Identification of dominant species common to kefir grains from seven origins for kefir grain reconstruction

Dominant species common to the kefir grains from 7 different origins were analyzed using culture-dependent methods using agar media that were able to grow and discriminate Lactobacillus kefiranofaciens ssp. kefiranofaciens. Identification of all isolates was performed on the basis of MALDI-TOF MS analysis and colony morphology. Lactic acid bacteria were present at 9 log cfu·g^-1 in all kefir grains. Lactobacillus kefiranofaciens was the predominant species (∼87%), followed by Lentilactobacillus kefiri (∼12%) in all grains. Within Lb. kefiranofaciens, Lb. kefiranofaciens ssp. kefirgranum (flocculent and nonflocculent strains) was most abundant (∼82%) and Lb. kefiranofaciens ssp. kefiranofaciens (kefiran producer) was present at ∼18%. Lentilactobacillus parakefiri, Leuconostoc mesenteroides, and Lactococcus lactis were also present in some grains as minor species. Yeasts were present at 6 to 7 log cfu·g^-1 and were highly diverse; only Kazachstania exigua was found in all grains. We attempted to reconstruct kefir grains using strains of the following major and minor common species: Lb. kefiranofaciens ssp. kefiranofaciens, Lb. kefiranofaciens ssp. kefirgranum, Len. kefiri, Len. parakefiri, Leu. mesenteroides, Lc. lactis, and K. exigua. We obtained reconstructed kefir grains that could grow logarithmically to some extent in milk. They were elastic, but each grain was smaller than a natural kefir grain. At least a combination of Lb. kefiranofaciens ssp. kefiranofaciens, Lb. kefiranofaciens ssp. kefirgranum, and Len. kefiri or Len. parakefiri could initiate grain formation.