Characterization of Two Multidrug-Resistant IncA/C Plasmids from the 1960s by Using the MinION Sequencer Device

Two A/C incompatibility group (IncA/C family) plasmids from the 1960s have been sequenced and classified into the A/C 2 type 1 group. R16a and IP40a contain novel antibiotic resistance islands and a complete GIsul2 genomic island not previously found in the family. In the 173.1-kb R16a, the 29.9-kb antibiotic resistance island (ARI) is located in a unique backbone position not utilized by ARIs. ARI R16a consists of Tn 1 , Tn 6020 , and Tn 6333 , harboring the resistance genes bla TEM-1D and aphA1b and a mer module, respectively; a truncated Tn 5393 copy; and a gene cluster with unknown function. Plasmid IP40a is 170.4 kb in size and contains a 5.6-kb ARI inserted into the kfrA gene. ARI IP40a carrying bla TEM-1D and aphA1b genes is composed of Tn 1 with a Tn 6023 insertion. Additionally, IP40a harbors single IS 2 , IS 186 , and Tn 1000 insertions scattered in the backbone; an IS 150 copy in GIsul2; and a complete Tn 6333 carrying a mer module at the position of ARI R16a . Loss of resistance markers in R16a, IP40a, and R55 was observed during stability tests. Every phenotypic change proved to be the result of recombination events involving mobile elements. Intramolecular transposition of IS copies that generated IP40a derivatives lacking large parts of the backbone could account for the formation of other family members, too. The MinION platform proved to be a valuable tool in bacterial genome sequencing since it generates long reads that span repetitive elements and facilitates full-length plasmid or chromosome assembly. Nanopore technology enables rapid characterization of large, low-copy-number plasmids and their rearrangement products.