Figure 3.

Phylogenetic analysis of the core gene SNP analysis for the USA subset of IncI2 plasmids. Rings (starting from inner to outer) correspond with: (1) designation of Group 1 or 2 based on core gene phylogeny, (2) year of isolation, (3) taxa, (4) presence or absence of mobile genetic elements and, (5) sub-cluster assigned using MOB-suite. All colors are indicated in the legends. Figure created using iTOL [19]. Analysis using MOB-suite confirmed that representatives from all 6 sub-nodes identified in the full dataset were also present in the smaller USA dataset. As the mcr-1 plasmids were primarily associated with AI350, it was not surprising that the USA plasmids were less prevalently found in the AI350 node (n = 10) in this dataset and were instead more commonly assigned to the AI345-347 nodes (n = 15, 33 and 35 respectively). Notably, IS elements continued to be absent from plasmids assigned to node AI345 and AI346, consistent with the results of the larger dataset. The plasmids carrying mcr-1 were divided between AI350 and AI347, suggesting separate acquisition/introduction events for mcr-1 plasmids in the USA. An examination of the association between the core SNP phylogeny and the assigned MOB nodes indicated that changes within the core genes could be occurring broadly within IncI2 plasmids, as opposed to being isolated to a closely related node. Similar to the pattern observed for IS elements, nodes AI348-350 were primarily associated with Group 2 (67%, 100% and 100%, respectively). Of the plasmids assigned to AI347, 46% were associated with Group 2, which is slightly higher than the observed association of IS elements with this node in the full dataset (35%). Despite a large number of representatives, only 6% of plasmids in node AI346 (2/33) were associated with Group 2. Interestingly, 27% of AI345 plasmids (4/15) were associated with Group 2, although there were no plasmids from this node found to carry any IS elements in either dataset.