Biological Strategies and Innovations in Pest Control and Fruit Storage in Apple Orchards: A Step Towards Sustainable Agriculture

Abstract

The production of apples plays a crucial role in global agriculture. In 2023, the world production of these fruits amounted to nearly 150 million tonnes, cultivated on 6.6 million ha. Today’s horticulture faces the difficult challenge of maintaining high productivity while simultaneously reducing negative environmental impact. Traditional methods based on chemical pesticides encounter increasing problems, such as biodiversity loss, toxic residues in food, development of pest resistance, and disrupted balance of ecosystems. Integrated Pest Management (IPM) responds to these challenges by combining biological and agrotechnical methods with selective use of chemicals. Biopesticides are a crucial component of IPM, and they include antagonist microorganisms, substances of natural origin, and other biological methods of control, which represent effective alternatives to conventional measures. Their development is driven by consumer requirements concerning food safety, as well as by the need to protect the environment. The aim of this article is to highlight current problems in apple production, describe microorganisms and natural substances used as biopesticides used for the protection of apple orchards, as well as present the characteristics of modern technologies used for biocontrol in apple orchards.

1. Introduction

The production of apples plays a crucial role in global agriculture [1]. According to the latest report of the Food and Agriculture Organisation of the United Nations developed in 2023, 147 million tonnes of apples over an area of 6.6 million ha were produced all over the world [2]. The greatest producers of this fruit are China (57.2% of the global production), the EU (13.1%), and the United States (5.8%) [2]. In Poland, which holds the first place in the EU, apple orchards cover over 150 thousand ha, generating an income estimated at approximately PLN 1.8 G a year [2].

However, today’s horticulture faces the double challenge of maintaining high productivity while simultaneously reducing negative environmental impact. Global problems in apple production are related to climate change, rising production costs, tightening food safety regulations, and changing consumer demands for healthier products [3]. The intensification of production in recent decades, based on new varieties and cultivation technologies, has enabled increasing yield while reducing in the area under crops. However, with the introduction of new varieties of apples and reduction in the use of pesticides, diseases and disorders like dry lenticel rot and white haze emerge, negatively affecting the quality of fruit and reducing their commercial value [4,5,6]. To obtain yields of high quality, effective management of biotic and abiotic factors is required [7]. The dominance of traditional methods for plant protection remains a challenge. Protection of apple crops in China, the U.S. and the EU is mainly based on chemical protection, which may lead to pesticide residues in the fruit and to the development of pathogen resistance, especially to fungicides: strobilurin and benzimidazole [8,9,10].

Traditional methods based on chemical pesticides encounter increasing problems, such as biodiversity loss, toxic residues in food, development of pest resistance, and disrupted balance of ecosystems [11]. In response to these challenges, IPM forms the basis of a sustainable approach by combining biological and agrotechnical methods, and selective use of chemical agents. Biopesticides are a crucial component here, and they include antagonist microorganisms, substances of natural origin, and other biological methods of control, which represent effective alternatives to conventional measures. Their development is driven by consumer requirements concerning food safety, as well as by the need to protect the environment [12,13,14].

The biological control represents a new alternative to synthetic pesticides in the integrated management of pathogens and pests in orchards [15,16]. Bacteria, yeasts, and yeast-like fungi require special attention as biocontrol agents (BCAs), because they can colonise the fruit surface and survive in stressful environmental conditions before and after the harvest (resistance to low availability of nutrients, a wide range of relative humidity, low temperatures, low oxygen levels, pH fluctuations, and UV radiation) [6]. Furthermore, they are adapted to the fruit microenvironment, which is characterised by high sugar levels, a high osmotic pressure, and a low pH; they also do not produce allergens or mycotoxins harmful to human health [16,17]. Most importantly, they do not negatively affect human health [18].

Treatments with bioformulations are applied in orchards before harvest, (pre-harvest), but they can also be applied post-harvest—during storage, because many microorganisms effectively control many apple diseases after harvest [19,20,21]. Recently, antagonists of microorganisms have been used with significant success to control diseases in storage [22]. However, the full potential of the use of biopesticides in horticulture requires an in-depth understanding of the mechanisms of their action and their effective integration with agrotechnical practices and IPM systems.

The term “sustainable agriculture” covers farming practices aimed at the production of high-quality and safe agricultural products that do not impair the natural environment and the social and economic conditions of farmers. Maintaining the ability of future generations to sustain their own needs, while ensuring inclusive economic growth remains the main objective for sustainable agriculture [23,24,25].

The aim of this review article is to highlight problems associated with production of apples, describe microorganisms and natural substances used in biopesticides used for the protection of apple orchards, as well as present the characteristics of modern technologies used for biocontrol in apple orchards.

2. Problems in Apple Orchards

Although apple orchards are one of the most important areas of horticulture in the world, this sector struggles with numerous challenges affecting the size and quality of yields [26]. Apple trees are highly susceptible to a wide range of pathogens and phytophages [27]. The most important threats include fungal and bacterial diseases that may result in significant economic losses, as well as various species of harmful insects that damage leaves, buds, and fruit, reducing their market value [18,28,29]. The additional problem is weed competition, which reduces the availability of water and nutrients for trees, negatively affecting their condition [30]. The complexity of these hazards requires a comprehensive strategy for orchard protection, by integrating diversified methods and technologies that are both chemical and biological. By combining these methods, agrophages can be effectively controlled, with simultaneous reduction in environmental impact.

2.1. Diseases

In general, the identification of disease status and spatial distribution represent the first and most important components of integrated disease management [31]. It is an important tactic for timely and accurate control in the integrated disease management approach [32]. Common diseases in apple orchards include anthracnose, apple scab, fire blight, powdery mildew, white haze, root rot, fruit rot, leaf defoliation, blossom blight, trunk, and viral diseases [33,34].

Anthracnose, caused by the fungus Cryptosporiopsis curvispora, attacks both apple trees and their fruit, and this affects the overall health of the plants [35]. The infection with anthracnose begins when tree stems receive large quantities of moisture following heavy rainfalls. Spores are disseminated with rain, mainly in autumn and spring, at the time when the infection is most active. The fungus attacks trees in autumn, but cankers may become visible only in spring [36]. The disease symptoms are cankers, which are small, round reddish spots on the bark that enlarge, cave in, and change their colour from orange to brown, with cracks at their edges. The infected fruit have areas of decay with a brown centre and a light rim. The tree can be infected through small wounds or even directly through an undamaged bark [33].

Apple scab, caused by the fungus Venturia inaequalis, represents the most serious disease affecting apple trees in the world, causing significant economic losses in orchard production, reaching even 30–40% without the appropriate control [37,38]. When sufficient measures for disease control are not implemented, the economic losses may increase up to 70% of the production value, potentially affecting 100% of crops [18]. Symptoms include black or brown lesions on leaves and on fruit, which can lead to fruit deformation. The infected fruit usually begins to rot within its infected area, and in severe cases, the fruit yield is negatively affected [39]. The life cycle of V. inaequalis covers both asexual and sexual phases, and it adopts to seasonal changes to survive and reproduce. Spring, when ejected ascospores germinate and form a mycelium, is the optimal time for infecting apple trees. In summer, the mycelium spreads the infection through the asexual cycle. In autumn, the fungus enters the phase of sexual reproduction, to increase its survival in winter [40]. Apple scab control, to a large extent, depends on fungicides and sprayings that are regularly applied every 7 to 10 days [41]. Its control frequently requires more than 12 treatments in one growth season [42]. Commercial orchards face the problem of the development of V. inaequalis resistance to fungicides. The double resistance to benzimidazole fungicides (MBC, e.g., thiophanate-methyl and quinone outside inhibitors (QoI, e.g., trifloxystrobin)) is common in over 50% of orchards in main production regions. The studies show that resistance to QoI and MBC fungicides can persist for decades, even after their use is discontinued [8,9].

