1. Introduction
The Persian walnut (
Juglans regia L.) is one of the most economically important cultivated nut tree species. Walnut tree cultivation is widely distributed and, on a global basis, walnuts rank first in nut production (4,498,442 Mt), following by cashews and almonds [
1]. During the last decade, worldwide walnut production has doubled, due to the recognition of its beneficial effects for consumers’ health. Various studies and reviews have highlighted the importance of walnut consumption as part of a regular, healthy diet, due to their content, including a wide range of nutrients and many phytochemical species that may be important to human health, such as a-linolenic acid and other PUFAs, phytosterols, minerals, γ-tocopherol, polyphenols and others, that exhibit antioxidant properties and offer protection against the development of many cardio-vascular related diseases, age-related neurological disorders, incidence of diabetes, and even some types of cancer [
2,
3,
4,
5,
6].
In Greece, walnut production ranks first among all nuts. With an average production of 31,040 tons (year 2019) [
1], Greece is the third largest walnut producer in the European Union after Romania and France. Despite the remarkable production of nuts, Greece imports nuts, mainly walnut kernels (4071 tons in 2019) [
1].
Walnut cultivation in Greece is characterized by two completely different situations. On the one hand, there are the new walnut orchards, which are increasing progressively, in which selected universal cultivars are cultivated, such as the lateral bearing cultivars, “Chandler”, “Lara” and “Pedro”, in the semi-mountainous and lowland areas, and the apical bearing cultivars, Franquette and Ronde de Montignac, in the mountainous and high-mountainous regions. On the other hand, there are the traditional, old walnut populations (seedlings), the cultivation of which has decreased over the years. Twenty-seven and seventy of these seedlings have been previously evaluated by Rouskas et al. (1997) [
7] and Rouskas and Zakynthinos (2001) [
8], respectively. Of the seedlings evaluated, a large number showed lateral bearing habit, and the majority were characterized as early-breaking material. According to the study authors, no genetic material was found that could be linked directly to the “great cultivar”, but there was very valuable genetic material that could be used as genitors in pre-planned crosses directed towards the genetic improvement of walnuts.
Nut production is progressively moving from a traditional situation to a new one involving an intensive approach which emphasises quality of the plant material (e.g., cultivars with lateral fruiting and rootstocks), increased plantation tree density, and mechanization of cultivation, harvesting and post-harvesting, which are some the features that are very important for the development of the Greek walnut industry today [
8]. Furthermore, the relief of the Greek mountains influences the climatic conditions and creates a variety of suitable regions for walnut cultivation [
9]. Therefore, there is a great need to develop new and improved walnut cultivars that best meet the needs of the growers, as well as the specific characteristics of the respective growing areas. This need worldwide, together with the fact that the walnut industry is faced with substantial losses due to pests and diseases affecting both cultivars and rootstocks, has driven the development of walnut breeding programs in the major producing countries. Many breeding programs for walnut varieties have been carried out in the USA, Turkey, Iran, France, and Hungary; some of these programs are still running [
10,
11,
12,
13,
14,
15]. The main goals of breeding in walnut production are high yield and kernel quality. Other characteristics, such as lateral bearing, nut and kernel weight, nut size, kernel percentage, shell thickness, kernel color and ease of kernel removal, are among the important objectives of walnut breeding, along with tolerance to biotic and abiotic factors [
7,
12,
13,
14,
16,
17,
18]. As a result of global warming and climate change, some other breeding objectives have also been considered, such as late leafing to avoid spring late frost, which is one of the most serious problems causing loss of production in walnuts. Late leafing is a desirable attribute for walnuts grown in regions prone to late spring frosts [
19].
