Pollination requirements of kiwifruit (Actinidia chinensis Planch.) differ between cultivars ‘Hayward’ and ‘Zesy002’猕猴桃（Actinidia chinensis Planch.）海沃德和阳光金果之间有所不同授粉要求
▲刚完成授粉的红心猕猴桃幼果
ABSTRACT
Pollination by insects is critical for the production of many crops worldwide. Crop cultivars vary in a number of traits, but their differing pollination requirements are often overlooked. Kiwifruit (Actinidia chinensis) is reliant on pollen movement between male and female plants, but there has been disagreement in the literature about what its pollination requirements are. Additionally, there is little information about how time-of-day might affect fruit and seed set, and how this may alter the efficacy of pollination management strategies. In this study, we compare the pollination requirements of A. chinensis var. deliciosa ‘Hayward’ (a hexaploid green-fleshed variety) and A. chinensis var. chinensis ‘Zesy002’ (a tetraploid gold-fleshed variety). We find that ‘Zesy002’ requires fewer pollen grains than ‘Hayward’ for full seed set. Kiwifruit appears to be equally able to set fruit at any time of the day, meaning that insects which forage outside the peak hours of 0900–1500 h may play an important supporting role in fruit production, and that artificial pollination could profitably be applied into the evening hours rather than being limited to the period of peak pollinator activity.
KEYWORDS: Kiwifruitpollen dose-responsepollen requirementcrop pollination
▲Kiwi Orchard Work
摘要
昆虫授粉对于全世界许多作物的生产至关重要。作物品种的许多性状各不相同，但它们不同的授粉要求常常被忽视。猕猴桃（Actinidia chinensis）依赖于雄性和雌性植物之间的花粉运动，但文献中对其授粉要求存在分歧。此外，关于一天中的时间如何影响果实和结籽，以及这如何改变授粉管理策略的功效的信息很少。在这项研究中，我们比较了中华木瓜的授粉要求。 deliciosa ‘Hayward’（六倍体绿肉品种）和 A. chinensis var. chinensis ‘Zesy002’（四倍体金肉品种）。我们发现“Zesy002”需要比“Hayward”更少的花粉粒才能获得完整的种子。猕猴桃似乎同样能够在一天中的任何时间结果，这意味着在 0900-1500 小时高峰时间之外觅食的昆虫可能在水果生产中发挥重要的支持作用，并且人工授粉可以有利地应用于水果生产。晚上时间，而不是仅限于授粉昆虫活动高峰期。
关键词: 猕猴桃花粉剂量反应 花粉需求 作物授粉
▲Kiwi insect pollination
Introduction
Pollination by insects is an important service to global agriculture, with one-third of crops relying to some extent on insects for pollination (Aizen et al. Citation2009), and worldwide dependence on insects for crop production increasing year-on-year (Garibaldi et al. Citation2011).
Kiwifruit was highlighted by Klein et al. (Citation2007) as being one of 13 major crops for which insect-vectored pollination is ‘essential’, and has an annual production of fruit over 4.25 million tonnes worldwide (FAOSTAT Citation2018). The plants are climbing vines, with large cream-coloured flowers (Schmid Citation1978). Female plants produce flowers with well developed anthers containing non-viable pollen, and neither sex produces any nectar (Schmid Citation1978).
▲Kiwi female flowers
介绍
昆虫授粉是全球农业的一项重要服务，三分之一的作物在一定程度上依赖昆虫授粉（Aizen 等人，Citation2009），全球作物生产对昆虫的依赖逐年增加（Garibaldi 等人）引文 2011）。
Klein 等人重点介绍了猕猴桃。 （引文 2007）被认为是昆虫传粉授粉“必不可少”的 13 种主要作物之一，全球水果年产量超过 425 万吨（粮农组织统计数据库引文 2018）。这些植物是攀缘藤本植物，开着大奶油色的花朵（Schmid Citation 1978）。雌性植物产生的花朵具有发育良好的花药，其中含有无活力的花粉，并且雌性植物均不产生任何花蜜（Schmid Citation1978）。
▲golden kiwi growing zone
Numerous studies have shown that fruit weight in kiwifruit is correlated with seed set (Brundell Citation1975; Schmid Citation1978; Hopping Citation1984; Vaissière et al. Citation1990; González et al. Citation1995a), and as such, it is directly affected by the amount of viable pollen received. Kiwifruit has been noted previously to have a substantial requirement in terms of numbers of pollinator visits and numbers of pollen grains, as it has many ovaries (24–40; McNeilage and Steinhagen Citation1998) which together have the ability to produce 1400–1500 seeds when fully pollinated (Hopping Citation1990). Flowers are syncarpous (Howpage et al. Citation1998), thus full pollination can occur if sufficient pollen is deposited on a single stigma.
