African Journal of
Microbiology Research

  • Abbreviation: Afr. J. Microbiol. Res.
  • Language: English
  • ISSN: 1996-0808
  • DOI: 10.5897/AJMR
  • Start Year: 2007
  • Published Articles: 5182

Full Length Research Paper

Effects of antifungal activity of essential oils, salts and antioxidants acids on pathogenic fungi and their application methods for controlling postharvest diseases in banana fruits

Zoeir H. A.
  • Zoeir H. A.
  • Botany Department, Faculty of Agriculture, Tanta University, Tanta, Egypt.
  • Google Scholar
El Zahaby H. M.
  • El Zahaby H. M.
  • Botany Department, Faculty of Agriculture, Tanta University, Tanta, Egypt.
  • Google Scholar
Ziedan E. H.
  • Ziedan E. H.
  • Plant Pathology Department, National Research Centre, Dokki, Cairo, Egypt.
  • Google Scholar
Maswada H. F.
  • Maswada H. F.
  • Botany Department, Faculty of Agriculture, Tanta University, Tanta, Egypt.
  • Google Scholar


  •  Received: 29 May 2017
  •  Accepted: 28 June 2017
  •  Published: 07 August 2017

 ABSTRACT

In vitro clove essential oil (1.0%), sodium carbonate (2.0%) and sorbic, salycilic and propinic acids (0.5%) completely (100%) inhibited mycelial linear growth of Colletotrichum musae and Fusarium moniliforme, which is the cause of postharvest diseases in banana fruits. In vivo different application methods, that is, spray, soaking and dusting were tested on crown rot, neck rot, finger rot and flower end rot under artificial infestation with causal pathogens. Soaking method significantly reduced major postharvest diseases incidence in banana fruits than spray and dusting treatments. Soaking banana fruits in clove suspension (2%) and sodium carbonate (4.0%) was the best treatment that completely inhibited (100%) crown rot and flower end rot diseases and significantly reduced finger rot and neck rot diseases. So, clove oil and sodium carbonate were the most promising agents for controlling major postharvest diseases of banana fruits as eco-friendly and alternative synthetic fungicides.

 

Key words: Banana, antioxidants, fungi, diseases, essential oil, salts.


 INTRODUCTION

Banana (Musa species) is an economic and important fruit crop grown worldwide in more than 120 countries throughout tropical and subtropical regions. It is a popular worldwide staple food for more than 400 million people (Zhang et al., 2005). Postharvest diseases destroy 10 to 30% of the total yield of crops during handling, transportation, storage and marketing (Agrios, 2005). Colletotrichum  musae  and  Fusarium  species,   that   is, Fusarium solani, Fusarium semitectum, Fusarium moniliforme and Fusarium musae are the major fungi causing postharvest complex diseases (fruit rot, crown rot, finger rot, cigar end rot) of banana fruits (Bhattacharyya and Chakraborty, 2007; Diedhiou et al., 2014; Khleekorn et al., 2015; Marin et al., 1996; Abd-Alla et al., 2014; Abdullah et al., 2016; Triesta et al., 2016). Application of cultural, physical and biological methods is an alternative to synthetic fungicides for controlling postharvest diseases of banana fruits (Dionisio and Natsuaki, 2007; Lassois et al., 2008). Recently, essential oils, antioxidants, organic and inorganic salts were used as fungicidal alternatives, that is, basil oil (Ocimum basilicum), cinnamon oil (Cinnamomum zeylanicum) and clove oil (Cymbopogon nardus). Low concentrations of essential oils were used against banana crown rot disease and their fungal pathogens; they were also used to maintain quality during storage (Ranasinghe et al., 2002; Anthony et al., 2003, 2004; Maqbool et al., 2010, 2011; Abd-Alla et al., 2014). Salts, that is, potassium sorbate, calcium propionate, sodium bicarbonate, sodium carbonate, and ammonium bicarbonate reduced crown rot incidence of banana fruits (Dionisio and Kobayashi, 2004). Antioxidants, that is, salicylic, citric, benzoic, ascorbic acids and hydroquinone have been used as protective and therapeutic treatments for plant diseases caused by a wide range of viral, fungal and bacterial pathogens (Mandal et al., 2009). Ascorbic acid, benzoic acid and butylated hydroxyl anisole (BHA) reduced mycelial growth of C. musae, the cause of anthracnose disease of banana fruits (Khan et al., 2001). Citric extract at 4% significantly reduced fruit rot incidence of banana, plus propionic acid (Ranasinghe et al., 2002; Cruz et al., 2013). The objectives of this study were to screen some essential oils, salts, and antioxidants against causal pathogens and their control of postharvest diseases of banana fruits.