Fire blight, caused by the bacterium Erwinia amylovora, mainly attacks apple and pear trees [43]. The main symptoms associated with fire blight include necrosis and ooze droplets, and the most susceptible plant organs are flowers, young leaves, actively growing shoots, and immature fruit. As the infection progresses, E. amylovora cells from flowers and/or green tissues reach lignified organs. The invasion of the perennial bark of branches, trunk, rootstock, and, occasionally, roots, by the pathogen usually leads to the development of fire blight cankers [44]. The importance of cankers in the fire blight disease cycle is their role as E. amylovora reservoirs and sources of inoculum. The pathogen overwinters in some cankers [45]. Insects, rain, wind, and contaminated tools for pruning may transfer the pathogen from affected plants onto healthy parts of the same or other plants, resulting in the fast spreading of the disease in orchards [46,47].

Powdery mildew, caused by the fungus Podosphaera leucotricha, manifests as white to grey powdery spots on various parts of plants, such as leaves, stems, flowers, and fruit [48,49]. Apart from these visible spots, the disease may cause other symptoms [33,50]. Attacked leaves may curl or deform, and this hinders their normal growth and forming [51]. Plants’ growth may be inhibited, resulting in a reduction in their height and overall development. Powdery mildew also affects the synthesis of chlorophyll, causing yellowing or browning of leaves [52]. In the advanced cases, the disease causes premature drying and falling of leaves, and deteriorating photosynthesis and nutrient transport. Fruit attacked by powdery mildew usually are of lower quality, and this may reduce the total yield of the plant [49].

White haze, a disease caused by excessive fungal growth on the apple surface, leads to deterioration in the fruit quality and reduction in their commercial value. This disease starts as a white or light grey layer of the fungus on the fruit skin, described for the first time in apples stored in a cold store [53], and later also observed in fruit in an orchard [54,55]. To this date, 11 species from the Basidiomycetous genera have been associated with white haze: Entyloma belangeri, E. davenportii, E. elstari, E. mali, E. randwijkense, Gjaerumia minor, Golubevia heteromorpha, G. mali, G. pallescens, Jamesdicksonia mali, and Tilletiopsis washingtonensis [53,54,55,56,57]. Knowledge on the biology of fungi associated with white haze is limited, and their epidemiology is currently being clarified [4]. No effective strategies for control, prevention, or treatment have been identified. High relative humidity during the growth season is favourable to a higher frequency of white haze occurrence, due to more intense rainfalls, longer periods of leaf wetting, and the use of meshes protecting against hailstones. Furthermore, low temperatures and leaf fertilisation may contribute to this disorder [55,58]. The implementation of correct agrotechnical practices, aimed at reduction in moisture retention in an orchard and avoidance of leaf fertilises, is the sole solution currently used to control white haze [6].

Other Diseases

Contrary to orchards in the U.S. and the EU, where it is one of the most dangerous bacterial disease, fire blight (E. amylovora) of apple trees does not occur in China because this pathogen is subject to the strict quarantine in China. Also, apple scab is very rarely found there. Fruit diseases, such as sooty blotch or bitter rot, are a marginal problem in China, mainly due to the common use of manual fruit bagging on the trees through the main part of their development. Several diseases of significant importance occur in Chinese apple orchards, which clearly differ from phytosanitary problems known from the U.S. or the EU. The most prominent of them are diseases of leaves, marssonina leaf blotch, alternaria, and glomerella leaf spot, and diseases of trunks, like apple valsa canker [59]. Marssonina leaf blotch, caused by Diplocarpon mali, is one of the most important leaf diseases in China. Its main symptom is multiple dark brown spots on leaves, which result in premature leaf falling. In moist years, this may result in defoliation exceeding 80% one month before the harvest, and this significantly reduces the yield and the quality of fruit. Marssonina leaf blotch is practically not found outside Asia [59,60]. Alternaria leaf spot, caused by Alternaria spp., also leads to the development of brown spots and premature falling of leaves, especially in warm and humid weather [61]. Both types of leaf spots result in weakening of trees, deterioration in fruit colour and taste, and a reduction in yield. Glomerella leaf spot (GLS), caused by Colletotrichum species, is a relatively new, but fast-spreading disease of leaves and fruit, which results in the development of necrotic spots, sudden falling of leaves, and minor damage to fruit in just a few days. GLS demonstrates a strong specificity depending on the variety; for example, “Fuji” is resistant, while “Gala” or “Golden Delicious” is highly susceptible to it. Today, this disease is found mainly in China, Brazil, Uruguay, and the U.S., but is practically unknown in Europe [59,62]. Apple valsa canker, caused by Cytospora mali, is the most serious disease affecting bark and wood in Chinese orchards. It results in the formation of wounds on trunks and branches, resulting in the death of tree parts. It is particularly dangerous in orchards with low soil organic matter content and in the case of potassium deficiencies [59,63]. All listed diseases negatively affect the cultivation of apples in China, leading to the weakening of trees, reduction in yield, and the deterioration of fruit quality, requiring intense plant protection activities. Their occurrence and significance differ significantly from diseases dominating in orchards in Europe and the U.S., where apple scab and fire blight represent more pressing problems, while diseases of leaves and bark are of lesser significance.

2.2. Insects

The double role of insects in orchards, as pests and as beneficial organisms, needs to be highlighted. Beneficial insects like pollinators and predators/parasitoids play a crucial role in pollination or pest control. Other insects damage trees and fruit, and therefore require a strategy for their control. Chemical control used to be a conventional strategy for the control of pest insects in apple orchards, and it frequently resulted in excessive use of pesticides, repeated increase in pest numbers, development of resistance to pesticides, and serious environmental pollution. These problems lead to the increase in the use of integrated strategies for pest control [64]. Today, the control of apple tree pests is based on the use of methods for orchard management that employ biological control [65]. Species composition of pest insects and their influence on the yield is diversified, and depends on the geographical region and climate conditions. Boltabaev et al. [66] identified six main pests of apple trees, whose presence and characteristics may differ from those occurring in orchards in different regions of the world. The most important pest insects affecting apple trees are apple maggot (Rhagoletis pomonella), codling moth (Cydia pomonella), European red mite (Panonychus ulmi), plum curculio (Conotrachelus nenuphar), and tarnished plant bug (Lygus lineolaris) [67].

Apple maggot (R. pomonella) originally fed on the fruit of the wild hawthorn (Crataegus spp.) but later became a primary pest of cultivated apples. The varieties that mature in the summer and early fall are particularly vulnerable, but hard winter apples are sometimes infested. Thin-skinned, sweet, and subacid varieties are most the susceptible, but acid varieties may also be attacked. The adult fly is black, with white bands on the abdomen (four in females and three in males), and its wings are conspicuously marked with four oblique black bands [68]. Adults emerge from the ground in early summer. Emergence continues for a month or longer, and many pupae may remain inactive and not emerge until the second year. Fruit is injured by maggots boring throughout them, forming irregular, winding tunnels which turn brown, often causing premature dropping of fruit. When the fruit is slightly infested, there may be no external indication of the maggots, but when the fruit ripens, the burrows show as dark, winding trails beneath the skin. The minute the egg punctures, distorted and pitted areas may show on the surface. Heavily infested fruit of early varieties will be reduced to a brown rotten mass filled with the larvae of the fly [69].

