Determination of fungal and bacterial species and their densities in fruit and airborne microbiota in citrus packing houses located in Hatay province

Abstract

Aims: This study aims to determine fungal and bacterial species and their densities in fruit and airborne microbiota in citrus packing houses located in Antakya, Dörtyol and Erzin districts of Hatay province.

Methods and Results: Investigation were conducted in citrus fruit packing houses in Hatay province in 2020-2021. In order to determine the airborne microbiota, the nutrient media were placed in 3 different regions of the packing houses (product entrance, fruit processing and product dispatch). Fruit-borne fungal and bacterial isolates were obtained from the deliberately selected rotten fruits. Fungal and bacterial isolates were identified by morphological, biochemical and MALDI-TOF proteomic analysis. Penicillium italicum, Penicillium digitatum, Aspergillus niger, Rhizopus oryzae, Mucor circinelloides, Xanthomonas hortorum, Pantoea eucrina, Leclercia adecarboxylata, Pseudescherichia vulneris, Bacelliotymegatibia vulnerica, Bacelliot, eucrina, Leclercia adecarboxylata and Pseudescherichia vulnerica were determined as airborne fungal and bacterial species. Penicillium digitatum, Penicillium italicum, Fusarium solani, Geotrichum citri-auranti, Alternaria alternata, Cladosporium herbarum, Phytophthora sp., Colletotrichum gloeosporioides, Geotrichum candidum, Erwinia herbicola, Xanthomonas hortorum were isolated and identified as fruit borne fungal and bacterial species on infected fruits.

Conclusions: It has been observed that the airborne microbiota density is quite higher in the fruit processing section of the packinghouses, while the microbiota density is low in the product acceptance (entry) and dispatch (exit) sections because of proper ventilation.

Significance and Impact of the Study: Airborne fungal and bacterial microbiota in the packinghouses can be easily transported by air movements and remain suspended in the air for a certain period of time, causing symptoms such as deterioration and rot in fruits. Since some of identified airborne bacterial and fungal species are known as allergens or human pathogens, necessary advices were given about the sanitation measures which should be taken into consideration in commercial citrus packinghouses.

Keywords

• Citrus
• packinghouses
• post-harvest diseases
• MALDI-TOF
• air/fruit borne microbiota.

Hatay ilinde yer alan turunçgil paketleme tesislerinde meyve ve hava kökenli mikrobiyata içerisindeki fungal ve bakteriyel türler ile yoğunluklarının belirlenmesi

Abstract

Amaç: Bu çalışmanın amacı, Hatay ilinin Antakya, Dörtyol ve Erzin ilçelerinde bulunan turunçgil meyve paketleme tesislerinde meyve ve hava kökenli mikrobiyata içerisinde yer alan fungal ve bakteriyel mikrobiyota türlerini ve yoğunluklarını belirlemektir.

Yöntem ve Bulgular: Hatay ilinde bulunan turunçgil meyve paketleme tesislerinde 2020-2021 aylarında incelemeler yapılmıştır. Hava kökenli mikrobiyotanın belirlenmesi için besi yerleri paketleme tesislerinin 3 farklı bölümüne (ürün kabul, ürün işleme ve ürün çıkışı) yerleştirilmiştir. Tesislerdeki meyve kökenli mikrobiyotanın belirlenmesi için, güdümlü olarak seçilen meyvelerden (bozulma, yumuşama, çürüme vb. gözlenen) ayrıca fungal ve bakteriyel izolatlar elde edilmiştir. Elde edilen fungal ve bakteriyel izolatların morfolojik, biyokimyasal ve MALDI-TOF proteomik yöntemler ile tanılanmışlardır. Yapılan izolasyonlar ve tanılamalar sonucunda Penicillium italicum, Penicillium digitatum, Aspergillus niger, Rhizopus oryzae, Mucor circinelloides, Xanthomonas hortorum, Pantoea eucrina, Leclercia adecarboxylata, Pseudescherichia vulneris, Bacillus megaterium, Lelliottia amnigena, Raoultella ornithinolytica, Klebsiella oxytoca, Bacillus subtilis, Erwinia herbicola hava kökenli fungal ve bakteriyel türler olarak belirlenmiştir. Penicillium digitatum, Penicillium italicum, Fusarium solani, Geotrichum citri-auranti, Alternaria alternata, Cladosporium herbarum, Phytophthora sp., Colletotrichum gloeosporioides, Geotrichum candidum, Erwinia herbicola, Xanthomonas hortorum ise enfekteli meyve kökenli fungal ve bakteriyel türler olarak belirlenmiştir.