 MATERIALS AND METHODS

Causal pathogenic fungi
 
Highly aggressive isolates causing major postharvest diseases of banana fruit, that is, C. musae and F. moniliforme isolates were provided from Botany Department, Faculty of Agriculture, Tanta University, Egypt.
 
Agents tested
 
The agents tested included commercial essential oils of cinnamon  oil (C. zeylanicum  Blume), thyme oil  (Thymus vulgaris L.), clove oil (Syzygium aromaticum), lemongrass oil (Cymbopogon citratus Stapf) and Black seed (Nigella sativa L.), antioxidants, that is, benzoic,  sorbic, malic, salicylic and propionic acids and salts, that is, sodium benzoate, sodium carbonate, sodium carbonate sodium chloride, and sodium hypochlorite. All agents were provided from Chemical Industrial Development Company (CID), Egypt.
 
Antifungal against causal organisms                                                                                                                       
 
Antifungal activity against mycelia linear growth of C. musae and F. moniliforme on potato dextrose agar (PDA) of essential oils, that is, cinnamon, clove, lemon grass, and black seed were tested at 0.0, 0.1, 0.5, and 1.0% concentrations. Antioxidants, that is, benzoic, sorbic, malic, salicylic and propionic acids concentrations (0.1, 0.5, and 1.0%) and salts, that is, sodium chloride, sodium hypochlorite, sodium benzoate, and sodium carbonate at 1.0, 2.0, and 4.0% were tested  at   different   concentrations   by   dissolving   the   requisite amounts in 0.5 ml of 0.1% Tween 80. Then, they were mixed completely in 100 ml of PDA medium before pouring in Petri dishes (9 cm-diameter). The control sets were prepared similarly using equal amounts of tween 80 on sterilized distilled water. Each plate was inoculated with 5 mm central disk for each fungus. Five plates were used as replicates and five plates free of each agent tested served as a control. The plates were incubated at 27±2°C for 6 days. Percentage of mycelial linear growth inhibition was calculated by the following formula (Skidmore and Dickinson, 1976) as follows:                                                                
 
Inhibition of fungal growth % = C - T /C×100
 
where C = the radial mycelia  growth in control and  T = the radial mycelia  growth in the treatment
 
Application methods on postharvest diseases incidence of banana fruits
 
Different treatment methods used for banana fruits, that is, spray, soaking and dusting were the most effective agents used for mycelial linear growth. Suspension of clove oil (1.0%) and propionic acid (0.5%) were used as spray and soaking for 5 min. Meanwhile, sodium carbonate (4.0%) and salicylic acid (0.5%) were used as spray, soaking and dust treatments on banana fruits before artificial infestation with pathogens.
 
Fruit samples
 
Banana fruits Cv. Balady were purchased from private orchard in El-Gharbeia Governorate Egypt at maturity stage. They were disinfected by double immersion in 2 of 70% ethanol for 5 min and allowed to dry at room temperature under sterile conditions. Fruits were separated in polyethylene bags previously disinfected with 70% ethanol and exposed to UV light for 20 min. They were infested with mixture spore (1:1). Suspension (1×106/ml) of either C. musae or F. moniliforme was used for inoculation of banana fruits freshly prepared from 7 days old (PDA) cultures.
 
