plants Article Potential Markers for Selecting Self-Eliminating Apple Genotypes Aurelijus Starkus, Birute Frercks *, Dalia Gelvonauskiene, Ingrida Mazeikiene , Rytis Rugienius, Vidmantas Bendokas and Vidmantas Stanys Lithuanian Research Centre for Agriculture and Forestry, Department of Orchard Plant Genetics and Biotechnology, Institute of Horticulture, Babtai, LT-54333 Kaunas, Lithuania; [email protected] (A.S.); [email protected] (D.G.); [email protected] (I.M.); [email protected] (R.R.); [email protected] (V.B.); [email protected] (V.S.) * Correspondence: [email protected]; Tel.:+370-37-555-253 Abstract: The heavy blooming of apple trees results in the inefficient usage of energy and nutritional material, and additional expenditure on fruitlet thinning is required to maintain fruit quality. A possible solution for controlling the fruit load on trees is the development of new cultivars that self-eliminate excess fruitlets, thus controlling yield. The aim of our study was to identify biological differences in apple cultivars in terms of blooming intensity and fruitlet load self-regulation. In total, 19 apple cultivars were studied in the years 2015–2017. The dynamics of fruitlet self-elimination, seed development in fruitlets and fruits, photosynthetic parameters, carbohydrates, and plant hormones were evaluated. We established that apple cultivars self-eliminating a small number of fruitlets need a lower number of well-developed seeds in fruit, and their number of leaves and area per fruit on a bearing branch are larger, compared to cultivars, self-eliminating large numbers of fruitlets. A higher Citation: Starkus, A.; Frercks, B.; carbohydrate amount in the leaves may be related to smaller fruitlet self-elimination. The amount Gelvonauskiene, D.; Mazeikiene, I.; of auxin and a high indole-3-acetic acid/zeatin ratio between leaves of cultivar groups with heavy Rugienius, R.; Bendokas, V.; Stanys, V. blooming were higher than in cultivars with moderate blooming. A lower amount of abscisic acid Potential Markers for Selecting was found in heavy-blooming cultivars during drought stress. All these parameters may be used as Self-Eliminating Apple Genotypes. markers for the selection of different apple genotypes that self-eliminate fruitlets. Plants 2021, 10, 1612. https:// doi.org/10.3390/plants10081612 Keywords: apple; carbohydrates; dynamics of fruitlet self-elimination; leaves; plant hormones; seeds Academic Editors: Gunars¯ Lacis¯ and Hilde Nybom 1. Introduction Received: 10 June 2021 Apple (Malus × domestica Borkh) is one the most important fruit trees in the temperate Accepted: 30 July 2021 Published: 5 August 2021 climate zone. Stable bearing and the production of high-quality fruits are crucial for growers. Only 5–10% of flowers grow to fruits; therefore, heavy blooming results in Publisher’s Note: MDPI stays neutral inefficient usage of energy and nutritional material [1]. To produce a consistent, high- with regard to jurisdictional claims in quality harvest and to eliminate tree biennial bearing in industrial orchards, fruitlet and published maps and institutional affil- fruit thinning is used; however, this process is costly and ineffective in some cases [2,3]. iations. The efficiency of chemical fruit thinning depends on the plant’s physiological condition and the environment [4]. Additionally, the chemicals used for thinning may negatively impact tree growth, fruit bearing, and the populations of pollinators. Apple bearing is determined by many factors: meteorological conditions, agrotechnical methods, and the biological parameters of a tree. One of the desired traits in future apple cultivars is the Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. genetically determined self-regulation of fruit load. The dropping processes of reproductive This article is an open access article organs occur in the region of the somatic body of plant known as the abscission zone (AZ). distributed under the terms and Abscission has been a useful process in evolution; it results in the development and conditions of the Creative Commons spreading of only high-quality fruits and seeds. The transport of indole-3-acetic acid (IAA) Attribution (CC BY) license (https:// through the AZ regulates the sensitivity of the AZ to ethylene [5,6]. AZ becomes sensitive creativecommons.org/licenses/by/ to ethylene when the source of IAA is removed, and abscission starts because of the action 4.0/). of cell membrane enzymes [7]. It has been established that carbohydrates are important Plants 2021, 10, 1612. https://doi.org/10.3390/plants10081612 https://www.mdpi.com/journal/plants Plants 2021, 10, 1612 2 of 16 in fruit drop [8–10]. A lack of carbohydrates increases the