Genel Yorum: Hava kökenli mikrobiyotanın paketleme tesislerinin ürün işleme bölümünde oldukça yoğun olduğu, ürün kabul (giriş) ve sevk (çıkış) alanlarında ise uygun hava sirkülasyonundan dolayı yoğunluğun az olduğu gözlenmiştir.

Çalışmanın Önemi ve Etkisi: Paketleme tesislerindeki hava kökenli fungal ve bakteriyel mikrobiyotanın hava hareketleri ile kolayca taşınabilmesi ve havada belli süre askıda kalması nedeniyle meyvelerde bozulma ve çürüme gibi belirtilerin çıkmasına neden olmaktadır. Tanılaması yapılan hava kökenli bazı bakteri türlerinin alerjen veya insan patojeni olmaları nedeniyle paketleme evlerine alınması gereken sanitasyon önlemleri hususunda gerekli bilgilendirmeler yapılmıştır.

Keywords

• Turunçgil
• paketleme tesi
• hasat sonu hastalıklar
• MALDI-TOF
• hava/meyve kökenli mikrobiyota

References

1. Akhtar N, Anjum T, Jabeen R (2013) Isolation and identification of storage fungi from citrus sampled from major growing areas of Punjab, Pakistan. Int. J. Agric. Biol. 15:1283-1288.
2. AKIB (2018) Akdeniz Yaş Meyve Sebze İhracatçıları Birliği, 2018 Çalışma Raporu. https://www.akib.org.tr/download/files/images/2019/1.%20Genel/2018%20yms%20%C3%A7al%C4%B1%C5%9Fma%20raporu.pdf (Erişim tarihi: 28.01.2022).
3. Akkoyun Bilgi A, Kiraz N (2019) Klinik Örneklerden İzole Edilen Aspergillus Türlerinin Tanımlanmasında Geleneksel Yöntemler, MALDI-TOF MS ve Dizi Analizi Yöntemlerinin Karşılaştırılması. Dicle Tıp Derg. 46 (3): 543 – 551.
4. Aktan Z, Soylu S(2020) Diyarbakır İlinde Yetişen Badem Ağaçlarından Endofit ve Epifit Bakteri Türlerinin İzolasyonu ve Bitki Gelişimini Teşvik Eden Mekanizmalarının Karakterizasyonu. KSÜ Tarım ve Doğa Derg. 23 (3): 641-654.
5. Anonim (2021) TUİK Bitkisel Üretim İstatistikleri. https://biruni.tuik.gov.tr/medas/?kn=92&locale=tr (Erişim Tarihi: 28.01.2022)
6. Anonymous (2020) FAOSTAT, Crops and livestock products. https://www.fao.org/faostat/en/#data/QCL/visualize (Erişim tarihi: 28.01.2022).
7. Bazioli JM, Belinato JR, Costa JH, Akiyama DY, de Moraes Pontes JG, Kupper KC, Augusto F, de Carvalho JE, Fill TP (2019) Biological control of citrus postharvest phytopathogens. Toxins (Basel) 11(8): 460.
8. Benli M (2003) Hasat Sonrası Fungal Hastalıklarla Kimyasal ve Biyolojik Mücadele. Orlab On-Line Mikrobiyoloji Derg. 1: 1-25.

