Genomically Informed Surveillance for Carbapenem-Resistant Enterobacteriaceae in a Health Care System

Abstract

Carbapenem-resistant Enterobacteriaceae (CRE) are an urgent public health concern. Rapid identification of the resistance genes, their mobilization capacity, and strains carrying them is essential to direct hospital resources to prevent spread and improve patient outcomes. Whole-genome sequencing allows refined tracking of both chromosomal traits and associated mobile genetic elements that harbor resistance genes. To enhance surveillance of CREs, clinical isolates with phenotypic resistance to carbapenem antibiotics underwent whole-genome sequencing. Analysis of 41 isolates of Klebsiella pneumoniae and Enterobacter cloacae, collected over a 3-year period, identified K. pneumoniae carbapenemase (KPC) genes encoding KPC-2, −3, and −4 and OXA-48 carbapenemases. All occurred within transposons, including multiple Tn4401 transposon isoforms, embedded within more than 10 distinct plasmids representing incompatibility (Inc) groups IncR, -N, -A/C, -H, and -X. Using short-read sequencing, draft maps were generated of new KPC-carrying vectors, several of which were derivatives of the IncN plasmid pBK31551. Two strains also had Tn4401 chromosomal insertions. Integrated analyses of plasmid profiles and chromosomal single-nucleotide polymorphism (SNP) profiles refined the strain patterns and provided a baseline hospital mobilome to facilitate analysis of new isolates. When incorporated with patient epidemiological data, the findings identified limited outbreaks against a broader 3-year period of sporadic external entry of many different strains and resistance vectors into the hospital. These findings highlight the utility of genomic analyses in internal and external surveillance efforts to stem the transmission of drug-resistant strains within and across health care institutions.

Enhanced Klebsiella pneumoniae Carbapenemase Expression from a Novel Tn 4401 Deletion

“…coli transformants containing Tn4401a and Tn4401h variants on native parent plasmid backgrounds showed meropenem and cefepime resistance MICs compared to E. coli transformants containing Tn4401b, consistent with the variations in bla KPC expression (i.e., Tn4401a ¡ Tn4401h ¡ Tn4401b). Previous studies have suggested that another mechanism, in addition to gene expression and copy number, namely, production of porin channel defects, is a primary contributor to the variability in susceptibility in testing KPC-producing isolates (3,11,20,(28)(29)(30). Here we excluded the impact of porin channel variation on susceptibility by evaluating plasmids bearing the Tn4401-bla KPC units in a fixed E. coli background.…”

“…To date, seven unique Tn4401 isoforms (Tn4401a to Tn4401g) have been identified (9)(10)(11)(12), with Tn4401a and Tn4401b being the most widespread (13). The original characterization of these isoforms demonstrated deletions in the noncoding region upstream of bla KPC or lacking genes or both as follows: a, Ϫ99 bp (14,15); b, no deletion (3); c, Ϫ215 bp (16); d, Ϫ68 bp (9) and Ϫ5.3 kb (9,17,18); e, Ϫ255 bp (17); f, truncated tnpA and lacking tnpR, ISKpn7 left, and Tn4401 IRL-1 (10); g, lacking tnpA, tnpR, and ISKpn7 left, with a Ϫ215-bp deletion in the noncoding region (Fig.…”

Genomic Characterization of Two KPC-Producing Klebsiella Isolates Collected in 1997 in New York City

“…This suggests that bla KPC -harboring pKp58_N-and pKm38_N-like plasmids were circulating in the Northeastern United States for almost 2 decades and may have contributed significantly to the spread of KPC at the early stages of the CRE epidemic. In addition, bla KPC -harboring IncN plasmids have been frequently reported elsewhere around the world and in other Enterobacteriaceae species (10,(25)(26)(27)(28)(29)(30), suggesting the IncN plasmid is an important vector in the dissemination of KPC. Interestingly, bla KPC in this study was identified in ST111 K. pneumoniae and K. michiganensis isolates.…”

Genetic environment of the KPC gene in Acinetobacter baumannii ST2 clone from Puerto Rico and genomic insights into its drug resistance

“…Although A. baumannii ST2 clone has been previously associated with the production of CHDLs (OXA-23, OXA-24/40 and OXA-58) and the VIM MBL, this is the first report of the identification of bla Location and characterization of the genetic environment of the bla KPC Southern blot hybridization with S1 nuclease and I-CeuI digestion revealed that the KPC gene was chromosomally encoded (data not shown). The chromosomal location of the KPC gene has been previously described from an A. baumannii from Puerto Rico (Martinez et al, 2014), one P. aeruginosa from Colombia (Villegas et al, 2007) and two K. pneumoniae from the USA (Chen et al, 2015;Pecora et al, 2015). In our isolate, KPC-2 gene was identified within a truncated version of the Tn4401e (255 bp deletion upstream of the KPC gene) in which the KPC gene is flanked by the ISKpn6 and ISKpn7 but lacks the transposase and resolvase genes (Figs 1 and 2a).…”