There are 21 species in the genus Dacrydium (https://www.conifers.org/po/Dacrydium.php). Their natural distribution ranges from New Zealand, New Caledonia, Fiji and the Solomon Islands through New Guinea, Indonesia, Malaysia and the Philippines, to Thailand and southern China (de Laubenfels 1969; Quinn 1982). Currently no information is available about when reproductive cones start to initiate for Dacrydium species. In New Zealand, D. cupressinum cone initiation is suggested to occur in late summer or autumn with pollination occurring in spring (Norton et al. 1988). D. pectinatum male cones initiate before April in Hainan Island, China, because male buds are distinguishable by early April (Fig. 1a). Different species and climates can be the contributing factors for the discrepancy observed. Buds will be collected in March and February for examination in order to determine the initiation period for D. pectinatum male strobili in the future. D. pectinatum male cones average 8.5 mm in length (Fig. 2), similar to de Laubenfels’s report for the same species (6–12 mm) (de Laubenfels 1988), while longer than D. Bidwillii’s (2 to 6 mm) (Young 1907). When more information become available for other Dacrydium species, it will be interesting to compare their reproductive buds and cones’ morphology and phenology. This is important for the diversity conservation.
Similar to other coniferous species such as Metasequoia glyptostroboides (also known as the dawn redwood, Chinese redwood and water fir), D. pectinatum has microsporophylls spirally arranged around a main axis, and each microsporophyll consists of a phylloclade at the apex and one or two microsporangia at the base (Fig. 3). Like M. glyptostroboides (Jin et al. 2012), D. pectinatum male cones are mainly located around the outer and sunlit parts of crown. This is advantageous for pollen dispersal by wind, which is a common in conifers (Leslie 2011a, Leslie 2011b). Our study shows that the development of D. pectinatum microspore can be divided into four stages: initiation and differentiation of microsporophyll primordia, microspore sac formation, division of pollen mother cells, and pollen grain formation. This process lasts for about 12 months. A similar study will be conducted on female buds and cones.
Plant hormones and their interplay have important roles in various aspects of plant growth, development, and reproductive processes. Major phytohormones include auxins, abscisic acid, cytokinins, ethylene gibberellin, brassinosteroids, jasmonates, and strigolactones (Davis 2009; Kazan and Lyons 2016). Effects of hormones and their balance on reproductive bud initiation and development seem to depend on species and sex. In female Gnetum parvifolium buds, level of GA3, zeatin riboside (ZR), and ABA declines, and IAA increases as development progresses. In contrast, these endogenous hormones have the opposite trends in male buds. As for ratios, female and male buds share similar trends for ABA/GA3 and ZR/GA3, and differ in ABA/IAA and ZR/IAA (Lan et al. 2018). High levels of GA3 are reported to be beneficial for male cone formation in G. parvifolium (Lan et al. 2018) and other conifers, such as Pinus (Niu et al. 2014), Douglas-fir (Kong et al. 2008), and white spruce (Greenwood et al. 2011). In our previous study with Metasequoia, higher levels of GA1 + 3 and lower levels of IAA and ABA were beneficial to male primordium initiation, while higher levels of IAA and GA1 + 3 and a lower level of ABA were favorable to female cone initiation (Liang and Yin 1994). In D. pectinatum, level of GA, IAA, ABA and JA and their ratios fluctuated during late August to late November when sporogenous tissues were actively differentiated, suggesting their involvement in make cone development.
It is noteworthy that there was a dramatic increase of JA in male buds collected in late October after microspore sac was formed (Fig. 5d). JA is well known for its roles in plant’s biotic responses, such as drought, salt stress, low temperatures (Piotrowska et al. 2009; Wasternack et al. 2014). More recently, there is increasing evidence indicating JA’s involvement in plant development and reproduction. According to the review by Huang et al and Yuan and Zhang (Huang et al. 2017; Yuan and Zhang 2015), JA is found in control of stamen development in Arabidopsis, inhibition of petal expansion in Arabidopsis, sex determination in maize, and control of stamen and spikelet development in rice, as well as regulate embryo/seed development and induction of leaf senescence. The actions of JA can be channeled through its signaling pathway. To our knowledge, no reports of similar information are available in a gymnosperm species. Therefore, the JA surge in D. pectinatum shall be further investigated.
In summary, D. pectinatum male buds become distinguishable in April in tropical montane rain forests and continue to differentiate and develop until the following February. The dynamic change of endogenous hormones suggests their roles in cone development. Cone induction with hormones may provide an alternate approach to address the seed shortage issue due to the species’ long juvenile phase. It is suggested that treatments for male cone induction should be applied no later than April before differentiation of vegetative and reproductive buds. This is the first report on the anatomical and endogenous hormone changes that occur during the development of D. pectinatum the male cone. Combining morphological analyses of reproductive development with transcriptome studies in the future may lead to the understanding of molecular mechanisms behind reproductive development in D. pectinatum.