and a small generative cell which forms by excentrically division of microspores nuclei. (I) Early bicellular pollen stage, tapetal layer almost disappear, endothecium layer cells enlarge remarkably in volume. Cy: Cytoplasm; En: Endothecium; Ep: Epidermis; ER: Endoplasmic reticulum; GN: Generative nucleus; GP: Germ pore; IW: Inner wall of cells; Mi: Mitochondrion; Ml: Middle layer; MM: Middle microspore; Nu: Nucleus; Pl: Plastid; PW: Pollen wall; SP: Spiked protuberance; Ta: Tapetum; Va: Vacuole; VN: Vegetative nucleus.

Figure 4

Pollen development of ‘Qx-097’ by paraffin section method. (A) Early microspores, tapetal layer cells condensed. (B) Middle microspores stage, showing germ pores, thickening cell walls and radial thinning of tapetal layer cells. (C) Late microspores stage, showing mononuclear microspores against the anther wall and degradation of tapetal layer. (D) Early bicellular pollen stage, showing nuclei divide excentrically to form a large central vegetative nucleus and a small generative nucleus, the tapetal cells only have residue and endothecium layer cells enlarge in volume. (E) Late bicellular pollen stage, contents such as starch began to accumulate in cytoplasm, but the two nuclei can be observed still and vacuoles gradually disappeared. (F) Mature pollen stage, contents such as starch fills the whole cell and the nuclei are hard to be observed. BP: Bicellular pollen; EM: Early microspore; GN: Generative nucleus; GP: Germ pore; LM: Late microspore; MaP: Mature pollen; MM: Middle microspore; Ta: Tapetum; TaD: Tapetum debris; VN: Vegetative nucleus.

Figure 5

Pollen abortion during pollen development of ‘Noa’. (A) Late microspore stage, showing rupture of cell wall and disintegration of cytoplasm (arrows). (B) Early microspore stage, showing rupture of the primary cell wall, shrinkage of cytoplasm and the nucleus which was in irregularly shaped. (C) Microspore in early stage of development, cell wall and cytoplasm were in degradation. (D) Distortion of cell wall of microspore and shrinkage of cytoplasm. (E-H) Pollen abortion in late pollen development stage (arrows).

Figure 6

Pollen abortion during pollen development of ‘Qx-115’ and ‘Kingfisher’. ‘Qx-115’ (A-B) and ‘Kingfisher’ (C-F). (A) Microspores at middle microspores stage of ‘Qx-115’, showing many broken, incomplete and abortive microspores. (B) Late microspores stage in ‘Qx-115’, it has few complete pollen grains in the anther. (C) Asymmetrical and abnormal dyad. (D) Asymmetrical and abnormal tetrad. (E) Degradation of tetrads, microspores can’t move apart and cytoplasm was degraded and in vacuolization. (F) Early microspores stage, showing pollen sac with no pollen grains of ‘Kingfisher’. AD: Abnormal dyad; AT: Abnormal tetrad; BM: Broken microspore; MD: Microspore in degradation; TaD: Tapetum debris.

Figure 7

Anatomical features of anther and Flower morphology during pollen dispersal of ‘Qx-097’, ‘Noa’ and ‘Qx-115’. Pollen dispersing of ‘Qx-097’, ‘Noa’ and ‘Qx-115’. (A, D, G) The section of anthers just in pollen dispersal and anther cracking which corresponds to the anthers in disk florets a, d and g (arrows) in picture of material object. (B, E, H) The section of anthers at the stage of pollen dispersal a lot which corresponds to the anthers in disk florets b, e and h (arrows) in picture of material object. (C, F, I) The section of anthers after pollen dispersal which corresponds to the anthers in disk florets c, f and i (arrows) in picture of material object. (J-O) Flower morphology of ‘Qx-097’, ‘Noa’ and ‘Qx-115’ during pollen dispersing.

Figure 8

Ultrastructure of endothecium layer cells at late stage of pollen development. (A) ‘U’ shaped thickening of endothecium layer cells (arrows) in ‘Qx-097’. (B) ‘U’ shaped thickening of endothecium layer cells (arrows) in ‘Noa’.