SPECIAL ISSUE: ORIGINAL ARTICLES
Evaluation of Self-fruitfulness with Hand-pollination in Crabapples, Malus spp.
2023 Volume 92 Issue 1 Pages 30-35
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2023 Volume 92 Issue 1 Pages 30-35
Based on hand pollination, 217 crabapple cultivars were examined to determine their self-fruitfulness potential. One hundred and six cultivars failed to self-pollinate for fruits set. More than 50% self-pollinated fruit were obtained from 29 cultivars. Especially, the fruit set of ‘Jiringo’, ‘Karafutozumi’, ‘Katherine Crab’, Malus hupehensis, M. scheideckeri, ‘Mary Porter Crab’, ‘Mokoto’, ‘Nepal Apple Collections No. 85-I-74-3 86090-3’, ‘Scheidecker Crab’, ‘Silver Moon Crab’, ‘Tachikaido’, and ‘Tea Crab 81105’ were more than 50% by self-pollination over two years. ‘Jiringo’, ‘Katherine Crab’, ‘Mary Porter Crab’, ‘Scheidecker Crab’, and ‘Tachikaido’ have the potential to be self-compatible cultivars because their self-pollinated fruits contain complete seeds. Non-pollinated fruits of M. scheideckeri, ‘Nepal Apple Collections No. 85-I-74-3 86090-3’, and ‘Tea Crab 81105’ may contain apomictic seeds. Moreover, self-pollinated fruits contained seeds and non-pollinated fruits contained no seeds, suggesting that ‘Karafutozumi’ and ‘Mokoto’ may be self-compatible and parthenocarpic. The study findings not only provide breeding material for self-compatible apple cultivars, but also have led to the discovery of new apomictic and parthenocarpic research materials.
Self-fertility is limited in apple cultivars by gametophytic self-incompatibility whereby the pollen tube growth is arrested. Even cultivars that appear to be self-compatible set more fruit with higher seed content when pollinated with a cross-compatible cultivar. Davary-Nejad et al. (1993) examined the effect of natural and artificial self-pollination on the fruit set and seed content, and found in both cases a lower level of fruit set (0.9 to 1.6%) and seed content (1.8 to 2.8 seeds/fruit) compared to open pollination (11.7% and 6.2 seeds/fruit, respectively). With the exception of a few cultivars, apple cultivars produce very limited fruit from self-pollination treatments or bagging without emasculation (Ishiyama et al., 1995; Komori et al., 1999; Saito et al., 1994). Reproducibility of self-fruitfulness depends on experimental material, but environmental factors are considerably larger than genetic factors (Saito et al., 1994). Gametophytic self-incompatibility is controlled by a single locus, the S locus, with multiple alleles (Frankel et al., 1977). Research on the S locus, an S gene coding S-RNase, of apples should aid in understanding this system (Broothaerts et al., 1995).
Japanese pear (Pyrus pyrifolia Nakai) is an important Rosaceous fruit tree in Japan, and Japanese pear natural mutant cultivar ‘Osanijisseiki’ was released as the first self-compatible cultivar of Japanese pear in 1979 (Furuta et al., 1980). ‘Osanijisseiki’ is self-compatible by deletion mutant S4 gene (Okada et al., 2008). Some apple cultivars have been classified as self-fertile (Crane, 1927). Pseudocompatibility was maximized by pollinating old flowers with large quantities of pollen, and maintaining a temperature of about 20°C during the period of pollen tube growth. Evidence suggests that, with some or all of the conditions, fertile flowers are able to set fruit through self-pollination (Williams and Maier, 1977). The ability of apomixis is characteristic of a number of Malus species, but does not appear to occur among the cultivated apples. The importance of apomixis in Malus species is that the seedlings of some are sufficiently uniform to be used as rootstock breeding (Schmidt, 1988). It is even possible in some apple cultivars and in certain conditions for fruit to develop parthenocarpically without fertilization and without seeds (Janick et al., 1996). The two apetalous apple cultivars ‘Spencer Seedless’ and ‘Ohio 3’ bore annual crops of parthenocarpic seedless fruits because insects do not visit the apetalous flowers (Chan and Cain, 1967). Cultivars that produce parthenocarpic fruits have been consistently used because of their ability to produce good yields in years when flowers are damaged by late spring frosts or when conditions are unfavorable for pollination. As described above, apple self-fruitfulness has three types: self-compatibility, in which self-pollen is fertilized to form seeds; apomixis, in which seeds are formed without fertilization; and parthenocarpy, in which fruit development occurs without fertilization and seeds are not formed.
Crabapples (Malus spp.) are small trees or shrubs in the rose family (Rosaceae), characterized by various types of flowers, colorful and small fruits (≤ 5cm), and various growth habits. They are also valued for their wide environmental adaptability, facilitating worldwide prominence in gardens and as landscape focal points (Höfer et al., 2014; Lisandru et al., 2017). Although nearly 700 cultivars have been named, constituting a genetic background derived from many species, little has been reported on their self-fruitfulness. Based on the results of hand pollination tests, we performed to characterize the self-fruitfulness of 217 crabapple cultivars.
