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Article

Identification of Apple Varieties Resistant to Fire Blight (Erwinia amylovora) Using Molecular Markers

1
Plant Protection and Quarantine Department, Kazakh National Agrarian Research University, Almaty 050010, Kazakhstan
2
TreeGene Molecular Genetics Laboratory, Almaty 050008, Kazakhstan
3
Institute of Plant Biology and Biotechnology, Laboratory of Molecular Biology, Almaty 050040, Kazakhstan
4
Institute of Plant Biology and Biotechnology, Breeding and Biotechnology Laboratory, Almaty 050040, Kazakhstan
*
Author to whom correspondence should be addressed.
Horticulturae 2023, 9(9), 1000; https://doi.org/10.3390/horticulturae9091000
Submission received: 1 August 2023 / Revised: 23 August 2023 / Accepted: 31 August 2023 / Published: 4 September 2023

Abstract

:
Fire blight of fruit crops is one of the most dangerous diseases for apple trees and other plants of the Rosaceae family, and in Kazakhstan, it is subject to quarantine. To study the spread of fire blight, a phytopathological evaluation of 59 apple varieties of domestic and foreign breeds was carried out in various regions of the south and southeast of Kazakhstan while also considering climatic conditions. The susceptibility of an apple tree to fire blight is influenced by the climatic conditions prevailing in a particular fruit region of Kazakhstan. Samples were collected from various varieties of apple trees with fire blight symptoms for molecular genetic analysis. The phytopathological evaluation and results of the PCR analysis made it possible to identify the causative agent of the disease and its spread to apple varieties in the main fruit regions of Kazakhstan. A molecular study of the resistance to the fire blight pathogen was carried out using the most effective molecular markers. A set of 10 (FBE-1_Y320; FBE-2_Y192; FBE-2_Y495; FBE-2_Y551; FB-MR5-K35; FB-MRS-R240; FB-MR5-R249; FB-MR5-rp16k15_M106; RLP1a; and RLP1b) SNPs was selected, including SNPs reported to be associated with three trait loci, as well as the two markers AE10-375 and GE-8019. Interestingly, the SNP analysis revealed that for all ten markers linked to fire blight resistance, the genotypes of all 59 apple cultivars were identical. No differences in the presence or absence of these markers were observed among the studied varieties. The 26 apple varieties of domestic and foreign breeds most resistant to fire blight were identified in the molecular analysis using the markers AE-375 and GE-8019. Among the studied 59 apple varieties, 23 varieties were identified using the AE-375 marker and 7 varieties with the GE-8019 marker. Samuret, Honeycrisp, Pinova, and Red Topaz were found to be resistant using markers AE-375 and GE-8019. The most promising apple varieties for further breeding for resistance to fire blight programs were selected.

1. Introduction

In Kazakhstan, as in most of the world’s countries, the most significant fruit crop is the apple tree (Malus domestica Borkh.), which has been a leader in the increase in fruit production. Orchard areas account for 47.18 thousand hectares in which pome and stone fruit crops are grown, and 75% of this area is used for apple orchards [1]. The State Register of breeding achievements approved for use in the Republic of Kazakhstan, for 2023, includes 73 varieties of apple trees of various ripening periods, 29% of which are varieties of local breeds, obtained from the Kazakh Research Institute of Horticulture, and 38% of which are varieties whereby the originator is not registered.
Fire blight, caused by the bacterium Erwinia amylovora, is one of the most dangerous diseases for Malus domestica and other species of the Rosaceae family. This disease is difficult to control because of the limited or completely abandoned application of antibiotics. Therefore, genetic resistance is thought to be the most sustainable approach to managing fire blight. E. amylovora is subject to quarantine in Kazakhstan. Fire blight, since 2010, has caused significant damage to the apple and pear orchards of Kazakhstan, causing large crop losses and the death of trees. According to Vanneste et al. [2], it affects more than 180 species of fruit and tree shrubs of the Rosaceae family around the world. It has been reported that approximately five hundred thousand fruit trees have been destroyed in Italy [2]. According to Norelli et al. [3], more than 240 hectares of apple trees in southwestern Michigan State, USA, have been affected, with significant economic losses, because of fire blight. In Turkey, fire blight was first detected in a pear orchard in 1985 [4]. Therefore, identifying varieties resistant to the disease as quickly as possible has become a major and worldwide issue [5].
In Kazakhstan, the first symptoms of fire blight were detected in 2008, and by 2010 it had begun to cause significant damage to apple and pear orchards in several areas of the Almaty Region. On some peasant farms, the proportion of affected trees in apple orchards reaches 50–60%, or more, with a high degree of disease symptom development [6,7]. According to the results of a study conducted by Gritsenko et al. [8] under such conditions in Kazakhstan, among the 30 varieties collected from three orchards in the Almaty Region, 23 varieties were infected by the pathogen, including 15 varieties without visible symptoms, and 8 varieties were infected by the pathogen [8].
Currently, a rather extensive range of fruit crops is recommended in Kazakhstan. The most important condition for increasing the economic efficiency of horticulture is the constant improvement of the varietal composition. New varieties should have advantages over existing analogs in terms of productivity, resistance to abiotic and biotic stressors, and fruit quality, and they should be distinguished by novelty, competitiveness, and quick return on investment [9]. The resistance of fruit crop varieties to the disease is the most important indicator that determines their market value. Despite numerous studies conducted in different countries, many issues related to the resistance of fruit crop varieties to fire blight have not been sufficiently studied. The creation of new varieties of perennial plants consists of several cycles of hybridization and can last, depending on the crop, an average of 20–25 years. Over the last decade, real prerequisites for the transition from traditional breeding techniques based on phenotypic traits to approaches that involve genomic characteristics, such as the identification of “trait–locus” associations, have appeared.
E. amylovora strains differ in virulence, which can lead to varying degrees of fire blight severity and complicate the determination of the genetic basis for the degree of susceptibility to the disease [10,11,12,13]. Most of the resistant forms are concentrated among the wild forms of the apple tree, such as Malus robusta, M. sublobata, M. atrosanguinea, M. prunifolia, and M. fusca. High resistance was also noted in other species: M. baccata [14], M. robusta var. persicifolia, and M. sieversii [15].
In the development of organic farming, the greatest priority with the most environmentally friendly direction is the cultivation of disease-resistant varieties and rootstocks of apple trees. An increased level of resistance to diseases is one of the most important requirements for modern varieties of agricultural plants, including fruit crops. Currently, the priority is to create varieties of fruit crops with genetic resistance to the most harmful diseases; in particular, fire blight. Resistance to this pathogen is an important goal of many apple breeding programs [16,17]. Therefore, optimal methods for assessing genetically determined resistance to pathogens will always be in demand. Breeding apple tree varieties with genetic resistance offers a solution to fire blight [18].
Based on the results of a study conducted in 2007, two dominant SCAR markers were created [19]. By inoculating plants with the pathogen, it was found that genotypes that carried both markers were more resistant than those that lacked them. According to research conducted by Drenova and others, in Russia, it was found that out of 54 analyzed samples, 11 varieties and 1 hybrid form (Avgusta, Zhelannoe, Zaslavskoye, Cinnamon, Orlinka, Skryzhapel, Svezhest, Stroevskoye, Apple Spas, Honey Crisp, Fire Sid, and 376-106) are potentially resistant to the disease and can be used as gene sources of the trait in breeding programs [6]. Omasheva et al. [20] identified apple varieties of Kazakh breeds, which were tested for the presence of alleles associated with fire blight resistance using AE10-375 and GE-8019 markers. According to the researchers, only two of the studied varieties (Alatau Column and Maximus) showed a positive result for both markers. The AE10-375 and GE-8019 markers flank the FBF7 QTL; the AE10-375 marker is located at a distance of 4 cm from the QTL peak, and the GE-8019 marker is located at a distance of 6 cm from the QTL peak. The CH-F7-Fb1 marker maps to the same side of the QTL as AE10-375 and is used to improve the reliability of molecular genetic analyses [21].
The aim of this research was to analyze domestic and foreign apple varieties using SNP and SCAR markers to identify genetic sources of fire blight resistance.

