Mycobiota and mycotoxins in Portuguese pork, goat and sheep dry-cured hams

  • Paula RodriguesEmail author
  • Diana Silva
  • Pedro Costa
  • Luís Abrunhosa
  • Armando Venâncio
  • Alfredo Teixeira
Original Article


The objectives of the present work were to survey, for the first time, the contamination of Portuguese fresh and dry-cured meat products with ochratoxin A (OTA) and aflatoxin B1 (AFB1), and to determine the fungi potentially responsible for this contamination. A total of 128 samples including pork fresh legs, dry-cured legs and shoulders, as well as goat and sheep dry-cured legs were analysed. Mycological analysis of these samples yielded a total of 630 fungal isolates. Penicillium sp. was the dominant fungal genus in all products (66% of all isolates). Penicillium nordicum and Aspergillus westerdijkiae were only rarely isolated from pork ham samples. In fresh pork meat, 40% of the samples were contaminated with OTA at levels below 1 μg/kg. In pork dry-cured legs with 20 to 25 months of ripening, 43% of the samples showed detectable contamination, while 18% of the shoulder hams were contaminated. OTA was not detected in any of the goat and sheep samples. OTA contamination does not seem to be a risk in small-piece and short-ripe products like goat and sheep legs, but affects longer ripe products like pork legs and shoulders. Although aflatoxigenic fungi were identified, AFB1 was not detected in any sample, and it should not be considered a risk in dry-cured hams.


