The lactococcal ICE-ome encodes a repertoire of exchangeable traits with potential industrial relevance.

Dairy industries apply selected lactococcal strains and mixed cultures to produce diverse fermented products with distinctive flavor and texture properties. Innovation of the starter culture functionality in cheese applications embraces natural biodiversity of the Lactococcus species to identify novel strains with alternative flavor or texture forming capacities and/or increased processing robustness and phage resistance. Mobile genetic elements (MGE), like integrative conjugative elements (ICEs) play an important role in shaping the biodiversity of bacteria. Besides the genes involved in the conjugation of ICEs from donor to recipient strains, these elements also harbor cargo genes that encode a wide range of functions. The definition of such cargo genes can only be achieved by accurate identification of the ICE boundaries (delimiting). Here, we delimited 25 ICEs in lactococcal genome sequences with low contig numbers using insertion-sites flanking single-copy core-genome genes as markers for each of the distinct ICE-integrases we identified previously within the conserved ICE-core genes. For ICEs in strains for which genome information with large numbers of contigs is available, we exemplify that CRISPR-Cas9 driven ICE-curing, followed by resequencing, allows accurate delimitation and cargo definition of ICEs. Finally, we compare and contrast the cargo gene repertoire of the 26 delimited lactococcal ICEs, identifying high plasticity among the cargo of lactococccal ICEs and a range of encoded functions that is of apparent industrial interest, including restriction modification, abortive infection, and stress adaptation genes.