ReseaRch PaPeR Journal of Agricultural and Marine Sciences 2023, 28(2): 38–44 DOI: 10.53541/jams.vol28iss2pp38-44 Received 20 November 2022 Accepted 9 May 2023 إمكانية إستخدام الزيت العطري املستخرج من Zataria multiflora للسيطرة على تعفن فاكهة الفراولة بعد احلصاد ماجدة بنت سعيد بن محد احلضرمية1 و زينب البلوشية1 و عيسى بن هاشل املهمويل1 و عبدهللا بن حممد السعدي1 و ريثيناسامي فيالزهاهن1* Potential use of Zataria multiflora essential oil to control postharvest Aspergillus flavus fruit rot of strawberry Majida Said Hamed Al-Hdhrami, Zainab Al-Balushi, Issa Hashil Al-Mahmooli, Abdullah Mohammed Al-Sadi and Rethinasamy Velazhahan* Rethinasamy Velazhahan1 *( ) velazhahan@squ.edu.om, Sultan Qaboos University, College of Agricultural and Marine Sciences, Department of Plant Sciences, P.O. Box 34, Al-Khoud, Muscat 123, Sultanate of Oman. Introduction Strawberry (Fragaria × ananassa Duch.) is one of the popular berries in the world. Strawberry fruits are preferred by consumers because of their attrac- tive colour, aroma, taste and nutritional qualities (Bhat et al., 2015). It is one of the principal fruit crops culti- vated in the greenhouses in Oman. Strawberry fruits are rich source of fiber and several phytochemicals, such as folic acid, vitamin C and other antioxidants (Chandler et al., 2012). It is being used as fresh fruit or as sliced and processed fruit (Chandler et al., 2012; Giampieri et al., 2012). These fruits are highly prone to attack by several fungal pathogens because of their succulent nature that can result in postharvest decay. The most common fun- gal pathogens that cause postharvest decay in strawber- ry fruits are Botrytis cinerea, Rhizopus stolonifer, Mucor spp., Colletotrichum acutatum, Geotrichum candidum, Penicillium expansum, Cladosporium spp., Alternar- ia spp. and Aspergillus spp. (Feliziani and Romanazzi, 2016; Ma et al., 2018; Palmer et al., 2019; Al-Rahbi et al., 2021). A few postharvest fungal pathogens of strawberry are known to produce foodborne mycotoxins (Hussein et al., 2020) and hence the quality of fruits after harvest is important for consumer’s safety. For instance, A. flavus produces aflatoxin, a naturally occurring carcinogen- ic mycotoxin (Khlangwiset et al., 2011) and Alternaria spp. produce foodborne mycotoxins including alterna- riol, alternariol methyl ether, tentoxin, tenuazonic acid, altenuene and altertoxins (Escriva et al., 2017). Posthar- vest diseases of strawberry can be managed by avoiding mechanical damage to fruits by following safe handling practices, cold storage of fruits, storage of fruits under modified storage atmosphere, application of chemical fungicides, biocontrol agents and natural products like plant extracts and essential oils, and physical method like UV-C treatment (Feliziani and Romanazzi, 2016). A. flavus is one of the most common postharvest fungal pathogens of fruits. Several studies reported the inhibitory activity of essential oils (EOs) from plants against A. flavus and aflatoxin production (Vilela et al., 2009; Kumar et al., 2010; Tian et al., 2012a; Kedia et al., 2014; Kedia et al., 2015; Kiran et al., 2016; Chaudhari et al., 2020). The EOs are volatile and odorous liquids extracted from plant tissues. The EOs pass through the cell wall of susceptible fungi because of their lipophilic nature and cause damage to the cytoplasmic membrane, Abstract. Postharvest fruit rot is a major problem in strawberry production chain worldwide. Aspergillus sp. is one of the major fungi associated with fruit rot of strawberry. In this study, an Aspergillus flavus was isolated from a rotten strawberry fruit. Based