2. Material and Methods

2.1. Sample Collection

For the timely detection of fire blight, regular examinations of apple plantations in the south and southeast of Kazakhstan were carried out during the growing season according to the methods for detecting and identifying the fire blight agent of fruit trees [22,23]. The monitoring of the diseases and sampling for the molecular genetic analysis was carried out in a total of 8 orchards, including the pomological orchard of the Kazakh Research Institute of Horticulture and commercial and farm orchards in the main commercial areas of horticulture: Turkestan (2 orchards), Zhambyl (1 orchard), and Almaty Regions (5 orchards). In this study, 59 apple varieties approved for use in the Republic of Kazakhstan were used, including 24 varieties of Kazakhstani breeds, as well as 35 varieties of foreign breeds. In addition, five varieties not approved for use in the Republic of Kazakhstan were studied. The samples were collected from various varieties of apple trees that displayed symptoms of fire blight, including browned and necrotic young shoots, as well as those with the characteristic shape of a “shepherd’s crook”, and samples were also taken from trees without visible symptoms. The selected samples of the 59 studied apple varieties were delivered to the laboratory and kept in a freezer (−80 °C) until the DNA was isolated (Table 1).
The disease distribution index (DDI) was assessed in mid-July using a 5-point scale according to the following classification: 0—asymptomatic; 1—sporadic symptoms of fire blight, with less than 10% of trees affected; 2—10% to 20% of trees affected by pathogen; 3—20% to 50% of trees affected by the pathogen; 4—50% to 100% of trees damaged by infection. The DDI was calculated by dividing the number of fire-blight-affected trees of 1.000 randomly examined trees in the field multiplied by 100 [72]:
DDI = (fire blight shoots/total shoots) × 100
The disease severity index (DSI) was calculated using the following formula:
D S I = Σ   c l a s s   o f   f r e q u e n c y ×   s c o r e   o f   r a t i n g   c l a s s     T o t a l   n u m b e r   o f   o b s e r v a t i o n s × m a x i m a l   d i s e a s e   i n d e x      
The DDI and DSI were evaluated using only three apple varieties, namely Golden Delicious, Idared, and Aport, because these varieties are more often infected by fire blight. Over the last 5 years, these varieties have increasingly become rapidly infected, and according to the results, a detailed analysis was conducted for these main susceptible varieties.

2.2. DNA Isolation

Total DNA was isolated with a silica gel membrane and resuspended in TE buffer (10 mM Tris; 0.1 mM EDTA). The DNA concentration was determined using a nanospectrophotometer and normalized to 20 ng/µL [19].

2.3. SNP Genotyping

Resistance to fire blight caused by E. amylovora was identified using 10 SNP molecular markers: FBE-1_Y320, FBE-2_Y192, FBE-2_Y495, FBE-2_Y551, FB-MR5-NZsnEH034548_K35, FB-MR5-NZsnEH034548_R240, FB-MR5-NZsnEH034548_R249, FB-MR5-rp16k15_M106, RLP1a, and RLP1b in the FBE, MR5, and RLP1 genes [73]. All 10 markers are true SNPs that have previously been successfully evaluated using direct sequencing technologies [73,74]. Primers and Taqman® (Life Technologies Corporation—Pleasanton, 6055 Sunol Boulevard, Pleasanton, CA, USA) probes were designed using the Custom Taqman® Assay Design Tool (Thermo Fisher Scientific, Waltham, MA, USA) [75]. The Taqman assay ID is provided in Table 2.
The DNA extracts (20 ng each) were mixed with an equal volume of TaqMan® OpenArray® Genotyping Master Mix (Life Technologies Corporation—Pleasanton, 6055 Sunol Boulevard, Pleasanton, CA, USA) according to the instructions and amplified using the QuantStudio™ 5 Real-Time PCR System (Thermo Fisher Scientific).
The results were analyzed using Quantstudio® Design and Analysis software and Taqman® Genotyper (Thermo Fisher Scientific). Because the assays were newly designed, each genotyping result was manually verified by viewing the real-time trace and fluorescence endpoints. Any manual changes were saved using the Taqman® Genotyper Software and exported as a genotype matrix for each individual sample.

2.4. SCAR Markers

In the present work, two markers associated with fire blight resistance, AE10-375 and GE-8019 [19], were used (Table 1). For each DNA sample, 60 ng DNA was amplified in a 25 µL reaction mix containing 1× Taq buffer (750 mM Tris HCl, pH 8.8, 200 mM (NH4)2SO4, 0.1% Tween 20), 2.5 mM MgCl2, 0.2 mM dNTPs, 0.2 mM of each of the respective primers, and 1 unit Taq polymerase (Thermo Scientific, USA). The PCR cycling conditions for every marker are described in Table 3.

3. Results

3.1. Monitoring

Observation of the disease distribution index (DDI) and disease severity index (DSI) of fire blight in apple trees in the main fruit regions of Kazakhstan.
The routine inspection of apple orchards, to determine the DDI and DSI of the disease, was carried out over two years (2021–2022) on several varieties on various farms in the south and southeast of Kazakhstan, including Almaty Region (Almalyk village, Baibulak village, Kyzylsharik village, Bayseit village, Koram village), Turkestan Region (Akzhar village, Shakpak baba village), and Zhambyl Region (Merke village) located in different climatic areas.
During the visual inspection of apple trees, the presence of symptoms characteristic of fire blight was noted: sudden wilting and drying of blooming flowers; drying of the tops of young shoots, the tips of which are bent in a hook-like manner; leaf necrosis; brown spots on unripe fruits, which gradually mummy; characteristic “marbling” on the cut of the bark; wedge-shaped ulcers on the bark; the release of exudate on affected organs (Figure 1 and Figure 2).
The study area’s apple commercial and farm orchards (Turkestan, Zhambyl, and Almaty Regions) are favorable for horticulture under conditions of irrigation. The main unfavorable factor for fruit crops, in particular apple trees, is the return of spring frosts in April and early May, which have become more frequent in recent decades.
Meteorological stations collecting climate data are located in the city of Talgar and the foothills, covering the orchards of the Almaty Region; in the city of Merke, covering the orchards of the Zhambyl Region; and in the city of Shakpak Baba, covering the orchards of the Turkestan Region. Merke, in the Zhambyl Region, is described as more arid with sharp temperature fluctuations between day and night [76]. The climatic conditions of the Turkestan Region are characterized by pronounced continentality, aridity, hot and dry summers, and mild winters. Data of the meteorological observations for the regions in the main fruit-growing area of Kazakhstan, for 2021 and 2022, are presented in Figure 3. Figure 4 presents the correlation between the disease severity index (DSI) and climatic conditions (temperature, precipitation, and humidity) by using linear regression analysis. According to the analysis, the DSI decreases with increasing temperature, whereas the DSI increases with increasing precipitation and humidity.
The weather conditions in 2021 had a number of differences from the long-term averages in 2022. In the first half of spring 2021, the weather was cool. At the end of April, frosts were observed to be reduced by −3.90 °C, and in the second half of spring, the weather was warm. In addition, for all regions, in April 2022, a slight change was found, on average, in the form of an increase in temperature by +4.2 °C compared to 2021. The humidity across all regions increased by at least 10% in late spring and early summer 2022, while the precipitation doubled. The summer was dry and hot, and the average daily air temperature in the summer months of 2021 was 2–4 °C higher than the long-term values, which adversely affected the vegetation and productivity of trees and also helped to stop the development of the disease.
The inspection of apple plantations in the main commercial area of horticulture (Almaty, Turkestan, and Zhambyl Regions) showed that fire blight was quite widespread. A focal distribution of the disease in the apple orchards of eight inspected farms was established. During a visual observation of the commercial orchards and farms among 59 varieties, symptoms of fire blight were found on most varieties: Idared, Aport, Golden Delicious, Gala, Granny Smith, Maksat, Voskhod, Renet Burkhardt, Starkrimson, Rubin, and Fuji. Disease symptoms were noted among the following foreign apple varieties: Pinova, Pink Lady, Rashida, Konfetnoye, Sinap Almaty, Red Topaz, Deljohns, Santana, and Wilton Star. Asymptomatic samples were collected in the laboratory for identification of the pathogen.
The study of plant reaction to the pathogen was focused on three of the most susceptible varieties Golden Delicious, Idared, and Aport, due to the susceptibility of these genotypes to fire blight in 2021 and 2022, whereas other varieties only experienced sporadic infections. Using these genotypes as examples, it was able to demonstrate the varying levels of resistance/susceptibility to fire blight in different regions of Kazakhstan. As a result of assessing the intensity of the development of fire blight on the apple varieties Idared, Golden Delicious, and Aport in different regions, certain zonal foci in the manifestation of the disease were noted. As can be seen from the data in Figure 5, the Idared variety was highly affected in all regions. The DDI of fire blight reached 36.0% with a DSI of 9.6%. The most severe damage to all varieties of apple trees with fire blight was noted in the plantations of the Almaty Region.
The DDI ranged from 22.0 to 36.0% and DSI from 5.2 to 9.6%. The disease’s development was weaker in the Zhambyl Region, which has a more arid climate, and an even lower DDI was noted in the Turkestan Region, where the prevalence of the disease was between 12.0 and 20.0% and the DSI from 3.0 to 5.2%. This is probably because of the different natural and climatic conditions of these regions. The climatic conditions of the Turkestan Region are characterized by pronounced continentality and aridity, which adversely affect the development of infection. At the same time, in the Almaty Region with higher air humidity, favorable conditions are created for the development and spread of the disease. The two-year monitoring period made it possible to identify apple varieties susceptible and resistant to fire blight under field conditions, taking into account the climatic conditions of the main fruit regions of Kazakhstan.