production of reactive oxygen species and abscisic acid, thus activating ethylene signaling pathways. As a result, fruit abscission is activated as well. Plants use photosynthesis to produce carbohydrates in leaves, which are used in the control of endogenic and environmental signals. However, the role of leaves in fruitlet drop is unclear. Studies were conducted on the creation of easily handled systems for the advance identification of abscised fruitlets and the planning of technological means for fruitlet load regulation [11]. The authors established that the measurement of fruit growth speed might be used for determining fruitlets that will drop during the June drop [11]. Lee et al. established that frost shock during early fruit development stages results in changes in the expression of abscisic acid synthesis genes and in the activation of AZ movement in the cell cytoplasm [12]. Several studies in model plants identifying genetic and physiological parameters of fruit drop were performed. Possible gene targets for future research and rearrangement using biotechnological methods were proposed [13]. Gene groups responsible for ethylene biosynthesis, auxin carriers, metabolism of carbohydrates, and hydrolytic ferments were established [14]. However, gene expression may be different and specific to various cells and tissues [15]. Knowledge of the physiological, genetic, and genomic aspects related to fruit abscission is sparse. The self-elimination of fruitlets seems to be an important trait, leading to yield stability and lower orchard management costs; therefore, cultivars with this trait are desired by industrial growers. Modern apple breeding programs often include fruitlet self-elimination in breeding schemes. The aim of our study was to evaluate the biological differences in apple cultivars in terms of fruitlet load self-regulation and identify features closely related to self-eliminating fruitlets. 2. Results 2.1. Evaluation of Dynamics of Blossom and Fruitlet Self-Elimination during Vegetation The apple tree load of blossoms and fruitlet self-elimination depend on the cultivar. Fluctuations in blooming are characterized by the variation coefficient, which ranges from 1.9% to 72.7% (Table1). Cultivars with heavy blooming belonging to groups I and II had a similar average blossom number per 1 m of branch during a 3-year period, without significant differences. These cultivars had twice as many blossoms compared to cultivars from groups III and IV, which had a moderate bloom. Cultivar groups I and III showed high self-elimination of fruitlets. However, after the first drop, they had a much higher percentage of remaining fruitlets (18.6–50.8% on average) compared to cultivars in groups II and IV, which showed moderate fruitlet self-elimination (7.7% and 20.1% on average, respectively). The fruitlet number on groups I and III trees decreased more than two-fold until July, whereas the number of fruitlets decreased by one-third on trees in groups II and IV. Therefore, the dynamics of fruitlet self-elimination in cultivar groups were different. Plants 2021, 10, 1612 3 of 16 Table 1. Dynamics of the number of fruitlets during apple tree vegetation in 2015–2017. 3-Year Average Number of Drop I Drop II Coefficient of Blossoms per 1 m of Number of Fruitlets per Number of Fruitlets per Cultivar Variation Branch 1 m of Branch (%) 1 m of Branch (%) (%) ± SD 1 2015 2016 2017 2015 2016 2017 Orlovim 79.4 ± 18.3 23.1 38.4 21.4 8.5 19.1 10.9 6.2 Ottawa 49.9 ± 12.0 24.0 59.7 33.5 34.9 23.7 19.0 17.1 Sonata 63.7 ± 10.1 15.9 22.8 16.4 18.8 8.1 6.4 5.6 Lord Lemburne 77.3 ± 56.2 72.7 16.0 9.7 5.8 6.0 4.6 3.1 Melrose 52.8 ± 15.4 29.2 25.5 27.0 11.0 6.3 18.6 4.8 Selena 81.3 ± 15.5 19.0 61.8 47.7 28.7 20.6 20.0 12.9 Makresa 61.3 ± 21.8 35.6 28.8 31.4 26.5 9.9 17.7 7.9 Montvilinis 88.2 ± 4.7 5.3 46.8 61.4 19.5 20.0 23.7 12.8 McIntosh 52.7 ± 16.3 30.9 9.8 13.2 2.7 3.4 0.6 2.7 Average of group I 67.4 ± 11.6 17.2 32.3 30.9 18.6 14.5 14.0 8.9 Yellow Arkad 46.4 ± 15.1 32.5 36.9 20.6 16.3 23.3 13.6 11.4 Discovery 88.4 ± 36.3 41.0 10.7 3.2 3.2 8.7 1.9 2.3 Albrechtapfel 96.2 ± 35.3 36.7 30.8 5.1 3.3 20.5 3.8 2.2 Osvald 57.7 ± 1.3 2.3 25.9 9.6 21.3 10.2 7.8 9.9 Voshod 67.8 ± 1.3 1.9 0 0.7 - 0 0.3 0 Average of group II 71.1 ± 14.1 19.9 20.1 7.7 8.9 12.8 5.4 5.1 Port Oxford beauty 44.1 ± 16.6 37.6 29.3 54.0 43.5 15.1 13.2 14.5 Aldas 29.8 ± 13.1 43.9 40.8 32.1 36.4 16.7 17.2 30.9 Sandov 36.2 ± 4.2 11.5 54.7 66.2 52.9 17.2 24.3 20.0 Average of group III 35.6 ± 5.3 14.8 49.6 50.8 38.8 20.2 18.5 16.3 Wonchester permine 44.5 ± 13.9 31.3 16.2 10.4 7.3 12.3 5.8 5.9 Hume 32.2 ± 5.5 17.0 15.9 5.5 11.1 9.6 1.6 11.1 Average of group IV 38.3 ± 9.3 24.2 16.0 8.4 8.6 10.8 4.1 7.8 1 Standard deviation.