Diversity of bacteria present in milk kefir grains using culture-dependent and culture-independent methods

In the present work bacteria associated with milk kefir grains from several Brazilian States, Canada and the United States of America under traditional conditions have, for the first time, been studied using a combination of pheno-and genotypic methods. Conventional culturing was performed and a total of 270 isolates were obtained from all samples. Isolates were identified using biochemical tests and partial sequence analysis of 16S rDNA. Denaturing gradient gel electrophoresis (DGGE) of partially amplified 16S rDNA followed by sequencing of the most intense bands showed that the dominant bacterium was Lactobacillus kefiri. PCR-DGGE revealed the presence of Gluconobacter japonicus and Lactobacillus uvarum which were no isolated. Conventional isolation revealed the presence of L. helveticus, L. kefiri and Acetobacter syzygii not identified among the sequenced DGGE bands. This study is the first to report the presence of Lactobacillus satsumensis and Acetobacter syzygii in milk kefir grains.