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Few last-line antimicrobial agents effectively treat infections caused by New Delhi metallo-[beta]-lactamase (NDM)-producing Enterobacterales (1). Cefiderocol is a novel synthetic conjugate siderophore cephalosporin that is more stable against [beta]-lactamase hydrolysis [...]

OXA-244, an R214G variant of OXA-48, is silently spreading worldwide likely because of difficulties in detection using classical screening media. Here, we characterized two clinical isolates of Escherichia coli and Citrobacter youngae that displayed reduced susceptibility to carbapenems but were lacking significant carbapenemase activity as revealed by negative Carba NP test results. However, positive test results were seen for OXA-48-like enzymes by lateral flow immunoassays. WGS revealed the presence of a blaOXA-181-like gene that codes for OXA-484, an R214G variant of OXA-181. BlaOXA-484 gene was located on a 58.4-kb IncP1-like plasmid (pN-OXA-484), that upon transfer into E. coli HB4 with impaired permeability, conferred carbapenem and temocillin resistance (MICs > 32 mg/L). E. coli TOP10 (pTOPO-OXA-484) revealed reduced MICs in most substrates as compared to E. coli TOP10 (pTOPO-OXA-181), especially for imipenem (0.25 mg/L versus 0.75 mg/L) and temocillin (16 mg/L versus 1028 mg/L). Catalytic efficiencies of OXA-484 were reduced as compared to OXA-181 for most ß-lactams including imipenem and temocillin with 27.5- and 21.7-fold reduction, respectively. Molecular modeling confirmed that the salt bridges between R214, D159, and the R1 substituent's carboxylate group of temocillin were not possible with G214 in OXA-484, explaining the reduced affinity for temocillin. In addition, changes in active site's water network may explain the decrease in hydrolysis rate of carbapenems. OXA-484 has weak imipenem and temocillin hydrolytic activities, which may lead to silent spread due to underdetection using selective screening media or biochemical imipenem hydrolysis confirmatory tests. [ABSTRACT FROM AUTHOR]

This article discusses a study on antimicrobial resistance (AMR) in Lebanon, specifically focusing on the presence of the blaNDM-5 gene in E. coli isolates. The study found that blaNDM-5-harbouring E. coli ST617 was present in Syrian refugees, their animals, and the camp environment, posing a health risk to both the refugees and the hosting community. The study emphasizes the importance of integrated surveillance programs and One Health studies to address AMR in vulnerable populations and highlights the role of war, mass migration, and displacement in the spread of AMR. Funding for the study was provided by various organizations, and ethical considerations were taken into account. [Extracted from the article]

Background As carbapenemase-producing Enterobacterales are increasingly reported worldwide, their rapid detection is crucial to reduce their spread and prevent infections and outbreaks. Lateral flow immunoassays (LFIAs) have become major tools for the detection of carbapenemases. However, as for most commercially available assays, only the five main carbapenemases are targeted. Objectives Here, we have developed and evaluated an LFIA prototype for the rapid and reliable detection of the increasingly identified GES-type β-lactamases. Methods The GES LFIA was validated on 103 well-characterized Gram-negative isolates expressing various β-lactamases grown on Mueller–Hinton (MH) agar, chromogenic, and chromogenic/selective media. Results The limit of detection of the assay was 106 cfu per test with bacteria grown on MH agar plates. GES LFIA accurately detected GES-type β-lactamases irrespective of the culture media and the bacterial host. The GES LFIA was not able to distinguish between GES-ESBLs and GES-carbapenemases. Because GES enzymes are still rare, their detection as an ESBL or a carbapenemase remains important, especially because extensive use of carbapenems to treat ESBL infections may select for GES variants capable of hydrolysing carbapenems. Conclusions The GES LFIA is efficient, rapid and easy to implement in the routine workflow of a clinical microbiology laboratory for the confirmation of GES-type β-lactamases. Combining it with immunochromatographic assays targeting the five main carbapenemases (KPC, NDM, VIM, IMP and OXA-48) would improve the overall sensitivity for the most frequently encountered carbapenemases and ESBLs, especially in non-fermenters. [ABSTRACT FROM AUTHOR]

Easy and robust antimicrobial susceptibility testing (AST) methods are essential in clinical bacteriology laboratories (CBL) in low-resource settings (LRS). We evaluated the Beckman Coulter MicroScan lyophilized broth microdilution panel designed to support Médecins Sans Frontières (MSF) CBL activity in difficult settings, in particular with the Mini-Lab. We evaluated the custom-designed MSF MicroScan Gram-pos microplate (MICPOS1) for Staphylococcus and Enterococcus species, MSF MicroScan Gram-neg microplate (MICNEG1) for Gram-negative bacilli, and MSF MicroScan Fastidious microplate (MICFAST1) for Streptococci and Haemophilus species using 387 isolates from routine CBLs from LRS against the reference methods. Results showed that, for all selected antibiotics on the three panels, the proportion of the category agreement was above 90% and the proportion of major and very major errors was below 3%, as per ISO standards. The use of the Prompt inoculation system was found to increase the MIC and the major error rate for some antibiotics when testing Staphylococci. The readability of the manufacturer's user manual was considered challenging for low-skilled staff. The inoculations and readings of the panels were estimated as easy to use. In conclusion, the three MSF MicroScan MIC panels performed well against clinical isolates from LRS and provided a convenient, robust, and standardized AST method for use in CBL in LRS. [ABSTRACT FROM AUTHOR]

