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First description of NDM-1-positive Klebsiella pneumoniae in the Tunisian community

Abstract : First description of NDM-1-positive Klebsiella pneumoniae in the Tunisian community Sir, Multidrug-resistant bacteria, especially carbapenemase-producing Enterobacteriaceae, are a major public health-threat worldwide. As part of a collaborative monitoring programme, our laboratory at the University of Bordeaux has received a collection of multidrug-resistant bacterial strains to further characterise their β-lactamase content. They were sent from private Tunisian diagnostic laboratories and were collected from community patients suffering from urinary tract infection. In this context, multidrug-resistant isolate 18TA was collected in January 2018 in Sfax region from the urine of a 45-year old female with no previous hospitalisation during the preceding month and no history of recent foreign travel. Strain 18TA had been initially classified as Enterobacter spp. by biochemical tests (API 10S gallery). Following matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF/MS) (Bruker Daltonics) and confirmation by PCR amplification and sequencing of 16S rDNA, strain 18TA was re-identified as Klebsiella pneumoniae. Multilocus sequence typing (MLST) (http:// bigsdb.pasteur.fr/klebsiella/klebsiella.html) indicated that strain 18TA belonged to Sequence Type (ST), ST147. Minimum inhibitory concentrations (MICs) of various anti-microbials were determined using a BD Phoenix TM 100 automated system (BD Diagnostic Systems, Le Pont-de-Claix, France) and the results were interpreted using BD EpiCenter TM software (BD Diagnostic Systems). The MICs for ciprofloxacin and colistin were also determined by the broth microdilution method according to European Committee on Antimicrobial Susceptibility Testing 2019 guidelines (https://www.sfm-microbiologie.org/2019/05/06/ casfm-eucast-2019-v2/). Strain 18TA was resistant to all tested β-lactams, including carbapenems (Table 1). The strain was also resistant to gentamicin, tobramycin, quinolones (nalidixic acid), fluoroquinolones (cipro-floxacin) and trimethoprim/sulfamethoxazole (SXT) and showed decreased susceptibility to tigecycline (MIC = 2 mg/mL). It remained susceptible to amikacin, fosfomycin and colistin (Table 1). The imipenem/ethylene diamine tetra-acetic acid (EDTA) combined disk diffusion test was positive since the inhibition zone increased by 7 mm with the imipenem/EDTA disk compared with the imipenem disk alone, suggesting the presence of a metallo-β-lactamase (MBL) [1]. In addition, the double-disk synergy test (between amoxicillin/clavulanic acid and broad-spectrum cepha-losporins) showed the presence of an extended-spectrum β-lactamase (ESBL)-producing phenotype (data not shown). The transferability of the β-lactam resistance determinant was assessed by conjugation assay using an azide-resistant (Az R) mutant of Escherichia coli C600 as the recipient strain. Selection was performed on Mueller-Hinton agar plates supplemented with sodium azide (300 mg/mL) and ertapenem (4 mg/mL). A transfer frequency of ca. 10-4 transconjugants per donor was observed. Comparison of MICs between the transconjugant (Tc-18TA) and its recipient strain (C600 Az R) showed increased resistance not only to the tested β-lactams but also to gentamicin, tobramycin and SXT (Table 1). Total genomic DNA of strain 18TA was screened using different multiplex PCR amplifications for various β-lactamase genes (bla TEM-like , bla SHV-like , bla OXA-1-like , bla CTX-M groups 1, 2, 9, 18 and 25, bla OXA-48-like , bla KPC and bla GES and the MBL genes bla VIM , bla IMP and bla NDM) as described previously [2]. Amplification results following agarose gel electrophoresis analysis showed the presence of group 1 bla CTX-M and bla NDM genes together with bla TEM-like , bla SHV-like and bla OXA-1-like genes. Except for the bla SHV gene that was found in strain 18TA but not in the transconjugant Tc-18TA and that was attributed to the chromosomally-encoded species-specific enzyme of K. pneumoniae, the four other β-lactamases were also found in transconjugant Tc-18TA (Table 1). Amplification of the entire bla genes was performed and subsequent sequencing ((Custom DNA sequencing; Eurofins Genomics GmbH, Ebersberg, Germany) showed the presence of the narrow-spectrum β-lactamase genes bla TEM-1B and bla OXA-1 associated with the bla CTX-M-15 ESBL gene and the bla NDM-1 MBL gene both in 18TA and Tc-18TA (Table 1). Furthermore, amplifications searching for aac(3)-IIa (gentamicin and tobramycin resistance) and sul1 and dfrA1 (sulfamethoxazole and trimetho-prim resistance, respectively) were positive both in 18TA and Tc-18TA. These genes were also present in Kp3771, a ST147 NDM-1-producing K. pneumoniae strain recently recovered from a patient hospitalised in an intensive care unit of University Hospital Tahar Sfar in Tunisia [3]. NDM-1-positive K. pneumoniae strains have been previously described in Tunisia, but only from hospitalised patients [1-5]. The current study reports the first description of K. pneumoniae carrying the carbapenemase NDM-1 in the Tunisian community http://dx.
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Amel Mhaya, Rahma Trabelsi, Fatima M’zali, Dominique Bégu, Slim Tounsi, et al.. First description of NDM-1-positive Klebsiella pneumoniae in the Tunisian community. Journal of Global Antimicrobial Resistance, Elsevier, 2020, 21, pp.49-50. ⟨10.1016/j.jgar.2020.02.023⟩. ⟨hal-02565943⟩

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