Traditionally viewed as a hospital pathogen, multidrug-resistant (MDR) Klebsiella pneumoniae is emerging in community settings, yet its population structure and spread are currently undefined. A nationwide genomic survey of 2006 ceftriaxone-nonsusceptible outpatient isolates from 42 US states reveals that the spread of resistance is primarily driven by the bla_CTX-M-15 extended-spectrum-β-lactamase (ESBL) gene across diverse K. pneumoniae lineages. This gene, harbored by a family of IncFIB(Kpn3) plasmids, has spread to hundreds of lineages, coupling antibiotic resistance with stress- and metal-tolerance traits that may enhance survival beyond human hosts. Nearly half of isolates formed transmission clusters spanning multiple states, underscoring active community dissemination. The majority originated from elderly women with urinary tract infections, where oral treatment options are increasingly ineffective. Capsule and O-antigen profiling reveal broad antigenic diversity, yet a manageable subset of serotypes could offer wide coverage for vaccine development. Our findings expose a rapidly evolving, plasmid-driven epidemic of community-associated MDR K. pneumoniae across the United States.

Klebsiella quasipneumoniae subsp. similipneumoniae is an important member of the K. pneumoniae species complex (KpSC) and is increasingly reported as multidrug-resistant (MDR) in healthcare- and community-associated infections. Since clinical laboratories do not routinely distinguish K. quasipneumoniae subsp. similipneumoniae from K. pneumoniae , national prevalence estimates, particularly for MDR, are lacking. In this study, a total of 2,006 community-associated MDR KpSC isolates were collected from 42 U.S. states, with 30 K. quasipneumoniae subsp. similipneumoniae isolates originating from 12 states identified using whole genome sequencing. All isolates were resistant to ceftriaxone and exhibited high rates of resistance to other antimicrobial agents, including ampicillin-sulbactam (56.7%, 17/30), levofloxacin (75.9%, 22/29), and trimethoprim-sulfamethoxazole (53.3%, 16/30). Notably, five isolates were also carbapenem-resistant. Genomic analysis resolved 10 sequence types (STs), with ST2059 ( n = 13) and ST414 ( n = 9) predominating. Ceftriaxone resistance in most isolates (90%, 27/30) was conferred by an extended-spectrum β-lactamase gene, predominantly bla _CTX-M-15 (73.3%, 22/30); the remaining isolates carried either a carbapenemase ( bla _KPC-3 ) or an AmpC β-lactamase ( bla _CMY-2 ). Nanopore sequencing identified bla _CTX-M-15 harbored on two types of IncFIB(Kpn3) antimicrobial resistance (AMR) plasmids, either with or without the conjugative tra gene cluster. Interestingly, the KL1 locus, associated with canonical hypervirulent K. pneumoniae strains, was detected in all ST2059 isolates. Further analysis of public genomic data showed that the KL1 locus is widely distributed across KpSC. KL1 phylogenetic analyses indicated frequent intrasubspecies recombination but limited intersubspecies exchange of KL1. The identification of the dominant MDR K. quasipneumoniae subsp. similipneumoniae KL1-ST2059 clone in the U.S. underscores the importance of ongoing genomic surveillance. IMPORTANCE: Klebsiella quasipneumoniae subsp. similipneumoniae is an underrecognized member of the Klebsiella pneumoniae species complex that is frequently misidentified in clinical laboratories, leading to an incomplete understanding of its role in antimicrobial resistance. In this study, we used large-scale genomic surveillance of community-associated multidrug-resistant isolates across the U.S. to identify this subspecies as a reservoir of clinically relevant resistance plasmids. Notably, we detected a widely distributed ST2059 lineage carrying the K1 capsular locus, a feature traditionally associated with hypervirulent K. pneumoniae . These findings highlight the convergence of resistance and virulence-associated traits in an overlooked species and underscore the need for genomic surveillance to monitor emerging high-risk lineages in community settings.

Cefiderocol (CFDC) is a siderophore-conjugated cephalosporin developed to overcome multidrug resistance in gram-negative bacteria. Despite its unique iron-dependent entry mechanism, CFDC resistance has emerged in Klebsiella pneumoniae , primarily driven by alterations in siderophore transport and β-lactamase evolution; however, the broader intrinsic resistome that supports CFDC tolerance remains incompletely defined. Here, we performed high-density transposon mutagenesis (Tn-Seq) in the epidemic ST258 K. pneumoniae to map the functional genetic landscape of CFDC susceptibility. Tn-Seq identified siderophore uptake components ( tonB and cirA ) as the dominant determinants of CFDC resistance. In contrast, disruption of genes involved in peptidoglycan recycling ( ampG and ldcA ), synthesis ( mrcB and lpoB ), and enterobacterial common antigen biosynthesis ( wzxE and wzyE ), as well as deletion of plasmid-encoded bla _KPC-3 , increased CFDC susceptibility. In a CFDC-resistant Δ tonB strain, targeting these envelope homeostasis pathways yielded only limited resensitization relative to the siderophore-competent parental strain. Deletion of bla _KPC-3 produced the greatest increase in susceptibility, reducing the CFDC MIC fourfold. This pattern is consistent with a model in which reduced CFDC influx in the Δ tonB background lowers intracellular drug exposure to levels at which the otherwise limited anti-CFDC activity of KPC β-lactamase becomes sufficient to drive resistance. Together, these data support a multi-layered functional network governing cefiderocol susceptibility in ST258 K. pneumoniae , in which iron-dependent drug uptake constrains intracellular exposure, while β-lactamase activity and intrinsic envelope homeostasis pathways further shape bacterial tolerance once cefiderocol enters the cell.

