Detection by Whole-Genome Sequencing of a Novel Metallo-ß-Lactamase Produced by Wautersiella falsenii Causing Urinary Tract Infection in Tunisia.

Wautersiella falsenii is a rarely non-fermenting Gram-negative bacterium and belongs to the Flavobacteriaceae family. This nosocomial pathogen can cause several human infections, especially among immunocompromised patients. Here, we describe the whole genome sequence of a clinical W. falsenii strain isolated from a urine sample of a 35-year-old woman with a urinary tract infection in Tunisia. We investigated its phenotype and genotype. After bacterial identification by the MALDI-TOF method, the whole-genome sequencing of this strain was performed. This isolate was not susceptible to various antibiotics, including ß-lactams, aminoglycosides, and quinolones. However, it remains susceptible to imipenem (MIC = 0.25 mg/l), ertapenem (MIC = 0.75 mg/l), and meropenem (MIC = 0.19 mg/l). Interestingly, the E-TEST® (MP/MPI) showed a reduced MIC of meropenem +/- EDTA (0.064 µg/ml). Besides, the color change from yellow to red in the ß CARBA test only after 24 hours of incubation can be interpreted in two ways. On the one hand, as a likely low expression of the gene encoding metallo-ß-lactamase. On the other hand, and more likely, it may be a false-positive result because, according to the test manufacturer's recommendations, the test should be read after 30 minutes. Perhaps, therefore, this gene is not expressed in the tested strain. Moreover, the whole-genome sequence analysis demonstrated the presence of a novel chromosomally located subclass B1 metallo-ß-lactamase EBR-like enzyme, sharing 94.92% amino acid identity with a previously described carbapenemase produced by Empedobacter brevis, EBR-1. The results also showed the detection of other antibiotic resistance genes and the absence of plasmids. So far, this study is the first report on the detection of W. falsenii in Tunisia. These findings prove that W. falsenii could be a potential reservoir of antibiotic resistance genes, e.g., ß-lactamases. Collaborative efforts and effective hygiene measures should be established to prevent the emergence of this species in our health care settings.

Occurrence of NDM-1 and VIM-2 Co-Producing Escherichia coli and OprD Alteration in Pseudomonas aeruginosa Isolated from Hospital Environment Samples in Northwestern Tunisia.

Hospital environments constitute the main reservoir of multidrug-resistant bacteria. In this study we aimed to investigate the presence of Gram-negative bacteria in one Northwestern Tunisian hospital environment, and characterize the genes involved in bacterial resistance. A total of 152 environmental isolates were collected from various surfaces and isolated using MacConkey medium supplemented with cefotaxime or imipenem, with 81 fermenter bacteria (27 Escherichia coli, and 54 Enterobacter spp., including 46 Enterobacter cloacae), and 71 non-fermenting bacteria (69 Pseudomonas spp., including 54 Pseudomonas aeruginosa, and 2 Stenotrophomonas maltophilia) being identified by the MALDI-TOF-MS method. Antibiotic susceptibility testing was performed by disk diffusion method and E-Test was used to determine MICs for imipenem. Several genes implicated in beta-lactams resistance were characterized by PCR and sequencing. Carbapenem resistance was detected among 12 isolates; nine E. coli (blaNDM-1 (n = 8); blaNDM-1 + blaVIM-2 (n = 1)) and three P. aeruginosa were carbapenem-resistant by loss of OprD porin. The whole-genome sequencing of P. aeruginosa 97H was determined using Illumina MiSeq sequencer, typed ST285, and harbored blaOXA-494. Other genes were also detected, notably blaTEM (n = 23), blaCTX-M-1 (n = 10) and blaCTX-M-9 (n = 6). These new epidemiological data imposed new surveillance strategies and strict hygiene rules to decrease the spread of multidrug-resistant bacteria in this area.