9. Carolis ED, Posteraro B, Lass-Flo C, Vella A, Florio AR, Torelli R, Girmenia C, Colozza C,Tortorano AM, Sanguinetti M, Fadda G (2012) Species identification of Aspergillus, Fusarium and Mucorales with direct surface analysis by Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry. Clinic. Microbiol. Infect. 18: 475–484.
10. Coates L, Johnson G (1989) Postharvest Diseases of Fruit and Vegetables. https://www.appsnet.org/Publications/Brown_Ogle/33%20Postharvest%20diseases%20(LMC&GIJ).pdf, pp 533-548.
11. Deng B, Wang WH, Deng LL, Yao SX, Ming J, Zeng KF (2018) Comparative RNA-seq analysis of citrus fruit in response to infection with three major postharvest fungi. Postharvest Biol. Technol., 146: 134-146. Dugan FM (2006) The Identification of Fungi: An Illustrated Introduction With Keys, Glossary, And Guide to Literature. APS Press, St Paul, MN. 184 pp.
12. El-Otmani M, Ait-OubahouL A, Zacarías L (2011) Citrus spp.: orange, mandarin, tangerine, clementine, grapefruit, pomelo, lemon and lime. In: Postharvest Biology and Technology of Tropical and Subtropical Fruits, (Eds. EM Yahia), Woodhead Publishing Limited, pp 437-514.
13. Fischer IH, Lourenço SA, Spósito MB, Amorim L (2009) Characterisation of the fungal population in citrus packing houses. Eur. J. Plant Pathol. 123:449–460.
14. Gomes AAM, Queiroz MV, Pereira OL (2015) Mycofumigation for the biological control of post-harvest diseases in fruits and vegetables: A review. Austin. J. Biotechnol. Bioeng. 2:1051.
15. Hocking AD (2014) Spoilage Problems: Problems Caused by Fungi. Encyclopedia of Food Microbiology, Second Edition (Eds. CA Batt, ML Tortorello), Academic Press, pp 471-481.
16. Ismail M, Zhang J (2004) Post–harvest citrus diseases and their control. Outlooks Pest. Manag. 15: 29–35. Jayasekara A, Daranagama A, Kodituwakku TD, Abeywickrama K (2022) Morphological and molecular identification of fungi for their association with postharvest fruit rots in some selected citrus species. J. Agricultural Sci. 17: 79-93. Kanashiro AM, Akiyama DY, Kupper KC, Fill TP (2020) Penicillium italicum: An Underexplored Postharvest Pathogen. Front. Microbiol. 11: 606852.
17. Khamis Y, Hashim AF, Margarita R, Alghuthaymi MA, Abd-Elsalam KA (2017) Fungicidal efficacy of chemically-produced copper nanoparticles against Penicillium digitatum and Fusarium solani on Citrus Fruit. Philipp. Agric. Sci. 100: 69-78.
18. Klein JD, Lurie S(1991) Postharvest heat treatment and fruit quality. Postharvest News Information 2: 15-19.
19. Ladaniya MS (2008) Postharvest Diseases and Their Management. In: Citrus Fruit: Biology, Technology and Evaluation (Ed. M Ladaniya), Academic Press, London, pp 417-449.
20. Li Y, Wang H, Zhao YP, Xu YC, Hsueh, PR, (2017) Evaluation of the Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry System for Identification of Aspergillus Species Directly from Growth on Solid Agar Media. Front Microbiol. 8: 1209.
21. Lugauskas A, Krikstaponis A, Sveistyte L (2004) Airborne fungi in industrial environments--potential agents of respiratory diseases. Ann. Agric. Environ. Med. 11(1): 19-25.
22. Maldonado MC, Corona J, Gordillo MA, Navarro AR (2009) Isolation and partial characterization of antifungal metabolites produced by Bacillus sp. IBA 33. Curr. Microbiol. 59: 646–650.
23. McKay AH, Förster H, Adaskaveg JE (2012) Efficacy and application strategies for propiconazole as a new postharvest fungicide for managing sour rot and green mold of citrus fruit. Plant Dis. 96: 235-242.
24. Palou L (2014) Penicillium digitatum, Penicillium italicum (Green Mold, Blue Mold). In: Postharvest Decay, (Ed. S Bautista-Banos), Academc Press, London. pp 45-102.
