Isolation and Identification of Representative Fungi from Shaoxing Rice Wine Wheat Qu Using a Polyphasic Approach of Culture-Based and Molecular-Based Methods

摘要

Ribosomal intergenic spacer analysis (RISA) and traditional culture-based methods were employed to examine the fungal community of Shaoxing rice wine wheat Qu. Results showed that RISA profiles of total DNA exhibited nine distinguishable bands. By comparison, the RISA fingerprints recovered from enrichment media gave variable patterns containing fewer bands. Bands corresponding to five fungal species detected by RISA of total DNA were also observed in RISA fingerprints from enrichment cultures. A total of twenty-four pure cultures were isolated with media of potato dextrose agar (PDA), Czapek (CPK) and wheat Qu extraction (WQE). A total of eight, eighteen and eleven fungal species and 7.9 × 10~5 cfu/g Qu, 3.0 × 10~5 cfu/g Qu, and 3.9 × 10~6 cfu/g Qu, were isolated using PDA, CPK and WQE media, respectively. Predominance of each representative species did vary with the culture medium. Comparison of ITS sequences of excised RISA bands and recovered pure isolates with those in GenBank database revealed that the representative fungal species in wheat Qu were Absidia corymbifera, Sac-charomyces cerevisiae, Rhizopus oryzae, Rhizomucor pusillus, Aspergillus oryzae, Candida tropicalis, Emericella nidulans, Clavispora lusitaniae, Aspergillus fumigatus, Aspergillus niger, Rhizopus microsporus, Pichia guilliermondii and Pichia anom-ala. Species of Saccharomyces cerevisiae, Candida tropicalis and Rhizopus oryzae detected in the original samples by analysis of RISA were not recovered in any of the three media. In addition, some species recovered from the enrichments were not detected by RISA analysis. Results obtained with culture-based or molecular-based methods clearly confirmed the importance of developing an integrated approach to gain a better understanding of the microbial community in wheat Qu.