Staphylococcus aureus remains a major public health concern due to its capacity for colonization, transmission, and antimicrobial resistance. Both methicillin-resistant (MRSA) and methicillin-susceptible S. aureus (MSSA) serve as reservoirs for resistance determinants, including those conferring reduced susceptibility to topical decolonization agents. We analyzed 2,330 S. aureus isolates collected between 2022 and 2025 from hospitals and community clinics in New Jersey and New York City, including 962 MRSA infection isolates and 1,368 nasal carriage isolates (310 MRSA and 1,058 MSSA) obtained from patients upon admission to intensive care, neonatal intensive care, and transplant units. Whole-genome sequencing and PCR characterized clonal lineages, plasmid structures, and resistance determinants, with focus on mupA , which mediates high-level mupirocin resistance. MRSA infection isolates were dominated by clonal complexes CC8 (63%), CC5 (21%), and CC30 (10%), whereas MSSA carriage isolates were more diverse, with CC398 most prevalent (15%). The mupA gene was detected in 26% of MRSA infection isolates, 50% of MRSA carriage isolates, and 36% of MSSA carriage isolates. Pairwise single-nucleotide polymorphism (SNP) analysis revealed most mupA variants differed by 0–2 SNPs across diverse lineages, consistent with recent horizontal dissemination. Long-read sequencing identified three major mupA -carrying plasmid groups—conjugative, mobilizable, and non-mobilizable—frequently co-harboring qacC , aadD1 , msr(A ), and mph(C ). Non-mobilizable plasmids were confined to CC8, while mobilizable and conjugative variants circulated across multiple lineages. The high prevalence of mupA -bearing plasmids co-associated with quaternary ammonium resistance genes raises concerns regarding empiric mupirocin- and chlorhexidine-based decolonization strategies. Genomic surveillance and rapid resistance screening may be critical to preserving topical agent efficacy.

Background: Fluoroquinolone prophylaxis during autologous stem cell transplantation (aSCT) reduces the risk of fever but raises the risk of bloodstream infection (BSI) with fluoroquinolone-resistant Enterobacterales (FRE). We performed a prospective cohort study to detect the presence and potential gain or loss of colonic FRE colonization using serial sampling before and after aSCT in a uniform population of patients with a diagnosis of multiple myeloma. Methods: Eligible subjects underwent aSCT after conditioning with dose-intense melphalan, 200 mg/m². Peri-anal swabs were obtained before aSCT, upon hospital discharge, and 12–16 weeks after transplantation. Samples were cultured in tryptic soy broth supplemented with either ciprofloxacin or ceftriaxone with subsequent plating onto selective chromogenic agar designed to facilitate recovery and differentiation of Enterobacterales. Results: FRE colonization on pre-transplant sampling was detected for 23 of 117 subjects (19.7%) and 29 of 98 (29.6%) subjects at hospital discharge after a course of fluoroquinolone (116/117 subjects) prophylaxis (p < 0.001) and 28 of 92 (30.4%) subjects at 12–16 weeks. Including all three sampling time points, 48 of 117 subjects (41.0%) tested positive for FRE colonization. In total, 58 of the 90 FRE isolates (64.4%) from 48 subjects expressed extended-spectrum beta-lactamase (ESBL). Three FRE-colonized subjects developed FRE BSI. Bloodstream isolates for two subjects were identical to the organisms identified on pre-transplant sampling. Conclusions: We hypothesize that fluoroquinolone prophylaxis of subjects with undetected low levels of FRE colonization allows the expansion of the FRE population, placing subjects at risk of BSI with fluoroquinolone-resistant (and ESBL-expressing) Enterobacterales. Pre-transplant testing for FRE colonization permits patient-specific design of prophylactic and empiric antibiotic regimens.

Cefiderocol (CFDC) is a siderophore-conjugated cephalosporin developed to overcome multidrug resistance in Gram-negative bacteria. Despite its unique iron-dependent entry mechanism, CFDC resistance has emerged in Klebsiella pneumoniae , primarily driven by alterations in siderophore transport and β-lactamase evolution; however, the broader intrinsic resistome that supports CFDC tolerance remains incompletely defined. Here, we performed high-density transposon mutagenesis (Tn-Seq) in the epidemic ST258 K. pneumoniae to map the functional genetic landscape of CFDC susceptibility. Tn-Seq identified siderophore uptake components ( tonB and cirA ) as the dominant determinants of CFDC resistance. In contrast, disruption of genes involved in peptidoglycan recycling ( ampG, ldcA ), synthesis ( mrcB, lpoB ) and enterobacterial common antigen (ECA) biosynthesis ( wzxE, wzyE ), as well as deletion of plasmid-encoded bla _KPC-3 , increased CFDC susceptibility. In a CFDC-resistant Δ tonB strain, targeting these envelope homeostasis pathways yielded only limited resensitization relative to the siderophore-competent parental strain. Deletion of bla _KPC-3 produced the greatest increase in susceptibility, reducing the CFDC MIC four-fold. This pattern is consistent with a model in which reduced CFDC influx in the Δ tonB background lowers intracellular drug exposure to levels at which the otherwise limited anti-CFDC activity of KPC β-lactamase becomes sufficient to drive resistance. Together, these data define a hierarchical genetic architecture for CFDC resistance in ST258 K. pneumoniae , in which iron-dependent drug uptake is primary, β-lactamase activity is secondary, and intrinsic envelope stress buffering shapes bacterial fitness once CFDC enters the cell.