3.2. SNP Genotyping

The study of genetic resistance using 10 SNP markers that 10 (100.0%) led to successful PCR amplification (Figure 6). Screening of a collection of apple varieties using markers in the FBE, MR5, and RLP1 genes made it possible to obtain clear reproducible results (Table 4). All ten markers (FBE-1_Y320; FBE-2_Y192; FBE-2_Y495; FBE-2_Y551; FB-MR5-K35; FB-MR5-R249; FB-MR5-rp16k15_M; FB-MRS-R240; RLP1a; and RLP1b), which were developed for fire blight resistance derived from Malus robusta “Robusta 5” (MR5/RLP1), were monomorphic for all varieties of apple genotyped in this study.
Remarkably, all 59 apple varieties exhibited identical genotypes for all ten markers associated with fire blight resistance. There was no variation observed among the studied varieties in terms of the presence or absence of these markers. This outcome indicates a lack of genetic diversity for fire blight resistance within the analyzed panel of apple varieties.

3.3. Molecular Screening Using SCAR Markers

The identification of variants of QTL FBF7 linked to fire blight resistance is commonly carried out using the SCAR markers AE10-375 and GE-8019. The resistant genotypes are indicated by the dominant alleles of markers AE10-375 and GE-8019, which have lengths of 375 bp and 397 bp, respectively [19,75]. The presence of resistance alleles for both markers has previously been revealed in the genotypes with high resistance to the pathogen compared with the genotypes bearing only the resistance allele for one of the two markers [19]. In the present research, we identified 23 varieties (Starkrimson, Aport, Fuji, Golden Delicious, Granny Smith, Red Delicious, Aigul, Jeromini, Kandil Sinap, Kandil Kirghiz, Starks Earliest, Scarlet Spur, Summer red, Nicole Grain, Gold spur, Honeycrisp, Camspur, Gala Anna, Jonagold, Synap Almatinsky, Wilton Star, Pinova, Red Topaz) and 7 varieties (Quinti, Summer red, Williams Pride, Honeycrisp, Elstar, Pinova, Red Topaz) carrying resistant alleles of AE10-375 and GE-8019, respectively, as shown in Table 4. However, according to DDI and DSI analyses, Golden Delicious, Idared, and Aport were infected by the fire blight pathogen in all evaluated orchards. Additionally, Honeycrisp, Red Topaz, Pinova, and Samuret varieties bear resistant alleles of both markers.

4. Discussion

The DDI and DSI of fire blight are highly dependent on environmental conditions in Almaty Region (Almalyk village, Baibulak village, Kyzylsharik village, Bayseit village, Koram village), Turkestan Region (Akzhar village, Shakpak baba village), and Zhambyl Region (Merke village) (e.g., temperature, humidity, and rainfall), host factors (e.g., tree vigor), E. amylovora strain virulence, various host–strain interactions, and host quantitative stability, which adds to the challenge of phenotypic resistance/susceptibility to fire blight [10,77,78]. Different phenotyping methods can produce variable and often uncorrelated results [79,80]. In addition, genetic markers can reduce the cost of breeding new varieties through early selection. The number of plants and duration of the evaluation can be reduced, and the varieties, even those with combined resistance, can become commercially available in a short time [81].
However, seedling phenotypic breeding is a cost-effective and relatively efficient approach used in apple tree breeding programs (e.g., WABP) to create breeding populations with low susceptibility to fire blight. The use of DNA information in breeding decisions (i.e., DNA-based breeding), which has become common for several traits (e.g., apple scab resistance and malic acid content) in apple trees [82], will enable more efficient and accurate breeding for fire blight resistance. The development of apple varieties with long-term fire blight resistance and superior fruit quality can be effectively achieved through DNA breeding; however, progress is hampered by the few DNA tests available that predict symptoms. In the short term, published information on the phenotypic resistance/susceptibility [80] and on the reduced and increased susceptibility alleles for several important parent plants (IBPs) and varieties [83] can be immediately applied to inform parental selection in apple tree breeding programs. The breeding-relevant QTLs that have been previously characterized can be targeted to develop DNA tests for breeders to pyramid favorable alleles and/or combine superior fruit quality with fire blight resistance. The introgression and pyramiding of favorable alleles can be accelerated by fast-cycle breeding techniques [18]. The objective of this research was to analyze domestic and foreign apple varieties using SNP and SCAR markers to identify genetic sources of fire blight resistance.
Khan et al. (2007) confirmed that genotypes carrying the markers AE10-375 and GE-8019 have, on average, higher resistance to fire blight compared to genotypes that do not contain the markers [19]. Previous studies have shown similar results [19,84]. Another study found that fire blight resistance in genotypes carrying AE10-375 and GE-8019 confirms their utility for MAS. The identified genotypes on the basis of this study especially varieties that contain AE10-375 and GE-8019 Honeycrisp, Red Topaz, Pinova, and Samuret will be useful for MAS and breeding programs. The advantage of determining resistance genes is their presence in large-fruited varieties of commercial fruit quality, which can be easily used to breed new varieties. Fire blight resistance scores under controlled conditions have been shown to correlate well with field resistance [85]. Many factors, such as host and environmental conditions, influence the DDI and DSI of the disease [86,87,88].
The identification of sources of resistance with high fruit quality (i.e., elites) for use as breeding parents is an important precursor to the development of breeding populations with low susceptibility to fire blight, suggesting moderate to high heritability of traits. As a result of the research, an assessment was made of the stability of currently grown varieties of domestic and foreign breeding in the main commercial farm area of horticulture (south and southeast Kazakhstan). Molecular diagnostic techniques based on DNA markers have revolutionized plant breeding processes by enabling more accurate and efficient selection of desirable traits such as yield forecast, fruit quality, and disease resistance. These techniques provide plant breeders with tools to identify and select plants with the desired traits at an early stage, saving time and resources compared to traditional breeding methods.
Most modern commercial apple varieties are susceptible to fire blight [89,90,91,92], and an updated comparison of resistance/susceptibility levels of 94 IBPs and varieties was provided. As in previous studies, for example [80,90,91,92], most apple varieties (e.g., Jonathan, Ginger Gold, Sansa, and Sweet Sixteen) have shown high or moderate susceptibility to fire blight [81]. Several varieties with medium to high resistance have been confirmed, with eight varieties (e.g., Dolgo, Enterprise, Frostbite, Kidd’s Orange Red, Tsugaru, Vista Bella, Wildung, and Williams’ Pride) being classified as highly resistant in one year and moderately resistant in another [80]. Artificial inoculation of seedlings from crosses with low susceptibility can be used to cull highly susceptible seedlings. Artificial seedling inoculation under greenhouse conditions often overestimates susceptibility and, thus, may not predict field performance [93].
Lyzhin and Saveleva [21] showed that most of the analyzed apple varieties (85.7%) had at least one marker out of three (AE10-375, GE-8019, and CH-F7-Fb1). Two out of three markers were present in five varieties (35.7%): variety Lobo has markers GE-8019 and AE10-375, and varieties Ligol, Skala, Fregat, and Fuji have markers AE10-375 and CH-F7-Fb1. The target fragments of the studied markers are absent in the varieties Antonovka ordinary and Galarina. Three markers were identified in the genome of Bylina, Rozhdestvenskoye, Uspenskoye, and Charodeyka varieties, indicating the presence of resistance to fire blight QTL FBF7. QTL flanking markers GE-8019 and AE10-375 (marker CH-F7-Fb1 was absent) were found in the variety Lobo. Presumably, this also indicates the presence of QTL FBF7 with a lower probability than the presence of three markers. After analyzing the varieties of apple trees for resistance to fire blight using molecular markers, the authors concluded that the markers GE-8019, AE10-375, and CHF7-Fb1, linked to QTL FBF7, were found in the varieties Bylina, Rozhdestvenskoye, Uspenskoye, and Charodeyka. They recommend these varieties for use in breeding for resistance to E. amylovora [21].
A previous study conducted in Kazakhstan by Nurtaza et al. [94] shows that resistant varieties to fire blight using both markers AE10-375 and GE-8019 have not been identified. However, they found that three genotypes from the wild Cherkesai population and five genotypes from Nur-Sultan contain fire blight-resistant alleles according to AE10-375 or GE-8019 [94]. Two SCAR marker alleles, AE10-375 and GE-8019, appear to be useful in marker-assisted breeding, but their ability to predict fire blight resistance is not ideal. The very large number of varieties that have only one of the two marker alleles suggests that the simultaneous occurrence of two marker alleles in the same variety, in some cases, is not associated with the presence of QTL. Similarly, the absence of two marker alleles does not necessarily indicate a lack of fire blight resistance. In our study, AE10-375 and GE-8019 alleles were not found in some genotypes. However, these genotypes (Pestrushka, Tulpan, Ainur, Egemen, Zarya Alatau, Esen, Sarkyt, Talgarskoe, Zharkyn, Modi, Braeburn, Mutsu, Redfree, Diana, July Chernenko, Maria Red, Red JonaPrince, and SQ159) were resistant to fire blight in the field. The level of relatedness to “Cox’s Orange Pippin” does not appear to have any effect on the strength of the association between DNA marker alleles and fire blight resistance. Another study used marker CH-F7-Fb1 to identify the resistance of rootstocks to fire blight [95]. The results of the molecular analysis and susceptibility of plants to metabolites of the causative agent of fire blight of fruit crops were compared. There was no clear relationship between the number of markers present and the degree of plant tissue necrosis in the tested forms. However, studies have shown that the presence of SCAR marker AE10-375 and microsatellite CH-F7-FB1 in the forms of 62-396 and 14-1 provides a phenotypic manifestation of resistance to E. amylovora metabolites [95]. However, more varieties need to be investigated before arriving at a general conclusion, which is confirmed by Nybom et al. [96].
Genotyping of genebank material is critical to ensure that genetic variation is maintained and also to make genebank material a useful resource for breeding. In the past, SSR markers have been widely used in apple germplasm research at national, regional, and European levels. More recently, SNP arrays have been used to determine apple diversity [97]. In a preliminary study, the gene sequence FBE, MR5, and RLP1 [75] was used which was found to be associated with fire blight resistance. In our study, all 10 pairs of primers and probes were designed based on this gene sequence and screened for 59 apple varieties of Kazakh and foreign breeding programs. Interestingly, all 59 apple varieties exhibited identical genotypes for all ten markers associated with fire blight resistance. There was no variation observed among the studied varieties in terms of the presence or absence of these markers. This outcome indicates a lack of genetic diversity for fire blight resistance within the analyzed panel of apple varieties. This unexpected lack of genetic diversity for fire blight resistance within the analyzed panel of apple varieties may have implications for future fire blight management in apple orchards. Noticeably, a few cases were identified where the SNP data were in conflict with data obtained from the presumed graft-wood source using SSR markers. Understanding the genetic basis of this uniformity is critical for developing effective strategies to enhance fire blight resistance in apple cultivars and maintaining the sustainability of apple production in the face of evolving pathogens.