All apple trees were maintained in the field of the Institute of Fruit Tree and Tea Science, NARO, Iwate, Japan. All trees were more than 10 years old, pruned to slender spindle and planted in single-row hedgerows 1 m apart with the rows spaced 4 m apart. One or two trees were used in this study.
Self-pollinationA total of 217 Malus germplasms, mainly from crabapple cultivars, were self-pollinated in mid- to late May from 2000 to 2003 and are listed in Table 1. Blossoms at the balloon stage were bagged, and after the petals expanded and the anthers dehiscenced, two flowers were self-pollinated per cluster by hand pollination and non-pollinated blossoms were removed. Hand pollination was conducted by rubbing anthesis flowers in the same bag. The flowers were bagged for preventing contamination with unknown pollen after hand pollination. Twenty flowers were tested for each crabapple cultivar.
Self-pollination tests with hand-pollination in crabapplesz.
Twenty-one crabapple cultivars, which had over 50% of fruit set by self-pollination in the first year, were self-pollinated again, and then emasculated and non-pollinated. Self-pollination was performed for 20 flowers according to the method described above. For non-pollination style, two flowers at the balloon stage were emasculated per cluster and other blossoms were removed. The flowers were bagged immediately after emasculation. Twenty flowers of ten clusters per crabapple cultivar were emasculated and bagged.
Fruit setting, harvested fruits, and seeds collectionAfter early fruit drop (June drop), fruit setting was counted, the fruits were harvested about 4 months after pollination or emasculation, and complete seeds that sank in water were collected from the harvested fruits. The number of complete seeds per fruit was calculated. For comparison, the number of loculi and seeds in the open-pollinated fruits of ‘Katherine Crab’, ‘Scheidecker Crab’, and ‘Tachikaido’ cultivars were counted.
Self-pollination results indicating lower values during a few years are shown in Table 1. A total of 106 of the crabapple cultivars (48.8% of total investigated) did not produce fruit by self-pollination. Fifty-one cultivars (23.5%) had 20% or more of the 20 fruits set by self-pollination. Over 50% self-pollinated fruits set were obtained from 29 cultivars, equivalent to 13.4% of the 217 tested cultivars. Most apple cultivars, (M. × domestica), are self-incompatible and do not bear fruit through self-pollination. In a previous study of 213 apple cultivars and strains discriminated to be parthenocarpic and self-compatible by bagging the balloon stage flowers without emasculation, 187 (87.8%) had 0% fruit set and 12 (5.6%) had fruiting rates of 20% or more (Komori et al., 1999). In another study, 21 of 27 apple cultivars set less than 20% of self-pollinated flowers without emasculation (Ishiyama et al., 1995). The crabapple cultivars in the present study showed higher self-fruitfulness than that previously reported in apple cultivars and strains. Especially, the percentages of fruit set for ‘Jiringo’, ‘Karafutozumi’, ‘Katherine Crab’, M. hupehensis, M. scheidecker, ‘Mary Porter Crab’, ‘Mokoto’, ‘Nepal Apple Collections No. 85-I-74-3 86090-3’, ‘Scheidecker Crab’, ‘Silver Moon Crab’, ‘Tachikaido’, and ‘Tea Crab 81105’ were more than 50% by self-pollination over two years. Moreover, ‘Daitou 2’, M. × gloviosa 91045, ‘Xifuhaitang’, and ‘Xizanhaitang’ could not set over 50% of self-pollinated fruits over a two-year period, but showed an average self-pollination fruiting rate of more than 50% over two years (Table 2).
Number of set fruits and seeds per fruit with self- and non-pollination testsz.
Compared to the previous results in apple cultivars, the crabapple cultivars that set more than 50% by self-pollination in this study are considered self-fruitful. ‘Jiringo’, ‘Katherine Crab’, ‘Mary Porter Crab’, ‘Scheidecker Crab’, and ‘Tachikaido’ set over 50% of self-pollinated fruits over two years, but could not bear fruit following non-pollination with emasculation (Table 2). Therefore, these cultivars are not parthenocarpic but instead self-fruitful. According to the criteria for self-compatibility of 3 or more seeds per fruit, established by Komori et al. (1999), ‘Jiringo’ and ‘Mary Porter Crab’ are self-compatible. The number of seeds in ‘Katherine Crab’, ‘Scheidecker Crab’, and ‘Tachikaido’ self-fruiting fruits (over 1.2 and less than 3 seeds per fruit) indicate a mixture of compatibility and incompatibility. The average number of loculi in the 10 fruits was 5.4, 5.3, and 5.2 for ‘Katherine Crab’, ‘Scheidecker Crab’, and ‘Tachikaido’, respectively. Since these cultivars have approximately five loculi, the results of seed numbers in self-fruiting fruits can be compared with those of apples cultivars that normally have five loculi. The number of seeds in the open-pollinated fruits of ‘Scheidecker Crab’ and ‘Tachikaido’ was also less than 3 seeds per fruit (Table 3). The number of seeds in self-pollinated fruits of ‘Katherine Crab’ requires investigation, but this cultivar may be self-compatible because the self-pollination fruit set rate was much higher than the standard determined by Komori et al. (1999). ‘Osanijisseiki’ is compatible with the pollen from ‘Nijisseiki’, an original cultivar of ‘Osanijisseiki’, whereas the pollen from ‘Osanijisseiki’ is incompatible with ‘Nijisseiki’. These reciprocal crosses between ‘Osanijisseiki’ and ‘Nijisseiki’ indicate that ‘Osanijisseiki’ is a stylar-part mutant (sm) (Sato, 1993). The deletion region of the mutant S4sm haplotype of ‘Osanijisseiki’ has been completely determined. Okada et al. (2008) determined that the S4sm haplotype lacks ORFs containing S4 RNase, suggesting the factors for self-compatibility in ‘Osanijisseiki’. Structural analysis of the S gene in crabapple cultivars with suggested self-compatibility may provide an opportunity to clarify the origin of self-fruitfulness.
