Extracellular and intracellular DNA for bacterial profiling of long-ripened cheeses. Issue 13 (25th June 2020)

• Record Type: Journal Article
• Authors: Bardelli, Tommaso; Rossetti, Lia; Zago, Miriam; Carminati, Domenico; Giraffa, Giorgio; Tidona, Flavio
• Is Part Of: FEMS microbiology letters. Volume 367:Issue 13(2020)
• Publication Date: 2020-06-25
• Subjects: extracellular DNA -- intracellular DNA -- EDTA solution buffer -- LAB profiling -- long-ripened cheeses; Microbiology -- Periodicals; 579
• DOI: 10.1093/femsle/fnaa095
• Languages: English
• ISSNs: 0378-1097

Abstract

ABSTRACT: A novel approach was developed to extract the extracellular DNA (eDNA), i.e. the free DNA outside the microbial cell, compared to the intracellular DNA (iDNA). The two DNA fractions were investigated in seven long-ripened cheeses. Among different buffer solutions tested, EDTA 0.5 M at pH 8 enabled a mild homogenization of cheese samples and the highest eDNA recovery. The extraction protocol was tested on single strains of lactic acid bacteria characterizing many Italian long-ripened cheeses, such as Streptococcus thermophilus, Lactobacillus helveticus, and Lactobacillus rhamnosus . The method resulted suitable for eDNA extraction because it minimized cell-lysis, avoiding the leakage of iDNA from the cells. The yields of eDNA, ranging from 0.01 to 0.36 µg g −1 cheese, were generally higher than the iDNA, indicating that autolytic phenomena prevailed over intact cells after 8–12 months of ripening. In four of the seven cheeses, the same LAB species were detected in the eDNA and iDNA fractions by length-heterogeneity PCR, while in the remaining three samples, a higher number of species was highlighted in the eDNA compared to the corresponding iDNA. The sequential extraction of eDNA and iDNA can be applied to obtain additional information on the composition of the bacterial community in long-aged cheeses. Abstract : Determination of extracellular and intracellular DNA to obtain a more complete bacterial profile of long-ripened cheeses.