▲中国四川省的红心猕猴桃果园
大量研究表明，猕猴桃的果实重量与结实率相关（Brundell Citation1975；Schmid Citation1978；Hopping Citation1984；Vaissière et al. Citation1990；González et al. Citation1995a），因此，它直接受到活菌数量的影响收到花粉。之前已经指出，猕猴桃对授粉媒介访问次数和花粉粒数量有很大的要求，因为它有许多子房（24-40；McNeilage 和 Steinhagen Citation1998），当授粉者到达时，它们总共有能力产生 1400-1500 粒种子。完全授粉（Hopping Citation1990）。花是合心皮的（Howpage et al. Citation1998），因此如果有足够的花粉沉积在单个柱头上，则可以发生完全授粉。
▲授粉良好的黄心猕猴桃品质
Kiwifruit can be challenging to pollinate, particularly if there are insufficient honey bee hives or more attractive forage is nearby (Macfarlane and Ferguson Citation1983; Pomeroy and Fisher Citation2002). Problems with the supply of honey bees in Italy has led to decades of artificial-only pollination, though it is somewhat less effective than insect pollination (Sáiz et al. Citation2019). In other countries, such as New Zealand, artificial pollination is used in addition to honey bees (Max and McMonagle Citation2014), a strategy that balances the risk of relying solely on honey bees (Willis and Kirby Citation2015) and the expense in human labour of artificial pollination .
奇异果授粉可能具有挑战性，特别是如果蜂箱不足或附近有更具吸引力的饲料时（Macfarlane 和 Ferguson Citation 1983；Pomeroy 和 Fisher Citation 2002）。意大利蜜蜂供应问题导致了数十年的纯人工授粉，尽管其效果略低于昆虫授粉（Sáiz 等人，引文 2019）。在新西兰等其他国家，除了蜜蜂之外，还使用人工授粉（Max 和 McMonagle Citation 2014），这种策略平衡了完全依赖蜜蜂的风险（Willis 和 Kirby Citation 2015）和人工授粉的人力成本。
▲Kiwi pollination▲
The use of artificial pollination has closely mirrored the times when bees are active in the field, as this has been assumed to be the most receptive period for kiwifruit flowers (Goodwin, pers. comm). However, as far as we are aware, no previous study has examined floral receptivity over the course of the day, so it may be that flowers are still receptive in the afternoon and evening, after honey bees have typically left the orchard (Goodwin Citation1987), which would expand the possible times to apply pollen, as well as the possible times other pollinating insects could contribute to pollination.
人工授粉的使用密切反映了蜜蜂在田间活跃的时代，因为这被认为是猕猴桃花最容易接受的时期（Goodwin，个人通讯）。然而，据我们所知，之前没有研究检查过一天中花朵的接受性，因此在蜜蜂通常离开果园之后，花朵可能在下午和晚上仍然具有接受性（Goodwin Citation1987） ，这将扩大施加花粉的可能时间，以及其他授粉昆虫有助于授粉的可能时间。
▲Photo of red kiwi fruit pollination
The literature provides suggestive evidence that there is between-cultivar variability in pollination requirements; although the number of pollen grains required to produce a marketable fruit can be deposited in a single honey bee visit (Donovan and Read Citation1992), green-fleshed kiwifruit may require up to 40 honey bee visits (Goodwin and Haine Citation1995), while yellow-fleshed kiwifruit may require as few as six (Goodwin et al. Citation2017). These studies did not record the time of day of the trials, and if there are diurnal patterns in receptivity this could be a factor that helps explain the wide variation in per-visit seed set figures. Determining between-cultivar differences in pollen requirements and the effect of the time-of-day on the ability to set seed would help inform both insect and artificial pollination practices, and also honey bee stocking rates, as well as define the window during which successful pollination can take place.
文献提供了暗示性证据，表明授粉要求存在品种间差异；尽管生产可销售水果所需的花粉粒数量可以在单次蜜蜂访问中沉积下来（Donovan 和 Read Citation 1992），但绿肉奇异果可能需要多达 40 次蜜蜂访问（Goodwin 和 Haine Citation 1995），而黄肉奇异果可能需要多达 40 次蜜蜂访问（Goodwin 和 Haine Citation 1995）。肉质奇异果可能只需六个（Goodwin 等人，Citation2017）。这些研究没有记录试验一天中的时间，如果接受性存在昼夜模式，这可能是有助于解释每次访问种子结实数的巨大差异的一个因素。确定品种间花粉需求的差异以及一天中的时间对结籽能力的影响将有助于为昆虫和人工授粉实践以及蜜蜂饲养率提供信息，并确定成功的窗口期可以进行授粉。
▲yellow kiwi packaging
Materials and methods
We examined pollination in two common kiwifruit cultivars in New Zealand, green-fleshed Actinidia chinensis var. deliciosa ‘Hayward’ and gold-fleshed A. chinensis var. chinensis ‘Zesy002’. The three trials below – pollination requirement of ‘Zesy002’ and ‘Hayward’ and the examination of fruit and seed set over the course of a 24-h day – were conducted between 2011 and 2019 using differing methodologies.
材料和方法
我们检查了新西兰两个常见猕猴桃品种（绿肉猕猴桃变种）的授粉情况。 deliciosa ‘Hayward’ 和金肉 A. chinensis var.中华草'Zesy002'。以下三项试验——“Zesy002”和“Hayward”的授粉要求以及一天 24 小时内果实和种子的检查——是在 2011 年至 2019 年期间使用不同的方法进行的。
▲红心猕猴桃
General methodology
All experiments were conducted in New Zealand during the Austral spring and summer (October through March) in kiwifruit orchards trained on a pergola system. When possible, pollination was studied during fine weather conditions (15°C–30°C, wind below 15 km/h).