25. Palou L, Smilanick JL, Droby S (2008) Alternatives to conventional fungicides for the control of citrus postharvest green and blue moulds. Stewart Postharvest Rev. 2: 2-15.
26. Palumbo JD, Baker JL, Mahoney NE (2006) Isolation of bacterial antagonists of Aspergillus flavus almonds. Microbial. Ecol. 52: 45–52.
27. Pao S, Brown GE (1998) Reduction of Microorganisms on Citrus Fruit Surfaces during Packinghouse Processing. J. Food Protect. 61: 903-906.
28. Prusky D (2011) Reduction of the incidence of postharvest quality losses, and future prospects. Food Secur. 3: 463–474.
29. Saito S, Xiao CL (2017) Prevalence of postharvest diseases of mandarin fruit in California. Plant Health Prog. 18: 204-210.
30. Samson RA, Pitt JL (2000) Integration of Modern Taxonomic Methods for Penicillium and Aspergillus Classification. CRC Press, Amsterdam, The Netherlands. 554 pp.
31. Schipper MAA (1978) On certain species of Mucor with a key to all accepted species. Stud. Mycol. 17:53-71.
32. Simmons EG (2007) Alternaria: An Identification Manuel. CBS Biodiversity Series Vol. 6. CBS Fungal Biodiversity Centre, Utrecht, The Netherlands. 780 pp.
33. Simsekli Y, Gücin F, Asan A (1999). Isolation and identification of indoor airborne fungal contaminants of food production facilities and warehouses in Bursa, Turkey. Aerobiologia 15: 225–231.
34. Snowdon A (1990) A Colour Atlas of Post-Harvest Diseases and Disorders of Fruits and Vegetables, Vol. 1: General Introduction and Fruits, Wolfe Scientific, London, 302 pp.
35. Strōm G, Blomquıst G (1986) Airborne Spores From Mouldy Citrus Fruit— A Potential Occupational Health Hazard. Ann. Occup. Hyg. 30: 455–460.
36. Subaşı OS, Uysal O, Ünlü M (2014) Mersin ili turunçgil ihracatı yapan paketleme tesislerinin yaşadıkları sorunlar ve çözüm önerileri. Alatarım 13: 37-43.
37. Uysal A, Kurt Ş, (2018) An Important Fungal Disease on Citrus Orchards in Erdemli: Anthracnose. International Erdemli Symposium, April 19-21, Mersin, Turkey. p. 670.
38. Uysal A, Kurt Ş, Akgül DS (2016) Akdeniz Bölgesi Limon Bahçelerinde Antraknoz Hastalığına Neden olan Colletotrichum gloeosporioides' in Patojenik ve Moleküler Karakterizasyonu. Uluslararası Katılımlı Türkiye VI. Bitki Koruma Kongresi, 5-8 Eylül, Konya, Türkiye. s. 597.
39. Uysal A, Kurt Ş, Soylu S, Soylu EM, Kara M (2019) Yaprağı Yenen Sebzelerdeki Mikroorganizma Türlerinin MALDI-TOF MS (Matris Destekli Lazer Desorpsiyon/İyonizasyon Uçuş Süresi Kütle Spektrometresi Tekniği Kullanılarak Tanılanması. YYÜ Tar. Bil. Derg., 29: 595-601.
40. Wang W, Liu S, Deng L, Ming J, Yao S, Zeng K (2018) Control of citrus post-harvest green molds, blue molds, and sour rot by the Cecropin A-Melittin hybrid peptide BP21. Front. Microbiol. 9: 2455.
41. Yiğiter B (2013) Natamisinin turunçgillerde küflenmeye karşı kullanılması. Yüksek Lisans Tezi, Süleyman Demirel Üniversitesi, Fen Bilimleri Enstitüsü, Gıda Mühendisliği ABD, 96 sayfa. Yıldız AO (2014) Hatay İli Soğuk Hava Depolarının Mevcut Durumu ve Sorunları. Yüksek Lisans Tezi, Mustafa Kemal Üniversitesi, Fen Bilimleri Enstitüsü, Bahçe Bitkileri ABD, 93 sayfa.
42. Yıldız F, Yıldız M, Kınay P, Delen N(2002) Altıntoplarda hasat sonrası Penicillium çürüklerinin biyolojik ve kimyasal kontrolü üzerinde incelemeler. II. Bahçe Ürünlerinde Pazarlama ve Muhafaza Sempozyumu, 24-27 Eylül 2002, Çanakkale, Bildirileri 278-284.
43. Zaheer I, Iftikhar S, Khurshid T, Ahmad KS, Gul MM (2019) Isolation and ITS-rDNA based molecular characterization of plant pathogenic fungal species in postharvest citrus fruits. Sydowia 71: 267-278.