5. Conclusions

In the course of this study, an observation of disease in 59 apple varieties in the general fruit regions of Kazakhstan was carried out, taking into account the climatic conditions. Based on the results obtained in the study, it was found that fire blight has become quite widespread. Conspicuously, the SNP analysis showed that all 59 apple varieties exhibited identical genotypes for all ten markers associated with fire blight resistance. No variation was observed among the studied varieties regarding the presence or absence of these markers. The 26 most resistant apple varieties of domestic and foreign breeds to fire blight were identified using the molecular screening markers AE-375 and GE-8019. Among the studied 59 apple varieties, 23 varieties were identified using the AE-375 marker and 7 varieties with the GE-8019 marker. The varieties Samuret, Honeycrisp, Pinova, and Red Topaz were found to be resistant using markers AE-375 and GE-8019. As a result of the analysis of the genotypes, out of 59 analyzed samples, 26 varieties have distinctive alleles that can be associated with resistance to the disease and can be used as sources of resistance in breeding programs.

Author Contributions

Data curation, M.S.; formal analysis, N.S. and D.G.; funding acquisition, G.K.; investigation, N.D., V.B. and N.K.; methodology, S.O. and N.K.; project administration, G.K.; resources, D.G.; software, M.S. and Z.S.; supervision, G.K. and Z.S.; validation, V.B.; visualization, Z.S.; writing—original draft, G.K. and Z.S.; writing—review and editing, Z.S. All authors have read and agreed to the published version of the manuscript.

Funding

This research and APC were funded by the Science Committee of the Ministry of Science and Higher Education of the Republic of Kazakhstan (Grant No. AP09259636, “Study of the genetic resistance of promising Apple varieties and rootstocks to a dangerous disease-fire blight using SNP markers”).

Data Availability Statement

The data used to support the findings of this study can be made available by the corresponding author upon request.

Conflicts of Interest

The authors declare there are no competing interests.