To exclude pollinators while conducting pollination experiments, we bagged flowers just prior to opening with grease-proof paper bags closed with wire ties.
一般方法
所有实验均在澳大利亚春季和夏季（十月至三月）期间在新西兰的奇异果园中进行，并采用凉棚系统进行训练。如果可能，在晴朗的天气条件（15°C–30°C，风速低于 15 km/h）下研究授粉。
为了在进行授粉实验时排除授粉媒介，我们在开放前用防油纸袋将花朵装袋，并用扎带封闭。
▲▲golden kiwifruit vine
Artificial pollination was conducted using a wet-spray method. The industry standard practice is to use a ratio of 4 g pollen to 1 L of PollenAid® (KiwiPollen, New Zealand) and water mixture. The pollen used was commercially harvested ‘Chieftain’ pollen. Flower buds were bagged 1–2 days before opening. On the day flowers opened they were un-bagged and the pollen solution applied using a pneumatic sprayer (Badger Air Brush Co. IL, USA). A single person pollinated all flowers for a given trial, delivering two ∼300 ms sprays. For trials where we assessed fruit set, flowers were then immediately re-bagged to prevent any other pollination.
采用湿喷法进行人工授粉。行业标准做法是使用 4 g 花粉与 1 L PollenAid®（KiwiPollen，新西兰）和水混合物的比例。使用的花粉是商业收获的“Chieftain”花粉。花蕾在开放前1-2天装袋。花朵开放当天，将其拆袋并使用气动喷雾器（Badger Air Brush Co. IL，USA）施用花粉溶液。在一次给定的试验中，一个人对所有花朵进行授粉，进行两次约 300 毫秒的喷雾。在我们评估坐果的试验中，花朵立即重新装袋以防止任何其他授粉。
▲猕猴桃甜度检测
To assess the number of pollen grains deposited on stigmas, we excised stigmas immediately following pollination and collected them in 1.5-mL Eppendorf tubes with 0.5 mL Alexander’s stain (Alexander Citation1980). This differentially stains staminate (male) and pistillate (female) pollen (purple-red and green-blue respectively; Goodwin and Perry Citation1992).
为了评估沉积在柱头上的花粉粒数量，我们在授粉后立即切除柱头并将其收集在带有 0.5 mL 亚历山大染色剂的 1.5 毫升 Eppendorf 管中（Alexander Citation1980）。这对雄花粉（雄性）和雌花粉（雌性）花粉（分别为紫红色和绿蓝色；Goodwin 和 Perry Citation 1992）进行了差异染色。
▲Golden Kiwi Orchard
Pollen dose–response relationship for ‘Hayward’
To determine how many pollen grains were required for full seed set in ‘Hayward’ (and thus maximal fruit weight), we conducted an artificial pollination trial in 2018–2019. The experiment was conducted in an orchard in the Waikato region of New Zealand near Cambridge. For our study, ‘Chieftain’ pollen was applied to 20 flowers at each of the following application rates: 0, 0.5, 1, 2, 4, 8 and 16 g/L. Counts of pollen tubes and corresponding seed numbers were obtained from the same flower, as opposed to separate groups of flowers as in the previous trial for ‘Zesy002’.
“海沃德”的花粉剂量-反应关系
为了确定“海沃德”完全结籽需要多少花粉粒（从而确定最大果实重量），我们在 2018 年至 2019 年进行了一次人工授粉试验。该实验是在新西兰剑桥附近怀卡托地区的一个果园进行的。在我们的研究中，“Chieftain”花粉按以下施用率施用于 20 朵花：0、0.5、1、2、4、8 和 16 g/L。花粉管计数和相应的种子数量是从同一朵花中获得的，而不是像之前“Zesy002”试验中那样从不同的花组中获得。
▲猕猴桃硬度检测仪器
Previous work has identified that it takes kiwifruit pollen 74 h to germinate and reach the ovary (Hopping and Jerram Citation1979; González et al. Citation1995b). By removing stigmas after 96 h, we allowed for the possibility of slower germination due to inclement weather. Each flower was tagged with an individual identification number, and so were the corresponding stigma samples, which were placed in 95% ethanol. Stigmas were then processed according to Martin (Citation1959) using aniline blue and fluorescence microscopy to identify and count the pollen tubes. This measure was compared with the seed number in the correspondingly tagged mature fruit. To count seeds, mature fruit were introduced to a ripening chamber containing ethylene. When soft, the fruit were peeled and the core with seeds spread flat inside a 50-μm polythene bag and photographed under controlled lighting conditions. The photographed seeds were then counted using the particle counting function in ImageJ (Rueden et al. Citation2017).