Codling moth (C. pomonella) is a major pest of pome fruit crops, which are primarily apples and pears [70]. It is found in most parts of the world where these fruits are grown. In orchards without pest management, C. pomonella infestation rates can reach 80% for apples [71]. The larvae of the codling moth bore into the fruit, causing it to rot and become unmarketable. The adult codling moth is a small moth, with greyish-brown forewings with a coppery band at the tip. The hindwings are paler. The larvae are white with a brown head. Female codling moths lay eggs on the surface of fruit, typically near the stem or calyx end. Once the eggs hatch, the larvae bore into the fruit, feeding on the flesh and seeds. This feeding activity results in the destruction of the fruit’s internal tissue, making it unsuitable for consumption or commercial sale [72].

European red mite (P. ulmi) is a pest of various trees and small fruit crops, including apples. P. ulmi overwinters as eggs deposited in the rough bark, cracks, and crevices of the limbs and twigs of trees in apple orchards [73]. Upon favourable conditions during the subsequent spring, these eggs hatch into larvae that develop to adults. Adult and immature P. ulmi causes damage by primarily feeding on the host leaves and sucking their cell contents, including chlorophyll, thus interfering with processes of photosynthesis and the production of carbohydrates [74]. Continuous feeding on leaves initially causes white stippling but the damaged areas of infested leaves quickly turn brown in what is known as ‘bronzing’ by apple growers. Depending on the population level and the tree physiological stage and status, P. ulmi infestation in apple leaves results in economic losses by inversely interacting with crop load and fruit quality, size, and yield, as well as the bloom and development of fruit spurs in the subsequent year [75].

Plum curculio (C. nenuphar) is a key pest of apple orchards. Overwintered adults move from wooded areas into cultivated crops around bloom, where they feed, mate, and reproduce throughout the early growing season. Feeding and oviposition (egg-laying) scars cause fruit drop, leading to significant yield losses, with an estimated damage of up to 85% in untreated orchards [76]. C. nenuphar adults are mottled brown, grey, and white, with distinct sets of ‘humps’ along the dorsal area of the elytra [77]. C. nenuphar injury can be classified into two distinct categories: adult feeding and oviposition. Since this insect feeds on the same hosts that it uses as oviposition sites, these injuries often co-occur on the same fruit, but are generally distinct in their appearance. Feeding injuries can appear as round holes, wherein the beetle inserts its rostrum as far as it can into the fruit, feeding on the flesh underneath abscessed skin. In some fruits, such as apple, this can result in feeding holes large enough for C. nenuphar adults to hide in, particularly when multiple feeding holes run together. Oviposition injury involves the female C. nenuphar first cutting a small oval incision in the surface of the fruit, then depositing an egg inside the hole and forming a scar [78].

Tarnished plant bug (L. lineolaris) is a polyphagous, sap-feeding true bug that causes significant damage in several crops. It feeds on developing apple flowers and fruit, inserting its mouthparts to extract plant juices and inject saliva. This feeding causes damage such as flower bud abscission, underdeveloped fruit, and malformations.

Each type of pest insect has different biological characteristics and ways of feeding, and this translates into greater problems in establishing a single effective method for their control. This diversity is reflected in the polymorphism of symptoms within the vegetative and generative organs of apple trees. Agrophages may attack leaves, stems, and fruit, leading to reduction in quality and quantity of the yield [79]. Apple tree pests, to a large extent, influence the effectiveness and profitability of horticultural production. Their composition and influence are strongly diversified regionally, and this results in the need for an individual approach for monitoring and controlling these organisms [80].

2.3. Weeds

Weeds in orchards pose a serious threat to the health and growth of trees, as they compete with them for essential resources such as water, nutrients, and light. They frequently gain an advantage, because their root system is usually denser and better developed than that of young orchard plants. In spring, damages caused by frost are greater on the soil covered with weeds than on the bare soil. Their presence may lead to the deterioration of the condition of plants, thereby increasing their susceptibility to diseases and pest attacks [81]. Furthermore, weeds may also provide good conditions for the development of certain orchard pests. Weed control in orchards requires appropriate methods, which minimise environmental impacts and maintain biological balance [82]. In practice, both mechanical weed removal and selective herbicides are used; however, increasingly greater attention is paid to the integrated management that promotes the preservation of the biodiversity of weeds of lower harmfulness that may support the orchard ecosystem, creating advantageous conditions for natural antagonists [83]. In orchards, the most common annual weeds are chickweed, common lamb’s quarters, common groundsel, and shepherd’s purse. Some perennial weeds, like quackgrass and dandelion, can also be problematic [84].

2.4. Fruit Storage

The question of apple storage is an important issue for orchard owners. Fruit is at risk of rotting, taste deterioration, and physical damage. These problems are frequently associated with factors such as incorrect temperature, humidity, and ventilation, as well as the presence of pathogens [85]. Storage diseases caused by fungal pathogens represent the most common problem in the storage of apples and other fruit [86]. The main storage diseases of apples are blue mould, bitter rot, and bull’s eye rot. Blue mould of apples is caused by a fungus, Penicillium expansum, which grows in temperatures as low as 1 °C. It manifests as circular, light tan to dark brown lesions that often have a soft, watery texture, and involve blue-green spore masses. A musty or earthy odour is also a key indicator. The disease is particularly problematic because it can render fruit inedible and can also lead to the production of the mycotoxin patulin, which is dangerous to human health [87,88,89]. Bitter rot of apple, caused by fungi Colletotrichum spp., leads to the development of dark, concave spots with pinkish coating. The spots expand and cause rotting of fruit. The disease develops particularly in high humidity [90]. Bull’s eye rot of apple caused by the Neofabraea species, is characterised by round, depressed lesions with a darker centre and light brown peripheral ring. It develops only after several weeks of storage [91].

3. Chemical Pesticides Applied in Apple Production and Pesticide Residues in Apples

Chemical pesticides form the basis of the conventional protection of apple orchards. They are routinely applied for a period of up to eight months in the year, to control numerous pests and diseases. In conventional systems, the strategy is based on the principle of maximising profit and minimising the risk of yield losses. Chemicals are the main, and sometimes the only, agents used to control pests, diseases, and weeds in production systems of this type. Orchards using pesticides are characterised by having the highest percentage of top quality fruit (85–90%) and the lowest level of losses caused by pests and diseases [92].

The intensity of pesticide use is greatly varied, depending on the crop. In the temperate zone, apples belong to crops with the most intense use of pesticides. In 2022, the number of notified active substances used in the production of apples amounted to 30 in Germany [93], 41 in Italy [94], and 49 in Austria [95]. The use of pesticides poses a significant threat to the agroecosystems of apple orchards, which are characterised by high biodiversity and represent an important habitat for many groups of organisms [96,97]. Pollinating insects, and the honeybee in particular, play a crucial role in the pollination of horticultural crops, directly influencing the yield and the quality of fruit [98,99]. Beneficial insects (predators, parasites), which are natural enemies of apple pests, play a key role in the protection of apple crops [79]. Birds represent an important link in biological pest control in apple orchards and vineyards, as they eat a wide range of phytophagous insects, and this translates into a natural, environmentally sustainable control of pest numbers [100,101]. Earthworms, through the process of the mineralisation of organic matter and its transformation into humus, significantly contribute to improvement in the physical and chemical properties of soil [102]. Their bioturbation activity contributes to the increase in soil porosity, intensifying its aeration and water infiltration. Chemical plant protection agents have numerous side effects, negatively affecting non-target organisms, including pollinating insects, birds, and soil fauna, and this leads to a decrease in biodiversity on the scale of the entire ecosystem [103,104,105,106,107,108,109].

