1. Introduction
Kiwifruit, a healthy, emerging, and typical third-generation fruit, has very high nutritious, medicinal and economical values [
1,
2]. The kiwifruit industry, a major industry for boosting agricultural modernization, alleviating poverty and vitalizing rural areas, has expanded rapidly in Guizhou Province, China, where it has more than 40,000 hm
2 of planting area [
2,
3]. Nonetheless, the soft rot disease of kiwifruit caused by
Botryosphaeria dothidea,
Phomopsis spp.,
Botrytis cinerea and
Cryptosporiopsis actinidiae, etc., is the most serious postharvest disease [
4,
5,
6,
7,
8,
9,
10,
11]. Koh et al. [
6] reported that
B. dothidea caused soft rot in Korea with 83.3% of average isolation rate. Our previous study also found that
B. dothidea and
Phomopsis spp. were the most frequent pathogens of kiwifruit in Guizhou with 89.93% and 10.07% of average isolation rates, respectively [
9]. Soft rot disease has had an annual incidence of about 30~50% in Guizhou Province since 2014, and seriously influences the yield and quality of kiwifruit, as well as frequently causes severe financial losses [
9,
10,
11]. Accordingly, it is of realistic significance to excogitate various candidate measures against kiwifruit soft rot for the sustainable development of its industry.
Currently, chemical control are still the most typical, frequent and effective strategies for managing plant fungal disease. For example, Koh et al. [
6] reported that some chemical synthetic fungicides including benomyl, thiophanate-methyl, tebuconazole, iprodione and flusilazole could be used as preventive fungicides against the postharvest rot diseases of kiwifruit. Considering the residual risks and pathogen resistance of chemical synthetic fungicides, however, reducing the application of chemical synthetic fungicides and their alternative or finding complementary approaches has been growing popular for plant disease management [
12,
13,
14]. We previously reported that some natural products including tetramycin, chitosan, ferulic acid, matrine and their combinations could effectively control kiwifruit soft rot and enhance their resistance, growth and quality [
9,
10,
11,
15,
16,
17]. Nevertheless, natural products are often slower to take effects compared to chemical fungicides for controlling plant diseases. Meanwhile, considering the perniciousness of kiwifruit’s soft rot, new candidate agronomic strategies need to be developed to help meet this challenge. Additionally, we also found that the co-application of chitosan and isopyrazam azoxystrobin effectively managed leaf spot disease in kiwifruit and reduced isopyrazam azoxystrobin application [
12]. Similarly, whether natural products can be used together with chemical fungicides to effectively reduce the application of chemical fungicides and control kiwifruit’s soft rot is extremely worth further exploring.
Tebuconazole, a broad-spectrum chiral triazole fungicide, is one of the most widespread sold pesticides in the world, and widely used for controlling many plant fungi diseases [
18,
19,
20]. Its action mechanism has been demonstrated to inhibit the sterol biosynthesis of plant pathogenic fungi [
21]. Ma et al. [
22] reported that the 50% effective concentration’s inhibition rate (EC
50 value) for tebuconazole on 105
B. dothidea isolates from a commercial pistachio orchard in California was 0.019~0.159 μg mL
−1. Oligosaccharins, a part of the cell walls of plants and fungi, is capable of regulating plant growth and acting as regulators of the plant response when infected by plant pathogens [
23,
24]. When plant pathogens attack plants, oligosaccharins can activate the generation of oligosaccharins by means of the enzymatic lysis of the cell wall of the attacked plant [
24,
25]. Recently, oligosaccharins has received considerable attention due to its ability to enhance antipathogenic compound synthesis and trigger defense responses in plants [
23,
24,
26,
27,
28]. For instance, Boyzo-Marín et al. [
27] reported that glutathione-oligosaccharins application led to the lowest incidence (0.66~13%) of downy mildew on blackberry in Mexico compared to that of other fungicides or control. The foliar application of oligosaccharins could effectively promote the stem elongation, biomass production and yield formation of tomatoes [
29,
30]. However, to date, there is little documentation available about oligosaccharins for controlling kiwifruit soft rot disease. In that case, it is worth further exploring whether oligosaccharins can promote tebuconazole to control kiwifruit soft rot and decrease tebuconazole application.