▲猕猴桃授粉染色
先前的工作已确定奇异果花粉需要 74 h 才能发芽并到达子房（Hopping 和 Jerram Citation1979；González 等人 Citation1995b）。通过在 96 小时后去除柱头，我们考虑到了由于恶劣天气而导致发芽速度减慢的可能性。每朵花都标有单独的识别号，相应的柱头样本也是如此，并将其放置在 95% 乙醇中。然后根据 Martin (Citation1959) 的方法，使用苯胺蓝和荧光显微镜对柱头进行处理，以识别和计数花粉管。将此测量值与相应标记的成熟果实中的种子数进行比较。为了计数种子，将成熟的果实放入含有乙烯的催熟室中。当果实变软时，将其去皮，将带有种子的果核平铺在 50 微米的聚乙烯袋中，并在受控照明条件下拍照。然后使用 ImageJ 中的粒子计数功能对拍摄的种子进行计数（Rueden 等人，Citation2017）。
▲sungold g3 kiwifruit vine
Pollen dose–response relationship for ‘Zesy002’
In 2011, we examined the relationship between the number of pollen grains, seeds and fruit weight of ‘Zesy002’ kiwifruit at a Bay of Plenty orchard near Paengaroa. The trial was conducted on 200 flowers spread over 15 vines, and 20 flowers were sprayed with a solution of ‘Chieftain’ pollen and PollenAssist at each of the following rates: 0.025, 0.05, 0.075, 0.1, 0.2, 0.3, 0.4, 0.5, and 0.6 g/L. The concentrations were applied in a random order. Ten of the flowers receiving each rate were re-bagged for assessment of seed set while the stigmas of the remaining 10 for each rate were excised for analysis of pollen deposition. To assess resulting fruit and seed set, kiwifruit were picked just prior to harvest and weighed. Seeds were counted by softening fruit tissue, straining out seeds, weighing the total dried seeds and then counting and weighing a subsample of at least 200 seeds. Differences between application rates were assessed with the Mann–Whitney U-test (MWU).
▲猕猴桃人工授粉方式
“Zesy002”的花粉剂量-反应关系
2011 年，我们在 Paengaroa 附近的丰盛湾果园检查了“Zesy002”奇异果的花粉粒数量、种子数量和果实重量之间的关系。该试验在分布在 15 根藤蔓上的 200 朵花上进行，其中 20 朵花按以下比例喷洒了“Chieftain”花粉和 PollenAssist 溶液：0.025、0.05、0.075、0.1、0.2、0.3、0.4、0.5、和0.6 g/L。以随机顺序应用浓度。将接受每个比率的十朵花重新装袋以评估结籽，同时切除每个比率的其余十朵花的柱头以分析花粉沉积。为了评估最终的果实和结籽，在收获前采摘猕猴桃并称重。通过软化果实组织、滤出种子、称重总干燥种子、然后对至少 200 粒种子的子样本进行计数和称重来对种子进行计数。使用曼-惠特尼 U 检验 (MWU) 评估施用量之间的差异。
▲sungold kiwi orchard
Stigmatic receptivity vs. time-of-day for ‘Hayward’
To determine if there was an optimal time to receive pollination, we artificially pollinated ‘Hayward’ kiwifruit flowers every 2 h between 0400 and 2200 h on 21–24 November 2013 and between 0600 and 2400 h on 27 November–2 December 2014 in a Waikato orchard. Pollen was applied at the standard rate of 4 g/L at each time point to previously bagged flowers, which were then re-bagged. Once mature, fruit were harvested, weighed and their seeds counted using the subsampling method described for ‘Zesy002’. The nonlinear effect was assessed with a generalised additive mixed model (GAMM) in the R package gamm4 (Wood and Scheipl Citation2009) with seed number as the response, time-of-day as the predictor, and year and sample date as nested random effects.
“海沃德”的耻辱接受度与一天中的时间
为了确定是否有最佳授粉时间，我们在2013年11月21日至24日的0400至2200 h之间以及2014年11月27日至12月2日的0600至2400 h之间每2 小时对“Hayward”奇异果花进行人工授粉。果园。在每个时间点将花粉以 4 g/L 的标准比例施用于先前装袋的花朵，然后将其重新装袋。成熟后，使用“Zesy002”描述的二次采样方法收获果实、称重并计算种子数量。使用 R 包 gamm4（Wood 和 Scheipl Citation2009）中的广义加性混合模型 (GAMM) 评估非线性效应，其中种子数作为响应，一天中的时间作为预测变量，年份和样本日期作为嵌套随机效应。
▲Kiwi pollen▲
Results
Pollen dose–response relationship for ‘Hayward’
After counting the pollen tubes in each stigma of each flower, we found that some flowers had over 15,000 tubes – more than 10 times the number of ovules. This high amount of pollination did not appear to have a negative impact on seed numbers. Seed numbers increased linearly at about 0.54 tubes per seed until 2000–2500 tubes, where additional pollen tubes no longer increased seed number (Figure 1). The probability of a fruit setting increased with greater numbers of pollen tubes (P = .009; generalised linear model), with 97% of flowers receiving over 2500 pollen tubes successfully setting fruit. We measured the total number of pollen grains in a small subset of our data, and they outnumbered pollen tubes, on average, by a factor of 5.3. This ratio translates to full fruit set being achieved when 10,600–13,250 pollen grains are deposited across the stigma and styles.