Control of apple tree diseases, to a large extent, involves the use of fungicides, with sprays routinely applied every 7 to 10 days from bud burst until the risk of scab subsidies [41]. Their control frequently requires more than 12 treatments in one growth season [42]. However, this high frequency of treatments increases input costs, reduces fungicide effectiveness, and increases the number of cases of developed resistance. Furthermore, they represent a serious hazard to human health and may cause degradation of the environment [18].

Studies conducted by Cai et al. [64] in the main regions of apple production in China showed that the average use of pesticides was at the level of 13 kg/ha. This amount significantly exceeds quantities used by other countries. For example, the amounts used are 2.54 kg/ha in the United States, 3.63 kg/ha in France, and 9.86 kg/ha in the Netherlands. The results of the studies show that 70.6% of apple farms use pesticides excessively, in amounts of 1.7 times higher than the economically optimal level. The excessive use of chemical pesticides in apple production has an adverse effect on food safety and human health. It is estimated that, in China, the excessive use of pesticides leads to a 100 thousand cases of accidental poisoning of farmers in a year, some of which are fatal [64]. Furthermore, higher pesticide residue levels raise growing consumer concern for the safety of eaten products.

In studies conducted in Lebanese orchards in 2012–2016, the residues of mainly two pesticide classes were found, i.e., insecticides (92% of samples) and fungicides (18% of samples). The detectable residues were determined in 77% of samples, with 61% exceeding the EU Maximum Residue Limits (MRLs). Particularly alarming were the cases involving chlorpyrifos (MRL of 10 μg/kg), as its levels reached 750 μg/kg (75 times the limit), and methidathion (MRL of 30 μg/kg) with determined levels of up to 234 μg/kg. In 22.6% of the samples, multiple residues were found (up to five substances simultaneously) [110]. These results indicate a real risk associated with the use of chemical pesticides in crops.

According to the European Food Safety Authority (EFSA), in 2022, quantitative results for a pesticide-active substance were obtained for 5704 (48.6%) out of 11,727 food samples. Among them, the presence of more than one pesticide was determined in 3760 (32.1%) of the samples. Food products with a higher rate of multiple residues included apples (18.6%), strawberries (17.5%), peaches (16.9%), tomatoes (14.2%), and lettuce (12.4%). For apples, some pesticide residues could cause acute risk to human health [111].

In conclusion, the negative consequences of pesticide use necessitate the development of alternative methods for pest control and environmentally friendly solutions. To meet these challenges, numerous studies were conducted to verify the possibility of using biological control agents to combat resistance, with the simultaneous task of minimising risks to human health and the environment [18,112].

4. Biopesticides

The limitations in pesticide use hold a prominent place in the international political agenda; for example, the European Green Deal with its “Field to Fork” and “Biodiversity” strategies, which aim, among others, to reduce by half the usage and associated risks of chemical pesticides in the European Union (EU) by 2030, and to expand organic farming to 25% of farmlands in the EU [112,113]. Biopesticides are a group of organisms or substances of a natural origin used to protect plants against agrophages—which consists of pathogens, weeds and pests—while minimising environmental impact and the influence on consumers’ health [114]. Basic mechanisms of their actions include competition for nutrients and living space, induced systemic resistance (ISR), mycoparasitism, antagonism, and secretion of lytic enzymes or antibiotics [115]. Key advantages of the use of biopesticides are the significantly reduced risk of ecotoxicity (e.g., safety to pollinators and beneficial organisms, minimised environmental impact, elimination of toxic residues in crops, compatibility with sustainable production systems), organic farming, lower risk of resistance development in pathogens, improvement in soil fertility and tree health, and the short or absence of waiting period [116].

Biopesticides and natural substances used in apple production are summarised in Table 1 and described in detail below.

4.1. Fungi, Yeast, and Bacteria Recommended for Apple Cultivation

4.1.1. Aureobasidium pullulans

A. pullulans is naturally present on apple trees without causing plant diseases; therefore, it is commonly used in the biological control of apple tree diseases, due to its antagonistic effect towards pests [22].

The main mechanisms of the effects of A. pullulans are competition for nutrients and space [117], production of siderophores [118], aureobasidin A [119], and biofilm formation [120]. Secretion of lytic enzymes, including exo- and endo-chitanases, β-1,3-glucanase and protease, also contributes to biological activity [121]. Furthermore, volatile organic compounds (VOCs) produced by A. pullulans showed antifungal activity against apple pathogens after harvest, both in vitro and in vivo [6,20,122,123]. The above mechanisms protect crops against diseases and also have an advantageous effect on plant growth and development. As they can produce siderophores that help to dissolve iron in the soil, they increase the availability of this element to plants [124].

A. pullulans is used to protect apples in storage, which reduces rotting; at the same time, it also supports a microbiome that hinders the development of diseases, offering an environmentally friendly strategy that enables the sustainable post-harvest control of diseases [125]. Remolif et al. [6] demonstrated that A. pullulans strains, AP2 and PL5, are effective in the controlling of white haze on apples caused by E. belangeri, Golubevia pallescens, and Tilletiopsis washingtonensis during storage.

Zeng et al. [126] used A. pullulans DSM 14940 and 14941 to control fire blight caused by E. amylovora, activating the systemic acquired resistance (SAR) in apple flowers. A. pullulans demonstrated effectiveness against P. expansum [19,125], B. cinerea [19,127], Colletotrichum acutatum [19], and Alternaria alternata [20]. A. pullulans is characterised by its ability to adapt to various environmental conditions, including fluctuations in temperature, humidity, and nutrients availability. It is also characterised by its resistance to drought and radiation, and it provides a great advantage when it is used as a component of bioformulations used in the agriculture [128,129]. In some cases, A. pullulans helps plants to cope with drought stress through the modulation of the microbiome of rhizosphere [130].

A. pullulans represents a safe and environmentally friendly alternative to synthetic fungicides used in traditional systems to control Botryosphaeraceae-induced diseases, such as benomyl or tebuconazole [131].

The only strains approved for use in the EU are A. pullulans DSM 14940 and DSM 14941 [132].

4.1.2. Pythium oligandrum

P. oligandrum represents a unique case in the Pythium genus, contrary to pathogenic species, and shows strong antagonistic effects against soil pathogens; therefore, it is one of the most promising bioformulations used in the sustainable cultivation of apple trees.

The mechanism most frequently used by P. oligandrum for biocontrol is mycoparasitism. This species produces enzymes—chitinases, cellulases, endo-β-1,3-glucanases, proteases, and phosphatases—which degrade pest cellular walls. P. oligandrum genome contains 114 glycoside hydrolases, of which 79 are secreted [133,134]. These processes can modify the surface structures of pest cells and support P. oligandrum in competing for living space and nutrients [135]. P. oligandrum may interact with plant roots, by the fast and mass colonisation of root tissues without interfering with the host’s cell wall or changing its cells. Instead, it induces defence mechanisms in plants, thus strengthening future plant reactions to pathogens [136].

P. oligandrum produces numerous elicitors, termed oligandrins, and cell wall protein fractions [136,137,138,139,140]. These proteins induce plant defence systems, thus initiating both local- and systemic-induced resistance against fungal, oomycete, and bacterial pathogens [141]. P. oligandrum contains other proteins that can act on the immune system of a plant [136]. Other proteins possibly involved in inducing defence reactions include necrosis- and ethylene-inducing peptide1-like proteins (NLPs), two of which activate defensin genes without causing the accumulation of reactive oxygen species [135,142,143].