Determination of postharvest disease incidence
 
Postharvest disease incidence was calculated as the number of infected fruits showing symptom of crown rot, neck rot, finger rot, and flower end rot as follows:
 
Disease % of diseased fruits = Number of diseased banana fruits/Total number of banana fruits × 100 
              
Disease severity was ranked by observing percentage of rotten symptom based on linear scale (0-4) as follows:
 
Disease severity (%) = Σ (n × r) × 100/N
 
where n= Number of fruits in each numerical disease grade;  r = Number of the disease grade and    N= Total number of inoculated fruits multiplied by the maximum numerical disease grade as follows:
 
0 = healthy fruit free rotten and discoloration;
1= 1-25% rotten and discoloration area;
2 = 26-50% rotten and discoloration area;
3= 51-75% rotten and discoloration area;
4 = 76-100% rotten and discoloration area.
 
Statistical analysis 
 
Data were analyzed with analysis of variance (ANOVA). Comparisons among means were made using Duncan’s multiple range test (Snedecor and Cocharn, 1980) at P = 0.05. 


 RESULTS

Laboratory study
 
Antifungal activity of essential oils against causal organisms
 
Data presented in Table 1  and  Figure  1  shows  that  all the different concentrations of essential oils tested significantly reduced mycelia linear growth of both tested fungi, that is, C. musae and F. moniliformae than the control. Data in Table 1 showed that increased essential oil concentrations led to the reduction of mycelial linear growth of two fungi tested. Clove oil (1.0%) was the best essential oil that completely inhibited (100%) linear growth of C. musae and F. moniliforme followed by black seed oil and then cinnamon oil. Clove essential oil included phenylpropanoids such as carvacrol, thymol, eugenol and cinnamaldehyde; its antimicrobial, antifungal, and antiviral activity (Chaieb et al., 2007). On the other hand, lemon grass oil was the least oil that reduced mycelial linear growth of two fungi tested. These results are in agreement with the results obtained in vitro, on fungi causing crown rot in banana fruits, that is, Lasiodiplodia theobromae, Fusarium proliferatum and C. musae. Ranasinghe et al. (2002) found that cinnamaldehyde (66.2%) is the major constituent of cinnamon bark oil, its fungistatic and fungicidal at 0.64 and 1.00 mg/ml of C. nardus and  0.2 to 0.6% (v/v) of O. basilicum. Anthony et al. (2004) and Maqbool et al. (2010) found that cinnamon oil suppressed mycelial growth and inhibited conidial germination (83.2%) of C. musae. Maqbool et al. (2011) reported that lemon grass at 0.05 and 0.4% had fungicidal activity against C. musae and Colletotrichum gloeosporioides, the causal organisms of banana and papaya anthracnose diseases, respectively.  Abd-Alla et al. (2014) found that cinnamon, thyme oils completely inhibited 100% mycelium growth and conidial germination of F. semitectum, the cause of crown rot of banana fruits. In addition, Idris et al. (2015) reported that basil, cinnamon and rosemary oil (0.1%) completely inhibited mycelia growth of C. musae.
 
 
Screening of salts against causal organisms
 
Data in Table 2 and Figure 2 showed that all salts tested significantly reduced linear growth of both fungi tested more than the control. In general, increased essential oils concentration increased their reduction of mycelial growth of the two fungi tested. Sodium carbonate was the best salt that completely (100%) inhibited the linear growth of C. musae at 2% and F. moniliforme at 4% followed by sodium hypochlorite at 4%. Data in Table 2 indicated that sodium bicarbonate and sodium benzoate significantly reduced linear growth of C. musae by 67.8 and 62.8, respectively. Data in Table 2 and Figure 2 showed that sodium carbonate and sodium hypochlorite completely (100) reduced mycelial linear growth of F. moniliforme at 4%, but sodium benzoate had moderate effect on mycelial growth of F. moniliforme followed by sodium bicarbonate. On the other hand, sodium chloride had the least effect on mycelial growth of two fungi. These results are in agreement with Dionisio and Koabyashi (2004) who found that organic and inorganic salts, that is, Na2CO3 (4 g/L), NaClO (5 g/L), NaHCO3, CaCl2, and NaCl (6 g/L) completely inhibited spore germination of fungi causing crown rot diseases in banana, that is, L. theobromae, Thielaviopsis paradoxa, C. musae, C. gloeosporioides, Fusarium verticillioides, and Fusarium oxysporum. Turkkan and Erper (2014) found that sodium metabisulfite completely inhibited mycelial growth of F. oxysporum.f.sp.capae, the cause of onion basal rot.
 