on the nucleotide sequence analysis of the internal transcribed spacer regions of rDNA, the fun- gus was confirmed as A. flavus. Pathogenicity of the isolated fungus was confirmed by artificially inoculating strawberry fruits under laboratory conditions. This strain was capable of producing aflatoxin B1 in vitro as determined by liquid chromatography-mass spectrometry analysis. Postharvest dip treatment of mature strawberry fruits with Zataria mul- tiflora essential oil (ZEO) (0.1%) completely suppressed A. flavus infection and prevented rotting of fruits. The results of this study suggest that ZEO can be used as a sustainable and safe alternative to chemical fungicides for the control of Aspergillus fruit rot of strawberry. Keywords: Aspergillus flavus, aflatoxin B1, essential oil, fruit rot, strawberry, Zataria multiflora امللخــص: تعــد مشــكلة تعفــن مثــار الفاكهــة مشــكلة رئيســية يف سلســلة إنتــاج الفراولــة يف مجيــع أحنــاء العــامل. يعــد فطــر أســرجيلس أحــد الفطــرايت الرئيســية املرتبطــة بتعفــن مثــار الفراولــة.يف هــذه الدراســة مت عــزل عزلــة مــن فطــر أســبريجيلس فليفــس مــن مثــار فراولــة فاســدة. اعتمــادا علــى دراســة القواعــد الوراثيــة للمنطقــة املصانــة ريبوســومي للحمــض النــووي مت أتكيــد الفطــر علــى أنــه أســبريجيلس فليفــس. و مت أتكيــد القــدرة اإلمراضيــة للفطــر عــن طريــق التلويــث اليــدوي لثمــار الفرولــة حتــت ظــروف خمريــة. أظهــرت الدراســة قــدرة هــذه الســاللة علــى إنتــاج األفالتوكســن )B1( يف املختــر مــن خــالل حتليــل الكروماتوغرافيــا الســائلة الطيــف الكتلــي. كمــا أشــارت الدراســة إىل أن غمــس مثــار الفراولــة الناضجــة بعــد احلصــاد يف الزيــت العطــري للزعــر O(ZEO) Zataria multiflora بنسبة 0.1 % يكبح عدوى الفطر ومينع تعفن الثمار متاما. تشري نتائج هذه الدراسة إىل أنه ميكن إستخدام الزيت العطري للزعر ZEO بديال آمنا ومســتداما للمبيدات الفطرية وذلك ملكافحة تعفن مثار الفراولة ابســبريجيلس. Zataria multiflora ، الزيت العطري ، تعفن الثمار ، فراولة ، B1 الكلمات املفتاحية: أسبريجيلس فليفس ، أفالتوكسن 39Research Paper Al-Hdhrami, Al-Balushi, Al-Mahmooli, Al-Sadi, Velazhahan coagulation of the cytoplasm and leakage of cellular macromolecules (Hyldgaard et al., 2012; Dwivedy et al., 2016). Gandomi et al. (2011) reported that Zataria mul- tiflora EO induced morphological and structural chang- es in A. flavus including loss of turgidity, vacuolization of cytoplasm and deformation of hyphae. An increase in the generation of reactive oxygen species (ROS) and mitochondrial membrane potential and decrease in AT- Pase and dehydrogenase enzyme activities, and ergos- terol content were reported in A. flavus due to the effect of Anethum graveolens (Dill) EO (Tian et al., 2012b). Chaudhari et al. (2020) demonstrated that Allspice (Pi- menta dioica) EO caused damage to plasma membrane of A. flavus that resulted in the leakage of cellular ions. Al-Harrasi et al. (2021) recently reported the antifungal activity against A. flavus and aflatoxin B1-detoxification potential of Zataria multiflora (Lamiaceae) essential oil (ZEO). In this study, A. flavus was isolated from diseased strawberry fruits and tested for its pathogenicity and af- latoxigenic potential. Furthermore, the efficacy of ZEO in the control of A. flavus- induced postharvest fruit rot of strawberry was studied. This is the initial study in Oman describing isolation of an aflatoxigenic strain of A. flavus from rotted strawberry fruits as well as demon- strating the effectiveness of ZEO in the management of A. flavus fruit rot of strawberry. Materials and Methods Collection of Fruits Strawberry fruits showing fruit rot symptoms were collected from a