结果
“海沃德”的花粉剂量-反应关系
在计算每朵花每个柱头上的花粉管后，我们发现有些花有超过 15,000 个花粉管，是胚珠数量的 10 倍多。如此大量的授粉似乎并未对种子数量产生负面影响。种子数量以每颗种子约 0.54 个管的速度线性增加，直到 2000-2500 个管，此时额外的花粉管不再增加种子数量（图 1）。花粉管数量越多，坐果的概率越高（P = .009；广义线性模型），接受超过 2500 个花粉管的花朵中有 97% 成功坐果。我们测量了一小部分数据中花粉粒的总数，它们的数量平均多于花粉管的 5.3 倍。当 10,600-13,250 个花粉粒沉积在柱头和花柱上时，这个比例就可以实现完整的坐果。
▲Golden Kiwi Orchard
Figure 1. The relationship between the number of pollen tubes in all styles of kiwifruit cultivar Actinidia chinensis var. deliciosa ‘Hayward’ and the number of seeds in the resulting fruit. The blue trend line is a generalised additive model with a smooth function applied to the log seed number.
图1 猕猴桃品种Actinidia chinensis var.各风格花粉管数关系deliciosa ‘Hayward’ 以及所得果实中种子的数量。蓝色趋势线是广义加性模型，其平滑函数应用于对数种子数。
▲新西兰rubyred宝石红奇异果
Figure 1. The relationship between the number of pollen tubes in all styles of kiwifruit cultivar Actinidia chinensis var. deliciosa ‘Hayward’ and the number of seeds in the resulting fruit. The blue trend line is a generalised additive model with a smooth function applied to the log seed number.
图1 猕猴桃品种Actinidia chinensis var.各风格花粉管数关系deliciosa ‘Hayward’ 以及所得果实中种子的数量。蓝色趋势线是广义加性模型，其平滑函数应用于对数种子数。
▲Golden Kiwi Orchard
The relationship between pollen tube number and fruit weight was strongly correlated (R2 = 0.69) until approximately 2000 pollen tubes, after which point the correlation was much weaker (R2 = 0.48; Figure 2). This weak relationship could be due to a number of factors we didn’t measure, including whether the flower was terminal or lateral, cane diameter, and plant stress (Richardson et al. Citation2019).
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花粉管数量与果实重量之间的相关性很强（R2 = 0.69），直到大约2000个花粉管为止，此后相关性就弱得多（R2 = 0.48；图2）。这种微弱的关系可能是由于我们没有测量的许多因素造成的，包括花是顶生的还是侧生的、茎直径和植物胁迫（Richardson et al. Citation2019）。
▲zespri sungold g3 kiwi
Figure 2. The relationship between the number of pollen tubes in all styles of kiwifruit cultivar Actinidia chinensis var. deliciosa ‘Hayward’ and the fruit weight of fruit that successfully set. The trend line is a generalised additive model with a smooth function applied to the log fruit weight.
图2 猕猴桃品种Actinidia chinensis var.各风格花粉管数的关系deliciosa ‘Hayward’ 和成功坐果的果实重量。趋势线是一个广义加性模型，具有应用于原木重量的平滑函数。
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Pollen dose–response relationship for ‘Zesy002’
Fruit were produced at each of the 10 pollen concentrations applied to ‘Zesy002’ flowers. The amount of pollen grains deposited on stigmas increased linearly with the rate of application (R2 = 0.97, Figure 3).
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“Zesy002”的花粉剂量-反应关系
在‘Zesy002’花上施加 10 种花粉浓度时，每种花粉浓度都会产生果实。沉积在柱头上的花粉粒数量随施用量线性增加（R2 = 0.97，图3）。
Figure 3. Pollen grains deposited on kiwifruit cultivar Actinidia chinensis var. chinensis ‘Zesy002’ stigmas at 10 different rates of pollen application. Boxes represent the middle 50% of the data, bars within boxes represent the median and whiskers are the spread of the data within 1.5× the interquartile range; data points outside this range are shown as dots.
图 3. 猕猴桃品种 Actinidia chinensis var. 上沉积的花粉粒。 chinensis ‘Zesy002’ 柱头在 10 种不同的花粉施用率下。方框代表中间 50% 的数据，方框内的条形代表中位数，须线代表数据在 1.5 倍四分位数范围内的分布；该范围之外的数据点显示为点。
▲Golden Kiwi Orchard
Fruit weight did not significantly increase with increasing pollen application (P = .409, linear model), though the weight of fruit produced under the two lowest application rates (0.025 and 0.05 g/L) was lower than that produced at higher application rates (P = .031; MWU). Fruit that were allowed to reach maturity had their seeds counted. Seed numbers increased with applied pollen until between 3500 and 4000 grains, when additional pollen applied had no net gain of seed number (Figure 4).
果实重量并没有随着花粉施用量的增加而显着增加（P = .409，线性模型），尽管在两个最低施用量（0.025和0.05 g/L）下产生的果实重量低于在较高施用量下产生的果实重量（ P = .031；对成熟的果实进行种子计数。种子数量随着施用花粉的增加而增加，直到 3500 至 4000 粒之间，此时施用额外的花粉没有种子数量的净增加（图 4）。
Figure 4. The relationship between the number of pollen grains on styles of kiwifruit cultivar Actinidia chinensis var. chinensis ‘Zesy002’ and the number of seeds in the resulting fruit. Points represent mean pollen grains deposited and mean seeds per fruit for each concentration. Bars represent standard error of the mean. The trend line is a generalised additive model with a smooth function applied to the log seed number.
