Evaluation of the Presence of Yeasts, Moulds and Aflatoxin M1 in Raw Sheep Milk in Mazandaran, Iran

Document Type : Research Paper


1 Department of Food Hgiene, Faculty of Veterinary Medicine, Amol University of Special Modern Technologies, Amol, Iran.

2 Department of Pathobiology, Faculty of Veterinary Medicine, Amol University of Special Modern Technologies, Amol, Iran.


Introduction: Aflatoxins are among the most important fungal mycotoxins with carcinogenic properties. Aflatoxin M1 can be transmitted to humans through milk and dairy products. This study proposed to appraise the fungal and aflatoxin M1 contaminations in sheep milk collcted from different farms in Mazandaran province. Methods: During July to September 2020, 70 samples of raw milk from sheep were collected in disparate regions of Northern Iran. All of the samples were analyzed in order to presence of fungal agents by culturing the raw milk onto Sabouraud dextrose agar (SDA) as well as AFM1 by competitive enzyme-linked immunosorbent assay (ELISA) technique. Results: Of the fungal agents identified, the most frequently moulds were related to Aspergillus spp. (38.8%), Cladosporium spp. (22.4%) and Penicillium spp. (13.9%); and yeasts dependent on Trichosporon spp. (47.1 %), Rhodotorula spp. (33.8%) and Candida spp. (14.7%). Of the 70 milk samples, all samples (100%) had AFM1 at condensation lower than the high level specified in European Union (EU) regulations, i.e., 50 ng/L.  The AFM1 contamination levels ranged from 1.15 to 48.50 ng/L with the average of 13.26 ± 4.87 ng/L. Conclusions: Contamination of sheep milk with mycotoxins, especially AFM1, can be a potential risk for the consumer; therefore, raw sheep milk should be checked for these toxins.