 
Effect of organic and antioxidant acids against causal organisms
 
Data in Table 3, Figures 3 and 4 shows that all organic and antioxidants acids tested significantly reduced linear growth of C. musae and F. moniliforme more than the control (untreated). In general, increased organic and antioxidants acids concentrations increased their reduction of mycelial growth of two fungi tested. Data in Table 3 clearly shows that propionic, salicylic and sorbic, and malic acids at 0.05 and 100% inhibited mycelial linear growth of fungi tested. Data in Table 3 shows that citric acid (1.0%) completely (100%) inhibited mycelial growth of C. musae and significantly reduced linear growth of F. moniliforme by 79.1%.
 
On the other hand, benzoic acid was the least agent that reduced linear growth of both fungi. Antioxidants safe natural phenolic compound chemicals, that is, salicylic, citric, benzoic, hydroquinone and ascorbic acids had been used as alternative pesticides to protective and therapeutic treatments of plant diseases caused by a wide range of viral, fungal and bacterial pathogens. Mandal et al. (2009) and Khan et al. (2001) found the greatest antifungal activity of BHA and benzoic acid against C. musae, the cause of banana fruit anthracnose disease.
 
 
 
 
Effect of treatment methods on postharvest diseases of banana fruit Cv. Balady
 
Different treatment methods, that is, spraying, soaking and dusting were tested against postharvest diseases of banana, such as, crown rot, neck rot, finger rot, and flower end rot. Data in Table 4 showed that clove oil (1.0%) and propionic acid (0.5%) were used as spray and soaking treatments for banana fruits. Meanwhile, spraying, soaking and dusting treatment were used with sodium carbonate (4.0%) and salicylic acid (0.5%). Data in Table 4 indicated that all the different treatments with clove oil, propionic acid, sodium carbonate and salicylic acid significantly reduced crown rot incidence of banana fruits more than the control. Soaking treatment significantly reduced crown rot incidence and disease severity with clove oil, propionic acid, sodium carbonate and salicylic acid compared to spray and dusting treatments. Data in Table 4 clear indicated that they were no significant differences between all the treatment methods (spray, soaking and dusting) on neck rot diseases incidence of banana fruit, except soaking treatment with sodium carbonate. Table 4 indicated that soaking treatment in clove oil and sodium carbonate significantly reduced finger rot of banana fruits more than propionic acid, salicylic acid as spray and dusting treatments.
 
 
Furthermore, data in Table 4 shows that there are no significant differences in percentage of flower end rot disease and disease severity between treatments tested and control, except soaking treatment. Clove oil was the best and significantly treatment that reduced flower end rot incidence at zero level on banana fruit. In general, soaking treatment with clove and sodium carbonate were the best treatments that reduced crown rot and neck rot. These results are in agreement with Anyhony et al. (2003) and Abd-Alla et al. (2014).
 
Effect of different treatments on the management of postharvest diseases of banana fruits
 
