Application of PCR-DGGE to the study of dynamics and biodiversity of yeasts and potentially OTA producing fungi during coffee processing

Introduction: Ochratoxin A (OTA) is a toxic secondary metabolite produced by fungi of the genera Aspergillus and Penicillium. It has been shown to have carcinogenic and immunotoxic properties in rats and to be responsible for human and animal kidney pathologies. OTA content in coffee was shown to be closely link to harvesting conditions, postharvest processing conditions and especially dry processing, storage and transportation conditions. Purpose: Fairly little is known about the conditions for contamination by fungi responsible for OTA production, their propagation and conditions for OTA production. Biodiversity and dynamics of fungal populations linked to OTA production could be studied by PCR-DGGE genetic fingerprinting with the aim to understand the effects of postharvest processing on the microbiota. Results: DGGE fingerprints analyzed by multivariate analysis showed an evolution of the fungal microflora of coffee during the different stages of the postharvest treatments (wet and dry processes) by extraction and amplification of 26S (yeasts) and 28S rDNA (filamentous fungi). PCR-DGGE stages were optimized: extraction and amplification, repeatability and sensitivity methodology applied to fungi were tested. Conclusion: PCR-DGGE is a rapid molecular technique to monitor the dynamics of coffee microbial populations (fungi and yeast). Significance of study: PCR-DGGE is a promising tool in order to investigate OTA production in coffee beans. (C) 2013 Elsevier Ltd. All rights reserved.