P. oligandrum also contributes to plant growth. It produces tryptamine—a precursor of auxin, and a growth phytohormone—which supports the development of roots and improves the overall condition of plants [144].

Being a strong mycoparasite, P. oligandrum was shown to coil around the hyphae of other oomycetes and fungi, such as Fusarium oxysporum, Phytophthora parasitica, Verticilium dahliae, Botrytis cinerea, Aphanomyces cochlioides, and even other Pythium species [135,142,145].

In the context of apple tree cultivation in a nursery, the use of P. oligandrum was highly beneficial for the development of the young trees of two apple varieties, “Szampion” and “Topaz”. The studies showed that this oomycete improved the percentage share of obtained one-year-old apple trees by 8.4–12.8%, depending on the variety used [146]. Another advantage of the use of P. oligandrum as a biopesticide is its ability to remain in the rhizosphere for a long time. The studies showed colonisation at a level of 50% or higher throughout the growth season (from April to September) [147].

Rubak et al. [148] proved that P. oligandrum contributes to the reduced rotting of apples in storage; when compared to samples not subjected to treatment, both stored at a lower temperature, and in a nitrogen atmosphere with ripening inhibitors.

Currently, two P. oligandrum strains—B301 and M1—are approved for use in the EU [132].

4.1.3. Candida oleophila

C. oleophila is used for biological plant protection mainly against apple tree diseases caused by B. cinerea and P. expansum. Its main effect involves competing with pathogens for nutrients and living space. This competition is of particular significance at sites of wounds, where pathogens imitate plant infections [149]. C. oleophila may induce the defence mechanisms of a plant, including the production of pathogenesis-related proteins and enzymes. This induced resistance may improve the plant’s resistance to successive attacks of pathogens [150,151].

C. oleophila produces and secretes extracellular lytic enzymes that degrade a cell wall, including exo-beta-1,3-glucanases, chitinases, and proteases. Exo-beta-1,3-glucanases showed an inhibitory effect on spore germination and germ-tube elongation [152].

Cai et al. [153] discovered that C. oleophila can control P. expansum in apples by competing for life space and nutrients, while Liu et al. [150] proved the ability of C. oleophila to inhibit B. cinerea by inducing expression of genes of proteins associated with pathogenesis in apples.

A crucial feature distinguishing C. oleophila from other microorganisms is its ability to grow at low temperatures, even 0 °C, and this is crucial for its use in the cold storage of apples. This thermophilic property enables yeasts to remain active in the conditions of apple storage, even for 12 months [154].

C. oleophila produces substances supporting plant growth, such as auxins and cytokinines [155].

The only strain approved for use in the EU is C. oleophila O [132].

4.1.4. Bacillus subtilis

Bacillus species are widely used for the biocontrol of plant diseases in the sustainable development of agriculture. Usually, Bacillus bacteria have a priority over other biocontrol agents, due to some of their basic properties, such as rapid replication, resistance to adverse environmental conditions, and development and longevity of spores [156,157].

Bacillus species isolated from the rhizosphere soil may be an effective stimulator of plant growth, inducing ISR and producing a wide range of antimicrobial compounds, such as antibiotics, lipopeptides, and enzymes [158]. Approved biological control agents containing B. subtilis have fungistatic and fungicidal effects [159].

The proposed modes of antagonistic actions of B. subtilis involve the production of either cyclic lipopeptides (LPs) (such as fengycin or plipastatin, surfactin, iturin, bacillomycin, mycosubtilin) or of lytic enzymes (such as chitinases, cellulase, endoglucanase, or hemicellulose), which suppresses pathogen growth [160]. Romero et al. [161] reported that LPs provide protection to plants both in pre- and post-harvest conditions, directly inhibiting the development of pathogenic fungi. B. subtilis produces high levels of antibiotics, especially iturin [162]. It competes with pathogens for life space and nutrients [163]. B. subtilis shows a strong, very stable formation of biofilm, and produces surfactin that has a very strong antibacterial effect [164].

Bacillus spp. support plant growth through phosphate solubilisation, nitrogen binding, and synthesis of phytohormones [165]. B. subtilis contributes to the induction of ISR in plants, involving the expression of specific genes and hormones [162]. Ethylene restricts the growth of roots and stems, helping to maintain plant homeostasis. The degradation of the ethylene precursor by bacterial ACC deaminase helps to alleviate plant stress and maintain normal growth in stressful conditions [166]. Certain VOCs produced by the B. subtilis strain also help plants in combating pathogens. Bacillus spp. also secrete exopolysaccharides and siderophores, which inhibit the movement of toxic ions and help to maintain ionic balance, promoting the movement of water in plant tissues [167].

B. subtilis strains are considered safe to be used in the food industry and are easy to store due to their ability to form endospores resistant to heat, UV radiation, organic solvents, and drought [168].

B. subtilis is used to protect plants against F. oxysporum, Alternaria solani, B. cinerea, Pseudomonas syringae, Pseudocercospora purpurea, Akaropeltopsis sp., V. inaequalis, Colletotrichum spp., and Rhizoctonia cerealis [156,168,169,170,171,172].

Currently, two B. subtilis strains, IAB/BS03 and RTI477, are approved for use in the EU [132].

4.1.5. Bacillus thuringiensis

B. thuringiensis is one of the most important and safest microbiological insecticides that is widely used in the biological protection of plants, including in apple orchards. This bacterium is distinguished from other microorganisms, especially by its ability to produce δ-endotoxins, particularly Cry proteins that are toxic to many insects [173,174]. The mode of B. thuringiensis effect is based on the lysis of epidermal cells in the insect midgut. When consumed by insect larvae, the digestive enzymes in the gastrointestinal tract activate the toxin, which results in the formation of pores in the membranes of intestinal cells, resulting in paralysis of the digestive tract, and eventually leading to larval death [175]. Cry proteins target a diverse range of insect species primarily of the order Lepidoptera (butterflies and moths), Coleoptera (beetles and weevils), and Diptera (flies and mosquitoes) [176]. Apart from Cry proteins, B. thuringiensis also synthesises vegetative insecticidal protein (Vip) and secreted insecticidal protein (Sip). They affect mainly species of Coleoptera and Lepidoptera [177].

Gumeniuk et al. [178] demonstrated that the use of formulations based on B. thuringiensis strains is an effective, environmentally and economically feasible agricultural measure to protect apple orchards against insect phytophages, such as Hyponomeuta malinellus, and C. pomonella, which resulted in 100% effectiveness in pathogens control, high yield efficiency, and product quality. When used for the protection of apple trees, B. thuringiensis is used to control C. pomonella, Operophtera brumata, Adoxophyes orana, Lymantria dispar, and Euproctis chrysorrhoea [179].

Currently, eight strains of B. thuringiensis, including subsp. aizawai ABTS-1857 and GC-91 strains, subsp. israelensis strain AM65-52, subsp. kurstaki ABTS-351, EG2348, PB 54, SA 11, and SA 12 strains, are approved for use [132].

