In silico characterization of IncX3 plasmids carrying bla OXA-181 in Enterobacterales.

Carbapenem-resistant Enterobacterales poses a global urgent antibiotic resistance threat because of its ability to transfer carbapenemase genes to other bacteria via horizontal gene transfer mediated by mobile genetic elements such as plasmids. Oxacillinase-181 (OXA-181) is one of the most common OXA-48-like carbapenemases, and OXA-181-producing Enterobacterales has been reported in many countries worldwide. However, systematic research concerning the overall picture of plasmids harboring bla OXA-181 in Enterobacterales is currently scarce. In this study, we aimed to determine the phylogeny and evolution of bla OXA-181-positive (gene encoding OXA-181) plasmids. To characterize the plasmids harboring bla OXA-181 in Enterobacterales, we identified 81 bla OXA-181-positive plasmids from 35,150 bacterial plasmids downloaded from the NCBI RefSeq database. Our results indicated that diverse plasmid types harbored bla OXA-181 but was predominantly carried by IncX3-type plasmids. We systematically compared the host strains, plasmid types, conjugative transfer regions, and genetic contexts of bla OXA-181 among the 66 bla OXA-181-positive IncX3 plasmids. We found that IncX3 plasmids harboring bla OXA-181 were mostly ColKP3-IncX3 hybrid plasmids with a length of 51 kb each and were mainly distributed in Escherichia coli and Klebsiella pneumoniae. Most of the IncX3 plasmids harboring bla OXA-181 were human origin. Almost all the bla OXA-181-positive IncX3 plasmids were found to carry genes coding for relaxases of the MOBP family and VirB-like type IV secretion system (T4SS) gene clusters, and all the 66 IncX3 plasmids were found to carry the genes encoding type IV coupling proteins (T4CPs) of the VirD4/TraG subfamily. Most IncX3 plasmids harbored both bla OXA-181 and qnrS1 in their genomes, and the two antibiotic resistance genes were found to a composite transposon bracketed by two copies of insertion sequence IS26 in the same orientation. Our findings provide important insights into the phylogeny and evolution of bla OXA-181-positive IncX3 plasmids and further address their role in acquiring and spreading bla OXA-181 genes in Enterobacterales.

In silico characterization of bla NDM-harboring plasmids in Klebsiella pneumoniae.

Klebsiella pneumoniae is a primary culprit of antibiotic-resistant nosocomial infections worldwide, and infections caused by NDM-producing strains are a major threat due to limited therapeutic options. The majority of bla NDM cases occur on plasmids; therefore, we explored the relationships between plasmids and bla NDM genes in K. pneumoniae by analyzing the variants of bla NDM, replicon types, conjugative transfer regions of 171 bla NDM-harboring plasmids from 4,451 K. pneumoniae plasmids. Of the nine identified bla NDM variants, bla NDM-1 (73.68%) and bla NDM-5 (16.37%) were the most dominant. Over half of the bla NDM-harboring plasmids of K. pneumoniae were classified into IncF plasmids. IncX3 single-replicon plasmids (46-57 kb) carried genes encoding relaxases of the MOBP family, T4CP genes of the VirD4/TraG subfamily, and VirB-like T4SS gene clusters, which were mainly geographically distributed in China. We found 10 bla NDM-harboring IncN plasmids (38.38-63.05 kb) carrying the NW-type origin of transfer (oriT) regions, genes coding for relaxases of MOBF family, genes encoding T4CPs of the TrwB/TraD subfamily, and Trw-like T4SS gene clusters, which were also mainly geographically distributed in China. Moreover, we identified 21 IncC plasmids carrying bla NDM-1 (140.1-329.2 kb), containing the A/C-type oriTs, genes encoding relaxases of MOBH family, genes encoding T4CPs belonging to TrwB/TraD subfamily, and Tra_F-like T4SS gene clusters. The bla NDM-harboring IncC plasmids were widely geographically distributed all over the world, mainly in the United States, China and Viet Nam. These findings enhance our understanding of the diversity of bla NDM-harboring plasmids in K. pneumoniae.

Comparative genomic analysis of plasmids harboring bla OXA-48-like genes in Klebsiella pneumoniae.

The emergence and spread of carbapenem-resistant Klebsiella pneumoniae (CRKP) is a serious medical problem worldwide. Acquired OXA-48-like carbapenemases encoded by plasmids are important causes of carbapenem resistance in K. pneumoniae. To explore the links between plasmids and bla OXA-48-like genes in K. pneumoniae, we systematically analyzed the variants of bla OXA-48-like plasmid replicon types, phylogenetic patterns, geographic distribution, conjugative transfer regions, and the genetic environments surrounding bla OXA-48-like of 191 bla OXA-48-like-harboring plasmids, which were identified from 4451 plasmids of K. pneumoniae downloaded from GenBank. Our results showed that seven different variants of bla OXA-48-like genes were identified from the 191 bla OXA-48-like-harboring plasmids in K. pneumoniae, with bla OXA-48, bla OXA-232, and bla OXA-181 being highly prevalent. In K. pneumoniae, bla OXA-48 was mainly carried by the composite transposon Tn1999.2 located on IncL/M-type conjugative plasmids, which were mainly geographically distributed in Switzerland, Germany, and China. In K. pneumoniae, the blaOXA-232 gene was mainly carried by 6.1-kb ColKP3-type mobilizable plasmids, which were mainly isolated in India. In K. pneumoniae, bla OXA-181 was mainly carried by a group of 50-kb ColKP3-IncX3 hybrid conjugative plasmids and a group of small ColKP3-type mobilizable plasmids with lengths of 5.9-9.3 kb, the former was sporadically discovered in China, South Korea, India, and Czech Republic, while the latter was almost all isolated in India. In addition, five bla OXA-245-harboring 65.9-kb IncL plasmids of K. pneumoniae isolated in Spain were found to have the genetic context of bla OXA-245 more complicated than that of bla OXA-48-harboring IncL/M-type plasmids, with two copies of IS1R inserted both upstream and downstream of bla OXA-245-lysR. These findings enhance our understanding of the genetic diversity of bla OXA-48-like-harboring plasmids in K. pneumoniae.