Efforts have been made by the Department of Nut Trees of Hellenic Agricultural Organization (HAO)—DIMITRA, from 1985 to the present day, to create new distinctive walnut cultivars, with high productivity and good fruit quality, suitable for the soil and climatic conditions in different regions of Greece, and resistant to pests and diseases, as well as to develop cultivars with late leafing, high lateral fruit bearing, and early harvest dates. Some of the grafted seedlings that have been evaluated [
7,
8] were selected because of their desirable nut traits and their resistance to walnut blight and anthracnose, as well as for their lateral bearing habit, etc. Their crosses with commercial cultivars (e.g., “Chandler”, “Gustine”, “Franquette”) have been established in the department’s collections and are now being assessed. In addition, crosses between well-known commercial cultivars (e.g., “Chandler”, “Hartley”, “Gustine”, “Pedro”, “Franquette”) have been performed to take advantage of each cultivar’s pertinent traits. Recently, through specific crosses, a lateral bearing cultivar with satisfactory traits, called “Ourania”, has been developed. It is a new walnut cultivar of Greek origin that was created by the cross between “Hartley” and “Gustine”. It is a mid-late cultivar, with similar pomological characteristics to “Hartley” but has a high lateral bearing habit. It is a walnut cultivar of great importance, not only for the region of Greece, but also for other countries, suitable for cultivation in semi-mountainous and mountainous regions, where the last spring frost occurs until mid-April [
20]. At the same time, from another cross between “Gustine” and “Pedro”, a new cultivar was created, named “Leto”. “Gustine” was selected as the maternal genetic material mainly because of its high lateral bearing habit and light kernel color, whereas “Pedro” was chosen for the good hull dehiscence and for the later bud break compared to “Gustine”. Thus, the aim of the present study was to provide insights into the characteristics of the newly created “Leto” cultivar and its performance. Comparisons of the most important phenological and pomological characteristics were made with its parental cultivars “Gustine” and “Pedro”, as well as with the best-selling walnut cultivar “Chandler”.
3. Discussion
In the Department of Nut Trees of HAO—DIMITRA, to create new walnut cultivars suitable for the various geographical and meteorological conditions of Greece, but also for other Mediterranean and Mediterranean-like areas, characteristics such as late leafing, high lateral fruit bearing, early harvest dates, resistance to pests and diseases, high yield, and good nut quality, were considered when deciding on crosses. Selected seedlings, and their crosses with commercial cultivars (e.g., “Chandler”, “Gustine”, “Franquette”), as well as crosses between widespread cultivated cultivars (e.g., “Chandler”, “Hartley”, “Gustine”, “Pedro”, “Franquette”), have been established in collections in the department and are now being evaluated. So far, a lateral bearing cultivar called “Ourania”, which emerged from a cross between “Hartley” and “Gustine”, has been described and suggested for cultivation in semi-mountainous and mountainous areas [
20]. In addition, the new “Leto” cultivar was created from the cross of “Gustine”, which was selected because of its high lateral bearing habit and light kernel color, and “Pedro”, which was selected because of good hull dehiscence and later bud break in relation to “Gustine”.
“Leto’s” phenological description shows that this new walnut cultivar has high lateral bearing habit, similar to “Chandler”, as well as the newly described “Ourania” [
20]. Lateral bud fruitfulness is a significant characteristic and criterion when choosing a cultivar for plantation and one of the most important traits in walnut breeding programs [
14,
21], as this habit makes a genotype more productive in terms of yield capacity [
22]. According to Hendricks et al. (1998) [
23], varieties with high (80–90%) lateral bud fruitfulness bear more heavily in the early years than varieties with low (0–10%) lateral bearing-habit. “Leto” presents low tree vigor, is a protandrous cultivar, and flowers at early age, as female flowers appear in the first year, while catkins appear in the third year of age, similar to parental cultivars. It is a mid-early cultivar and it has an earlier bud break in relation to its paternal “Pedro”, as well as to “Chandler” and “Ourania”. According to Hendricks et al. (1998) [
23], date of leafing and blooming are important considerations when choosing a cultivar. Early-leafing cultivars are susceptible to late spring frosts, thus reducing tree fruiting and yield, or they are susceptible to rain-related disease problems, such as walnut blight. For example, very early cultivars, such as “Serr”, could be cultivated in regions with a mild Mediterranean climate, while late cultivars, such as “Franquette”, could be cultivated in mountainous areas, where the last frosts often occur in late April. According to climatic diversity and the late spring frosts in Greece, four regions for walnut cultivation can be distinguished. The first includes the areas where the last spring frosts occur in the first fifteen days of March. The second region includes areas where the last spring frosts take place in late March. The third and fourth regions include areas where the last spring frosts occur in the first fifteen days of April and late April, respectively [
8]. Thus, the newly created cultivar “Leto” is suitable for cultivation in regions where the last spring frosts occur in late March to early April, while “Ourania” is a mid-late cultivar and is suggested for cultivation in semi-mountainous and mountainous regions, where the last spring frosts occur until mid-April [
20]. “Leto” female blooming starts and ends later than its maternal “Gustine” and earlier than “Chandler”, whereas male blooming starts and ends earlier than its paternal cultivar “Pedro”, and “Chandler”.