  1. Mollayusefian I, Ranaei V, Pilevar Z, Cabral-Pinto MM, Rostami A, Nematolahi A, Khedher KM, Fakhri Y, Khaneghah AM. The concentration of aflatoxin M1 in raw and pasteurized milk: A worldwide systematic review and meta-analysis. Trends Food Sci Technol. 2021;115:22-30.
  2. Bilandžić N, Varenina I, Solomun Kolanović B, Božić Luburić Đ, Varga I, Želježić B, Cvetnić L, Benić M, Tanković S, Cvetnić Ž. Occurrence of aflatoxin M1 in raw cow, goat and sheep milk during spring and autumn in Croatia during 2016. Toxin Reviews. 2017;36(4):290-6.
  3. Min L, Fink-Gremmels J, Li D, Tong X, Tang J, Nan X, Yu Z, Chen W, Wang G. An overview of aflatoxin B1 biotransformation and aflatoxin M1 secretion in lactating dairy cows. Animal Nutrition. 2021;7(1):42-8.
  4. Abdel-Azeem AM, Abdel-Azeem MA, Abdul-Hadi SY, Darwish AG. Aspergillus: Biodiversity, ecological significances, and industrial applications. Recent Advancement in White Biotechnology through Fungi. Springer, Cham.2019; 121-79.
  5. Marchese S, Polo A, Ariano A, Velotto S, Costantini S, Severino L. Aflatoxin B1 and M1: Biological properties and their involvement in cancer development. Toxins. 2018;10(6):214.
  6. Cammilleri G, Graci S, Collura R, Buscemi MD, Vella A, Macaluso A, Giaccone V, Giangrosso G, Cicero A, Lo Dico GM, Pulvirenti A. Aflatoxin M1 in cow, sheep, and donkey milk produced in Sicily, Southern Italy. Mycotoxin Res. 2019;35(1):47-53.
  7. Van Asselt ED, Van der Fels‐Klerx HJ, Marvin HJ, Van Bokhorst‐van de Veen H, Groot MN. Overview of food safety hazards in the European dairy supply chain. Compr Rev Food Sci Food Saf. 2017;16(1):59-75.
  8. European Union. Laying down themethods of sampling and analysis for theofficial control of the levels of mycotoxins in foodstuffs. Official J Eur Union L. 2006; 70: 20-3.
  9. Min L, Li D, Tong X, Sun H, Chen W, Wang G, Zheng N, Wang J. The challenges of global occurrence of aflatoxin M1 contamination and the reduction of aflatoxin M1 in milk over the past decade. Food Control. 2020;117:107352.
  10. Waqas M, Pervaiz W, Zia KM, Iqbal SZ. Assessment of aflatoxin B1 in animal feed and aflatoxin M1 in raw milk samples of different species of milking animals from Punjab, Pakistan. J Food Saf. 2021;41(3):e12893.
  11. Caboni P, Murgia A, Porcu A, Manis C, Ibba I, Contu M, Scano P. A metabolomics comparison between sheep's and goat's milk. Int Food Res J. 2019;119:869-75.
  12. Hashemi M. A survey of aflatoxin M1 in cow milk in Southern Iran. J Food Drug Anal. 2016;24(4):888-93.
  13. Ansari F, Pourjafar H, Christensen L. A study on the aflatoxin M1 rate and seasonal variation in pasteurized cow milk from northwestern Iran. Environ Monit Assess. 2019;191(1):1-6.
  14. Hajmohammadi M, Valizadeh R, Naserian A, Nourozi ME, Rocha RS, Oliveira CA. Composition and occurrence of aflatoxin M1 in cow's milk samples from Razavi Khorasan Province, Iran. Int J Dairy Technol. 2020;73(1):40-5.
  15. Spanamerg A, Fraga CF, Ferreiro L, Aguinsky MS, Sanches EMC, Roehe C, Lautert C, and Santurio JM. Yeasts in the raw ewe’s milk. Acta Sci Vet, 2014;42: 1-5.
  16. Barnett JA, Payne RW, Yarrow D. Characteristics and identification. United States. 2000; 3: 658.
  17. Delavenne E, Mounier J, Asmani K, Jany JL, Barbier G, Le Blay G. Fungal diversity in cow, goat and ewe milk. Int JFood Microbiol. 2011; 151(2):247-51.
  18. Panelli S, Brambati E, Bonacina C, Feligini M. Updating on the fungal composition in Sardinian sheep's milk by culture-independent methods. J Dairy Res. 2014;81(2):233-7.
  19. Callon C, Duthoit F, Delbès C, Ferrand M, Le Frileux Y, De Crémoux R, Montel MC. Stability of microbial communities in goat milk during a lactation year: molecular approaches. Syst Appl Microbiol. 2007;30(7):547-60.
  20. O'Brien M, O'Kiely P, Forristal PD, Fuller HT. Fungi isolated from contaminated baled grass silage on farms in the Irish Midlands. FEMS Microbiology Letters. 2005;247(2):131-5.
  21. Vacheyrou M, Normand AC, Guyot P, Cassagne C, Piarroux R, Bouton Y. Cultivable microbial communities in raw cow milk and potential transfers from stables of sixteen French farms. Int JFood Microbiol. 2011;146(3):253-62.
  22. Njage PM, Dolci S, Jans C, Wangoh J, Lacroix C, Meile L. Characterization of yeasts associated with camel milk using phenotypic and molecular identification techniques. Res J Microbiol. 2011;6(9):678.
  23. Movassagh MH. Presence of Aflatoxin M1 Ewe’s Milk in the Northwest Region of Iran. J Appl Biol Sci. 2009;3(3):17-9.
  24. Bahrami R, Shahbazi Y, Nikousefat Z. Aflatoxin M1 in milk and traditional dairy products from west part of Iran: occurrence and seasonal variation with an emphasis on risk assessment of human exposure. Food Control. 2016;62:250-6.
  25. Malissiova E, Tsakalof A, Arvanitoyannis IS, Katsafliaka A, Katsioulis A, Tserkezou P, Koureas M, Govaris A, Hadjichristodoulou C. Monitoring Aflatoxin M1 levels in ewe's and goat's milk in Thessaly, Greece; potential risk factors under organic and conventional production schemes. Food Control. 2013;34(1):241-8.
  26. Armorini S, Altafini A, Zaghini A, Roncada P. Occurrence of aflatoxin M1 in conventional and organic milk offered for sale in Italy. Mycotoxin Res. 2016;32(4):237-46.
  27. Rubio R, Licón CC, Berruga MI, Molina MP, Molina A. Occurrence of aflatoxin M1 in the Manchego cheese supply chain. J Dairy Sci. 2011;94(6):2775-8.
  28. Bognanno M, La Fauci L, Ritieni A, Tafuri A, De Lorenzo A, Micari P, Di Renzo L, Ciappellano S, Sarullo V, Galvano F. Survey of the occurrence of Aflatoxin M1 in ovine milk by HPLC and its confirmation by MS. Mol Nutr Food Res. 2006;50(3):300-5.
  29. Virdis S, Scarano C, Spanu V, Muritto G, Spanu C, Ibba I, De Santis EP. A survey on aflatoxin M1content in sheep and goat milk produced in Sardinia region, Italy (2005-2013). Ital J Food Saf. 2014; 3(4): 4517.
  30. Bilandžić N, Božić Đ, Đokić M, Sedak M, Kolanović BS, Varenina I, Tanković S, Cvetnić Ž. Seasonal effect on aflatoxin M1 contamination in raw and UHT milk from Croatia. Food Control. 2014;40:260-4.
  31. Hassan HF, Kassaify Z. The risks associated with aflatoxins M1 occurrence in Lebanese dairy products. Food Control. 2014;37:68-72.
  32. Hussain I, Anwar J, Asi MR, Munawar MA, Kashif M. Aflatoxin M1 contamination in milk from five dairy species in Pakistan. Food Control. 2010;21(2):122-4.
  33. Ayhan F, Sinan I, Fusun T. Survey of the occurrence of aflatoxin M1 in cheeses produced by dairy ewe’s milk in Urfa city, Turkey. Ankara Üniv. Vet Fakültesi Dergisi, 2010; 57: 197-9.
  34. Molle G, Decandia M, Cabiddu A, Landau SY, Cannas A. An update on the nutrition of dairy sheep grazing Mediterranean pastures. Small Ruminant Res. 2008; 77: 93-112.
  35. Tajkarimi M, Aliabadi FS, Nejad MS, Pursoltani H, Motallebi AA, Mahdavi H. Seasonal study of aflatoxin M1 contamination in milk in five regions in Iran. Int JFood Microbiol . 2007;116(3):346-9.