图4 猕猴桃品种Actinidia chinensis var. 花粉粒数与花柱的关系chinensis ‘Zesy002’ 以及所得果实中的种子数量。点代表每个浓度下沉积的平均花粉粒和每个果实的平均种子。条形代表平均值的标准误差。趋势线是广义加性模型，其平滑函数应用于对数种子数。
▲New Zealand Kiwi Orchard
Stigmatic receptivity vs. time-of-day for ‘Hayward’
There was no clear effect of time-of-day on seed set for the 2013 and 2014 data together, as over-dispersion could not be reduced, but there was an evening peak in 2013 (P < .001, GAMM), and an early morning peak in 2014 (P < .001, GAMM; Figure 5). The models of fruit weight with the full dataset were also over-dispersed when using time as a predictor variable and year and date as nested random effects.
“海沃德”的耻辱接受度与一天中的时间
2013 年和 2014 年的数据一起，一天中的时间对结籽没有明显影响，因为过度分散无法减少，但 2013 年有一个晚高峰（P < .001，GAMM），并且2014 年清晨高峰（P < .001，GAMM；图 5）。当使用时间作为预测变量以及年份和日期作为嵌套随机效应时，具有完整数据集的水果重量模型也过度分散。
▲zespri sungold kiwi
Figure 5. Relationship between seed set and time of day for kiwifruit cultivar Actinidia chinensis var. deliciosa ‘Hayward’. Boxes represent the middle 50% of the data, bars within boxes represent the median and whiskers are the spread of the data within 1.5× the interquartile range; data points outside this range are shown as dots.
图 5. 猕猴桃品种 Actinidia chinensis var. 的结籽率与一天中的时间之间的关系美味的“海沃德”。方框代表中间 50% 的数据，方框内的条形代表中位数，须线代表数据在 1.5 倍四分位数范围内的分布；该范围之外的数据点显示为点。
Discussion
Kiwifruit requires cross pollination in order to produce a marketable crop, which is delivered either by insects or artificial pollination in commercial orchards. We found that green-fleshed ‘Hayward’ had higher pollination requirements than gold-fleshed ‘Zesy002’, and identified a possible reason for the variability amongst measures of pollen requirement in kiwifruit.
讨论
猕猴桃需要异花授粉才能生产出适销对路的作物，这种作物可以通过昆虫或商业果园的人工授粉来实现。我们发现绿肉“Hayward”比金肉“Zesy002”具有更高的授粉要求，并确定了猕猴桃花粉需求测量差异的可能原因。
▲猕猴桃开花授粉
The pollination requirements of ‘Hayward’ have been reported, yielding a constellation of numbers which vary from 1000 to 15,000 grains per flower (Hii Citation2004). We found that ‘Hayward’ requires 2000–2500 pollen tubes to enter the style for maximum seed set, with a ratio of about 2 pollen tubes for each seed. This measurement is within the 1870–2960 range proposed by Hopping and Jerram (Citation1979). Given a typical pollen viability of 65%–86% (Hopping Citation1984), this means that 2300–3800 pollen grains must be deposited on the stigmatic surface to achieve full pollination. This contrasts markedly with the 10,600–13,250 pollen grains we estimate must be deposited on the stigma and style by artificial pollination to achieve full pollination.
据报道，“Hayward”的授粉要求每朵花的授粉数量从 1000 到 15,000 粒不等（Hii Citation2004）。我们发现，‘海沃德’需要 2000-2500 个花粉管才能进入花柱，以达到最大结籽率，每颗种子大约有 2 个花粉管。该测量值位于 Hopping 和 Jerram 提出的 1870-2960 范围内（Citation1979）。鉴于典型的花粉活力为 65%–86%（Hopping Citation1984），这意味着必须将 2300–3800 个花粉粒沉积在柱头表面才能实现完全授粉。这与我们估计必须通过人工授粉将 10,600-13,250 个花粉粒沉积在柱头和花柱上才能实现完全授粉形成鲜明对比。
▲New Zealand Kiwi Orchard
Indeed, the magnitude of difference in the apparent pollen requirement of ‘Hayward’ reported in the literature seems to be a result of differing methodologies to measure this requirement. One group of estimates is centred around 1000–3000, made up of studies counting pollen tubes (Hopping and Jerram Citation1979; Sheat et al. Citation1982; Stevens and Forsyth Citation1982), and another around 10,000–15,000, counting pollen washed off stigmas and styles; (Hopping and Hacking Citation1983; Hopping Citation1984; Goodwin and Haine Citation1995). These different approaches to assessing the pollen requirement of kiwifruit are each useful in different contexts; for artificial pollination, the larger number is likely a better assessment as it includes the significant pollen wastage by depositing pollen all along the style in addition to the stigma, with only one-third to one-fourth of the pollen germinating down the style (Hopping and Hacking Citation1983). The smaller number may be more appropriate for assessing pollen deposited by insects, which is preferentially deposited on stigmas due to their sticky surface and electrostatic forces (Thomson and Eisenhart Citation2003; Hii Citation2004).