Objectives: Detection of carbapenemase-producing Enterobacterales (CPEs) is sometimes difficult with AmpC-hyperproducing Enterobacterales (AHEs), as they may falsely be classified as CPEs. Here, we present a rapid Carbapenem Inactivation Method (rCIM) optimized for AmpC producers (rCIM-A) that allows rapid and easy discrimination between AHEs and CPEs.Methods: Enterobacterales (n = 249), including natural AmpC producers, AHEs, CPEs and non-carbapenemase-producing carbapenem-resistant control strains were evaluated, using Carba NP, rCIM and rCIM-A. The rCIM-A differs from the rCIM by the addition of cloxacillin (400 μg/mL) to the initial antibiotic incubation step.Results: The rCIM-A yielded a sensitivity and specificity of 84.26% (95% CI: 76.00%-90.55%) and 99.29% (95% CI: 96.11%-99.98%), respectively, while those of the rCIM were 86.11% (95% CI: 78.13%-92.01%) and 80.85% (95% CI: 73.38%-86.99%), respectively; those of Carba NP were lower at 84.04% (95% CI: 75.05%-90.78%) and 91.37% (95% CI: 85.41%-95.46%), respectively, due to indeterminate results. The rCIM-A was capable of discriminating between AHEs and true CPEs, but still failed to identify OXA-23-producing Proteus mirabilis isolates and remained only partially reliable for identifying IMI-like producers and a few MBL (2 NDM-1, 1 LMB-1, 1 TMB-1 and 1 IMP-13) producers. One chromosomally encoded AmpC variant, MIR-10, gave repeatedly positive results using all three tests and was thus considered a false positive.Conclusions: Specificity for AHEs greatly improved with the rCIM-A without altering the test performance for the other resistance mechanisms. It may replace the rCIM as a cheap, easy, rapid and accurate CPE detection test. [ABSTRACT FROM AUTHOR]

Two AMR isolates were found in the soapy water used for cleaning medical devices: one I Acinetobacter indicus i expressing I bla i SB NDM-1 sb and I bla i SB OXA-235 sb genes and one I Enterobacter chengduensis i expressing a I bla i SB CTX-M-15 sb gene. MDR Gram-negatives constitute a global public health concern, as they may cause severe, difficult-to-treat and often deadly infections, such as bloodstream infections and pneumonia. Carbapenem-resistant (CR) I Acinetobacter i spp. I Pseudomonas i spp. and Enterobacterales are among the most worrisome.[1] Even though these bacteria are increasingly described worldwide, epidemiological studies are very scarce from the African continent, except in Northern countries. [Extracted from the article]

OXA-48 has rapidly disseminated worldwide and become one of the most common carbapenemases in many countries with more than 45 variants reported with, in some cases, significant differences in their hydrolysis profiles. The R214 residue, located in the ß5-ß6 loop, is crucial for the carbapenemase activity, as it stabilizes carbapenems in the active site and maintains the shape of the active site through interactions with D159. In this study, we have characterized a novel variant of OXA-48, OXA-933 with a single D159N change. To evaluate the importance of this residue, point mutations were generated (D159A, D159G, D159K, and D159W), kinetic parameters of OXA-933, OXA-48 D159G, and OXA-48 D159K were determined and compared to those of OXA-48 and OXA-244. The bla OXA-933 gene was borne on Tn 2208 , a 2,696-bp composite transposon made of two IS 1 elements surrounded by 9 bp target site duplications and inserted into a non-self-transmissible plasmid pOXA-933 of 7,872 bp in size. Minimal inhibitory concentration values of E. coli expressing the bla OXA-933 gene or of its point mutant derivatives were lower for carbapenems (except for D159G) as compared to those expressing the bla OXA-48 gene. Steady-state kinetic parameters revealed lower catalytic efficiencies for expanded spectrum cephalosporins and carbapenems. A detailed structural analysis confirmed the crucial role of D159 in shaping the active site of OXA-48 enzymes by interacting with R214. Our work further illustrates the remarkable propensity of OXA-48-like carbapenemases to evolve through mutations at positions outside the β5-β6 loop, but interacting with key residues of it., Competing Interests: The authors declare no conflict of interest.