Comparison of methodologies for the extraction of bacterial DNA from mussels-relevance for food safety

Abstract

The control of the microbiological quality of bivalve molluscs assumes particular importance because they are among the most produced seafood products and mostly consumed as a whole, raw, or lightly cooked. The composition of the bacterial community associated with bivalves depends mostly on the microbiology of the surrounding environment at growing sites. Once the relationship between microbiology of bivalves and environment is established, a better classification and monitoring of the shellfish beds and evaluation of depuration strategies can be achieved. In this work, we tested if the methods of DNA extraction commonly used for the culture-independent microbiological analysis of sediment and water could be used directly, or with modifications, for the analysis of bacteria in mussels. The commercial kits Genomic DNA Purification Kit (MBI Fermentas, Vilnius, Lithuania), UltraClean(TM) Soil DNA Isolation Kit (MOBIO Laboratories, Inc., Carlsbad, CA) and the method described by Griffiths and collaborators for DNA/RNA co-extraction were compared. The efficiency of extraction was assessed by DNA fluorescence and the denaturing gradient gel electrophoresis gel patterns of 16S ribosomal RNA gene fragments were used to compare the reproducibility and representativeness of the extraction methods. Results showed that the DNA/RNA co-extraction method with modifications was the most suitable. However, the results must be interpreted in the light of the purpose of the study and the relevance of maximizing extraction yield or diversity estimate, without compromising reproducibility. To our knowledge, this was the first attempt to transpose the procedure currently used for DNA extraction from sediments or waters, to the analysis of whole mussels.