4.2. Natural Substances

4.2.1. Laminarin

Laminarin is a polysaccharide from a brown sea alga Laminaria digitata, which represents one of the most promising natural substances used as an inducer of natural resistance mechanisms in plants. This property can be used in the sustainable protection of apple orchards [180]. It also has antifungal properties. Laminarin effectiveness is comparable to a synthetic fungicide captan and has promising results in reducing the level of infection in leaves and fruit in two varieties of apples [181]. This polysaccharide activates natural plant defence mechanisms by stimulating the production of defence compounds and strengthening structural barriers against pathogens. The mode of laminarin action is based on the recognition of this polysaccharide as a pathogen-related molecule by plant receptors, and this triggers the signalling cascade, leading to the activation of genes responsible for the synthesis of proteins of pathogenesis or secondary metabolites with antimicrobial properties [182]. This substance is the most effective in a period of secondary infections with apple scab, when the quantity of chemical fungicides applied can be successfully reduced. In practice, laminarin needs to be combined with other fungicides at moments of high and moderate risks of infection, to ensure very good disease control in fruit [183]. In the study conducted in Belgium, it was demonstrated that residues of only two fungicides (captan and dithianon) at lower concentrations were found in a sample containing synthetic pesticides with laminarin added, while four different pesticide residues were found in the sample containing only chemical fungicides [183]. Additionally, laminarin does not leave its residues on fruit, due to its biodegradable characteristics [184], and this is important for orchard owners striving to meet increasingly strict commercial standards.

4.2.2. Orange Oil

Orange oil acts through a physical mode of action, desiccating the cuticle of mites and soft-bodied insects, resulting in their sudden death by exposing them to the loss of body fluids. It also affects winged insects, destroying their protective layer and wing tension, so they cannot fly. As a biofungicide, it dries protective membranes of fungal structures, inhibiting the spreading of the infection without damaging healthy tissues. Its insecticidal effect may be associated with the inhibition of acetylcholinesterase and the activity of the sodium–potassium pump in insects [185,186,187]. Orange oil can also act as a repellent or disturb the behaviour of certain insects, hampering their feeding and reproduction on the apple trees [186,188].

4.3. Pheromones

The application of insect sex pheromones is a well-established method in pest management in apple orchards, as it interferes with mating behaviour and can ultimately lead to long-term decreases in pest populations [189]. Artificially produced pheromones used for mating disruption are generally highly species-specific and have not been associated with significant negative effects on non-target organisms [190,191].

Pheromone traps are used to monitor the flight dynamics of pest moths, helping growers understand when pests are active and aiding in timely control decisions [192]. Trapping C. pomonella is an essential component of crop protection. The moth counts are used to determine when it first arrives in the crop area, the population dynamics, and when it is active. This information allows improved selection and timing of control measures. Consequently, pheromone-baited traps for monitoring C. pomonella have become commonplace in orchards [193].

Pheromone-based lures and traps are widely implemented across different crops, relying on the fact that many agricultural pests depend on sex pheromones to locate mates and ensure reproductive success. Synthetic female sex pheromones, in particular, can attract and concentrate males around the pheromone source [194].

Pheromone-based strategies offer a viable alternative to traditional foliar insecticides, providing a tool for organic apple production with benefits for worker safety and non-target effects [195].

4.4. Natural Enemies of Pest Insects on Apple

Natural enemies of apple pests include beneficial insects such as ladybugs, lacewings, parasitic wasps, hoverflies, and pirate bugs, as well as general predators like spiders and insectivorous birds [79]. Both the adult and larval stages of ladybugs feed on pests, particularly aphids. Wasps, such as Aphelinus mali, lay their eggs inside host pests like the woolly apple aphid, with the larvae killing the host [65]. The most important natural enemies of woolly apple aphid (Eriosoma lanigerum) are the parasitoid Aphelinus mali (Haldeman) and earwigs, but also other enemies like spiders (Araneae), ladybird beetles (Coleoptera: Coccinellidae), lacewings (Neuroptera: Chrysopidae), and hoverflies (Diptera: Syrphidae) could reduce its population [196].

Birds such as tits, swallows, and sparrows also play an important role as predators of insect pests, the most important of which are Great Tits and Blue Tits. Tits help to reduce the population of fruit-damaging pests, such as the Codling moth, thereby contributing to a healthier and more abundant apple harvest [197].

In order to increase biodiversity and create favourable conditions for the development of beneficial insects and natural enemies of pests in apple production, it is recommended to use flowers and cover plants and also create favourable conditions for insectivorous birds (nest boxes) [79].

Table 1. Biopesticides and natural substances used in apple production.

Active Substance	Application [198]	Mode of Action
A. pullulans	FIRE BLIGHT (E. amylovora) BULL’S EYE ROT (Neofabraea spp.)	antagonistic effect, ability to colonise various environments, endophytic characteristic, competition for nutrients, production of antibiotic compounds and induction of a systemic resistance in a host plant [133,136,142]; production of secondary metabolites, including pullulan (α-glucan) and β-glucans that strengthen plant cell membranes, induce ISR, and inhibit development of pathogens [131].
P. oligandrum	BULL’S EYE ROT (Neofabraea spp.) BLUE MOULD (P. expansum)	antagonistic effect, mycoparasitism, antibiotism by secreting substances toxic to pathogens, competition for nutrients and living space; stimulation of plant defence mechanisms and growth promotion through production of biologically active compounds (glycoprotein–oligandrin-inducing ISR, and tryptamine, a compound belonging to auxins that contributes to the development of the plant root system, supporting plant growth) [147].
C. oleophila	GREY MOULD (B. cinerea) BLUE MOULD (P. expansum)	production of the enzyme, exo-β,1-3 glucanase, that degrades fungal cell walls and reduces spore and mycelium growth [152]; other antagonistic mechanisms; competition for nutrients and space [153]; induction of the defence mechanisms of a plant and antibiosis [150,151].
B. subtilis	FIRE BLIGHT (E. amylovora)	ability to biosynthesise a complex set of secondary metabolites, including cyclic lipopolypeptides, polyketones, and antibiotics that determine its antagonistic effects; ISR induction strengthens long-term resistance of crops [158]; it also shows properties promoting plant growth, acting as a bacteria supporting plant development and facilitates phosphate dissolving [165]; produces siderophores of the catechol type, which are associated with affinity to iron and improves its supply to plants [167].
B. thuringiensis	TORTRIX MOTH (Tortricidae) APPLE ERMINE MOTH (H. malinellus) WINTER MOTH (O. brumata) AND OTHER LEAF-EATING CATERPILLARS SAN JOSE SCALE (Diaspidiotus perniciosus) CODLING MOTH (C. pomonella)	presence of δ-endotoxins, especially Cry proteins, which give it its insecticidal properties [173,174]; synthesis of insecticidal protein (Vip) and secretion of the insecticidal protein (Sip) [177]; lysis of epidermal cells in the midgut of the insect. When consumed by insect larvae, the digestive enzymes in the gastrointestinal tract activate the toxin, and this results in formation of pores in membranes of intestinal cells, then in paralysis of the digestive tract, and eventually to larval death [173].
laminarin	FIRE BLIGHT (E. amylovora) APPLE SCAB (V. inaequalis) BULL’S EYE ROT (Neofabraea spp.)	activates natural plant defence mechanisms by stimulating production of defence compounds and strengthening structural barriers against pathogens [182].
orange oil	POWDERY MILDEW (P. leucotricha) CODLING MOTH (C. pomonella) APPLE SUCKER (Psylla mali) APPLE PSYLLID (Cacopsylla mali)	physical mode of action—desiccation of the cuticles of mites and soft-bodied insects, leading to their sudden death by exposing them to a loss of body fluids. It also affects winged insects by disrupting their protective covering and wing tension, rendering them unable to fly [185]; as a biofungicide, it dries out fungal structures’ protective membranes, halting the spread of infection without harming healthy tissues [186]; insecticidal activity could be linked to inhibition of acetylcholinesterase and sodium–potassium pump activities in insects [187].