Regarding tree size, “Leto” has a lower crown radius and trunk circumference compared to its parental cultivars and “Chandler”, a trait that make this new cultivar suitable for dense plantations as well as in unfavorable areas where mechanical harvesting is not possible.
“Leto” nut harvest occurs in late September, similar to “Pedro”, later than “Gustine”, but earlier than “Chandler” and “Ourania”. The harvest date is another important phenological parameter that determines the economic potential of a cultivar, which directly affects the spot value of the marketed products. Although this applies to walnut, an early harvest date can also be viewed as a protective measure against early-autumn frost events [
24]. Early harvest dates offer walnut producers the opportunity to enter the walnut market earlier and thus achieve higher market prices. Another, benefit of an early harvest date is the reduced need for a walnut drying process, which saves energy and reduces drying costs. Therefore, the early harvest date of “Leto” can be considered another promising feature of the phenological traits of this cultivar.
“Leto” walnuts show easy hull dehiscence, similar to “Chandler”, intermediate sunburn susceptibility of the hull, in contrast to its parental cultivars which presented high sunburn susceptibility, intermediate susceptibility to codling moth, and lower susceptibility to walnut blight than “Gustine”. However, the “Ourania” cultivar presented lower sunburn susceptibility of the hull and lower susceptibility to walnut blight [
20].
Pomological assessment helps to determine fruit quality and has therefore been widely used in many breeding studies to identify fruit cultivars with superior characteristics. According to the current results, the nuts of the “Leto” cultivar are medium in size and have an ovate shape and their dimensions, shell texture, color, and integrity, are similar to those of the maternal cultivar “Gustine”. Similar to the parental cultivar, the shell is thicker than “Chandler”. The most important characteristics when considering a walnut cultivar are the nut weight, the kernel weight, the kernel percentage, the kernel color, the kernel fill, and the easy removal of the kernel halves [
24]. The mean dried nut weight (in shell) of “Leto” is similar to its parental cultivars, but lower to both “Chandler” and “Ourania” [
20]. However, the mean kernel weight is high, similar to the reference cultivar “Chandler”. The latter leads to a high kernel percentage of “Leto” nuts (49.37 ± 2.81%), similar to “Chandler”, and higher than “Pedro”. “Ourania” showed a lower kernel percentage (46.92 ± 0.82%) [
20]. The kernel percentage is considered a significant trait for the evaluation of a cultivar with regard to its productivity [
23]. Other positive, desirable nut characteristics of “Leto” are its light kernel color, well kernel filling, and easy removal of the kernel halves.
4. Materials and Methods
4.1. Materials and Experimentation Site Description
The study was carried out in the Department of Nut Trees of the Institute of Plant Breeding and Genetic Resources of HAO-DIMITRA. In an effort to obtain new productive and high-quality walnut genotypes, a cross between “Gustine” (maternal parent) and “Pedro” (paternal parent) by researcher Dimos Rouskas was made in 1986 as part of a European program (Noix UE, 1986). The certified parental cultivars were obtained from the Institut National de la Recherche Agronomique (INRA, Bordeaux). “Gustine” was chosen mainly for its high lateral bearing habit and light kernel color, while “Pedro” was chosen for its good hull dehiscence and the fact that the bud break occurs a week later than “Gustine”. The genotype was grafted onto seedling rootstock of the Lozeronne cultivar and the plants were planted in 1990 at a planting distance of 9 × 9 m. Collections of the parental cultivars “Gustine” and “Pedro”, as well as of “Chandler”, of similar age, were established in the same experimental orchard of the Department of Nut Trees and were included in the trial as reference cultivars.
The study area was located in Lamia, Fthiotida of Greece at the coordinates 38°49′36″ N 22°26′27″ E and at an elevation of 15 m and a slope of approximately 1%. In general, the same standard cultivation practices (e.g., fertilization, irrigation, pruning, pest and disease control) and care were applied to all trees of all cultivars studied. Irrigation was carried out through pipes with four micro drippers of 30 L/h around each tree. The applied pruning system was cup-shaped with a central axis for all trees of all cultivars examined.
