Assessment of the bacterial diversity of treated and untreated milk during cold storage by T-RFLP and PCR-DGGE methods

Abstract

The evaluation of the effect of treatments on the dynamics of milk microbiota by appropriate and reliable methods is critical for a better control of the microbiological quality of milk. The aim of this study was to compare the diversity and the alterations of the microbiota in treated and untreated milk samples during cold storage at 4 °C or 8 °C up to 7 days. Three treatments were applied to raw milk, carbon dioxide addition (CO2), thermization (TH), and microfiltration (MF), and untreated raw milk (UT) was used as a control. Two molecular fingerprinting methods, terminal restriction fragment length polymorphism (T-RFLP) and denaturing gradient gel electrophoresis (DGGE) were used. According to the T-RFLP and polymerase chain reaction–denaturing gradient gel electrophoresis (PCR-DGGE) profiles, the dominant bacteria in milk were presumably affiliated with the psychrotolerant genera Streptococcus, Staphylococcus, Pseudomonas, and Aerococcus. At day 7, phylotypes related to Pseudomonas spp., Trichococcus floculiformis and Prevotella spp. were dominant in UT and CO2 milk samples but not in TH milk samples while Staphylococcus spp. and Stenotrophomonas spp. were specifically dominant in TH and MF milk samples, respectively. Principal component analysis of T-RFLP profiles revealed the strong effect of treatment on the dynamics of specific bacteria. PCR-DGGE profiles revealed the dominant bacteria associated with the effect of treatments (TH, CO2, and MF) on milk microbiota, which can then be used as markers for the quality of treated milk. The T-RFLP method had a higher resolution than PCR-DGGE in this study and can thus be used as an accurate tool for analysis of the bacterial community in milk.