References

  1. Strategic Planning and Reforms Agency of the Republic of Kazakhstan. Home Page. Available online: https://stat.gov.kz/official/industry/14/statistic/7 (accessed on 15 June 2023).
  2. Vanneste, J.L. What is Fire Blight? Who is Erwinia amylovora? How to Control It. In Fire Blight. The Disease and Its Causative Agent, Erwinia Amylovora; CABI Publishing: Wallingford, UK, 2000; pp. 1–6. [Google Scholar] [CrossRef]
  3. Norelli, J.L.; Jones, A.L.; Aldwinckle, H.S. Fire Blight Management in The Twenty First Century: Using New Techniques That Enhance Host Resistance in Apple. Plant Dis. 2003, 87, 756–765. [Google Scholar] [CrossRef] [PubMed]
  4. Oktem, Y.E.; Benlioğlu, K. Yumuşak Çekirdekli Meyve Ağaçlarında Görülen Ateş Yanıklığı Hastalık Etmeni (Erwinia amylovora (Burr.)). Üzerinde Çalışmalar 1988, 4, 69–74. (In Turkish) [Google Scholar]
  5. Öztürk, G.; Basım, E.; Basım, H.; Emre, R.A.; Karamürsel, Ö.F.; Eren, İ.; İşçi, M.; Kaçal, E. Kontrollü Melezleme Yoluyla Ateş Yanıklığı (Erwinia amylovora) Hastalığına Karşı Dayanıklı Yeni Armut Çeşitlerinin Geliştirilmesi: İlk Meyve Gözlemleri. In Proceedings of the 6th Bahçe Bitkileri Congress, Şanlıurfa, Turkey, 4–8 October 2011. [Google Scholar]
  6. Drenova, N.V.; Isin, M.M.; Dzhaymurzina, A.A.; Zharmukhamedova, G.A.; Aitkulov, A.K. Bacterial burn of fruit crops in the Republic of Kazakhstan. J. Plant Quar. Sci. Pract. 2013, 1, 39–43. (In Russian) [Google Scholar]
  7. Azhimakhan, M.A.; Varitsev, Y.A.; Drenova, N.V.; Khasanov, V.T.; Dzhaymurzina, A.A.; Tuleeva, A.K.; Umiralieva, Z.Z. Development of enzyme-linked immunosorbent diagnostics of a test system to detect the causative agent of bacterial burn of fruit crops (Erwinia amylovora). Biotechnol. Theory Pract. 2016, 28–34. [Google Scholar]
  8. Gritsenko, D.A.; Nizamedinova, G.K.; Khamdieva, O.K.; Dinasilov, A.S. Identification of a bacterial burn by molecular gene-tic methods. News Natl. Acad. Sci. Repub. Kazakhstan Agric. Sci. Ser. 2017, 6, 109–115. [Google Scholar]
  9. Saveliev, N.N.; Savelyeva, N.I. Application of the achievements of genetics in the breeding of fruit crops: The contribution of the Michurin branch of the Vavilov Society of Geneticists and Breeders. Plant Genet. Breed. 2016, 20, 555–562. [Google Scholar] [CrossRef]
  10. Lee, S.A.; Ngugi, H.K.; Halbrendt, N.O.; O’Keefe, G.; Lehman, B.; Travis, J.W.; Sinn, J.P.; Mc Nellis, T.W. Virulence characte-ristics accounting for fire blight disease severity in apple trees and seedlings. Phytopathology 2010, 100, 539–550. [Google Scholar] [CrossRef] [PubMed]
  11. Emeriewen, O.F.; Wöhner, T.; Flachowsky, H.; Peil, A. Malus hosts—Erwinia amylovora interactions: Strain pathogenicity and resistance mechanisms. Front. Plant Sci. 2019, 10, 551. [Google Scholar] [CrossRef]
  12. Norelli, J.L.; Aldwinckle, H.S. Differential susceptibility of Malus spp. Cultivars Robusta 5, Novole, and Ottawa 523 to Erwinia amylovora. Plant Dis. 1986, 70, 1017–1019. [Google Scholar] [CrossRef]
  13. Norelli, J.L.; Holleran, H.T.; Johnson, W.C.; Robinson, T.L.; Aldwinckle, H.S. Resistance of Geneva and other apple rootstocks to Erwinia amylovora. Plant Dis. 2003, 87, 26–32. [Google Scholar] [CrossRef]
  14. Peil, A.; Wöhner, T.; Hanke, M.V.; Flachowsky, H.; Richter, K.; Wensing, A.; Kilian, A. Comparative mapping of fire blight resistance in Malus. Acta Hortic. 2013, 1056, 47–517. [Google Scholar] [CrossRef]
  15. Fazio, G.; Aldwinckle, H.; Robinson, T. Unique characteristics of Geneva® apple rootstocks. Encontro Nac. Sobre Frutic. Clima Temperado 2013, 1, 23. [Google Scholar]
  16. Kellerhals, M.; Schütz, S.; Patpcchi, A. Breeding for host resistance to the fire blight. J. Plant Pathol. 2017, 99, 37–43. Available online: https://www.jstor.org/stable/45156717 (accessed on 15 June 2023).
  17. Peil, A.; Emeriewen, O.F.; Khan, A.; Kostick, S.; Malnoy, M. Status of fire blight resistance breeding in Malus. J. Plant Pathol. 2021, 103, 3–12. [Google Scholar] [CrossRef]
  18. Kostick, S.A.; Teh, S.L.; Evans, K.M. Contributions of Reduced Susceptibility Alleles in Breeding Apple Cultivars with Durable Resistance to Fire Blight. Plants 2021, 10, 409. [Google Scholar] [CrossRef]
  19. Khan, M.A.; Durel, C.E.; Duffy, B.; Drouet, D.; Kellerhals, M.; Gessler, C.; Patocchi, A. Development of molecular markers linked to the ‘Fiesta’ linkage group 7 major QTL for fire blight resistance and their application for marker-assisted selection. Genome 2007, 50, 568–577. [Google Scholar] [CrossRef]
  20. Omasheva, M.Y.; Pozharskiy, A.S.; Maulenbay, A.D.; Ryabushkina, N.A.; Galiakparov, N.N. SSR genotyping of Kazakhstan apple varieties and identification of alleles associated with resistance to the most dangerous pathogens. Biotechnol. Theory Pract. 2016, 2, 168. [Google Scholar] [CrossRef]
  21. Lyzhin, A.; Saveleva, N. Identification of QTL FBF7 fire blight resistance in apple varieties germplasm. BIO Web Conf. 2021, 34, 02002. [Google Scholar] [CrossRef]
  22. International Standard GOST 33829-2016; Plant Protection. Requirements for the Production of Products of Plant Origin at Risk of Developing a Phytosanitary Condition. Standardinform: Moscow, Russian, 2016.
  23. Interstate Standard for Phytosanitary Measures (ISPM 27); Diagnostic Protocols for Regulated Pests. International Plant Protection Convention (IPPC). Food and Agriculture Organization of the United Nations: Rome, Italy, 2016.
  24. Garden Focused. Home Page. Available online: https://www.gardenfocused.co.uk/contactus.php (accessed on 15 June 2021).
  25. Keepers Nursery. Home Page. Available online: http://surl.li/hrfcb (accessed on 22 June 2023).
  26. Rennie Orchards. Home Page. Available online: https://rennieorchards.com/gala-apple-tree/ (accessed on 20 June 2023).
  27. Livingasia. Home Page. Available online: https://livingasia.online/amp/2016/09/10/apples-almaty/# (accessed on 26 June 2023).
  28. The Arbor Day Foundation. Home Page. Available online: https://www.arborday.org/trees/treeguide/TreeDetail.cfm?itemID=2514 (accessed on 20 June 2023).
  29. Plant Me Green. Home Page. Available online: https://www.plantmegreen.com/products/golden-delicious-apple-tree (accessed on 20 June 2023).
  30. Orange Pippin. Home Page. Available online: https://www.orangepippin.com/varieties/apples/granny-smith (accessed on 22 June 2023).
  31. FGBNU VNIISPK. Home Page. Available online: https://vniispk.ru/varieties/suislepskoe-suisleper-malinovka (accessed on 5 July 2023).
  32. Anig Lider Innovation. Home Page. Available online: http://anig.kz/ru/catalog/yablonya-renet-burhardta-limonka_151 (accessed on 13 July 2023).
  33. Yastrebov, I.I. Brief Description of Apple Varieties. Available online: https://www.dachacha.ru/u/245.html (accessed on 20 July 2023).
  34. The Arbor Day Foundation. Home Page. Available online: https://www.arborday.org/trees/treeguide/TreeDetail.cfm?ItemID=721 (accessed on 26 June 2023).
  35. Nurmuratuly, T.; Madenov, E.D.; Nurtazina, N.Y. Genofond of local and Old varieties of apple, pear, apricot, and grapes in the South and Southeast of Kazakhstan. Russ. J. Genet. 2012, 54, 176–187. [Google Scholar]
  36. Anig Lider Innovation. Home Page. Available online: https://www.anig.kz/ru/catalog/yablonya-saltanat_199 (accessed on 15 June 2023).
  37. Sadovnik Ingo. Home Page. Available online: https://sadovnik.info/sort-yabloni-zarya-alatau.html (accessed on 15 June 2023).
  38. Specialty Produce. Home Page. Available online: https://specialtyproduce.com/produce/Modi_Apples_14446.php (accessed on 15 June 2023).
  39. Garden Focused. Home Page. Available online: https://www.gardenfocused.co.uk/fruitarticles/apples/variety-braeburn.php (accessed on 15 June 2023).
  40. Tsesmelis. Home Page. Available online: https://tsesmelis.gr/en/portfolio/jeromin-apple-variety-2/ (accessed on 20 June 2023).
  41. Salt Spring Apple Company. Home Page. Available online: https://www.saltspringapplecompany.com/quinte (accessed on 15 June 2023).
  42. Cummins Nursery. Home Page. Available online: https://www.cumminsnursery.com/buy-trees/product-detail.php?type=tree&id=2886 (accessed on 15 June 2023).
  43. Minneopa Orchards. Home Page. Available online: https://minnetonkaorchards.com/mutsu-apple-tree/ (accessed on 15 June 2023).
  44. Soldatov, I.V. The Best Local/Old-Time Varieties of Apple Trees in Kyrgyzstan. Available online: http://centralasia.bioversityinternational.org/fileadmin/templates/centralasia.net/upload/Resources/TRG/2614-0020.pdf (accessed on 15 June 2023).
  45. Orange Pippin. Home Page. Available online: https://www.orangepippin.com/varieties/apples/pinklady (accessed on 15 June 2023).
  46. Victoriana Nursery Gardens. Home Page. Available online: https://www.victoriananursery.co.uk/Apple-Tree-Starks-Earliest/ (accessed on 15 June 2023).
  47. Dalival. Home Page. Available online: https://www.dalival.com/pommes/scarlet-spur-evasni-ru/ (accessed on 15 June 2023).
  48. Seedlings Maryanivka. Home Page. Available online: https://sadzhanciderev.com.ua/p722676977-sazhentsy-yablon-samared.html (accessed on 15 June 2023).
  49. Roots to Fruits Nursery. Home Page. Available online: https://rootstofruitsnursery.com/products/redfree (accessed on 15 June 2023).
  50. Orange Pipin. Home Page. Available online: https://www.orangepippintrees.com/trees/apple-trees/williams-pride (accessed on 15 June 2023).