supermarket in Muscat, Sultanate of Oman and transferred to the laboratory in an ice box and used within 24 h after collection for isolation of fungus. Isolation of Fungus Diseased tissues (5×5 mm, approx.) from the fruits were cut with a sterile scalpel and surface-disinfected with NaOCl solution (1%) for 2 min, rinsed 2-3 times with sterile distilled water (SDW), blot-dried on sterile What- man No. 1 filter paper and then placed on potato dex- trose agar (PDA) medium (Oxoid Ltd., UK) in 9-cm di- ameter Petri dishes. The plates were sealed with parafilm and incubated at 27°C for 3-4 days. The pure culture of the fungus was obtained by single spore isolation meth- od and the culture was maintained on PDA slants at 4°C. Pathogenicity Test Healthy strawberry fruits were surface sterilized with 70% ethanol for 5 s, rinsed twice with SDW and air- dried under aseptic condition. On the surface of each fruit, minute wounds were created by using a sterile in- oculation needle. Twenty µl of spore suspension (1×106 spores/ml) of the fungal isolate prepared from a 5-day- old PDA culture was applied on the wounded site and incubated for 7-10 days at 25°C. Strawberry fruits ap- plied with SDW served as control. Disease severity was measured using a 0-8 scale developed for gray mold of strawberry, where 0 indicated no disease and 8 in- dicated 87.6-100% of fruit decayed (Chen et al., 2018). Aflatoxin B1 Production by Fungal Isolate Agar disc (6 mm diameter) of the fungus taken from a 7-day-old PDA culture was transferred to 200 ml of ster- ile SMKY medium (200 g sucrose, 0.5 g magnesium sul- fate, 0.3 g potassium nitrate, and 7.0 g yeast extract in 1 l of distilled water) (Tiwari et al., 2022) in a 500 ml-conical flask and incubated at 27°C. After 14 days of incubation the culture was filtered through Whatman No.1 filter pa- per and the culture filtrate was collected. In a sterile 1.5 ml centrifuge tube, 500 µl of the fungal culture filtrate and 500 µl of chloroform were added and vortexed for a few seconds. The tubes were then centrifuged at 12000 g at room temperature (25±2°C) and the chloroform layer was transferred to a new 1.5 ml tube and dried completely by using a water bath at 60°C. The residue was dissolved in 250 µl of analytical grade methanol and analyzed by liquid chromatography-mass spectrometry (LC-MS). LC-MS Analysis Analysis of Aflatoxin B1 was performed using an Agi- lent LC/MS/MS (Agilent Technologies, Inc., CA, USA) system, equipped with a high-performance auto sampler (G4226A), quaternary pump (G4204A), thermostated column compartment (G1316C) and Agilent 6460 Tri- ple Quad Mass Spectrometer. The analyte separation was achieved using Symmetry C18; 5 µm, 3 mm × 150 mm column (Waters), with a mobile phase of water (A) and acetonitrile (B). Both the eluents were with 0.1% for- mic acid under gradient condition (eluent A 10-70% in 0-1 min, 70-95% in 1-2 min, hold at 95% for 4 min, 95- 70% in 6-7.5 min, 70-10% in 7.5-8 min and hold at 10% for 1 min) with a flow rate of 0.3 ml per min. The column oven temperature was maintained at 45°C. Standard Aflatoxin B1 (Sigma-Aldrich, MO, USA) solution was used to optimize all MS parameters. The optimized MS parameter values were: gas temperature 300°C, gas flow 3 L min-1, nebulizer pressure 50 PSI, sheath gas heater 375°C, sheath gas flow 10 L min-1, capillary voltage 3500 V, scan range 100 to 3000 m/z and positive polarity. MS Data acquisition and analysis were performed using Ag- ilent MassHunter Software. Molecular Characterization of the Fungus The genomic DNA extraction from the fungal isolate was carried out using