Penicillium Aspergillus Ochratoxin A Aflatoxins Food safety Meat products 


Funding information

Work was funded by the Portuguese PRODER research Project number 020260013013 “New goat and sheep processed meat products” and BISOVICAP—Processing meat from pigs, sheep and goats, to produce new products. Ham and pâté, Project PROTEC, SI I&DT—Projects in Co-Promotion, no. 21511. The authors received support from the Laboratory of Carcass and Meat Quality of Agriculture School of Polytechnic Institute of Bragança ‘Cantinho do Alfredo’ and from MARCARNE network, funded by CYTED (ref. 116RT0503). The authors also received financial support from the Foundation for Science and Technology (FCT, Portugal) and FEDER under Programme PT2020 to CIMO (UID/AGR/00690/2019). CEB also received support from FCT under the scope of the strategic funding of UID/BIO/04469/2019 unit, and BioTecNorte operation (NORTE-01-0145-FEDER-000004), funded by the European Regional Development Fund under the scope of Norte2020—Programa Operacional Regional do Norte.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Alapont C, López-Mendoza MC, Gil JV, Martínez-Culebras PV (2014) Mycobiota and toxigenic Penicillium species on two Spanish dry-cured ham manufacturing plants. Food Addit Contam part A 31:93–104CrossRefGoogle Scholar
  2. Alves A, Phillips AJL, Henriques I, Correia A (2007) Rapid differentiation of species of Botryosphaeriaceae by PCR fingerprinting. Res Microbiol 158:112–121CrossRefGoogle Scholar
  3. AOAC (1990) AOAC official methods of analysis (15th ed.). Washington, DC: Association of Official Analytical ChemistsGoogle Scholar
  4. Asefa DT, Gjerde RO, Sidhu MS, Langsrud S, Kure CF, Nesbakken T, Skaar I (2009) Moulds contaminants on Norwegian dry-cured meat products. Int J Food Microbiol 128:435–439CrossRefGoogle Scholar
  5. Battilani P, Pietri VA, Giorni P, Formenti S, Bertuzzi T, Toscani T, Virgili R, Kozakiewicz Z (2007) Penicillium populations in dry-cured ham manufacturing plants. J Food Prot 70:975–980CrossRefGoogle Scholar
  6. Bogs C, Battilani P, Geisen R (2006) Development of a molecular detection and differentiation system for ochratoxin A producing Penicillium species and its application to analyse the occurrence of Penicillium nordicum in cured meats. Int J Food Microbiol 107:39–47CrossRefGoogle Scholar
  7. Canel RS, Wagner JR, Stenglein SA, Ludemann V (2013) Indigenous filamentous fungi on the surface of Argentinean dry fermented sausages produced in Colonia Caroya (Córdoba). Int J Food Microbiol 164:81–86CrossRefGoogle Scholar
  8. Chiavaro E, Lepiani A, Colla F, Bettoni P, Pari E, Spotti E (2002) Ochratoxin A determination in ham by immunoaffinity clean-up and a quick fluorometric method. Food Addit Contam 19:575–581CrossRefGoogle Scholar
  9. Comi G, Orlic S, Redzepovic S, Urso R, Iacumin L (2004) Moulds isolated from Istrian dried ham at the pre-ripening and ripening level. Int J Food Microbiol 96:29–34CrossRefGoogle Scholar
  10. Dall’Asta C, Galaverna G, Bertuzzi T, Noseriti A, Pietri A, Dossena A, Marchelli R (2010) Occurrence of ochratoxin A in raw ham muscle, salami and dry-cured ham from pigs fed with contaminated diet. Food Chem 120:978–983CrossRefGoogle Scholar
  11. EC (2006a) Commission Regulation (EC) No 401/2006 of 23 February laying down the methods of sampling and analysis for the official control of the levels of mycotoxins in foodstuffs. Off J Eur Union L70/12-L70-34Google Scholar
  12. EC (2006b) Commission Regulation (EC) No 1881/2006 of 19 December 2006, setting maximum levels for certain contaminants in foodstuffs. Off J Eur Union L364/5-L364-24Google Scholar
  13. EFSA-Q-2003-039 (2004) Opinion of the scientific panel on contaminants in food chain on a request from the commission related to ochratoxin A (OTA) as undesirable substance in animal feed. EFSA (European Food Safety Authority). EFSA J 101:1–36Google Scholar
  14. Ferrara M, Perrone G, Gallo A, Epifani F, Visconti A, Susca A (2015) Development of loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Penicillium nordicum in dry-cured meat products. Int J Food Microbiol 202:42–47CrossRefGoogle Scholar