事实上，文献中报道的“海沃德”表观花粉需求的差异程度似乎是衡量这一需求的不同方法的结果。一组估计集中在 1000-3000 个左右，由对花粉管进行计数的研究组成（Hopping 和 Jerram Citation 1979；Sheat 等人 Citation 1982；Stevens 和 Forsyth Citation 1982），另一组估计大约 10,000-15,000 个，对从柱头和花柱上冲走的花粉进行计数; （Hopping and Hacking Citation 1983；Hopping Citation 1984；Goodwin and Haine Citation 1995）。这些评估猕猴桃花粉需求的不同方法在不同的情况下都有用。对于人工授粉，较大的数字可能是更好的评估，因为它包括除柱头外沿整个花柱沉积花粉所造成的显着花粉浪费，只有三分之一到四分之一的花粉沿着花柱发芽（跳花）和黑客引用 1983）。较小的数字可能更适合评估昆虫沉积的花粉，由于其粘性表面和静电力，花粉优先沉积在柱头上（Thomson 和 Eisenhart Citation2003；Hii Citation2004）。
▲Large packaged product kiwi pollen大包装商品花粉
In comparison to the 10,600–13,250 pollen grains on the stigma and style required to produce full seed set in ‘Hayward’, ‘Zesy002’ requires only 3500–4000 pollen grains. That green-fleshed kiwifruit has a higher pollen requirement than gold kiwifruit is reflected in the number of honey bee visits required to fully set seed, with yellow-fleshed varieties A. chinensis var. chinensis ‘Hort16A’ and ‘Zesy002’ requiring ∼5 (Goodwin et al. Citation2013) and 6 (Goodwin et al. Citation2017) visits, respectively, for maximum seed set compared with over ∼40 for green-fleshed ‘Hayward’ (Goodwin and Haine Citation1995). Their maximum seed numbers, 600–700 for ‘Hort16A’ (Goodwin et al. Citation2013; Seal et al. Citation2013) and 500–700 for ‘Zesy002’, are also much less than the 1400–1500 typically observed for ‘Hayward’ (Hopping Citation1984) The underlying reason for the difference in pollen requirement between ‘Hayward’ and ‘Gold3’ may be due to the latter having fewer ovules (Alan Seal, pers. comm.); this does not hold true for all gold-fleshed cultivars, however, with ‘Hayward’ (up to 1716; Seal and McNeilage Citation1988) having comparable numbers to ‘Hort16A’ (∼1650; Seal et al. Citation2013).
与‘Hayward’中产生完整种子所需的柱头和花柱需要 10,600-13,250 个花粉粒相比，‘Zesy002’只需要 3500-4000 个花粉粒。绿肉猕猴桃比黄金猕猴桃需要更高的花粉量，这体现在完全结籽所需的蜜蜂访问次数上，黄肉品种 A. chinensis var. chinensis 'Hort16A' 和 'Zesy002' 分别需要约 5 次（Goodwin 等人 Citation2013）和 6 次（Goodwin 等人 Citation2017）访问才能获得最大种子集数，而绿肉 'Hayward'（Goodwin 和海恩引文 1995）。它们的最大种子数“Hort16A”为 600-700 个（Goodwin 等人 Citation2013；Seal 等人 Citation2013），“Zesy002”为 500-700 个，也远低于“Hayward”通常观察到的 1400-1500 个（ Hopping Citation1984）“Hayward”和“Gold3”花粉需求差异的根本原因可能是后者的胚珠较少（Alan Seal，个人通讯）；然而，这并不适用于所有金肉品种，“Hayward”（截至 1716 年；Seal 和 McNeilage Citation 1988）的数量与“Hort16A”（∼1650；Seal 等人 Citation 2013）相当。
▲猕猴桃雄花采摘运输
We found that ‘Hayward’ remained receptive throughout the day and night, with some between-year variation. This opens the possibility for complementary pollination from both artificial pollination and alternate floral visitors outside of the late morning to early afternoon peak activity period of honey bees. While honey bees may leave the orchard in the afternoon due to relatively low pollen supply (Goodwin Citation1987), pollen is still available in the orchard and insects with lower pollen requirements than honey bees may assist in transfer in the late afternoon and evening and as flowers open during the night and early morning (Thakur and Rathore Citation1991; Goodwin et al. Citation2013). Other insect species have been observed foraging on kiwifruit in the late afternoon and into the evening (Macfarlane and Ferguson Citation1983; Miñarro and Twizell Citation2015; Howlett et al. Citation2017), which may be providing this service already, and there is the possibility to apply artificial pollination outside the typical window of honey bee foraging – increasing flexibility around a very busy time of year.
