In France, OXA-48\u2013like enzymes are the most prevalent carbapenemases in Enterobacterales (2<\/a>). However, in recent years, the prevalence of metallo-\u03b2-lactamases, particularly New Delhi metallo-\u03b2-lactamase (NDM), increased exponentially (3<\/a>), and 86 NDM variants had been identified by April 2025 (http:\/\/www.bldb.eu<\/a>). Diverse plasmid types, such as IncF, IncA\/C, IncL\/M, IncH, IncN, and IncX3, carry blaNDM genes (3<\/a>,4<\/a>). We characterized NDM-producing Enterobacterales circulating in France during 2021\u20132023 to decipher the prevalence of NDM variants among Enterobacterales species.<\/p>\n Figure 1<\/p>\n Figure 1<\/a>. Variants and species involved in emergence and polyclonal dissemination of blaNDM-7\u2013carrying InX3 plasmid in Enterobacter cloacaecomplex, France, 2021\u20132023. A) Distribution of NDM variants among CPE….<\/p>\n Among 11,825 carbapenemase-producing Enterobacterales (CPE) isolates received at the French National Reference Center for Antimicrobial Resistance (F-NRC) during January 1, 2021\u2013December 31, 2023, we recovered and genetically characterized 3,367 NDM-producing isolates (Appendix 1<\/a>). We found that the number of NDM producers increased from 22.5% (592\/2,582) of all CPE isolates in 2021 to 27.1% (1,135\/4,187) in 2022 and 32.4% (1,640\/5,056) in 2023 (Figure 1<\/a>, panel A; Appendix 2<\/a>). Among isolates, NDM-1, NDM-5, NDM-7, and NDM-14 represented 98% of all NDM enzymes (Figure 1<\/a>, panel B). <\/p>\n We identified NDM-type carbapenemases in 10 different genera (Appendix 1<\/a>), but the distribution of NDM variants varied drastically within Enterobacterales species (Figure 1<\/a>, panel B). We identified NDM-1 mainly in Klebsiella pneumoniae (43%) and NDM-5 predominantly in Escherichia coli (67%), and we found that NDM-14 was nearly exclusively associated with K. pneumoniae (98%). Unexpectedly, Enterobacter cloacae complex (ECC) accounted for 41% (107\/261) of the NDM-7 producers.<\/p>\n Figure 2<\/p>\n Figure 2<\/a>. Distribution of STs among 107 Enterobacter cloacae complex isolates used in a study of emergence and polyclonal dissemination of blaNDM-7\u2013carrying InX3 plasmid, France, 2021\u20132023. ST,…<\/p>\n Among the 107 NDM-7\u2013producing ECC isolates, we identified 32 different sequence types (STs), the most prevalent of which were ST873 (22.4%), ST135 (10.2%), ST145 (10.2%), ST683 (9.3%), ST252 (6.5%), and ST32 (5.6%) (Figure 2<\/a>). Those STs corresponded to 8 different Enterobacter species, including 46.7% of E. hormaechei subspecies hoffmanii (mostly ST135, ST145, and ST683), 22.4% E. quasihormaechei (all ST873), 7.4% E. hormaechei subsp. steigerwaltii, 6.5% (7\/107) E. xianfangensis, 6.5% E. asburiae, 5.6% E. kobei (all ST32), 2.8% E. cloacae, 0.9% E. hormaechei subsp. oharae, and 0.9% E. hormaechei subsp. hormaechei. Phylogenetic analysis confirmed the polyclonal dissemination of NDM-7\u2013producing ECC, including in the 4 major STs (ST873, ST145, ST135, and ST683) comprising several isolates (Appendix 1<\/a> Figure 1).<\/p>\n Among the 107 NDM-7\u2013producing ECC isolates, short-read sequencing enabled us to identify 12 different plasmid replicases (Appendix 2<\/a>). Long-read sequencing performed on 30 representative NDM-7\u2013producing ECC showed that blaNDM-7 was carried on an IncX3-type plasmid in all isolates (Appendix 1<\/a>). Of note, 93.3% (28\/30) of the IncX3 plasmids were 46,161-bp long, and 6.7% (2\/30) were 49,830-bp long (Appendix 1<\/a> Figure 2). The size difference corresponded to the Tn5403 transposon. Those 2 plasmids shared 99.9% nucleotide identity with the 46,161-bp blaNDM-5\u2013carrying plasmid pEC21Z078-46K (GenBank accession no. CP10126), previously described in an E. coli isolate from China (5<\/a>).<\/p>\n To compare the blaNDM-7\u2013carrying plasmids with those harboring blaNDM-5 in Enterobacter spp., we performed long-read sequencing on 19 additional NDM-5\u2013ECC isolates (Appendix 2<\/a>). We found that 96% of those isolates also harbored an IncX3 plasmid. Among them, 84.2% (16\/19) harbored an IncX3 plasmid almost identical to one of the blaNDM-7\u2013carrying plasmids (except the 2 single-point mutations between blaNDM-5 and blaNDM-7); 12 harbored the 46,161-bp plasmid and 4 harbored the 49,830-bp plasmid. Among the other 3 NDM-5\u2013producing ECC isolates, we localized blaNDM-5 on different IncFII-type plasmids: a 177,571-bp and a 96,517-bp plasmid similar to pABC143C-NDM (GenBank accession no. KY130431.1), and a 187,303-bp plasmid similar to p60214CZ (GenBank accession no. CP085746.1).