  51. Garden Web. Home Page. Available online: http://gardenweb.ru/diana-sort-yabloni (accessed on 15 June 2023).
  52. Jparker’s. Home Page. Available online: https://www.jparkers.co.uk/apple-golden-spur-0001160c (accessed on 15 June 2023).
  53. Stark Bros. Home Page. Available online: https://www.starkbros.com/products/fruit-trees/apple-trees/honeycrisp-apple (accessed on 15 June 2023).
  54. B2B.Trade. Home Page. Available online: https://b2b.trade/product/agrofirma-doneckaya-dolina-yablonya-red-chif-kamspur (accessed on 15 June 2023).
  55. Sad Sezon. Home Page. Available online: https://sadsezon.com/sad/plodovie/yabloni/sorta/k/korej.html (accessed on 15 June 2023).
  56. Nature Hills. Home Page. Available online: https://www.naturehills.com/anna-apple-tree (accessed on 15 June 2023).
  57. FGBNU VNIISPK. Home Page. Available online: https://vniispk.ru/varieties/iulskoe-chernenko (accessed on 15 June 2023).
  58. Orange Pippin. Home Page. Available online: https://www.orangepippin.com/varieties/apples/elstar (accessed on 15 June 2023).
  59. Sortover.ru. Home Page. Available online: https://sortoved.ru/yablonya/sort-yabloni-dzhonagold.html (accessed on 15 June 2023).
  60. Johan Nicolai. Home Page. Available online: https://johan-nicolai.com/en/varieties/d/detail/elstar-elrosa (accessed on 10 June 2023).
  61. Mary, E.E. Gardening Know How. Home Page. Available online: https://www.gardeningknowhow.com/edible/fruits/apples/prima-apple-information.htm (accessed on 10 June 2023).
  62. Celtic Orchards. Home Page. Available online: https://www.theapplefarm.com/celtic/redelstar.htm (accessed on 10 June 2023).
  63. OOO Firma Ltd. Home Page. Available online: https://ltdsad.ru/catalog/brebur-marija-red/ (accessed on 10 June 2023).
  64. New England Apple. Home Page. Available online: https://newenglandapples.org/apples/jonaprince/ (accessed on 10 June 2023).
  65. Alma Global. Home Page. Available online: https://www.almaglobal.kz/sinap-kandil-almatinskij (accessed on 15 June 2023).
  66. Johan Nicolai. Home Page. Available online: https://johan-nicolai.com/en/varieties/d/detail/wilton-s-star-red-jonaprince-select (accessed on 10 June 2023).
  67. Fresh Plaza. Home Page. Available online: https://www.freshplaza.com/north-america/article/9156688/overview-global-apple-market/ (accessed on 10 June 2023).
  68. Orange Pippin. Home Page. Available online: https://www.orangepippin.com/varieties/apples/santana (accessed on 10 June 2023).
  69. Minneopa Orchards. Home Page. Available online: https://minnetonkaorchards.com/pinova-apple-tree/ (accessed on 10 June 2023).
  70. Orange Pippin. Home Page. Available online: https://www.orangepippin.com/varieties/apples/topaz (accessed on 10 June 2023).
  71. Fresh Plaza. Home Page. Available online: https://www.fruit-inform.com/ru/technology/experience/175413 (accessed on 10 June 2023).
  72. Beer, S.V.; Schwager, S.J.; Norelli, J.L.; Aldwinckle, H.S.; Burr, T.J. Development of a Risk-Assessment System for Fire Blight. In Proceedings of the Sixth International Conference on Plant Pathogenic Bacteria, College Park, MA, USA, 2–7 June 1985; p. 4. [Google Scholar]
  73. Jansch, M.; Broggini, G.A.L.; Weger, J.; Bus, V.G.M.; Gardiner, S.E.; Bassett, H.; Patocchi, A. Identification of SNPs linked to eight apple disease resistance loci. Mol. Breed. 2015, 35, 45. [Google Scholar] [CrossRef]
  74. Gardiner, S.E.; Norelli, J.L.; de Silva, N.; Fazio, G.; Peil, A.; Malnoy, M.; Horner, M.; Bowatte, D.; Carlisle, C.; Wiedow, C.; et al. Putative resistance gene markers associated with quantitative trait loci for fire blight resistance in Malus ‘Robusta 5’ accessions. BMC Genet. 2012, 13, 25. [Google Scholar] [CrossRef] [PubMed]
  75. Chagne, D.; Vanderzande, S.; Kirk, C.; Profitt, N.; Weskett, R.; Gardiner, S.; Peace, C.; Volz, R.; Bassil, N. Validation of SNP markers for fruit quality and disease resistance loci in apple (Malus × domestica Borkh.) using the OpenArray® platform. Hortic. Res. 2019, 6, 30. [Google Scholar] [CrossRef] [PubMed]
  76. Meteorological and Hydrological Databases. Home Page. Available online: https://www.kazhydromet.kz/en/meteo_db (accessed on 10 June 2023).
  77. Van der Zwet, T.; Halbrendt, O.N.; Zeller, W. Fire Blight: History, Biology and Management; APS Press: Saint Paul, MI, USA, 2016. [Google Scholar] [CrossRef]
  78. Brown, S. Apple. In Fruit Breeding, Handbook of Plant Breeding; Springer: Berlin/Heidelberg, Germany, 2012; pp. 329–367. [Google Scholar]
  79. DuPont, S.T.; Munir, M.; Cox, K.; Johnson, K.; Peter, K.; Baro, A. Evaluation of pruning therapies in apple trees with fire blight. J. Plant Pathol. 2023. [Google Scholar] [CrossRef]
  80. Kostick, S.A.; Norelli, J.L.; Evans, K.M. Novel metrics to classify fire blight resistance of 94 apple cultivars. Plant Pathol. 2019, 68, 985–996. [Google Scholar] [CrossRef]
  81. Baumgartner, I.; Franck, L.; Silvestri, G.; Patocchi, A.; Duffy, B.; Frey, J.; Kellerhals, M. Advanced strategies for breeding fire blight resistant high-quality apples. In Proceedings of the 14th International Conference on Organic Fruit-Growing, Hohenheim, Germany, 22–24 February 2010. [Google Scholar]
  82. Peace, C.P. DNA-informed breeding of rosaceous crops: Promises, progress and prospects. Hortic. Res. 2017, 4, 17006. [Google Scholar] [CrossRef]
  83. Kostick, S.A.; Teh, S.; Norelli, J.L.; Vanderzande, S.; Peace, C.; Evans, K.M. Fire blight QTL analysis in a multi-family popula-tion identifies a reduced-susceptibility allele in ‘Honeycrisp’. Hortic. Res. 2021, 8, 28. [Google Scholar] [CrossRef]
  84. Sehic, J.; Nybom, H.; Garkava-Gustavsson, L.; Patocchi, A.; Kellerhals, M.; Duffy, B. Fire blight (Erwinia amylovora) resistance in apple varieties associated with molecular markers. Int. J. Hortic. Sci. 2009, 15, 53–57. [Google Scholar] [CrossRef]
  85. Quamme, H.A.; van der Zwet, T.; Dirks, V. Relationship of fire blight resistance of young pear seedlings inoculated in the greenhouse to mature seedling trees naturally infected in the field. Plant Dis. Report. 1976, 60, 660–664. Available online: https://www.jstor.org/stable/41992429 (accessed on 15 June 2023).
  86. van der Zwet, T.; van der Zwet, K.T.; Keil, H.L. Fire blight, a bacterial disease of rosaceous plants. In Agriculture Handbook; U.S. Department of Agriculture: Washington, DC, USA, 1979. [Google Scholar]
  87. van der Zwet, T.; Beer, S.V. Fire blight—Its Nature, Prevention, and Control: A Practical Guide to Integrated Disease Management; USDA: Washington, DC, USA, 1999. [Google Scholar] [CrossRef]
  88. Brisset, M.N.; Paulin, J.P. Plant mechanisms interfering with fire blight infection. Acta Hortic. 2006, 704, 483–487. [Google Scholar] [CrossRef]
  89. van der Zwet, J.; Hanssen, V.G.; Zwietering, P.J.; Muijtjens, P.J.; van der Vleuten, A.M.; Metsemakers, C.P. Workplace learning in general practice: Supervision, patient mix and independence emerge from the black box once again. Med. Teach. 2010, 32, e294–e299. [Google Scholar] [CrossRef]
  90. Aldwinckle, H.S.; Gustafson, H.L.; Forsline, P.L. Evaluation of the core subset of the USDA apple germplasm collection for resistance to fire blight. Acta Hortic. 1999, 489, 269–272. [Google Scholar] [CrossRef]
  91. Le Lezek, M.; Paulin, J.P.; Lecomte, P. Shoot and blossom susceptibility to fireblight of apple cultivars. Acta Hortic. 1987, 217, 311–315. [Google Scholar] [CrossRef]
  92. Mohan, S.K.; Fallahi, E.; Bijman, V.P. Evaluation of apple varieties for susceptibility to Erwinia amylovora by artificial inocula-tion under field conditions. Acta Hortic. 2002, 590, 373–375. [Google Scholar] [CrossRef]
  93. Hampson, C.R.; Sholberg, P.L. Estimating combining ability for fire blight resistance in apple progenies. Acta Hortic. 2008, 793, 337–343. [Google Scholar] [CrossRef]
  94. Nurtaza, A.; Pozharskiy, A.; Dyussembekova, D. Conservation of Malus Niedzwetzkyana Dieck Ex Koehne Genotypes from Kazakhstan Resistant to Scab and Fire Blight Diseases. Res. Sq. 2022, 1–18. [Google Scholar] [CrossRef]
  95. Shamshin, I.N.; Maslova, M.V.; Drenova, N.V.; Dubrovsky, M.L.; Parusova, O.V. Assessment of fire blight resistance in apple clonal rootstocks using molecular markers. Proc. Appl. Bot. Genet. Breed. 2021, 181, 185–191. [Google Scholar] [CrossRef]
  96. Nybom, H.; Mikicinski, A.; Garkava-Gustavsson, L.; Sehic, J.; Lewandowski, M.; Sobiczewski, P. Assessment of fire blight tole-rance in apple based on plant inoculations with Erwinia amylovora and DNA markers. Trees 2012, 26, 199–213. [Google Scholar] [CrossRef]
  97. Sätra, S.A.J.; Troggio, M.; Odilbekov, F.; Sehic, J.; Mattisson, H.; Hjalmarsson, I.; Ingvarsson, P.K.; Garkava-Gustavsson, L. Genetic Status of the Swedish Central collection of heirloom apple cultivars. Sci. Hortic. 2020, 272, 109599. [Google Scholar] [CrossRef]
Figure 1. Symptoms of fire blight on the Idared apple variety (Merke, Zhambyl Region): (A) drying of the tops of young shoots and a shepherd’s crook; (B) release of bacterial exudate.
Figure 1. Symptoms of fire blight on the Idared apple variety (Merke, Zhambyl Region): (A) drying of the tops of young shoots and a shepherd’s crook; (B) release of bacterial exudate.
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Figure 2. Symptoms of fire blight on the Golden Delicious apple variety (Baidibek bi village, Enbekshikazakh District): (A) withering and drying of blossoming flowers and young leaves; (B) affected leaves on young shoots.
