Reduction of postharvest diseases of loquat fruit by serine protease and possible mechanisms involved
Introduction
Loquat (Eriobotrya japonica) is a subtropical evergreen fruit tree originated in China, and its fruit is one of the fruit with Chinese characteristics (Liu et al., 2016). Zhejiang, Suzhou and Fujian are famous for being the three major production areas in China (Lin, 2010). Loquat pulp is rich in potassium, calcium, iron, phosphorus, vitamins and dietary fiber, etc., which has high nutritional and medicinal value (Hamada et al., 2004). However, loquat is vulnerable to mechanical damage, physiological changes, and microbial infection, which will cause fruit decay and deterioration, and affect the postharvest quality of loquat (Cao et al., 2014b). The anthracnose rot caused by Colletotrichum acutatum is the main disease of loquat after harvest (Cao et al., 2008). The rot of loquat fruit is one of the main reasons affecting its edible quality (Cao et al., 2014b). Therefore, taking safe and effective measures to control postharvest diseases of loquat fruit is of great significance to improve fruit quality, prolong fruit storage period and promote economic development of loquat fruit industry.
At present, physical and chemical methods are mainly used to control the decline of postharvest fruit quality. Physical methods mainly include low temperature storage (Pace & Cefola, 2021), controlled atmosphere storage (Caleb et al., 2013), heat treatment (Liu et al., 2010) and UV-C irradiation (Smilanick, 2004), while chemical methods mainly include chemical fungicides and other chemicals. 1-methylcyclopropene (1-MCP) (Liguori et al., 2014), methyl jasmonate (Cao et al., 2014a) , volatile essential oils of myrtle leaves (Bahadirli et al., 2020) , 0.5% Nigella sativaoil and 0.5% propolis extract (Kahramanoglu, 2020) can effectively control the decline of fruit quality after harvest. Chemical fungicides such as benzimidazole fungicides (MBCs), methoxyacrylate fungicides (QoIs), and dimethyl imide fungicides (DCFs) can also be used for controlling postharvest diseases (Ishii et al., 2016). However, widely used chemical reagents will enhance the drug resistance of pathogenic fungi, and pollute the environment and endanger human health (Cao et al., 2014b).
Therefore, more researchers have turned their attention to safe and environmentally friendly biological control methods. Debaryomyces hansenii KI2a, Wickerhamomyces anomalus BS91 (Grzegorczyk et al., 2017) and Yarrowia lipolytica (Zhu et al., 2019) have been proved to reduce the incidence and severity of postharvest diseases of fruits. Bacillus amyloliquefaciens SF14 and SP10 (Lahlali et al., 2020) and Bacillus subtilis SL-44 (Wu et al., 2019) have also been proved to inhibit the deterioration of harvested fruit. The use of microorganisms will always bring about safety problems, and the secondary metabolites produced by microorganisms have attracted researchers' attention because of their high safety and environmental protection. Exopolysaccharide from Pythium arrhenomanes (Yuan et al., 2020), anti-fungal protein PgAFP from Penicillium chrysogenum CECT 20922 (Delgado et al., 2019), and volatile organic compounds from Enterobacter asburiae Vt-7 (Gong et al.,2019) have all been proved to inhibit the growth of postharvest pathogenic fungi.
The serine protease was isolated and purified from the fermentation broth of Bacillus amyloliquefaciens in our laboratory. It was proved that it could inhibit postharvest diseases of loquat fruit (Yan et al, 2021). However, the mechanisms of the serine protease in relieving postharvest diseases of loquat are still unclear. Therefore, in this paper, the antifungal mechanisms of serine protease in vitro, the effects of the serine protease on ROS metabolism, the activity of resistance-related enzymes, the quality of resistance-related substances and the expression level of resistance-related genes in loquat fruit were studied, and the possible mechanisms of serine protease in alleviating postharvest diseases of loquat fruit was discussed.