In the present work, the toxicities of ten synthetic fungicides and oligosaccharins on B. dothidea were firstly determined. Then, the field control effects of tebuconazole + oligosaccharins, tebuconazole and oligosaccharins to manage soft rot disease in kiwifruit were investigated. Subsequently, the influences of tebuconazole + oligosaccharins, tebuconazole and oligosaccharins on the resistance, yield and quality of kiwifruit were evaluated. This finding excogitates a green, practicable and candidate agronomic measure for managing kiwifruit soft rot and decreasing chemical synthetic fungicide application.
4. Discussion
Tebuconazole is one of the chiral triazole fungicides, and it has a broad-spectrum systemic activity on many plants’ pathogenic fungi; it also has the three functions of protection, treatment and eradication [
18,
19,
20]. Ma et al. [
22] reported that the EC
50 value of tebuconazole against 105
B. dothidea isolates from a pistachio orchard was 0.019~0.159 μg mL
−1. The results here showed that tebuconazole displayed a toxicity against
B. dothidea RF-1 soft rot pathogen with 0.87 mg kg
−1 of EC
50 value, which was higher by 1.23, 1.37, 1.39, 3.03, 5.29, 6.39, 10.79, 22.11, 25.05 and 255.56 folds compared to flutriafol, epoxiconazole, trifloxystrobin tebuconazole, difenoconazole, fluazinam, oxine-copper, eugenol, azoxystrobin, pyrimethanil and oligosaccharins, respectively. This result is consistent with previous research. However, oligosaccharins had a relatively inferior toxicity on
B. dothidea RF-1 with an EC
50 value of 222.34 mg kg
−1.
Oligosaccharins can be used as a biostimulant for triggering the plant’s defense responses and enhancing the plant’s antipathogenic compound synthesis [
23,
24,
26,
27,
28]. Boyzo-Marín et al. [
27] found that glutathione-oligosaccharins application had the lowest incidence (0.66~13%) of downy mildew in blackberry compared to that of other fungicides. In this study, tebuconazole + oligosaccharins, tebuconazole and oligosaccharins could significantly (
p < 0.01) decrease fruit incidence rate, the field control effect of tebuconazole + oligosaccharins 5000-fold liquid on soft rot was 84.83%, which significantly (
p < 0.01) exceeded the 72.05% of the tebuconazole 2500-fold liquid, 52.59% of the oligosaccharins 2500-fold liquid, 62.17% of the tebuconazole 5000-fold liquid, and 33.52% of the oligosaccharins 5000-fold liquid. Oligosaccharins exhibited an inducing control effect on soft rot disease, which, mixed together with tebuconazole, could more effectively control soft rot disease compared to tebuconazole or oligosaccharins alone, as well as effectively reduce tebuconazole application. The excellent controlling effect of tebuconazole + oligosaccharins on soft rot was possibly originated from the therapeutic, preventive and eradication characteristics of tebuconazole, and the inducing effect of oligosaccharins.