the Plant/Fungi DNA Isolation Kit (Norgen Biotek Corp., Canada) as per the manu- facturer’s instructions. The amplification of the internal transcribed spacer (ITS) regions of the rDNA gene was done as suggested by Karthikeyan et al. (2009). Briefly, the PCR reaction was performed in a 25 µl reaction mix- ture consisting of a PuReTaq Ready-To-Go PCR bead 40 SQU Journal of Agricultural and Marine Sciences, 2023, Volume 28, Issue 2 Potential use of Zataria multiflora essential oil to control postharvest Aspergillus flavus fruit rot of strawberry (GE Healthcare, UK), 1 µl each of ITS4 and ITS5 prim- ers (White et al., 1990) and 2 µl (100 ng) of the purified DNA and 21 µl of SDW. The thermal profile for PCR was as described by Halo et al. (2018). The amplification was confirmed by running 5 µl of the PCR product on 1% agarose gel in Tris-borate-EDTA (TBE) buffer followed by observation under UV light using a GeneFlash gel im- aging system (Syngene, Cambridge, UK). The PCR prod- uct (~700 bp) was sequenced at Macrogen, Seoul, Korea. A homology search of the obtained nucleotide sequence was performed using the BLAST program (www.ncbi. nlm.nih.gov/BLAST) in order to identify the fungus. Efficacy of ZEO in Controlling Aspergillus Fruit Rot of Strawberry ZEO extracted previously (Al-Harrasi et al., 2021) was used in this study. Healthy strawberry fruits of uniform size and maturity were selected, and surface sterilized by dipping them in 1% NaOCl for 1 min and then washed twice in SDW. Then, they were dipped in ZEO (0.1% in distilled water) for 5 s and then air-dried in a laminar flow chamber for ~10 min. The fruits were then inocu- lated with A. flavus spore suspension as described ear- lier and incubated for 7 days at 25 °C. Strawberry fruits applied with SDW and inoculated with A. flavus served as control. Three fruits were used for each treatment and each treatment was replicated thrice. Statistical Analysis All experiments were performed in triplicate. The fruit rot disease severity data were expressed as mean ± stan- dard deviation (SD). Results and Discussion In this study, A. flavus (isolate STR10) was isolated from strawberry fruits showing the symptoms of fruit rot and pure culture was obtained (Figure 1). To con- firm the identity the fungus, the ITS region of rDNA was amplified by PCR using ITS4 and ITS5 primers and the amplified product was sequenced. BLAST analy- sis of the obtained nucleotide sequence showed 100% sequence similarity with that of A. flavus strain acces- sions MT635198, MT509715, MN893386, MN893385, MN533904 and MN533814 in the GenBank database. The nucleotide sequence of A. flavus isolate STR10 from this study was deposited in the GenBank with the acces- sion number OL437469. The pathogenicity of the isolate was confirmed by artificial inoculation of strawberry fruits. The inoculated fruits exhibited the fluffy mycelial growth and symptoms of fruit rot 5-8 days after inocula- tion (disease severity score 6.4 ± 0.9) whereas the control fruits remained symptomless (disease severity score 0) (Figure. 2). A. flavus has been reported to cause posthar- vest rots in several fruits including grapes (Ghuffar et al., 2020), jujube (Singh and Sumbali, 2000), peach (Micha- ilides and Thomidis, 2007), kiwi (Zhu et al., 2022) and Figure 1. Growth of Aspergillus flavus isolated from decayed strawberry fruit on potato dextrose agar medium 41Research Paper Al-Hdhrami, Al-Balushi, Al-Mahmooli, Al-Sadi, Velazhahan lemon (Kotan et al., 2009). Several strains of A. flavus isolated from rotten fruits were shown to produce afla- toxins. The aflatoxins are