Healthy ripe banana fruits Cv. Balady were soaked for 5 min, in each suspension of clove oil (2.0%), sodium carbonate (6.0%), sorbic acid (2.0%), propionic acid (1.0%) and salicylic acid (1.0%) before artificial infestation with causal pathogens (Figure 5). Data in Table 5 showed that banana fruits soaked for 5 min in clove oil suspension (2.0%), sodium carbonate (6.0%), sorbic acid (2.0%), propionic acid (1.0%) and salicylic acid (1.0%) significantly reduced postharvest diseases of banana fruits, that is, crown rot, neck rot, finger rot and flower end rot more than the control. Soaking banana fruit in clove suspension was the best and significantly  treatment  that completely suppressed (100%) crown rot and flower end rot incidence of banana fruits and significantly reduced finger rot. On the other hand, salicylic acid followed by propionic acid was the lowest treatments that reduced crown, neck and finger rots. These results are in line with that of Ranasinghe et al. (2002) study who found that, spraying embul banana with emulsions of  cinnamom   oil prior storage controlled crown rot stored up to 14 days at ambient temperature (28±20°C) and 21 days at 40°C in modified atmosphere. Also, banana fruits treated with emulsions of cinnamon oil combined with modified atmosphere packaging extended the storage life of Embul banana up to 21 days in a cold room and 14 days at  28  ±  2°C   without   affecting the organoleptic and physico-chemical properties (Ranasinghe et al., 2005; Maqbool et al., 2010; Abd-Alla et al., 2014). Idris et al. (2015) found that treatment of banana fruits Cvs. Cavendish and Williams treated with essential oil (0.20%) of basil, cinnamon and rosemary essential oils reduced anthracnose of banana fruits after 19 days of storage. Singh and Tripathi (2015) showed that, C. zeylanicum oil treated banana fruits showed enhancement storage life up to 4 days. Dionisio and Kobayashi (2004) reported that dipping banana fruits in NaClO or NaHCO3 for 10 to 15 min reduced the incidence of crown rot 17 days after harvest. Kazemi et al. (2013) reported that dipping of pomegranates  fruits  for  4 min  in  4%  calcium   chloride solution combined with sodium hypochlorite (10%) was the best treatment to enhance postharvest factors of pomegranates. Cruz et al. (2013) reported that citric extract at 4% significantly reduced fruit rot incidence of banana by 19.44% more than 90.16% in the control.
 


 CONCLUSION

Clove essential oil, sodium carbonate and sorbic acid were the most promising eco-friendly, antifungal and alternative synthetic fungicides against pathogenic fungi and   their  ability  to   deteriorate   banana  fruits   during storage, marketing and transportation.


 CONFLICT OF INTERESTS

The authors have not declared any conflict of interests.



 REFERENCES

Abd-Alla MA, El-Gamal NG, El-Mougy NS, Abdel-Kader MM (2014). Post-harvest treatments for controlling crown rot disease of Williams banana fruits (Musa acuminata L.) in Egypt. Plant Pathol. Q. 4(1):1-12.

 

Abdullah Q, Mahmoud A, Al-Harethi A (2016). Isolation and identification of fungal postharvest rot of some fruits in Yemen. PSM Microbiol. 1(1):36-44.

 
 

Agrios GN (2005). Plant Pathology, 5thed Academic Press, USA. 922p.

 
 

Anthony S, Abeywickrama K, Wijeratnam SW (2003).The effect of spraying essential oils of cymbopogon nardus, cymbopogan flexuosus and Ocimum basilicum on postharvest diseases and storage life of Embul banana. J. Hortic. Sci. Biotechnol. 78:780-785.
Crossref

 
 

Anthony S, Abeywickrama K, Dayananda R, Wijeratnam SW, Arambewela L (2004). Fungal pathogens associated with banana fruit in Sri Lanka, and their treatment with essential oils. Mycopathologia 157:91-97.
Crossref

 
 

Bhattacharyya A, Chakraborty S (2007). Effect of inorganic salts on Colletotrichum musae and Fusarium solani - causal organisms of crown rot disease of banana. Assam Agricultural University, AICRP on Fruits, Jorhat, India, 290 p.

 
 

Chaieb K, Hajlaoui H, Zmantar T, Kahla-Nakbi AB, Rouabhia M, Mahdouani K, Bakhrouf A (2007).The chemical composition and biological activity of clove essential oil, Eugenia caryophyllata (Syzigium aromaticum L. Myrtaceae): a short review. Phytother. Res. 21(6):501-506.
Crossref

 
 

Cruz M, Estrada KRF, Clemente E, Itako AT, Stangarlin JR, Cruz M (2013). Plant extracts for controlling the post-harvest anthracnose of banana fruit. Rev. Bras. Plant. Med. 15(4):727-733.
Crossref

 
 

Diedhiou MP, Zakari HA, Mbaye N, Rokhaya F, Samb LP (2014). Control methods for postharvest diseases of banana (Musa sinensis) produced in Senegal. Int. J. Sci. Environ. Technol. 3:1648-1656.