Soil examination (at 0–30 cm, 30–60 cm, and 60–90 cm depth) provided the following characteristics: (a) mechanical soil composition: clay 52–56%, silt 32–36%, sand 8–14%, soil characterization as clayey [
25], (b) water saturation 65–70%, electrical conductivity 0.55–0.59 mS/cm, total salts 0.2%, pH 7.7 [
26], (c) calcium carbonate 11.0% and 2.8% organic matter.
4.2. Meteorological Data
The meteorological data were collected from the meteorological station, which is set up in the institute’s facilities. For the years examined (2010–2019), meteorological data (e.g., average monthly temperature, minimum and maximum temperature, average monthly precipitation and relative humidity), for the studied years (2010–2019), were recorded and mean values (±SD) were calculated, as shown in
Figure 3 and
Figure 4.
4.3. Tree and Phenological Characteristics Observation
Tree and phenological characteristics (
Table 1,
Table 2,
Table 3 and
Table 4) of the new cultivar “Leto” as well as its parental cultivars and “Chandler”, which were included in the trial, were collected, from ten (10) trees per cultivar, for ten consecutive years from 2010 to 2019, and were evaluated according to International Plant Genetic Resources Institute (IPGR, 1994) [
27] criteria.
4.4. Nut Traits Measurements and Observation
For the evaluation of the pomological characteristics (nut characteristics) of “Leto” and the other three reference cultivars (
Table 5 and
Table 6), 20 nuts per cultivar were collected at harvest time, each year, between 2010 and 2019 and assessed according to the criteria of IPGR (1994) [
27] and UPOV-TG/125/6 (1999) [
28]. Nut height, nut width, nut thickness and nut roundness index ((nut width + nut thickness)/2) × ut height) were defined according to the criteria of UPOV (1999) [
28] and the measurements were carried out with a digital caliper of 0.01 mm. In addition, in shell nut weight and kernel weight were determined with a digital scale (0.01 g). The kernel weight to in shell nut weight ratio was calculated as kernel percentage (%). The nut shape and shell characteristics of the examined cultivars were determined according to the criteria of IPGR (1994) [
27], as well as UPOV (1999) [
28], as indicated in the tables. The color of the kernel was evaluated according to the criteria of UPOV (1999) [
28].
4.5. Assessment of Susceptibility to BIOTIC and Abiotic Factors
The susceptibility of hulls of cultivars to sunburn was studied for two years (2018–2019) under natural conditions on summer days at temperatures above 42 °C. Nuts that were not covered by tree foliage and exposed to sunlight were examined for sunburn symptoms, and the degree of susceptibility was calculated (%). In order to categorize the sunburn sensitivity of each cultivar according to IPGR (1994), the following categorization was carried out: susceptibility scale 1: 0–20%, 3: 21–40%, 5: 41–60%, 7: 61–80% and 9: 91–100%. The susceptibility of nuts of cultivars to codling moth (Cydia pomonella L.) and walnut blight (Χanthomonas arboricola pv. juglandis) was investigated for two years (2018–2019). The susceptibility to both factors was assessed under usual phytosanitary orchard control. At the time of harvest, 100 nuts were randomly selected from each cultivar and the number of infected nuts was determined and the percentage (%) was calculated. In order to categorize the degree of susceptibility of each cultivar to codling moth and walnut blight according to IPGR (1994), the following categorization was made: susceptibility scale 0: no signs of susceptibility, 1: 0–20%, 3: 21–40%, 5: 41–60%, 7: 61–80% and 9: 91–100%.
4.6. Statistical Analysis
To compare the bud break and flowering dates of the examined cultivars, calendar days were used (from 1 January as day 1). The data are expressed as mean date ± standard deviation (SD). Data from tree and nut measurements are also expressed as means ± standard deviation (SD). The statistical analysis of all these data was performed with the Statistical Package for the Social Sciences (2011) [
29].
5. Conclusions
“Leto” is a walnut cultivar of great interest, due to the combination of its characteristics, i.e., small tree size, high lateral fruiting, mid-early vegetation season and excellent fruit characteristics, comparable to those of one of the top walnut cultivars “Chandler”. The main advantage of this cultivar is the small size of the tree, which allows easier harvesting, for example, in cases of small available areas (2–8 acres), and in semi-mountainous and disadvantaged areas, where there is no possibility of mechanization of harvesting. The “Leto” cultivar enables significantly denser plantings per acre compared to its competitor “Chandler”. It is suitable as the main cultivar, where the last spring frost occurs in late March to early April, making it a promising cultivar not only for Greece but also for other regions with similar climatic conditions.