carcinogenic, mutagenic and immunosuppressive compounds produced by A. flavus and A. parasiticus as secondary metabolites when the fungi grow on susceptible crops and food matrices un- der favourable conditions. Among the different forms of aflatoxins, aflatoxin B1 (AFB1) is considered as the most dangerous and toxic to humans and animals (Velazha- han, 2017). Saleem (2017) reported the production of aflatoxins by A. flavus and A. parasiticus from straw- berry and the concentration of aflatoxins ranged from 23.6 to 71.1 ppb. Hussein et al. (2020) reported that A. flavus was recovered from 53.3%, of strawberry samples collected from Qena city, Egypt and the amount of af- latoxin produced by A. flavus was 3.5 ppb. The strain of A. flavus, STR10, isolated from infected strawberry fruits in this study was capable of producing aflatoxin B1 to a level of 4063 ± 993 ppb under in vitro conditions. Hence, control of Aspergillus fruit rot is of paramount importance to preserve the quality of harvested fruits for consumer’s safety especially if the fruits are intended to be used as dry fruits. The natural products such as plant extracts and EOs have been suggested as an alter- native to synthetic chemical fungicides for the control of postharvest diseases of fruits (Shao et al., 2013; Hos- seini et al., 2020; Jahani et al., 2020; Raveau et al., 2020). The results of the present study revealed that post- harvest dip treatment of strawberry fruits with ZEO (0.1%) completely prevented A. flavus infection under laboratory conditions (Figure 3). The fungal mycelium covered 65-75% of the fruit surface within 5-7 days af- ter inoculation in the untreated control (disease severity score 6.1 ± 0.8); whereas the ZEO-treated fruits showed no visible symptoms (disease severity score 0). The effec- tiveness of plant-derived EOs in controlling postharvest fruit rots have been reported in previous studies (Nik- khah et al., 2017; Hosseini et al., 2020). Garlic and rose- mary EOs have been reported to control anthracnose (Colletotrichum nymphaeae) of strawberry (Hosseini et al., 2020). El-Mogy and Alsanius (2012) reported the suppression of Botrytis cinerea fruit rot of strawberry by Cassia EO. Alizadeh-Salteh et al. (2010) demonstrat- ed the antifungal activity of Shiraz thyme EO against Rhizopus stolonifer the causal agent of Rhizopus rot of peach fruits. Mohammadi et al. (2015) demonstrated the effectiveness of combined application of chitosan with Z. multiflora or Cinnamomum zeylanicum EOs in the control of B. cinerea rot in strawberry. A few stud- ies demonstrated the synergistic antimicrobial effect of EOs (Sharma and Sharma, 2011; Nguefack et al., 2012; Nikkhah et al., 2017). Nikkhah et al. (2017) demonstrat- ed that pear fruits treated with cinnamon + rosemary + thyme or thyme + cinnamon EOs showed higher reduc- tion in the lesion size of rot induced by Botrytis cinerea and Penicillium expansum than single EO treatments Figure 2. Aspergillus flavus infection on artificially inoculated strawberry fruits Surface-disinfected strawberry fruits were inoculated with A. flavus spore suspension (1×106 spores/ml) and incubated at 25 °C. The pictures were taken 5, 8 and 10 days after inoculation (DAI). 