Phenolics and flavonoids themselves can not only be used as antagonists of pathogens, but also as precursors of lignin biosynthesis, resulting in host cell lignification and further producing disease resistance [
31]. Soluble protein is the basis of material and energy metabolism, while MDA is an index to reflect membrane lipid peroxidation [
11,
31]. SOD is a key protective enzyme for scavenging free radicals in plants, and POD plays a role in catalyzing H
2O
2 decomposition in the final step of lignin biosynthesis [
31,
32]. PPO can catalyze the formation of lignin and other phenolic oxidation products to form a protective barrier against the invasion of pathogens, and can also play a direct role in disease resistance by forming quinones [
31]. PAL is one of the enzymes for controlling the biosynthesis of phenolics and flavonoids [
11,
31]. In fact, oligosaccharins originates from the hydrolysis of chitosan [
32]; while chitosan can induce protein increase, MDA reduces and triggers the defense enzyme activity [
33,
34,
35,
36,
37]. In our previous studies, we reported that chitosan, tetramycin + chitosan and chitosan + ferulic acid could effectively improve the phenolics, flavonoids and soluble protein of kiwifruit, inhibit its MDA and enhance its SOD, POD, PPO and PAL activities [
9,
10,
11,
16]. The present results show that the co-application of oligosaccharins and tebuconazole notably enhanced the phenolics, flavonoids and soluble protein content of kiwifruit, reduced their MDA content and reliably promoted their SOD, POD, PPO and PAL activities. These results also emphasize that oligosaccharins had a notable potential for enhancing kiwifruit’s disease resistance, and when used together with tebuconazole notably enhanced the improving effects of oligosaccharins or tebuconazole on the disease-resistant substances and resistant enzyme activities of kiwifruit, thereby providing more benefit for enhancing the disease resistance of fruits.
A good growth determines the fruit yield, quality and commodity of kiwifruit. Costales et al. [
29] demonstrated that foliar application of oligosaccharins effectively promoted the biomass production and yield formation of tomatoes. Hernández et al. [
24] indicated that oligosaccharins stimulated the photosynthesis of tomatoes and improved their fruit production. In grapes, oligosaccharides resulted in an increase of its color intensity and anthocyanin [
38]. He et al. [
39] demonstrated that chitosan oligosaccharides application improved the quality of strawberry fruits. Although not specifically oligosaccharins, we also found that its parent chitosan could effectively improve kiwifruit’s yield and quality [
33,
34,
35,
36,
37]. In this study, tebuconazole + oligosaccharins notably (
p < 0.05) enhanced the growth and quality of kiwifruit. Meanwhile, this improving effects of tebuconazole + oligosaccharins were more obvious than those of oligosaccharins or tebuconazole alone. This probably derived from their labor division: tebuconazole protected kiwifruit from a pathogen infection, oligosaccharins induced its disease resistance and enhanced its growth, yield and quality.
Recently, a growing attention has been focused on reducing the application of chemical fungicides and natural products as the adjuvants or alternative approaches of chemical fungicides for plant disease management [
12,
40]. In China, the recommended concentration of tebuconazole on banana, apple, pear and other fruit trees is a 1250~7500-fold dilution liquid. To date, however, there are no registration and application of tebuconazole for controlling kiwifruit diseases in China. In the present study, oligosaccharins used together with tebuconazole more effectively controlled soft rot in kiwifruit and promoted its resistance, growth and quality compared with tebuconazole or oligosaccharins alone. Meanwhile, the tebuconazole + oligosaccharins 5000-fold dilution liquid was equivalent to tebuconazole 10,000 + oligosaccharins 10,000-fold dilution liquid, hence oligosaccharins used together with tebuconazole effectively decreased tebuconazole application compared with the tested concentration (2500- or 5000-fold) or the recommended concentration (1250~7500-fold) of tebuconazole. The field concentration of the tebuconazole 10,000-fold dilution liquid was relatively low. Moreover, oligosaccharins is a nontoxic natural product widely used in food, medicine, agriculture, cosmetics fields and so on [
23,
24]. Furthermore, the total time of safe interval and soft ripening periods of kiwifruit was more than 80 days. Accordingly, the potential safety risks of tebuconazole + oligosaccharins are very small and almost nonexistent. This study highlights that a 80% tebuconazole WG + 5% oligosaccharins AS 5000-fold dilution liquid can be proposed as an effective candidate practice for managing kiwifruit soft rot and reducing the application of chemical fungicides.