Novel Variant of New Delhi Metallo-Beta-Lactamase (blaNDM-60) Discovered in a Clinical Strain of Escherichia coli from the United Arab Emirates: An Emerging Challenge in Antimicrobial Resistance

Abstract

Background/Objectives: Carbapenem resistance poses a significant health threat. This study reports the first detection and characterization of a novel variant of New Delhi metallo-β-lactamase (bla_NDM-60) in Escherichia coli from the United Arab Emirates (UAE), including its genetic context and relationship to global strains. Methods: NDM-60-producing E. coli was isolated from a rectal swab during routine screening. Characterization involved whole-genome sequencing, antimicrobial susceptibility testing, and comparative genomic analysis with 66 known NDM variants. Core genome analysis was performed against 42 global E. coli strains, including the single other reported NDM-60-positive isolate. Results: The strain demonstrated extensive drug resistance, including resistance to novel β-lactam/β-lactamase inhibitor combinations, notably taniborbactam. NDM-60 differs from the closely related NDM-5 by a single amino acid substitution (Asp202Asn) and two amino acid substitutions (Val88Leu and Met154Leu) compared to NDM-1. NDM-60 is located on a nonconjugative IncX3 plasmid. The strain belongs to sequence type 940 (ST940). Phylogenetic analysis revealed high diversity among the global ST940 strains, which carry a plethora of resistance genes and originated from humans, animals, and the environment from diverse geographic locations. Conclusions: NDM-60 emergence in the UAE represents a significant evolution in carbapenemase diversity. Its presence on a nonconjugative plasmid may limit spread; however, its extensive resistance profile is concerning. Further studies are needed to determine the prevalence, dissemination, and clinical impact of NDM-60. NDM evolution underscores the ongoing challenge in managing antimicrobial resistance and the critical importance of vigilant molecular surveillance. It also highlights the pressing demand to discover new antibiotics to fight resistant bacteria.

Keywords: Escherichia coli; NDM-60; New Delhi metallo-β-lactamase; United Arab Emirates; carbapenem resistance; whole-genome sequencing.

Figure 1

Phylogenetic analysis of bla_NDM alleles. The circular dendrogram depicts the evolutionary relationships among different NDM gene alleles, with each branch representing a distinct allele. NDM-60 is included in the red-colored cluster and marked with a star.

Figure 2

Protein homology model showing the differences between NDM variants: NDM-1 (A), NDM-5 (B), and NDM-60 (C). The protein backbone is shown as a ribbon/line. The three amino acid substitutions (Val88Leu in yellow, Met154Leu in blue, and Asp202Asn in green) in NDM-60 compared to NDM-1 are shown and labeled (C vs. A). Val88Leu and Met154Leu were also found in NDM-5 compared to NDM-1 (B vs. A).

Figure 3

Gel electrophoresis and Southern blot analysis of plasmid profiles from E. coli strains EC961 (harboring NDM-60) and BAA-2469 (harboring NDM-1). Plasmid gel image (A) shows the results of agarose gel electrophoresis, displaying plasmid profiles for each isolate, with band sizes indicated. E. coli strain 39R861 was used as a size marker control (A). (B,C) Southern blots of the same gel, hybridized with a probe specific for NDM gene (B), then hybridized with a probe specific to the IncX3 plasmid replicon (C) to localize the IncX3-bearing plasmid. Yellow arrows highlight bands where successful hybridization occurred, confirming the presence of NDM-carrying plasmids at anticipated sizes in the analyzed strains and confirming that the same plasmid in EC961 carried the IncX3 plasmid replicon. BAA-2469 is lacking IncX3 based on ATCC records; thus, it was used as a negative control in plasmid replicon experiment (C), while it worked as a positive control for the NDM gene (B).

Figure 4

Comparative genomic analysis of Escherichia coli strain EC961 (SAMN43761667) against strain RIVM_C055236 (SAMN39950546) (carrying NDM-60 gene) and MG1655 E. coli reference strain (NC_000913.3). The innermost ring (black) displays the GC content, while the next ring (green and purple) represents the GC skew (green for positive skew, and purple for negative skew). The outer colored rings depict the sequence identity compared to the reference genomes, with colors indicating 100% identity. The NDM-60 gene, associated with carbapenem resistance, is highlighted in red on the outermost ring, along with key annotated genes. These labeled genes are positioned according to their location on the genome, facilitating the visualization of genomic similarities and differences between the strains. Black-colored genes represent antibiotic resistance and survival traits.

Figure 5

Comparative genomic analysis of NDM gene context across various Escherichia coli isolates. The figure displays the genetic environment surrounding different NDM alleles, including NDM-5, NDM-17, NDM-20, NDM-35, NDM-55, NDM-56, NDM-57, and NDM-60. Key genes associated with the NDM gene are highlighted. The red and gray shadings indicate regions of sequence homology and potential gene rearrangements, respectively. Abbreviations: CutA: periplasmic divalent cation tolerance protein; DsbD: cytochrome c-type biogenesis protein DsbD, protein-disulfide reductase; Isomerase: Phosphoribosylanthranilate isomerase; Bleomycin: bleomycin-resistance protein; InsH-IS5: transposase InsH for insertion sequence element IS5; MEP: mobile element protein; QacEΔ1: small multidrug resistance (SMR) efflux transporter; QacEΔ1, quaternary ammonium compounds; UmuD: error-prone repair protein; Sul2: dihydropteroate synthase type-2-sulfonamide resistance protein.

Figure 6

Core genome SNP-based phylogenetic tree of E. coli strains belonging to ST940 (n = 42) in comparison with strain EC961 from the same ST. The tree highlights evolutionary relationships based on SNPs and is color-coded according to cgST type. Detailed annotations include strain name, geographic origin, year of isolation, source (human/animal/environment), OH serotype, and cgST type. The presence of beta-lactamase genes, including CMY, TEM, CTX-M, DHA, EC, OXA-1, and carbapenemases (NDM, and OXA-48-like), is indicated with circles: black representing presence and white indicating absence. Our strain EC961 from the UAE is marked with a black star.