42 SQU Journal of Agricultural and Marine Sciences, 2023, Volume 28, Issue 2 Potential use of Zataria multiflora essential oil to control postharvest Aspergillus flavus fruit rot of strawberry under laboratory conditions. A few essential oil-based commercial products such as Promax™, Sporan™, EcoP- COR and EcoTrol are being used as food preservatives to prevent fungal contamination (Dwivedy et al., 2016). Conclusion The results of this study suggest that ZEO can be used as a safe method for the control of Aspergillus flavus fruit rot of strawberry. The U.S. Food and Drug Administration (FDA) listed a number of EOs under “Generally Recog- nized as Safe (GRAS)” category. Further research is need- ed to test the effectiveness of ZEO on other postharvest fruit rot fungal pathogens of strawberry, to elucidate the mechanisms of antifungal action of ZEO and to deter- mine if ZEO affects sensory quality of strawberry fruits. Acknowledgments The authors thank the Central Analytical and Applied Re- search Unit (CAARU), SQU for LC-MS analysis of AFB1 References Al-Harrasi MMA, Al-Sadi AM, Al-Sabahi JN, Al-Farsi K, Waly MI, Velazhahan R. (2021). Essential oils of Heliotropium bacciferum, Ocimum dhofarense and Zataria multiflora exhibit aflatoxin B1 detoxification potential. All Life 14: 989-996. Al-Rahbi BAA, Al-Sadi AM, Al-Mahmooli IH, Al-Maawali SS, Al-Mahruqi NMT, Velazhahan R. (2021). Meyerozyma guilliermondii SQUCC-33Y suppresses postharvest fruit rot of strawberry caused by Alternaria alternata. Australasian Plant Pathology 50: 349-352. Alizadeh-Salteh S, Arzani K, Omidbeigi R, Safaie N. (2010). Essential oils inhibit mycelial growth of Rhi- zopus stolonifer. European Journal of Horticultural Science 75: 278-282. Bhat R, Geppert J, Funken E, Stamminger R. (2015). Consumers perceptions and preference for strawber- ries-a case study from Germany. International Jour- nal of Fruit Science 15: 405-424. Chandler CK, Folta K, Dale A, Whitaker VM, Her- rington M. (2012). Strawberry. In: Badenes M, Byrne D, editors. Fruit Breeding. Handbook of Plant Breed- ing, vol 8. Springer, Boston, MA, USA. p. 305-325. Figure 3. Effect of postharvest dip treatment of strawberry fruits with Zataria multiflora essential oil (ZEO) on Aspergillus flavus fruit rot ZEO-treated or sterile distilled water-treated (control) strawberry fruits were inoculated with A. flavus spore suspension (1×106 spores/ml) and the disease severity was measured using a 0-8 scale after incubation at 25 °C for 7 days. Data are mean ± SD. 43Research Paper Al-Hdhrami, Al-Balushi, Al-Mahmooli, Al-Sadi, Velazhahan Chaudhari AK, Singh VK, Dwivedy AK, Das S, Upad- hyay N, Singh A, Dkhar MS, Kayang H, Prakash B, Dubey NK. (2020). Chemically characterised Pimen- ta dioica (L.) Merr. essential oil as a novel plant based antimicrobial against fungal and aflatoxin B1 con- tamination of stored maize and its possible mode of action. Natural Product Research 34: 745-749. Chen PH, Chen RY, Chou JY. (2018). Screening and eval- uation of yeast antagonists for biological control of Botrytis cinerea on strawberry fruits. Mycobiology 46: 33-46. Cox SD, Mann CM, Markham JL, Bell HC, Gustafson JE, Warmington JR, Wyllie SG. (2000). The mode of antimicrobial action of the essential oil of Melaleuca alternifolia (tea tree oil). Journal of Applied Microbi- ology 88: 170-175. Dwivedy AK, Kumar M, Upadhyay N, Prakash B, Dubey NK. (2016). Plant essential oils against food borne fungi and mycotoxins. Current Opinion in Food Sci- ence 11: 16-21. El-Mogy MM, Alsanius BW. (2012). Cassia oil for con- trolling plant and human pathogens on fresh straw- berries. Food Control 28: 157-162. Escriva L, Oueslati S, Font G, Manyes L. (2017). Alter- naria mycotoxins in food and feed: an overview. Jour- nal of Food Quality 2017: 1-20 (Article 1569748). Feliziani E, Romanazzi G. (2016). Postharvest decay of strawberry fruit: Etiology, epidemiology, and disease management. Journal of Berry Research 6: 47-63. Gandomi H, Misaghi A, Basti AA, Hamedi H, Shirvani