Number of seeds per fruit with open-pollination tests.
M. scheideckeri, ‘Nepal Apple Collections No. 85-I-74-3 86090-3’, and ‘Tea Crab 81105’ produced fruits with over 2 complete seeds per fruit by emasculation and non-pollination (Table 2). This suggests ability of apomixis in those crabapples. Saito et al. (1993) reported that M. scheideckeri, maintained at the Hirosaki University Fujisaki farm and is identical to M. hupehensis based on morphological characteristics and isozyme analysis, is one of the apomictic Malus species suggesting that M. scheideckeri used in this study may also be apomictic M. hupehensis. Since the common name of M. hupehensis is Tea crabapple (Benson et al., 2001; Olien, 1987), ‘Tea Crab 81105’ may have characteristics of apomixis. ‘Nepal Apple Collections No. 85-I-74-3 86090-3’ was obtained from seeds contained in fruit collected in Nepal. Because ‘Nepal Apple Collections No. 85-I-74-4 86090-4’, set 75% of self-pollinated fruits in Table 1, was obtained from the same fruits, ‘Nepal Apple Collections No. 85-I-74-4 86090-4’ is required to confirm the possibility of encompassing apomixis characteristics by emasculation and non-pollination. Pollen of ‘Silver Moon Crab’ could not be visually confirmed at self-pollination; hence, ‘Silver Moon Crab’ may be male-sterile. This suggests that fruits obtained by self-pollination in ‘Silver Moon Crab’ included apomictic seeds. M. corronaria, M. hupehensis, M. sargentii, M. sieboldii, M. sikkimensis, and M. toringoides are known as an apomictic Malus species (Dermen, 1936; Janick et al., 1996; Olien, 1987; Sax, 1959). M. sikkimensis and M. toringoides respectively set 70% and 45% of self-pollinated fruits in a single-year (Table 1) and may include apomictic fruit set.
‘Karafutozumi’ and ‘Mokoto’ produced fruits containing seeds by self- and non-pollination. Most of the non-pollinated fruits in ‘Karafutozumi’ and ‘Mokoto’ contained no seeds (Table 4). Apple cultivar ‘Megumi’ produced seed-containing fruits by self-pollination using stored pollen and seedless fruits by non-pollination with emasculation, indicating that ‘Megumi’ exhibits self-fruiting ability through self-compatibility and parthenocarpy (Saito et al., 1978). Self-pollinated fruits contained seeds and non-pollinated fruits contained no seeds, suggesting that ‘Karafutozumi’ and ‘Mokoto’ may be self-compatible and parthenocarpic. The autotetraploid apple cultivars showed self-fertility, and their pollen was compatible with the pistils of their original diploid cultivars (Adachi et al., 2009). ‘Karafutozumi’ and ‘Mokoto’ are classified under M. baccata, and some M. baccata cultivars are known to be tetraploid (Way et al., 1991). Almost all of the crabapple cultivars that exhibited high self-fruiting potential were of unknown origin, and future studies of ploidy would help to elucidate the factors of high self-fruitfulness. A few seeds found in non-pollinated fruits of ‘Karafutozumi’ and ‘Mokoto’ suggested that these cultivars may be apomictic.
Number of seeds in each harvested fruit with self- and non-pollination with emasculation tests.
‘Alps Otome’, ‘Hanyae-Hanakaidou’, ‘Red Jade’, and ‘Red Splendor Crab’ had over 50% fruit set by self-pollinated in the first year, but less than 10% in the second year (Table 2). It may be that the second-year flowers of these cultivars were not in good condition, thereby warranting future re-examination of self-pollination tests.
Saito et al. (2007) carried out self-pollination of ‘Fuji’ using X-ray irradiated pollen, and many progenies from the self-pollination of ‘Fuji’ were obtained from fruits containing seeds through application of embryo culture. These self-pollinated plants were grown in a green house, but some plants showed abnormal phenotypic appearances. The progeny of ‘Megumi’ showed self-fruiting rates of 30% to 50%, indicating that self-fruiting ability is inherited (Ishiyama et al., 1981). However, no practical self-compatible apple cultivars like the Japanese pear ‘Osanijisseiki’ have been established. The results from the present study not only provide breeding material for self-compatible apple cultivars, but also have led to the discovery of new apomictic and parthenocarpic research materials.
