Spread of carbapenemase-producing Morganella spp from 2013 to 2021: a comparative genomic study.

BACKGROUND: Morganella spp are opportunistic pathogens involved in various infections. Intrinsic resistance to multiple antibiotics (including colistin) combined with the emergence of carbapenemase producers reduces the number of active antimicrobials. The aim of this study was to characterise genetic features related to the spread of carbapenem-resistant Morganella spp. METHODS: This comparative genomic study included extensively drug-resistant Morganella spp isolates collected between Jan 1, 2013, and March 1, 2021, by the French National Reference Center (NRC; n=68) and European antimicrobial resistance reference centres in seven European countries (n=104), as well as one isolate from Canada, two reference strains from the Pasteur Institute collection (Paris, France), and two colistin-susceptible isolates from Bicêtre Hospital (Kremlin-Bicêtre, France). The isolates were characterised by whole-genome sequencing, antimicrobial susceptibility testing, and biochemical tests. Complete genomes from GenBank (n=103) were also included for genomic analysis, including phylogeny and determination of core genomes and resistomes. Genetic distance between different species or subspecies was performed using average nucleotide identity (ANI). Intrinsic resistance mechanisms to polymyxins were investigated by combining genetic analysis with mass spectrometry on lipid A. FINDINGS: Distance analysis by ANI of 275 isolates identified three groups: Morganella psychrotolerans, Morganella morganii subspecies sibonii, and M morganii subspecies morganii, and a core genome maximum likelihood phylogenetic tree showed that the M morganii isolates can be separated into four subpopulations. On the basis of these findings and of phenotypic divergences between isolates, we propose a modified taxonomy for the Morganella genus including four species, Morganella psychrotolerans, Morganella sibonii, Morganella morganii, and a new species represented by a unique environmental isolate. We propose that M morganii include two subspecies: M morganii subspecies morganii (the most prevalent) and M morganii subspecies intermedius. This modified taxonomy was supported by a difference in intrinsic resistance to tetracycline and conservation of metabolic pathways such as trehalose assimilation, both only present in M sibonii. Carbapenemase producers were mostly identified among five high-risk clones of M morganii subspecies morganii. The most prevalent carbapenemase corresponded to NDM-1, followed by KPC-2, and OXA-48. A cefepime-zidebactam combination was the most potent antimicrobial against the 172 extensively drug-resistant Morganella spp isolates in our collection from different European countries, which includes metallo-ß-lactamase producers. Lipid A analysis showed that the intrinsic resistance to colistin was associated with the presence of L-ARA4N on lipid A. INTERPRETATION: This global characterisation of, to our knowledge, the widest collection of extensively drug-resistant Morganella spp highlights the need to clarify the taxonomy and decipher intrinsic resistance mechanisms, and paves the way for further genomic comparisons. FUNDING: None.

Evaluation of the French novel disc diffusion-based algorithm for the phenotypic screening of carbapenemase-producing Enterobacterales.

OBJECTIVES: The early identification of carbapenemase-producing Enterobacterales (CPE) is required to prevent their spread and initiate proper therapy. Accordingly, it is crucial to develop efficient algorithms using susceptibility testing results to discriminate non-carbapenemase producers (non-CPE) from those that require complementary tests. In 2022, to adapt its recommendations to the evolution of CPE epidemiology (increased prevalence of OXA-244 producers), the Antibiogram Committee of the French Society of Microbiology (CA-SFM) proposed a new algorithm for the screening of CPE. We compared this algorithm to the former algorithm (2015-2021). METHODS: From July 2022 to January 2023, all nonduplicate enterobacterial isolates referred to French National Reference Centre for carbapenemase detection (n = 518) were subjected to the former CA-SFM algorithm (2015 to 2021) using inhibition diameters of ertapenem, ticarcillin-clavulanate, temocillin and meropenem or imipenem, and the novel CA-SFM algorithm (since 2022) using inhibition diameters of ceftazidime-avibactam, temocillin, and meropenem or imipenem. RESULTS: Sensitivity, specificity, negative predictive value, and positive predictive value were of 80.8% (CI95 76.3%-84.6%), 66.2% (58.1%-73.5%), 59.3% (51.5%-66.6%), and 85.0% (80.7% - 88.5%) for the old CA-SFM algorithm and 97.8% (95.5%-99.0%), 45.5% (37.5%-53.7%), 89.7% (80.3%-95.2%), and 80.9% (76.9%-84.4%) for the novel CA-SFM algorithm. DISCUSSION: The novel CA-SFM algorithm possesses the best performance for the screening of CPE particularly in countries with a high prevalence of OXA-48-like producers.

Cholesterol accumulation induced by acetylated LDL exposure modifies the enzymatic activities of the TCA cycle without impairing the respiratory chain functionality in macrophages.

The unregulated uptake of modified low-density lipoproteins (LDL) by macrophages leads to foam cell formation, promoting atherosclerotic plaque progression. The cholesterol efflux capacity of macrophages by the ATP-Binding Cassette transporters depends on the ATP mitochondrial production. Therefore, the mitochondrial function maintenance is crucial in limiting foam cell formation. Thus, we aimed to investigate the mechanisms involved in the mitochondrial dysfunction that may occur in cholesterol-laden macrophages. We incubated THP-1 macrophages with acetylated LDL (acLDL) to obtain cholesterol-laden cells or with mildly oxidized LDL (oxLDL) to generate cholesterol- and oxidized lipids-laden cells. Cellular cholesterol content was measured in each condition. Mitochondrial function was evaluated by measurement of several markers of energetic metabolism, oxidative phosphorylation, oxidative stress, mitochondrial biogenesis and dynamics. OxLDL-exposed macrophages exhibited a significantly reduced mitochondrial respiration and complexes I and III activities, associated to an oxidative stress state and a reduced mitochondrial DNA copy number. Meanwhile, acLDL-exposed macrophages featured an efficient oxidative phosphorylation despite the decreased activities of aconitase, isocitrate dehydrogenase and α-ketoglutarate dehydrogenase. Our study revealed that mitochondrial function was differently impacted according to the nature of modified LDL. Exposure to cholesterol and oxidized lipids carried by oxLDL leads to a mitochondrial dysfunction in macrophages, affecting the mitochondrial respiratory chain functional capacity, whereas the cellular cholesterol enrichment induced by acLDL exposure results in a tricarboxylic acid cycle shunt while maintaining mitochondrial energetic production, reflecting a metabolic adaptation to cholesterol intake. These new mechanistic insights are of direct relevance to the understanding of the mitochondrial dysfunction in foam cells.