ZR. (2011). Effect of Zataria multiflora Boiss. essen- tial oil on colony morphology and ultrastructure of Aspergillus flavus. Mycoses 54: e429-e437. Ghuffar S, Irshad G, Ahmed MZ, Zeshan MA, Ali R, Haq EU, Anwaar HA, Abdullah A, Ahmad F, Haque K. (2020). First report of Aspergillus flavus causing fruit rot of grapes (Vitis vinifera) in Pakistan. Plant Disease 104: 3062. Giampieri F, Tulipani S, Alvarez-Suarez JM, Quiles JL, Mezzetti B, Battino M. (2012). The strawberry: Com- position, nutritional quality, and impact on human health. Nutrition 28: 9-19. Halo BA, Al-Yahyai RA, Al-Sadi AM. (2018). Aspergillus terreus inhibits growth and induces morphological abnormalities in Pythium aphanidermatum and sup- presses Pythium-induced damping-off of cucumber. Frontiers in Microbiology 9: 1-12 (Article 95). Hosseini S, Amini J, Saba MK, Karimi K and Pertot I. (2020). Preharvest and postharvest application of garlic and rosemary essential oils for controlling an- thracnose and quality assessment of strawberry fruit during cold storage. Frontiers in Microbiology 11: 1-15 (Article 1855). Hussein MA, El-Said AH, Yassein AS. (2020). Mycobiota associated with strawberry fruits, their mycotoxin po- tential and pectinase activity. Mycology 11: 158-166. Hyldgaard M, Mygind T, Meyer RL. (2012). Essential oils in food preservation: mode of action, synergies, and interactions with food matrix components. Frontiers in Microbiology 3: 1-24 (Article 12). Jahani M, Beheshti M, Aminifard MH, Hosseini A. (2020). Effects of essential oils to control Penicillium sp. in in vitro and in in vivo on grapevine (Vitis Vi- nifera L.) fruit. International Journal of Fruit Science 20: 812-826. Karthikeyan M, Sandosskumar R, Mathiyazhagan S, Mohankumar M, Valluvaparidasan V, Kumar S, Velazhahan R. (2009). Genetic variability and afla- toxigenic potential of Aspergillus flavus isolates from maize. Archives of Phytopathology and Plant Protec- tion 42: 83-91. Kedia A, Prakash B, Mishra PK, Dubey NK. (2014). An- tifungal and antiaflatoxigenic properties of Cuminum cyminum (L.) seed essential oil and its efficacy as a preservative in stored commodities. International Journal of Food Microbiology 168: 1-7. Kedia A, Prakash B, Mishra PK, Dwivedy AK, Dubey NK. (2015). Trachyspermum ammi L. essential oil as plant based preservative in food system. Industrial Crops and Products 69: 104-109. Khlangwiset P, Shephard GS, Wu F. (2011). Aflatoxins and growth impairment: a review. Critical Reviews in Toxicology 41: 740-755. Kiran S, Kujur A, Prakash B. (2016). Assessment of preservative potential of Cinnamomum zeylanicum Blume essential oil against food borne molds, aflatox- in B1 synthesis, its functional properties and mode of action. Innovative Food Science and Emerging Tech- nologies 37: 184-191. Kotan R, Dikbas N, Bostan H. (2009). Biological control of post harvest disease caused by Aspergillus flavus on stored lemon fruits. African Journal of Biotech- nology 8: 209-214. Kumar A, Shukla R, Singh P, Dubey NK. (2010). Chem- ical composition, antifungal and antiaflatoxigenic activities of Ocimum sanctum L. essential oil and its safety assessment as plant based antimicrobial. Food and Chemical Toxicology 48: 539-543. Ma W, Zhang Y, Wang C, Liu S, Liao X. (2018). A new dis- ease of strawberry fruit rot, caused by Geotrichum can- didum in China. Plant Protection Science 54: 92-100. Michailides T, Thomidis T. (2007). First report of Asper- gillus flavus causing fruit rots of peaches in Greece. Plant Pathology 56: 352. Mohammadi A, Hashemi M, Hosseini SM. (2015). The control of Botrytis fruit rot in strawberry using com- bined treatments of Chitosan with Zataria multiflora or Cinnamomum zeylanicum essential oil. Journal of Food Science and Technology, 52: 7441-7448. 