Typing of mecD Islands in Genetically Diverse Methicillin-Resistant Macrococcus caseolyticus Strains from Cattle

Since its first description in 2017, the methicillin resistance gene mecD has been detected in M. caseolyticus strains from different cattle sources and countries. Our study provides new insights into the molecular diversity of mecD-carrying M. caseolyticus strains by using two approaches to characterize mecD elements: (i) multiplex PCR for molecular typing of McRImecD and (ii) read mapping against reference sequences to identify McRImecD types in silico. In combination with multilocus sequence typing, this approach can be used for molecular characterization and surveillance of M. caseolyticus carrying mecD. ABSTRACT Macrococcus caseolyticus belongs to the normal bacterial flora of dairy cows and does not usually cause disease. However, methicillin-resistant M. caseolyticus strains were isolated from bovine mastitis milk. These bacteria had acquired a chromosomal island (McRImecD-1 or McRImecD-2) carrying the methicillin resistance gene mecD. To gain insight into the distribution of McRImecD types in M. caseolyticus from cattle, 33 mecD-containing strains from Switzerland were characterized using molecular techniques, including multilocus sequence typing, antibiotic resistance gene identification, and PCR-based McRImecD typing. In addition, the same genetic features were analyzed in 27 mecD-containing M. caseolyticus strains isolated from bovine bulk milk in England/Wales using publicly available whole-genome sequences. The 60 strains belonged to 24 different sequence types (STs), with strains belonging to ST5, ST6, ST21, and ST26 observed in both Switzerland and England/Wales. McRImecD-1 was found in different STs from Switzerland (n = 19) and England/Wales (n = 4). McRImecD-2 was only found in 7 strains from Switzerland, all of which belonged to ST6. A novel island, McRImecD-3, which contains a complete mecD operon (mecD-mecR1m-mecIm [where the subscript m indicates Macrococcus]) combined with the left part of McRImecD-2 and the right part of McRImecD-1, was found in heterogeneous STs from both collections (Switzerland, n = 7; England/Wales, n = 21). Two strains from England/Wales carried a truncated McRImecD-3. Phylogenetic analyses revealed no clustering of strains according to geographical origin or carriage of McRImecD-1 and McRImecD-3. Circular excisions were also detected for McRImecD-1 and McRImecD-3 by PCR. The analyses indicate that these islands are mobile and may spread by horizontal gene transfer between genetically diverse M. caseolyticus strains. IMPORTANCE Since its first description in 2017, the methicillin resistance gene mecD has been detected in M. caseolyticus strains from different cattle sources and countries. Our study provides new insights into the molecular diversity of mecD-carrying M. caseolyticus strains by using two approaches to characterize mecD elements: (i) multiplex PCR for molecular typing of McRImecD and (ii) read mapping against reference sequences to identify McRImecD types in silico. In combination with multilocus sequence typing, this approach can be used for molecular characterization and surveillance of M. caseolyticus carrying mecD.