Figure 2. Symptoms of fire blight on the Golden Delicious apple variety (Baidibek bi village, Enbekshikazakh District): (A) withering and drying of blossoming flowers and young leaves; (B) affected leaves on young shoots.
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Figure 3. Agroclimatic indicators for April–September according to the weather stations of the Almaty, Zhambyl, and Turkestan Regions, 2021–2022.
Figure 3. Agroclimatic indicators for April–September according to the weather stations of the Almaty, Zhambyl, and Turkestan Regions, 2021–2022.
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Figure 4. The correlation between the disease severity index (DSI) and climatic conditions by using linear regression analysis.
Figure 4. The correlation between the disease severity index (DSI) and climatic conditions by using linear regression analysis.
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Figure 5. Infection of apple trees with the pathogen under the conditions of the south and southeast of Kazakhstan in 2021 and 2022. Note: DDI—The disease distribution index; DSI—The disease severity index.
Figure 5. Infection of apple trees with the pathogen under the conditions of the south and southeast of Kazakhstan in 2021 and 2022. Note: DDI—The disease distribution index; DSI—The disease severity index.
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Figure 6. Allelic discrimination of the variety samples for the following markers: (A) FBE-1_Y320; (B) FBE-2_Y192; (C) FBE-2_Y495; (D) FBE-2_Y551; (E) FB-MR5-K35; (F) FB-MR5-R249; (G) FB-MR5-rp16k15_M; (H) FB-MRS-R240; (I) RLP1a; (J) RLP1b. Variety samples are marked in blue.
Figure 6. Allelic discrimination of the variety samples for the following markers: (A) FBE-1_Y320; (B) FBE-2_Y192; (C) FBE-2_Y495; (D) FBE-2_Y551; (E) FB-MR5-K35; (F) FB-MR5-R249; (G) FB-MR5-rp16k15_M; (H) FB-MRS-R240; (I) RLP1a; (J) RLP1b. Variety samples are marked in blue.
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Table 1. Characteristics of the apple varieties.
Table 1. Characteristics of the apple varieties.
#GenotypeOriginLocation *Reference
1IdaredCross between Jonathan and Wagener varieties, first raised in Idaho, USA. It was first released to the public in 1942 after a ten-year breeding program.
Included in the State Register of the Republic of Kazakhstan since 1998.
1; 4; 6[24]
2StarkrimsonPollination: Starkrimson is self-sterile and requires a pollinator to produce a crop. Origin: USA, 1870.
Included in the State Register of the Republic of Kazakhstan since 1988.
6; 8[25]
3GalaDeveloped in New Zealand, it is the result of breeding among some of the most historically important varieties such as Cox’s Orange Pippin, Golden Delicious, and Kidd’s Orange Red. Included in the State Register of the Republic of Kazakhstan since 2011.4; 6[26]
4AportAn ancient variety from Russia. The originator is not registered. Crossing them with the local wild Sievers apple tree made the new variety famous. Included in the State Register of the Republic of Kazakhstan since 1965.4; 7[27]
5FujiBred by crossing Red Delicious × Rale Jennet. Originator: Tohoku Science Station, Marioka, Japan. Included in the State Register of the Republic of Kazakhstan since 2011.1; 6[28]
6Golden DeliciousBred by crossing Grimes Golden × Golden Reinette. The originator is not registered. One of the symbols of West Virginia, USA. Included in the State Register of the Republic of Kazakhstan since 1965.1; 6[29]
7Granny SmithBred by crossing a French wild apple tree with a local Australian apple tree.
Originator: Maria Ann Smith, Australia. Included in the State Register of the Republic of Kazakhstan since 2011.
6[30]
8SuislepperFrom Baltic countries, it is an old variety of folk selection. Included in the State Register of the Republic of Kazakhstan since 1965.7[31]
9Renet BurhardtaA variety of Crimean origin. Originator: State Nikitsky Botanical Garden, Ukraine. Included in the State Register of the Republic of Kazakhstan since 1965.4; 7[32]
10PestrushkaAn ancient South Kazakhstan variety. The originator is not registered. Included in the State Register of the Republic of Kazakhstan since 1965.4[33]
11Red Delicious (5)Red-fruited clone of the variety Delicious. Originator: private nursery in Wilsburg, USA. Included in the State Register of the Republic of Kazakhstan since 2011.5[34]
12Tulpan (8)Variety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Autumn maturity. Passes the State variety test.8[35]
13AinurVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Winter maturity. Included in the State Register of the Republic of Kazakhstan since 2011.1; 7[35]
14AigulBred by crossing Aport × Golden Delicious.
Variety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture.
7[35]
15SaltanatVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Bred by selection of seedlings Renette Burchardt from free pollination. Included in the State Register of the Republic of Kazakhstan since 1980.7[36]
16VoskhodVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Winter maturity. Included in the State Register of the Republic of Kazakhstan since 2011.1[35]
17EgemenVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Winter maturity. Included in the State Register of the Republic of Kazakhstan since 2019.1[35]
18Zarya AlatauVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Obtained from seedlings of the variety Renet Orleans from free pollination. Included in the State Register of the Republic of Kazakhstan since 1974.1[37]
19MaksatVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Autumn maturity. Included in the State Register of the Republic of Kazakhstan since 2011.1[35]
20EsenVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Bred by crossing varieties Zailiyskoye and Aport. Passes the State variety test.1[35]
21SarkytVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Bred by crossing varieties Zarya Alatau and Aport. Passes the State variety test.1[35]
22TalgarskoeVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Winter maturity. Included in the State Register of the Republic of Kazakhstan since 2004.1[35]
23ZharkynVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Passes the State variety test.1[35]
24DanalykVariety of selection of the Kazakh Research Institute of Fruit Growing and Viticulture. Winter maturity. Included in the State Register of the Republic of Kazakhstan since 2020.1[35]
25ModiCreated at the Consorzio Italiano Vivaisti in Italy by researchers in the 2000s and then introduced to the USA in 2014.
Bred by crossing varieties Gala and Liberty.
8[38]
26BraeburnThe New Zealand variety was obtained via free pollination of Lady Hamilton in 1952.1; 3; 8[39]
27JerominiFrench variety. Derived from varieties Erovan and Early Red.4[40]
28QuintiCreated in Canada as a result of crossing Crimson Beauty × Red Melba in 1964, summer ripening.5; 8[41]
29Kandil SinapAn old Crimean variety of autumn ripening. Obtained by random mutation when sowing seeds of the related variety “Sary-sinap”.8[42]
30MutsuCreated in Japan in 1948. Obtained as a result of crossing Golden Delicious and Indo-Japanese varieties of apples.8[43]
31Kandil KirghizAn ancient variety in Kyrgyzstan with a winter ripening period.8[44]
32Pink LadyAn Austrian variety, bred by the English gardener John Cripps by crossing the Australian variety Lady Williams and Golden Delicious.4; 6[45]
33Starks EarliestDeveloped in Idaho, USA, in 1938 by the Stark brothers of Missouri.8[46]
34Scarlet SpurAmerican winter variety. Mutant Oregonspur® Trumdor.6[47]
35Summer redSummer variety of apples of Canadian selection. The maximum height of the tree is 2.5–3.5 m, belonging to the medium height group.6[48]
36Nicole GrainNo data.--
37RedfreeCreated by the Purdue, Rutgers, and University of Illinois Agricultural Experimental Station in the 1980s, the Redfree apple was designed to be a disease-resistant summer apple. Obtained as a result of crossing a domestic apple tree and a clone of Malus floribunda 821.3[49]
38Williams PrideWas developed by the well-respected “co-op” breeding program of Purdue, Rutgers, and Illinois (PRI) universities in the 1970s, and originally known as Co-op 23. Its ancestry is similar to that of many of the PRI varieties, including Rome Beauty and Malus floribunda, but its early-season character probably comes from Mollie’s Delicious and Julyred.3[50]
39DianaVariety of Ukrainian selection, winter ripening.3[51]
40Gold spurObtained from the variety Golden Delicious, winter term
maturation.
4[52]
41HoneycrispOriginates from Excelsior, Minnesota in 1974. Crossed between Mekaun and Honeygold.4[53]
42CamspurA Starkimson mutant discovered in 1967 in Washington, DC. Maturing time—Winter. Derevo: average, crown Compact. Power Supporters: Idared, Granny Smith, Everest, Professor Sprenger.4[54]
43KoreaJapanese selection at the Aomori experimental station, obtained by hybridization of Golden Delicious and Indus apple trees.4[55]
44Gala AnnaProduced in Italy as a Gala clone.5[56]
45July ChernenkoAn ancient variety of Russian selection. Obtained from crossing varieties Anis velvet and Papirovka.1[57]
46ElstarReceived in 1954–1955 in the Netherlands. Created by crossing varieties Ingrid Maria and Golden Delicious.1[58]
47JonagoldBred in the USA at the Geneva Experimental Station by crossing varieties Golden Delicious and Jonathan.1[59]