44 SQU Journal of Agricultural and Marine Sciences, 2023, Volume 28, Issue 2 Potential use of Zataria multiflora essential oil to control postharvest Aspergillus flavus fruit rot of strawberry Nguefack J, Tamgue O, Dongmo JL, Dakole CD, Leth V, Vismer HF, Zollo PA, Nkengfack AE. (2012). Syner- gistic action between fractions of essential oils from Cymbopogon citratus, Ocimum gratissimum and Thymus vulgaris against Penicillium expansum. Food Control 23: 377-383. Nikkhah M, Hashemi M, Najafi MBH, Farhoosh R. (2017). Synergistic effects of some essential oils against fungal spoilage on pear fruit. International Journal of Food Microbiology 257: 285-294. Palmer MG, Mansouripour SM, Blauer KA, Holmes GJ. (2019). First report of Aspergillus tubingensis causing strawberry fruit rot in California. Plant Disease 103: 2948. Raveau R, Fontaine J, Sahraoui ALH. (2020). Essen- tial oils as potential alternative biocontrol products against plant pathogens and weeds: A review. Foods 9: 1-31. Saleem AR. (2017). Mycobiota and molecular detection of Aspergillus flavus and A. parasiticus aflatoxin con- tamination of strawberry (Fragaria ananassa Duch.) fruits. Archives of Phytopathology and Plant Protec- tion 50: 982-996. Shao X, Wang H, Xu F, Cheng S. (2013). Effects and possible mechanisms of tea tree oil vapor treatment on the main disease in postharvest strawberry fruit. Postharvest Biology and Technology 77: 94-101. Sharma M, Sharma R. (2011). Synergistic antifungal ac- tivity of Curcuma longa (turmeric) and Zingiber of- ficinale (ginger) essential oils against dermatophyte infections. Journal of Essential Oil Bearing Plants 14: 38-47. Singh YP, Sumbali G. (2000). Natural incidence of toxi- genic Aspergillus flavus strains on the surface of pre-harvest jujube fruits. Indian Phytopathology 53: 404-406. Tian J, Ban X, Zeng H, He J, Chen Y, Wang Y. (2012b). The mechanism of antifungal action of essential oil from dill (Anethum graveolens L.) on Aspergillus fla- vus. PLoS One 7: 1-10. Tian J, Huang B, Luo X, Zeng H, Ban X, He J, Wang Y. (2012a). The control of Aspergillus flavus with Cin- namomum jensenianum Hand.-Mazz essential oil and its potential use as a food preservative. Food Chemistry 130: 520-527. Tiwari S, Upadhyay N, Singh BK, Singh VK, Dubey NK. (2022). Facile fabrication of nanoformulated Cin- namomum glaucescens essential oil as a novel green strategy to boost potency against food borne fungi, aflatoxin synthesis, and lipid oxidation. Food and Bi- oprocess Technology 15: 319-337. Velazhahan R. (2017). Bioprospecting of medicinal plants for detoxification of aflatoxins. International Journal of Nutrition, Pharmacology, Neurological Diseases 7: 60-63. Vilela GR, de Almeida GS, D’Arce MABR, Moraes MHD, Brito JO, da Silva MFDG, Silva SC, de Stefano Piedade SM, Calori-Domingues MA, da Gloria EM. (2009). Activity of essential oil and its major com- pound, 1, 8-cineole, from Eucalyptus globulus La- bill., against the storage fungi Aspergillus flavus Link and Aspergillus parasiticus Speare. Journal of Stored Products Research 45: 108-111. White TJ, Bruns T, Lee S, Taylor J. (1990). Amplifica- tion and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ (eds) PCR protocols: a guide to methods and applications. Academic Press, San Di- ego, p.315-322. Zhu G, Wang X, Chen T, Wang S, Chen X, Song Z, Shi X, Laborda P. (2022). First Report of Aspergillus flavus Causing Fruit Rot on Kiwifruit in China. Plant Dis- ease. 106: 1990.