 
 

Dionisio G, Kobayashi T (2004). Inhibitory influence of inorganic salts on banana postharvest pathogens and preliminary application to control crown rot. J. Gen Plant Pathol. 70:61-65.
Crossref

 
 

Dionisio GA, Natsuaki KT (2007). Control of crown rot-causing fungal pathogens of banana by inorganic salts and a surfactant. Crop Prot. 26(11):1667-1673.
Crossref

 
 

Idris MF, Ibrahim MA, Forsido FS (2015). Essential oils to control Colletotrichum musae in vitro and in vivo on banana fruits. Am-Eur. J. Agric. Environ. Sci. 15(3):291-302.

 
 

Kazemi F, Jafararpoor M, Golparvar A (2013). Effects of sodium and calcium treatments on the shelf life and quality of pomegranate. Intl. J. Farm Allied Sci. 2(S2):1375-1378.

 
 

Khan SHJ, Aked J, Magan N (2001). Control of the anthracnose pathogen of banana (Colletotrichum musae) using antioxidants alone and in combination with thiabendazole or imazalil. Plant Pathol. 50(5):601:608.

 
 

Khleekorn S, McGovern J, Wongrueng S (2015). Control of the banana anthracnose pathogen using antagonistic microorganisms. J. Agric. Technol. 11(4):965-973.

 
 

Lassois L, Bellaire L, Jijakli MH (2008). Biological control of crown rot of bananas with Pichia anomala strain K and Candida oleophila strain. Biol. Control 45:410-418.
Crossref

 
 

Mandal S, Mallick N, Mitra A (2009). Salicylic acid induced resistance to Fusarium oxysporum f.sp.lycopersici in tomato. Plant Physiol. Biochem. 47:642-649.
Crossref

 
 

Maqbool M, Ali A, Peter G, Alderson PG (2010). Effect of cinnamon oil on incidence of anthracnose disease and postharvest quality of bananas during storage. Int. J. Agric. Biol. 12:516-520.

 
 

Maqbool M, Ali A, Peter G, Aldersona PG, Mohamed MTM, Siddiqui Y, Zahida N (2011). Postharvest application of gum Arabic and essential oils for controlling anthracnose and quality of banana and papaya during cold storage. Postharvest Biol. Technol. 62:71-76.
Crossref

 
 

Marin H, Sutton B, Blankenship M, Swallow H (1996). Pathogenicity of fungi associated with crown rot of banana in Latin America on grande naine and disease resistant hybrid banana. Plant Dis. 80(5):525-528.
Crossref

 
 

Ranasinghe L, Jayawardena B, Abeywickrama K (2005). An integrated strategy to control post- harvest decay of Embul banana by combining essential oils with modified atmosphere packaging. Intl. J. Food Sci. Technol. 40:97-103.
Crossref

 
 

Ranasinghe L, Jayawardena B, Abeywickrama K (2002). Fungicidal activity of essential oils of Cinnamomum zeylanicum (L.) and Syzygium aromaticum (L.) Merr et L M Perry against crown rot and anthracnose pathogens isolated from banana. Lett. Appl. Microbiol. 35:208-211.
Crossref

 
 

Singh R, Tripathi P (2015). Cinnamomum zeylanicum essential oil in the management of anthracnose of banana fruits. J. Innov. Pharm. Biol. Sci. 2(3):290-299.

 
 

Skidmore AM, Dickinson CH (1976). Colony interactions and hyphal interference between Septoria nodorum and phylloplane fungi. Trans. Br. Mycol. Soc. 66:57-64.
Crossref

 
 

Snedecor GW, Cochran WG (1980). Statistical Methods. 7th Ed. Iowa State Univ. Press, Ames. pp. 476-480.

 
 

Triesta D, Pierard D, De Cremer K, Hendrickx M (2016). Fusarium musae infected banana fruits as potential source of human Fusariosis : may occur more frequently than we might think and hypotheses about infection. Commun. Integr. Biol. 9(2):e1162934.
Crossref

 
 

Turkkan M, Erper I (2014). Evaluation of antifungal activity of sodium salts against onion basal rot caused by Fusarium oxysporum f. sp. cepae. Plant Prot. Sci. 50(1):19-25.

 
 

Zhang P, Whistler RL, BeMiller JN, Hamaker BR (2005). Banana starch: Production, physicochemical properties, and digestibility-a review. Carbohyd. Polym . 59:443-458.
Crossref

 

 




          */?>