<\/p>\n To investigate the origin of the blaNDM-7 IncX3 plasmid, we further explored 79 full-length sequences of NDM-1\u2013producing ECC for IncX3 plasmids. Among the 11 different replicases identified, IncX3 was in only 14% of NDM-1\u2013producing ECC. Long-read whole-genome sequencing of those 11 IncX3-positive NDM-1\u2013producing ECC identified the blaNDM-1 gene on IncX3 plasmids of 39,582-bp (n = 7) and 44,682-bp (n = 2) length (Appendix 1<\/a> Figure 2). Both of those plasmids were close to blaNDM-5\u2013 and blaNDM-7\u2013carrying IncX3 plasmids but showed notable differences in the genetic region surrounding the blaNDM gene (Appendix 1<\/a> Figure 2). Despite detection of an InX3 plasmid, the other 2 strains harbored blaNDM-1, either directly localized on the chromosome or on a 51,088-bp IncFII plasmid that had no similarity to plasmids available in GenBank.<\/p>\n The close genetic context of blaNDM genes localized on IncX3 plasmids showed that both blaNDM-7 and blaNDM-5 were flanked upstream by \u0394Tn2\u2013\u0394IS3000\u2013ISAba125\u2013IS5 and downstream by genes encoding bleMBL, PAI, ccdA, IS26, UmuD protein, ISKox3, a resolvase encoding gene, and Tn5403 (Appendix 1<\/a> Figure 2). Several elements were truncated or absent in the blaNDM-1 genetic environment compared with blaNDM-5 and blaNDM-7, including truncation of ISAba125 and deletions of IS5, ISKox3, and Tn5403.<\/p>\n Finally, to decipher whether the IncX3 plasmids were more prevalent in ECC, we looked for IncX3 replicase in 20,028 multidrug-resistant Enterobacterales genomes in the F-NRC database collected since 2022. Of those, 11.9% (n = 2,393) of isolates carried the IncX3-encoding replicase gene, 27.5% (n = 658) of which were Klebsiella spp., 27.3% (n = 653) were Citrobacter spp., 26.8% (n = 642) were E. coli, and 17.7% (n = 423) were ECC (Appendix 1<\/a> Figure 3).<\/p>\n During 2021\u20132023, F-NRC received increasing numbers of carbapenem-resistant Enterobacterales isolates: 2,582 in 2021, 4,187 in 2022, and 5,056 in 2023. In addition, the number of NDM-producing isolates more than tripled during that timeframe (Figure 1<\/a>).<\/p>\n Although >86 NDM variants have been reported globally in the Beta-Lactamase DataBase (http:\/\/bldb.eu<\/a>), 4 main variants are dominant in Europe: NDM-1, NDM-5, NDM-7, and NDM-14. In France, our results demonstrated that NDM-1 remains predominantly associated to K. pneumoniae, among which ST147 has been described as highly prevalent (48%) (6<\/a>,7<\/a>). The blaNDM-1 gene is carried on various plasmid types, including IncX3 (8<\/a>), IncFIB, IncHI1B, and IncL, that harbor both resistance and virulence factors (9<\/a>). In France, NDM-1 producers have been progressively replaced by NDM-5 producers since 2023. NDM-5 and NDM-7 differ from NDM-1 by substitutions responsible for an enhanced carbapenemase activity (7<\/a>). In Europe, dissemination of NDM-5 is mostly caused by E. coli (10<\/a>), which represented 70% of NDM-5 producers in our study. As previously described (11<\/a>), our results confirmed that blaNDM-5 gene dissemination is mainly mediated by IncX3 plasmids. Our results confirmed that NDM-14\u2013producing K. pneumoniae ST147 became established after it emerged in France in 2022 (12<\/a>).<\/p>\n Our results highlight the emergence of NDM-7 in ECC, as noted in 41% of the isolates in our study. However, we did not identify any exclusive or particularly dominant clone involved in the dissemination of NDM-7 ECC, except ST873, which was slightly overrepresented. Of note, ECC isolates have been reported to trigger dissemination of Verona-integron\u2013encoded metallo-\u03b2-lactamases in France, of which ST893 is the most prevalent ST (13<\/a>). Our results also raise concerns about E. quasihormaechei ST873 as a high-risk clone for acquiring multidrug resistance, particularly carbapenemase resistance.<\/p>\n In summary, we demonstrated that the IncX3 blaNDM-7\u2013carrying plasmids most probably derived of IncX3 blaNDM-5\u2013carrying plasmids that were derived from IncX3 blaNDM-1\u2013carrying plasmids and contain several additional features, including IS5, ISKox3, and Tn5403. Despite the high transfer frequency (14<\/a>) and low fitness cost (15<\/a>) of IncX3 plasmids, their implication in the dissemination of blaNDM remain unequal between Enterobacterales species. The strong association between IncX3 blaNDM-7\u2013carrying plasmids and ECC underscores the need for enhanced surveillance to monitor spread of antimicrobial resistance.<\/p>\n![\"Variants](\"https:\/\/www.newsbeep.com\/il\/wp-content\/uploads\/2025\/09\/25-0830-F1-tn.jpg\")
<\/p>\n![\"Distribution](\"https:\/\/www.newsbeep.com\/il\/wp-content\/uploads\/2025\/09\/25-0830-F2-tn.jpg\")
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