Diversity and host association of cfr-carrying plasmids of multidrug-resistant microorganisms from the Netherlands

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) carrying a plasmid-localized cfr-gene were identified in the Netherlands. Expression of this gene causes reduced binding of phenicols, lincosamides, oxazolidinones, pleuromutilins and streptogramin A antimicrobials to ribosomes. To gain insight into the introduction and spread of cfr-genes in the Netherlands, cfr-carrying plasmids from isolates obtained in national surveillance programs were analyzed alongside internationally retrieved cfr-carrying plasmids. Isolates carrying a cfr-gene were selected from isolates sequenced through Dutch national surveillance programs (2016-2024), and relatedness was assessed with wgMLST. Short-read and long-read sequence data were used for hybrid assembly and plasmid reconstructions. International cfr-carrying plasmids were retrieved from the PLSDB plasmid database. Diversity of Dutch cfr-carrying plasmids was characterized based on resistome, %G+C content and size. Relatedness between plasmids was investigated with Mash and Pling. One Proteus mirabilis, 12 LA-MRSA and one non-LA-MRSA from Dutch national surveillance contained cfr-carrying plasmids. All isolates except one LA-MRSA were human-derived and unrelated. From PLSDB, 218 cfr-carrying plasmids were included. Resistomes, %G+C content and size of Dutch cfr-carrying plasmids were diverse. Both Mash and Pling mostly clustered plasmids according to host genus, except for one cluster. Dutch plasmids linked to plasmids from China, Denmark, the United States, South-Korea, Germany and Poland. Across all Dutch plasmids, the cfr-gene was flanked by transposable elements. The diversity among cfr-carrying plasmids in unrelated Dutch surveillance isolates and their relatedness to plasmids from countries in different global regions suggest multiple independent introductions into the Netherlands, although horizontal dissemination by transposons may contribute to their spread.