Metagenomic analysis of 130 years old Danish starter culture material including sequence analysis of the genome of a Lactococcus cremoris starter

Two unopened Danish dairy starter culture bottles from the 1890s were analyzed to investigate their microbiome and genetic characteristics, offering insights into historical dairy production. Using culture-independent methods, DNA was extracted and sequenced via Nanopore-based Barcode-Amplified Random Sequencing. Metataxonomic profiling revealed contamination with Cutibacterium acnes, while the 1893 sample showed significant Lactococcus cremoris DNA (76.7 % genome coverage), compared to minimal coverage (<1 %) in the 1899 sample. Bioinformatic analysis identified functional genes in L. cremoris associated with dairy adaptation, including casein degradation, lactose metabolism, and acetoin/diacetyl production, which contribute to buttery flavors. These findings are of interest, considering mild flavor profile preferred in Danish butter for the English market during the 19th century. Additionally, a conserved lactococcin gene (lcn) was detected in both the 1893 sample and a modern isolate of L. Cremoris, suggesting its persistence over time. This study sheds light on the microbiome and functional properties of historical starter cultures.