Section snippets
Preparation of serine protease
Bacillus amyloliquefaciens MG-3 was supplied by the Food Storage and Preservation Laboratory of Fujian Agriculture and Forestry University. MG-3 was cultured according to the methods described by Yan et al. (2020) After centrifugation at 10000 × g for 20 min, the supernatant was filtered with a 0.22 μm filter and stored at 4 °C. After precipitation with 40-60 % ammonium sulfate, dialysis and filtration with 0.22 μm filter, the protein was separated and purified by DEAE-650C and Sephacryl S-200,
Analyses of SP against C. acutatum in vitro
As shown in Fig. 1A, the antifungal rate showed positively correlated with the concentration of SP, the coefficient of determination R2 was 0.99762 (y = 1.24567x - 0.48794). According to the equation, it could be calculated that the IC50 of SP was 40.58 mg•L−1 and the MIC was 80.6 mg•L−1. After 0.5 mL of C. gloeosporioides spores with a concentration of 106 spores•mL−1 and 0.5 mL of PDB medium containing SP were co-cultured for 11 d, there was no hypha growth in both liquid medium and solid
Discussion
C .acutatum infection can cause anthracnose, which leads to postharvest decay and poor quality of loquat, and affects the shelf life and commodity value of loquat. It has been studied that low-dose UV-C treatment (Liguori et al., 2014), calcium treatment (Cao et al., 2014a), Bacillus suspension treatment (Lahlali et al., 2020; Wu et al., 2019) and other methods have been used to inhibit the infection of postharvest pathogens of loquat. In recent years, there is more and more evidence that
Conclusion
In conclusion, SP could control anthracnose caused by C. acutatum in loquat fruit, and its possible mode was to inhibit the germination and growth of C. acutatum, improve the activities of ROS scavenging enzymes and the contents of ROS non-enzyme scavenging materials, and maintain the stability of ROS metabolism. Besides, its possible mechanisms included increasing the contents of total phenolics and flavonoid and enhancing the activities of resistant enzymes; inducing the expression of genes
Credit authorship contribution statement
FenYan: Supervision, Resources, Conceptualization, Project administration, Writing - review & editing.
Dan Zhang: Investigation, Data curation, Formal analysis, Writing - original draft & review.
Xue Wang: Investigation.
Cong Liu: Writing - review & editing.
Fan Zhang: Investigation.
Declaration of Competing Interest
The authors declare that they have no conflict of interest.
Acknowledgements
This research was financially supported by the Program of Finance Department of Fujian Province in China (Grant Nos.2015-1297, 2018-983, 2020-822), the Science and Technology Planning Project of Fujian Province in China (Grant Nos.2020N0001).
References (57)
- et al.
Biocontrol activity of an alkaline serine protease from Aureobasidium pullulans expressed in Pichia pastoris against four postharvest pathogens on apple
Int. J. Food Microbiol.
(2014) - et al.
Effect of MeJA treatment on polyamine, energy status and anthracnose rot of loquat fruit
Food Chem.
(2014) - et al.
Antioxidant enzymes and fatty acid composition as related to disease resistance in postharvest loquat fruit
Food Chem.
(2014) - et al.
Energy regulated enzyme and non-enzyme-based antioxidant properties of harvested organic mung bean sprouts (Vigna radiata)
LWT-Food Sci. Technol.
(2019) - et al.
Evaluation of the activity of the antifungal PgAFP protein and its producer mould against Penicillium spp. postharvest pathogens of citrus and pome fruits
Food Microbiol.
(2019) - et al.
Postharvest oxalic acid treatment induces resistance against pink rot by priming in muskmelon ( Cucumis melo L.) fruit
Postharvest Biol. Technol.
(2015) - et al.
Postharvest ASM dipping and DPI pre-treatment regulated reactive oxygen species metabolism in muskmelon (Cucumis melo L.) fruit
Postharvest Biol. Technol.
(2015) - et al.
Postharvest biocontrol ability of killer yeasts against Monilinia fructigena and Monilinia fructicola on stone fruit
Food Microbiol.
(2017) - et al.
Benzo-(1, 2, 3)-thiadiazole-7-carbothioic acid s-methyl ester (BTH) promotes tuber wound healing of potato by elevation of phenylpropanoid metabolism
Postharvest Biol. Technol.
(2019) - et al.
Biocontrol activity and putative mechanism of Bacillus amyloliquefaciens (SF14 and SP10), Alcaligenes faecalis ACBC1, and Pantoea agglomerans ACBP1 against brown rot disease of fruit
Microb. Pathog.
(2020)