Table 1. Biopesticides and natural substances used in apple production.

Active Substance	Application [198]	Mode of Action
A. pullulans	FIRE BLIGHT (E. amylovora) BULL’S EYE ROT (Neofabraea spp.)	antagonistic effect, ability to colonise various environments, endophytic characteristic, competition for nutrients, production of antibiotic compounds and induction of a systemic resistance in a host plant [133,136,142]; production of secondary metabolites, including pullulan (α-glucan) and β-glucans that strengthen plant cell membranes, induce ISR, and inhibit development of pathogens [131].
P. oligandrum	BULL’S EYE ROT (Neofabraea spp.) BLUE MOULD (P. expansum)	antagonistic effect, mycoparasitism, antibiotism by secreting substances toxic to pathogens, competition for nutrients and living space; stimulation of plant defence mechanisms and growth promotion through production of biologically active compounds (glycoprotein–oligandrin-inducing ISR, and tryptamine, a compound belonging to auxins that contributes to the development of the plant root system, supporting plant growth) [147].
C. oleophila	GREY MOULD (B. cinerea) BLUE MOULD (P. expansum)	production of the enzyme, exo-β,1-3 glucanase, that degrades fungal cell walls and reduces spore and mycelium growth [152]; other antagonistic mechanisms; competition for nutrients and space [153]; induction of the defence mechanisms of a plant and antibiosis [150,151].
B. subtilis	FIRE BLIGHT (E. amylovora)	ability to biosynthesise a complex set of secondary metabolites, including cyclic lipopolypeptides, polyketones, and antibiotics that determine its antagonistic effects; ISR induction strengthens long-term resistance of crops [158]; it also shows properties promoting plant growth, acting as a bacteria supporting plant development and facilitates phosphate dissolving [165]; produces siderophores of the catechol type, which are associated with affinity to iron and improves its supply to plants [167].
B. thuringiensis	TORTRIX MOTH (Tortricidae) APPLE ERMINE MOTH (H. malinellus) WINTER MOTH (O. brumata) AND OTHER LEAF-EATING CATERPILLARS SAN JOSE SCALE (Diaspidiotus perniciosus) CODLING MOTH (C. pomonella)	presence of δ-endotoxins, especially Cry proteins, which give it its insecticidal properties [173,174]; synthesis of insecticidal protein (Vip) and secretion of the insecticidal protein (Sip) [177]; lysis of epidermal cells in the midgut of the insect. When consumed by insect larvae, the digestive enzymes in the gastrointestinal tract activate the toxin, and this results in formation of pores in membranes of intestinal cells, then in paralysis of the digestive tract, and eventually to larval death [173].
laminarin	FIRE BLIGHT (E. amylovora) APPLE SCAB (V. inaequalis) BULL’S EYE ROT (Neofabraea spp.)	activates natural plant defence mechanisms by stimulating production of defence compounds and strengthening structural barriers against pathogens [182].
orange oil	POWDERY MILDEW (P. leucotricha) CODLING MOTH (C. pomonella) APPLE SUCKER (Psylla mali) APPLE PSYLLID (Cacopsylla mali)	physical mode of action—desiccation of the cuticles of mites and soft-bodied insects, leading to their sudden death by exposing them to a loss of body fluids. It also affects winged insects by disrupting their protective covering and wing tension, rendering them unable to fly [185]; as a biofungicide, it dries out fungal structures’ protective membranes, halting the spread of infection without harming healthy tissues [186]; insecticidal activity could be linked to inhibition of acetylcholinesterase and sodium–potassium pump activities in insects [187].

5. Technological Innovations in Biocontrol Applications

Recently, new technological innovations used in the biocontrol of apple orchards have been developed. The most important innovations are reviewed below.

5.1. RNAi

5.1.1. RNAi-Based Pest Control

The RNA interference (RNAi) technology is a modern method for pest control that uses natural mechanisms for gene silencing by sequence-specific inhibition of translation or RNA degradation, controlled by small RNA [199,200]. In this method, insects are given specially designed molecules of double-stranded RNA (dsRNA), which initiate the process of decomposition of specific mRNA molecules in their cells. As a consequence, the body cannot produce crucial proteins, and this results to pest weakening, which frequently leads to death. RNA acts with high precision—it attacks only selected genes and is not harmful to other organisms. Out of the three natural pathways of these mechanisms in arthropods, the siRNA path is most commonly used in practice. Firstly, RNAi-based formulations are already registered in the U.S., and their usage is a part of the strategy for sustainable plant protection [201,202].

RNAi was effectively applied in different arthropod species, including apple tree pests C. pomonella, Halyomorpha halys, and Tetranychus urticae [203,204,205,206].

One of the methods for dsRNA application onto a tree is its injection into the trunk, and this enables the vascular system of the tree to transport dsRNA to leaves and other tree parts, where it can be taken in by pests. The studies showed that dsRNA can persist in apple tree crowns for longer, for at least 141 days [207].

In 2011, the Arctic^® apple variety was successfully grown using RNAi technology, by silencing the polyphenol oxidase gene, which is the enzyme responsible for the browning of apples. As a result, apples stayed white after cutting and were not discoloured after bruising. Granny Smith, Golden Delicious, and Arctic Fuji varieties of the Arctic apple are currently being grown in commercial orchards. As of 2020, there were 1350 acres (550 ha) of Arctic apple orchards in Washington State, with 7.7 million kg of fruit harvested in 2021 [208,209].

5.1.2. RNAi Modifying Apple Tree Characteristics

The RNAi technology was used to silence genes associated with apple tree dormancy. This resulted in delayed ageing of leaves and their falling in autumn, no dormancy of buds in winter, and continuous growth of new leaves regardless of the season for over 3 years. The premature flowering was observed, but morphology of leaves, fertility, and fruit development were normal [210].

5.1.3. The Importance of RNAi in Sustainable Agriculture

The main advantage of RNAi is the specificity of this technique. RNA can influence specific genes, ensuring greater precision when compared to broad-spectrum pesticides. Another advantage of RNAi is the potential reduction in the use of synthetic pesticides [199,211].

According to Germing et al. [212], 275 patent applications concerning crop protection-based RNAi were filed in 2003–2023, and this reflects significant interest in this subject. The top jurisdictions for patent filing are the United States, followed by Brazil, Argentina, India, and China, reflecting the current focus on the Americas and Asia. The number of scientific papers on RNAi-based crop protection increased steadily in 2003–2016. In 2016–2019, that number remained stable. In the successive period, that number increased again, exceeding 30 publications in 2021.

However, RNAi technology poses some challenges, including concerns about its environmental impact. The long-term environmental impact of RNAi technology in agriculture requires further studies. From the regulatory perspective, RNAi technologies need to be thoroughly tested and undergo registration processes to assess their safety, environmental impact, and efficacy, for which different frameworks are established in different countries and regions. In the EU, these products are classified as a new class of active substance and undergo a two-stage approval process conducted by EFSA and individual member states. Although no specific guideline documents are available yet for RNAi-based products, adjustments for requirements concerning data are being developed. Another challenge is their social acceptance, due to concerns about genetic engineering and potential unforeseen ecological consequences. Ultimately, the management of these challenges and regulatory risk is of fundamental significance for the safe and effective implementation of RNAi in agriculture [211,212,213,214].

5.2. CRISPR and Gene Editing

The CRISPR technology offers an effective and precise solution, enabling the increase in plant protection against hazards such as diseases, pests, and environmental stresses. By enabling targeted modifications of plant genes, CRISPR can improve resistance to diseases, improve resistance to abiotic stress, and increase yield and quality of crops [215].

