Biological control of Alternaria solani by tomato epiphyte yeasts and investigation of some their biocontrol mechanisms

Document Type : Complete paper

Authors

1 Department of Plant Protection, Faculty of Agronomy, Sari Agricultural Sciences and Natural Resources University, Sari, Iran.

2 Department of Plant Protection, Faculty of Agronomy, Sari Agricultural Sciences and Natural Resources University, Sari, Iran

3 Plant Protection Research Department ,West Azarbaijan Agricultural and Natural Resources Research Center, AREEO, Urmia, Iran

4 Forest and rangelands Department,West Azarbaijan Agricultural and Natural Resources Research Center, AREEO, Urmia, Iran

Abstract

Early blight disease (is caused by Alternaria solani Sor) is a limiting factor in tomato production and various studies showed that yeasts are potential for biocontrol of these diseases. The present study aims is to demonstrate the antagonistic effect of the tomato epiphytic yeasts on the mycelial development of A. solani and study their mode of action. In vitro, seventeen epiphytic yeast strains of tomato were evaluated against A. solani. Results showed that 15 isolate are able to control of A. solani. In the postharvest experiments, the abilities of yeast strains to reduce the severity of A. solani on tomato fruit rot were conducted. In vitro two isolates of Aureobasidium pullulans (N3 and N4) reduced the fruit rot over than 85%. Results of the study on biocontrol mechanisms showed that all tested isolates except isolates N14 and N15 were able to significantly inhibit the growth of A. solani through the formation of volatile compounds (VOCs). Study on the ability of isolates to produce enzymes, sidrophore and formation of biofilm showed that five isolates were able to produce pectinase, four isolates were able to produce amylase, five isolates produced protease, five isolates were able to produce siderophore and seven isolates were able to form biofilm. In conclusion, our results confirm the potential of A. pullulans (N3, N4) as biocontrol agents against A. solani and suggested it as a suitable option for controlling postharvest diseases of tomatoes.

Keywords

Main Subjects

Extended Abstract

(Solanum lycopersicum L.) is the world’s second most important vegetable crop, with a total production of 182 million tons in 2018. The main postharvest fungal diseases of tomato fruit caused by various pathogenic fungi include Alternaria alternate, Alternaria solani, Botrytis cinerea and Rhizopus stolonifer. A. solani is a saprophytic pathogen of tomato causing postharvest black rot at high frequency. Different strategies as fungicides, resistant hybrids, and cultural practice are generally used as commercial strategies to prevent postharvest diseases. However, the use of fungicides intensively may help pathogen to be resistant as well as fungicides released to the environment. However, it may represent environmental and health hazards. Therefore, its application is becoming more limited to the consumer’s anxieties and the human health importance  In organic farming, in which chemical pesticides are prohibited, other additional control methods, such as biological methods, need to be investigated. Microbial biocontrol agents (BCA) are sustainable tools in crop protection against damage caused by plant pathogens. In integrated pest management, the use of effective biocontrol agents is considered first before chemical pesticides are applied. In recent years, some antagonists have been successfully applied in the biocontrol of postharvest diseases of tomato fruit. Aureobasidium pullulans, a cosmopolitan yeast-like fungus, is found to be an effective biological agent for controlling postharvest rots of cherry tomatoes. The assays indicated production of volatile organic compounds (VOCs), competition for nutrients, biofilm formation, production of sidrophore and Enzymes as mechanisms of action of yeast isolates. Collectively, our results showed the potential of yeast isolates that were isolated from tomato phyllosphere s of tomato to control the mycelial development of A. solani and some mechanisms of action of these yeasts. The aim of this study was to investigate the efficacy of tomato epiphytic yeast strains in reducing the postharvest tomato fruit disease caused by A. solani and determine the mode of action of these strains.

Among 17 yeast strains tested against A. solani using in vitro dual culture assays. Based on the results, 15 yeast isolates showed inhibitory activity against A. solani at dual culture assays. The growth of A. solani mycelium was significantly inhibited by yeast strains compared to the control (p = 0.05). On the 14 day of the assay, the N4 and N3 strains inhibited the mycelium growth by 70.76% and 70.33, respectively. In the present investigation, post-harvest treatment of tomato fruits with the native antagonists isolated from tomato significantly reduced the severity of Alternaria fruit rot. The differences between isolates in isolates efficacies were statistically significant (P < 0.05). Among the antagonistic microorganisms tested, A. pullulans was the most effective for controlling Alternaria fruit rot of tomato fruits. Postharvest treatment of tomato fruits with isolates N4 and N3 reduced the fruit rot lesion size by 88.23% and 86.83%, respectively, compared to untreated control.

Production of diffusible and volatile substances was conducted on agar plates. Production of diffusible substances was evaluated using the dual culture method. For evaluation of volatile antifungal metabolites by yeast isolates, the two plates were placed “mouth to mouth”, wrapped to gather with parafilm. In both assays plates were incubated for 14 days at 24 °C. After incubation, colony diameter was used to calculate inhibition. All experiments were performed in triplicate. For postharvest assays, artificial wounds (5 mm deep and 4 mm wide) were performed using a sterile tool on superficially sanitized pears (3 wounds per fruit). Yeast suspension (20 μL, 1 × 108 cells/mL) of respective yeast was individually inoculated into the wounds. Inoculated fruits were placed on tray packs in boxes and incubated for 21 days at 4 °C and seven days at 24 °C.

 In order to characterize the capability of the selected yeasts to produce and secrete fungal cell wall lytic enzymes (esterase, protease, amylase, and pectinase), qualitative tests were performed on solid media. Production of lytic exoenzymes by yeast strains was determined. Results showed that six different isolates could produce protease, amylase and pectinase. Amylase production was not observed in any of the strains. Also seven isolate were able to formation biofilm and five isolates were able to production of sidrophore.

In summary, our data show that the yeast strain A. pullulans (isolates N3, N4) has potent antifungal activity as demonstrated by in vitro inhibitory activity against A. solani. Further, postharvest treatment of tomato fruits with A. pullulans significantly reduced the Alternaria fruit rot. The present investigation was carried out under high inoculum conditions. Under normal postharvest conditions, treatment of tomatoes with A. pullulans would be expected to give better protection against fruit rot because of usually low inoculum levels. Furthermore, we demonstrated the production of VOCs, protease and Amylase enzyms, Sidrophore and biofilm formation by A. pullulans. The antagonistic activity of A. pullulans might be due to the synergistic effects of diffusible and volatile antimicrobial compounds.

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BIOLOGICAL CONTROL OF PESTS AND PLANT DISEASES
Volume 12, Issue 1
April 2024
Pages 95-110

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