  15. Frisvad JC, Samson RA (2004) Polyphasic taxonomy of Penicillium subgenus Penicillium — a guide to identification of food and air-borne terverticillate Penicillia and their mycotoxins. Stud Mycol 49:1–173Google Scholar
  16. Iacumin L, Chiesa L, Boscolo D, Manzano M, Cantoni C, Orlic S, Comi G (2009) Moulds and ochratoxin A on surfaces of artisanal and industrial dry sausages. Food Microbiol 26:65–70CrossRefGoogle Scholar
  17. Iacumin L, Milesi S, Pirani S, Comi G, Chiesa LM (2011) Ochratoxigenic mold and ochratoxin a in fermented sausages from different areas in northern Italy: occurrence, reduction or prevention with ozonated air. J Food Saf 31:538–545CrossRefGoogle Scholar
  18. IARC (1993) Some naturally occurring substances: food items and constituents, heterocyclic aromatic amines and mycotoxins. In: International Agency for Research on Cancer, World Health Organization (ed) IARC monographs on the evaluation of carcinogenic risks to humans. Lyon, France: IARC Press 56:599Google Scholar
  19. Klich MA (2002) Identification of common Aspergillus species. CBS, The NetherlandsGoogle Scholar
  20. Larsen TO, Svendsen A, Smedsgaard J (2001) Biochemical characterization of ochratoxin A-producing strains of the genus Penicillium. Appl Environ Microbiol 67:3630–3363CrossRefGoogle Scholar
  21. Ludemann V, Pose G, Pollio ML, Segura J (2004) Determination of growth characteristics and lipolytic and proteolytic activities of Penicillium strains isolated from Argentinean salami. Int J Food Microbiol 96:13–18CrossRefGoogle Scholar
  22. Markov K, Pleadin J, Bevardi M, Vahčić N, Sokolić-Mihalak D, Frece J (2013) Natural occurrence of aflatoxin B1, ochratoxin A and citrinin in Croatian fermented meat products. Food Control 34:312–317CrossRefGoogle Scholar
  23. Martín A, Córdoba JJ, Núñez F, Benito MJ, Asensio MA (2004) Contribution of a selected fungal population to proteolysis on dry-cured ham. Int J Food Microbiol 94:55–66CrossRefGoogle Scholar
  24. Martín A, Córdoba JJ, Aranda E, Córdoba MG, Asensio MA (2006) Contribution of a selected fungal population to the volatile compounds on dry-cured ham. Int J Food Microbiol 110:8–18CrossRefGoogle Scholar
  25. Meftah S, Abid S, Dias T, Rodrigues P (2018) Effect of dry-sausage starter culture and endogenous yeasts on Aspergillus westerdijkiae and Penicillium nordicum growth and OTA production. LWT Food Sci Technol 87:250–258CrossRefGoogle Scholar
  26. Merla C, Andreoli G, Garino C, Tosi G, Guglielminetti ML, Moretti A, Biancardi A, Arlorio M, Fabbi M (2018) Monitoring of ochratoxin A and ochratoxin- producing fungi in traditional salami manufactured in Northern Italy. Mycotox Res 34:107–116CrossRefGoogle Scholar
  27. Ministero della Sanità (1999) Circolare 09.06.1999. In M. D. Sanità (Ed.), Circolare 09.06.1999 (vol. 135) 11.06.1999. Gazzetta Ufficiale Repubblica ItalianaGoogle Scholar
  28. Montanha FP, Anater A, Burchard JF, Luciano FB, Meca G, Manyes L, Pimpão CT (2018) Mycotoxins in dry-cured meats: a review. Food Chem Toxicol 111:494–502CrossRefGoogle Scholar
  29. Nuñez F, Rodríguez MM, Bermúdez ME, Córdoba JJ, Asensio MA (1996) Composition and toxigenic potential of the mold population on dry-cured Iberian ham. Int J Food Microbiol 32:185–197CrossRefGoogle Scholar
  30. Nuñez F, Westphal CD, Bermúdez E, Asensio MA (2007) Production of secondary metabolites by some terverticillate penicillia on carbohydrate-rich and meat substrates. J Food Prot 70:2829–2836CrossRefGoogle Scholar
  31. Paulos K, Rodrigues S, Oliveira AF, Leite A, Pereira E, Teixeira A (2015) Sensory characterization and consumer preference mapping of fresh sausages manufactured with goat and sheep meat. J Food Sci 80:S1568–S1573CrossRefGoogle Scholar
  32. Pereira E, Barros L, Antonio AL, Cabo Verde S, Santos-Buelga C, Ferreira ICFR, Rodrigues P (2017) Is gamma radiation suitable to preserve phenolic compounds and to decontaminate mycotoxins in aromatic plants? A case-study with Aloysia citrodora Paláu. Molecules 22(3):347CrossRefGoogle Scholar
  33. Petzinger E, Ziegler K (2000) Ochratoxin A from a toxicological perspective. J Vet Pharmacol Ther 23:91–98CrossRefGoogle Scholar