This technology was used to improve crops such as wheat, rice, maize, potato, barley, tomato, soybean, grapevine, rapeseed, watermelon, citrus, banana, cucumber, and cassava [216,217,218,219,220,221,222,223,224,225,226,227,228]. Following mutations introduced into the plant genome, plants gained resistance to various pests, including those commonly found in apple orchards.

A number of reports were published concerning the modification of apples to achieve desirable characteristics, such as improved biosynthesis of carotenoids [229], increased anthocyanine content [230], early flowering [231], and resistance to fire blight [232], or Botryosphaeria dothidea [233].

However, no regulatory framework is available for gene editing based on the CRISPR technique focusing on issues of safety to humans and the environment, with an innovative potential for next-generation cultivation based on genome editing to support sustainable agriculture.

5.3. Nanotechnology

Nanotechnology offers new mechanisms for delivery and new agrochemical agents that improve crop yield while reducing the use of synthetic pesticides.

5.3.1. Encapsulation for Harvested Fruit Protection in Storage

Encapsulation, especially nanoencapsulation, increases stability and enables the controlled, long-term release of active substances. This technique involves enclosing a substance (the core) within a nanoscale material (the shell), offering protection from degradation and enabling its controlled delivery [234].

Literature reports are available concerning the use of nanoencapsulation; for example, essential oils can be used as edible biopolymer-based coatings that act as agents prolonging crop shelf life.

The nanoencapsulation of essential oils is a safe, natural alternative to synthetic substances, which was used in numerous fruit and vegetable crops. Essential oils have antibacterial and antioxidative effects, maintain sensory properties, prevent loss of crispiness, and lead to a change in colours and taste in storage. In the apple crops, tarbush extract from Flourensia cernua [235] and lemongrass essential oil [236] were used.

5.3.2. Nanopesticides—Pest Control

A new technology, named nanobiotechnology, was introduced to the sector of biopesticides. In this context, the development of non-toxic and environmentally friendly nanopesticides is of crucial importance. Metallic particles with relatively better anti-pathogenic, antifungal, and antibacterial effects are used in these nanopesticides, and this represents the main application of this technology in the protection of plants against pests [237,238].

5.3.3. Nanotechnology Application for Mitigating Abiotic Stresses in Plants

Plant development and productivity are negatively impacted by abiotic stress [239,240,241]. For plants, two most important stressors are salt and drought [242]. The use of nanotechnology gained great publicity due to promising results in this sector. Nanoparticles are used to control the activity of antioxidative enzymes, like catalase (CAT), super oxide dismutase (SOD), and peroxidase, which are very effective in overcoming effects of drought in plants. Applications of nano-silicon dioxide particles on plants resulted in an increase in their chlorophyll contents, fresh and dry leaf weights, accumulation of proline, and regulation of antioxidant enzymes under saline conditions [243].

5.3.4. Nanotechnology in Sustainable Agriculture

The issues of biological safety play a crucial role in the responsible use of synthetic and yet organic nanoparticles in agriculture and the food industry. The EU introduced regulations for food additives using nanotechnology [244], and the U.S. Food and Drug Agency (FDA) [245] monitors both food packaging and food products containing nanoparticles. However, many countries do not have necessary legal regulations in this respect. Therefore, comprehensive regulatory frameworks are required for legal and safe development of nanotechnology, with detailed regulations and advanced methods for toxicological assessment [246]. A detailed risk assessment, responsible production of nanoparticles, thoroughly planned distribution, regular monitoring, and effective inspection procedures should form an integral part of these regulations. When these guidelines are followed, nanoparticles can be safely and sustainably used in agriculture, respecting both the environment and human health.

5.4. Drones and Precision Spraying Systems for Targeted Delivery

Drones and precision agriculture enable the implementation of effective plant protection systems to prevent and control plant diseases and pests, and at the same increase yield and support sustainable development.

The research presents a novel approach to enhancing the cultivation of orchard crops by combining deep-stream algorithms with drone technology. A thermal camera monitors the condition of plants, the estimated yield, fertilisation management, and the mapping of irrigation. This camera supports early diagnosis of infected plants by recording fluctuations in the temperature, enabling the fast performance of procedures and optimising the allocation of resources. The optical RGB camera supports this functionality, measuring vegetation indices, evaluating quality of fruit, or even detecting weeds [247,248,249]. These cameras are installed on unmanned aerial vehicles (UAVs), helping farmers to monitor their crops in a more punctual way [250]. There are several literature reports on the use of these cameras in the cultivation of apple trees. Apolo–Apolo et al. [251] used UAV in apple orchards, using thermal and RGB cameras to detect damages caused by low temperature, evaluate fruit setting, predict yield, and monitor flowering stages. Similarly, He et al. [252] and Jemaa et al. [253] used the same systems to evaluate yield and monitor the health of apple trees, while Chandel et al. [254] used cameras to estimate the irrigation schedule for apple orchards. All these analyses were conducted with high precision of 89–92%.

Precision spraying enables precise delivery of plant protection products to specific areas, minimising quantities of chemical waste, reducing the environmental impact, and maximising the effectiveness of these treatments. Introduction of precision spraying is not only beneficial for the environment, but also economically advantageous for the farmers. Precision spraying promotes sustainable development of the environment [255]. It uses advanced sensor technologies, variable rate application (VRA) systems, automated spraying technologies, and site-specific spraying, which drive the revolution in crop protection [256,257,258].

6. Perspectives and Recommendations

Biopesticides and biological pest control methods represent an effective alternative to chemical plant protection products in apple orchards. The main advantages of using biopesticides are undoubtedly environmental and consumer protection, as well as the preservation and enhancement of biodiversity. The main disadvantages result from the limited availability of biological agents, higher production and application costs, and precise operation, requiring detailed knowledge and correct application.

Chemical control provides rapid, effective, and broad-spectrum pest control but can harm non-target organisms, develop pesticide resistance, and leave harmful residues. Biological control is slower but more targeted, offering long-term pest control with minimal environmental damage, lower toxicity, and increased food safety by avoiding hazardous residues. The most effective approach often depends on the specific pest and infestation severity, and biological control holds great promise in IPM programmes due to its sustainability [259].

Unfortunately, the number of microbiological agents for the protection of apple trees approved for use and available in the market is low. No microbiological agents are available for controlling the most important diseases of apple trees, such as apple scab (only natural product laminarin) or powdery mildew (only orange oil). Therefore, new microorganisms that are effective against pathogens, such as bacteria, fungi, and yeasts, need to be investigated. Furthermore, technological innovations in biocontrol application are also available, which certainly will be used increasingly often in the future.

Progress in microbiology, chemistry, and nanotechnology enables the development of pesticides of the new generation that combine high effectiveness with minimal environmental impact, e.g., smart formulations or nanocapsules reacting to environmental factors and activating in response to environmental conditions.

Clearly, the future of agriculture is associated with automation and digital decision making, e.g., precise application of pesticides only onto infected trees, as well as the use of artificial intelligence models analysing weather data as an early warning of potential infectors.

The transformation of the plant protection system towards the biological one requires not only technologies, but also appropriate legal frameworks. The path for biopesticide registration needs to be shortened and simplified. Subventions for orchard owners for the use of biopesticides or helpful organisms will also be helpful, as well as counselling programmes and training in biological protection.

The implementation of biological strategies for orchard protection is a part of the objectives for sustainable agriculture, ensuring a balance between production efficiency, environmental protection, and consumers’ health.