我们发现“海沃德”在白天和晚上都保持着接受性，但年份之间存在一些差异。这为人工授粉和在上午晚些时候到下午早些时候蜜蜂活动高峰期之外的交替花客进行补充授粉提供了可能性。虽然由于花粉供应相对较少，蜜蜂可能会在下午离开果园（Goodwin Citation1987），但果园中仍然有花粉，并且花粉需求量低于蜜蜂的昆虫可能会在下午晚些时候和晚上协助转移，并作为花朵夜间和清晨开放（Thakur 和 Rathore Citation 1991；Goodwin 等 Citation 2013）。已观察到其他昆虫物种在下午晚些时候和晚上以奇异果为食（Macfarlane 和 Ferguson Citation 1983；Miñarro 和 Twizell Citation 2015；Howlett 等人 Citation 2017），这些昆虫可能已经提供了这项服务，并且有可能申请在蜜蜂典型的觅食窗口之外进行人工授粉——增加了一年中非常繁忙的时间段的灵活性。
▲猕猴桃雄花展示
Our results on full pollination consider seed set alone as a measure of full pollination. Other fruit attributes, such as fruit weight (McKay et al. Citation1979; Hopping Citation1984; Costa et al. Citation1993; Howpage et al. Citation2001; Goodwin et al. Citation2013) and dry matter (Buxton Citation2005) are also correlated with pollination, often reaching asymptotes before full seed set is achieved. However, the response of these other factors is also influenced by non-pollination related variables like irrigation (Chartzoulakis et al. Citation1993), plant age (Hopping Citation1984) and disease status (Mauri et al. Citation2016). Other factors this study did not address are pollen and cultivar ploidy. ‘Hayward’ is hexaploid and ‘Zesy002’ is tetraploid – both were pollinated with ‘Chieftain’ pollen, which is hexaploid. Mismatches in ploidy can cause deficiencies in seed development (Wu Citation2014; Seal et al. Citation2016), and fruit size (Seal et al. Citation2013, Citation2016). However, the slope of the effect of additional seeds on fruit weight appears to change little across genotypes (Seal et al. Citation2013), and we observed seed numbers and fruit weights similar to open pollinated orchards for both varieties, giving us confidence that full pollination was achieved. If using hexaploid pollen to pollinate the tetraploid ‘Zesy002’ had a detrimental effect on seed set, we would have over-estimated how many pollen grains it requires – so the difference between it and ‘Hayward’ would be even greater than reported.
我们关于完全授粉的结果将单独的结籽视为完全授粉的衡量标准。其他果实属性，例如果实重量（McKay 等人 Citation1979；Hopping Citation1984；Costa 等人 Citation1993；Howpage 等人 Citation2001；Goodwin 等人 Citation2013）和干物质（Buxton Citation2005）也通常与授粉相关。在获得完整的种子集之前达到渐近线。然而，这些其他因素的响应也受到非授粉相关变量的影响，如灌溉（Chartzoulakis 等人 Citation1993）、植物年龄（Hopping Citation1984）和疾病状态（Mauri 等人 Citation2016）。这项研究没有涉及的其他因素是花粉和品种倍性。 “Hayward”是六倍体，“Zesy002”是四倍体——两者都是用六倍体“Chieftain”花粉授粉的。倍性不匹配会导致种子发育缺陷（Wu Citation2014；Seal 等人 Citation2016）和果实大小缺陷（Seal 等人 Citation2013、Citation2016）。然而，额外种子对果实重量的影响斜率似乎在不同基因型之间几乎没有变化（Seal 等人，Citation2013），并且我们观察到两个品种的种子数量和果实重量与开放授粉果园相似，这使我们有信心充分授粉已实现。如果使用六倍体花粉为四倍体“Zesy002”授粉对结实产生不利影响，我们就会高估它需要多少花粉粒，因此它与“Hayward”之间的差异将比报道的还要大。
▲新西兰的猕猴桃雄花园Kiwi Male Garden in New Zealand
Understanding variety-specific pollination requirements has a number of potential advantages both for growers and agriculture more broadly. Knowing that ‘Zesy002’ has a lower pollination requirement than ‘Hayward’, it is possible to reduce hive stocking numbers from the recommended 8 hives/ha for green-fleshed varieties (Goodwin Citation2012) and do artificial pollination at lower rates than presently practised to both save costs and increase the efficiency of pollen and bee hive use across the landscape. There is also scope for increased reliance on alternative pollinators, which, while they may be less efficient than honey bees, would be doing proportionally more work per visit on cultivars with lower pollination requirements. Better cultivar-specific pollination management provides growers with insurance against pollinator losses and the potential to obtain increases in production by delivering the optimal amount of pollen to their crop – ultimately safeguarding food production both locally and regionally.
了解特定品种的授粉要求对于种植者和更广泛的农业来说具有许多潜在的优势。知道“Zesy002”的授粉要求比“Hayward”低，因此可以将绿肉品种的蜂箱饲养数量从建议的 8 个蜂箱/公顷（Goodwin Citation2012）减少，并以低于目前实践的比率进行人工授粉。既可以节省成本，又可以提高花粉和蜂巢在整个景观中的使用效率。对替代授粉媒介的依赖也有可能增加，虽然它们的效率可能低于蜜蜂，但每次访问对授粉要求较低的品种来说，它们的工作量会成比例地增加。更好的针对特定品种的授粉管理为种植者提供了防止授粉媒介损失的保险，并有可能通过向作物提供最佳数量的花粉来增加产量，最终保障当地和区域的粮食生产。
▲正在采摘猕猴桃雄花
▲sungold g3 kiwifruit packing
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