  34. Pitt JI, Hocking AD (2009) Fungi and food spoilage, 3rd edn. Springer 519 ppGoogle Scholar
  35. Pleadin J, Perši N, Kovačevic D, Vahčić N, Scortichini G, Milone S (2013) Ochratoxin A in traditional dry-cured meat products produced from sub-chronic-exposed pigs. Food Addit Contam part A 30:1827–1836CrossRefGoogle Scholar
  36. Pleadin J, Staver MM, Vahčić N, Kovačevic D, Milone S, Saftić L, Scortichini G (2015) Survey of aflatoxin B1 and ochratoxin A occurrence in traditional meat products coming from Croatian households and markets. Food Control 52:71–77CrossRefGoogle Scholar
  37. Rodrigues P, Santos C, Venâncio A, Lima N (2011) Species identification of Aspergillus section Flavi isolates from Portuguese almonds using phenotypic, including MALDI-TOF ICMS, and molecular approaches. J Appl Microbiol 111:877–892CrossRefGoogle Scholar
  38. Rodrigues P, Venâncio A, Lima N (2013) Incidence and diversity of the fungal genera Aspergillus and Penicillium in Portuguese almonds and chestnuts. Eur J Plant Pathol 137:197–209CrossRefGoogle Scholar
  39. Rodrigues P, Venâncio A, Lima N (2018) Toxic reagents and expensive equipment: are they really necessary for the extraction of good quality fungal DNA? Lett Appl Microbiol 66:32–37CrossRefGoogle Scholar
  40. Rodríguez A, Rodríguez M, Martín A, Nuñez F, Córdoba JJ (2012a) Evaluation of hazard of aflatoxin B1, ochratoxin A and patulin production in dry-cured ham and early detection of producing molds by pPCR. Food Control 27:118–126CrossRefGoogle Scholar
  41. Rodríguez A, Rodríguez M, Martín A, Delgado J, Córdoba JJ (2012b) Presence of ochratoxin A on the surface of dry-cured Iberian ham after initial fungal growth in the drying stage. Meat Sci 92:728–734CrossRefGoogle Scholar
  42. Samson RA, Houbraken J (eds.) (2011) Phylogenetic and taxonomic studies on the genera Penicillium and Talaromyces. Stud Mycol 70:183Google Scholar
  43. Samson RA, Varga J (eds.) (2007) Aspergillus systematics in the genomic era. Stud Mycol 59:206Google Scholar
  44. Samson RA, Hoekstra ES, Frisvad JC (2004a) Identification of the common food- and airborne fungi. In: Samson RA, Hoekstra ES, Frisvad JC (eds) Introduction to food- and airborne fungi, 7th edn. CBS, The Netherlands, pp 1–282Google Scholar
  45. Samson RA, Hoekstra ES, Lund F, Filtenborg O, Frisvad JC (2004b) Methods for the detection, isolation and characterisation of food-borne fungi. In: Samson RA, Hoekstra ES, Frisvad JC (eds) Introduction to food- and airborne fungi, 7th edn. CBS, The Netherlands, pp 283–297Google Scholar
  46. Samson RA, Varga J, Frisvad JC (eds) (2011) Taxonomic studies on the genus Aspergillus. Stud Mycol 69:97Google Scholar
  47. Schmidt-Heydt M, Abdel-Hadi A, Magan N, Geisen R (2009) Complex regulation of the aflatoxin biosynthesis gene cluster of Aspergillus flavus in relation to various combinations of water activity and temperature. Int J Food Microbiol 135:231–237CrossRefGoogle Scholar
  48. Sonjak S, Ličen M, Frisvald JC, Gunde–Cimerman N (2011) The mycobiota of three dry-cured meat products from Slovenia. Food Microbiol 28:373–376CrossRefGoogle Scholar
  49. Sørensen LM, Jacobsen T, Nielsen PV, Frisvad JC, Koch AG (2008) Mycobiota in the processing areas of two different meat products. Int J Food Microbiol 124:58–64CrossRefGoogle Scholar
  50. Teixeira A, Fernandes A, Pereira E, Manuel A, Rodrigues S (2017) Effect of salting and ripening on the physicochemical and sensory quality of goat and sheep cured legs. Meat Sci 134:163–169CrossRefGoogle Scholar
  51. Vipotnik Z, Rodríguez A, Rodrigues P (2017) Aspergillus westerdijkiae as a major ochratoxin A risk in dry-cured ham based-media. Int J Food Microbiol 241:244–251CrossRefGoogle Scholar

Copyright information

© Society for Mycotoxin (Research Gesellschaft für Mykotoxinforschung e.V.) and Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Centro de Investigação de Montanha (CIMO), ESA, Instituto Politécnico de BragançaCampus de Santa ApolóniaBragançaPortugal
  2. 2.CEB – Centre for Biological EngineeringUniversidade do MinhoBragaPortugal

Personalised recommendations