48ElrosaWilfred and Annet Zweeren, Netherlands, 1999.1[60]
49PrimaEarly autumn variety of American selection. Created by crossing varieties: M. floribunda 821 and the varieties Welsey, Melba, Rum Beauty, Golden Delicious, and their derivatives.1[61]
50Red ElstarDutch selection, winter ripening.1[62]
51Maria RedDutch selection, winter ripening.1[63]
52Red JonaPrince (1)It is one of the clones of Jonagold, isolated in Belgium, winter ripening.1[64]
53Synap AlmatinskyAn ancient variety, comes from the Crimean Sinap known since the 19th century. Winter maturity.1[65]
54Wilton StarThe Netherlands. The Wilton’s Star Red Jonaprince Select is a new mutation from Red Jonaprince, which colors darker red.2[66]
55DeljoncaThe Deljonca varieties in the German growing areas. Formed by crossing varieties Delbard Florina × Stark Jongrimes.2[67]
56Santana (2)Bred by the International Plant Research Institute Wageningen (P.R.I. Wageningen), the Netherlands.2[68]
57PinovaWinter variety of German selection is obtained by crossing varieties Golden Delicious and Clivia.2; 7[69]
58Red Topaz (2)Winter variety of Czech selection by crossing varieties Vanda and Rubín in 1984.2[70]
59SQ159A variety of apples for long storage, suitable for organic production. Developed by PRI Wageningen (Netherlands).2[71]
* (1) Regional Branch of the Kazakh Research Institute of Horticulture, Almalyk village, Talgar District of Almaty Region; (2) UOH Agrouniversitet, Baibulak village, Talgar District of Almaty Region; (3) Agricultural Innovations Limited, Kyzylsharik village, Enbekshikazakh District of Almaty Region; (4) Peasant farm “Mahmut”, Bayseit village, Yenbekshikazakh District of Almaty Region; (5) Sady Zhetysu Trade LLP, Koram village, Yenbekshikazakh District of Almaty Region; (6) Amangeldi LLP, Akzhar village, Kazygur District of Turkestan Region; (7) LLP “Kentau”, Shakpak baba village, Tulkibas District of Turkestan Region; (8) peasant farm “Devrosh”, Merke village, Zhambyl Region.
Table 2. Description of 10 single nucleotide polymorphisms (SNPs) selected to study fire blight resistance.
Table 2. Description of 10 single nucleotide polymorphisms (SNPs) selected to study fire blight resistance.
SNP MarkerLinkage GroupGene/Locus
Name
SNP IDTaqman Assay
ID
SNP
Type
Reference
FBE-1_Y32012FBEFBE-1_Y320AH704YYC/T[73]
FBE-2_Y19212FBEFBE-2_Y192AH89246C/T[73]
FBE-2_Y49512FBEFBE-2_Y495AHABIAZC/T[73]
FBE-2_Y55112FBEFBE-2_Y551AHBKGG7C/T[73]
FB-MR5-K353MR5FB-MR5-NZsnEH034548_K35AH0JFXMG/T[73]
FB-MR5-R2403MR5FB-MR5-NZsnEH034548_R240AH1SD3UA/G[73]
FB-MR5-R2493MR5FB-MR5-NZsnEH034548_R249AH21B92A/G[73]
FB-MR5-rp16k15_M1063MR5FB-MR5-rp16k15_M106AH4AAGAA/C[73]
RLP1a3RLP1RLP1aAH5I8MIC/A[74]
RLP1b3RLP1RLP1bAH6R6SQA/T[74]
Table 3. Marker, F-forward and R-reverse primer sequences, and PCR cycling program.
Table 3. Marker, F-forward and R-reverse primer sequences, and PCR cycling program.
Gene, LocusMarkerPrimer Sequence (5′–3′)PCR Cycling Program
F7 QTLAE10-375CTGAAGCGCACGTTCTCC-F
CTGAAGCGCATCATTTCTGATAG-R
1 cycle95 °C—3 min, 35 cycles (95 °C—40 s; 60 °C—40 s; 72 °C—60 s), 1 cycle 72 °C—10 min.
F7 QTLGE-8019TTGAGACCGATTTTCGTGTG-F
TCTCTCCCAGAGCTTCATTGT-R
1 cycle 95 °C—3 min, 35 cycles (95 °C—40 s; 60 °C—40 s; 72 °C—60 s), 1 cycle 72 °C—10 min.
The amplification results were analyzed using electrophoresis in 1.5% agarose gel in TAE buffer. Statistical analysis. The linear regression analysis was performed using Phyton version 3.10.0 software.
Table 4. Results of the SNP and SCAR analysis of apple varieties.
Table 4. Results of the SNP and SCAR analysis of apple varieties.
GenotypesSNP IDSCAR Markers
FBE-1_Y320FBE-2_Y192FBE-2_Y495FBE-2_Y551FB-MR5-K35FB-MRS-R240FB-MR5-R249FB-MR5-rp16k15_M106RLP1aRLP1bAE-375GE-8019
Idared
(1, 4, 6)
T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Starkrimson (6, 8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Gala (4, 6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Aport (4, 7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Fuji (1, 6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Golden Delicious (1, 6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Granny Smith (6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Suislepper (7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/AN/AN/A
Renet Burhardta (4, 7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Pestrushka (4)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Red Delicious (5)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Tulpan (8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Ainur (1, 7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Aigul (7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Saltanat (7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/AN/AN/A
Voskhod (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Egemen (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Zarya Alatau (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Maksat (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Esen (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Sarkyt (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Talgarskoe (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Zharkyn (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Danalyk (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/AN/AN/A
Modi (8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Braeburn (1, 3, 8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Jeromini (4)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Quinti (5, 8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A01
Kandil Sinap (8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Mutsu (8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Kandil Kirghiz (8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Pink Lady (4, 6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/AN/AN/A
Starks Earliest (8)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Scarlet Spur (6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Summer red (6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A11
Nicole Grain (6)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Redfree (3)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Williams Pride (3)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A01
Diana (3)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Gold spur (4)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Honeycrisp (4)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A11
Camspur (4)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Korea (4)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/AN/AN/A
Gala Anna (5)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
July Chernenko (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Elstar (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A01
Jonagold (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Elrosa (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Prima (1)T/T T/T T/TG/GG/GC/C A/A00
Red Elstar (1)T/TC/CNo AMPT/TT/TG/GG/GC/CC/CA/AN/AN/A
Maria Red (1)T/TC/CNo AMPT/TT/TG/GG/GC/CC/CA/A00
Jonaprints (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Synap Almatinsky (1)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Wilton Star (2)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A10
Deljonca (2)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/AN/AN/A
Santana (2)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
Pinova (2,7)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A11
Red Topaz (2)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A11
SQ159 (2)T/TC/CT/TT/TT/TG/GG/GC/CC/CA/A00
1—Amplification for AE10-375 and GE-8019; 0—no amplification; N/A—nonamplified. (1) Regional Branch of the Kazakh Research Institute of Horticulture, Almalyk village, Talgar District of Almaty Region; (2) UOH “Agrouniversitet”, Baibulak village, Talgar District of Almaty Region; (3) Agricultural Innovations Limited Company, Kyzylsharik village, Yenbekshikazakh District of Almaty Region; (4) Peasant farm “Mahmut”, Bayseit village, Yenbekshikazakh District of Almaty Region; (5) Sady Zhetysu Trade LLP, Koram village, Yenbekshikazakh District of Almaty Region; (6) Amangeldi LLP, Akzhar village, Kazygur District of Turkestan Region; (7) Kentau LLP, Tulkibas District of Turkestan Region; (8) Peasant farm “Devrosh”, Merke village, Zhambyl Region.
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Kairova, G.; Daulet, N.; Solomadin, M.; Sandybayev, N.; Orkara, S.; Beloussov, V.; Kerimbek, N.; Gritsenko, D.; Sapakhova, Z. Identification of Apple Varieties Resistant to Fire Blight (Erwinia amylovora) Using Molecular Markers. Horticulturae 2023, 9, 1000. https://doi.org/10.3390/horticulturae9091000

AMA Style

Kairova G, Daulet N, Solomadin M, Sandybayev N, Orkara S, Beloussov V, Kerimbek N, Gritsenko D, Sapakhova Z. Identification of Apple Varieties Resistant to Fire Blight (Erwinia amylovora) Using Molecular Markers. Horticulturae. 2023; 9(9):1000. https://doi.org/10.3390/horticulturae9091000

Chicago/Turabian Style

Kairova, Gulshariya, Nurzhan Daulet, Maxim Solomadin, Nurlan Sandybayev, Shynggys Orkara, Vyacheslav Beloussov, Nazym Kerimbek, Dilyara Gritsenko, and Zagipa Sapakhova. 2023. "Identification of Apple Varieties Resistant to Fire Blight (Erwinia amylovora) Using Molecular Markers" Horticulturae 9, no. 9: 1000. https://doi.org/10.3390/horticulturae9091000

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