Emergence of OXA-48-Producing Escherichia coli Clone ST38 in Fowl

LETTER

The bla_OXA-48 gene, which encodes a carbapenem-hydrolyzing class D β-lactamase, was first described in Klebsiella pneumoniae isolates from Turkey (1). Escherichia coli strains producing OXA-48 were reported in several countries, especially in the Mediterranean basin region (2), including Lebanon (3). Different E. coli clones have successfully emerged in the world; examples are E. coli O15:K52:H1-D of sequence type 393 (ST393) (4), E. coli ST131 (5), and E. coli ST38, which is characterized by the production of the OXA-48 β-lactamase and is related to strains of the Mediterranean basin countries (2, 6). Here we report the detection of an E. coli ST38 OXA-48 producer in fowl from Lebanon.

E. coli CMUL 64 was recovered in December 2013 from the fowl species Gallus domesticus by means of a rectal swab during a study aiming to detect carbapenemase-encoding genes in fowls in Lebanon. Forty samples were collected from eight livestock farms. Carbapenem-producing isolates were selected in MacConkey medium supplemented with ertapenem (1 mg/liter). One positive carbapenemase-producing isolate was detected, and it was isolated from an alive and nonsick fowl that had not had any previous use of antibiotics or contact with any other animals.

Susceptibility testing was performed by disk diffusion testing (Sanofi-Diagnostic Pasteur, Marnes-la-Coquette, France). MICs were determined by Etest (bioMérieux, Marcy l'Etoile, France) on Mueller-Hinton agar plates. The isolate, E. coli CMUL 64, showed an extended-spectrum β-lactamase (ESBL) phenotype and was resistant to all penicillins, cefotaxime, and co-trimoxazole, intermediately susceptible to ceftazidime, aztreonam, gentamicin, tobramycin, and amikacin, and susceptible to nitrofurantoin and fluoroquinolones (7). Furthermore, this isolate showed reduced susceptibility to carbapenems. The MICs of imipenem, meropenem, and ertapenem were, respectively, 2, 0.25, and 2 μg/ml. Production of carbapenemase was verified using the biochemical Carba NP test (8).

PCR was conducted using previously described primers (9) to confirm the presence of β-lactamases genes. E. coli CMUL 64 carried, in addition to the bla_OXA-48 gene, a bla_CTX-M gene together with a bla_TEM gene. Sequencing showed that the two ESBLs were TEM-1 and CTX-M-14. Mating-out assays performed as previously described (1) allowed us to obtain E. coli transconjugants producing the CTX-M-14 or OXA-48 β-lactamase. These transconjugants harbored two plasmids (50 and approximately 62 kb) encoding the ESBL or the OXA-48 β-lactamase (Table 1). To further characterize the plasmid bearing the bla_OXA-48 gene, we performed whole-genome sequencing of E. coli CMUL 64 using Illumina MiSeq and analyzed the genetic platform of the bla_OXA-48 gene. The genetic organization of the bla_OXA-48 gene, bracketed with transposons, was found to be 100% similar to the K. pneumoniae E71T plasmid bearing bla_OXA-48 (GenBank accession number KC335143), as shown in Fig. 1. However, we were unable to type the plasmid to a known incompatibility group, including the L/M incompatibility group, as all the results for the typing were negative. We intend to fully characterize this plasmid. Phylogenetic grouping performed by PCR as previously described (10) showed that this isolate belongs to group B2, corresponding to an extraintestinal and virulent strain. Multilocus sequence typing was performed as previously described (11) and showed that this isolate belonged to ST38. E. coli isolates belonging to ST38 producing the OXA-48 β-lactamase had previously been detected in the Mediterranean basin countries, including Lebanon (2). All these isolates were clinically isolated from humans. Recently, E. coli OXA-48 was also detected in dogs (9) in Germany.

E. coli OXA-48 ST38 represents an emerging clone that is disseminating in the Mediterranean basin countries; however, this clone has never been described in animals in that region. Here we report the detection of E. coli OXA-48 clone ST38 in fowl.

The emergence of carbapenemase-producing Gram-negative bacteria from animals assumes that the dissemination of this clone in the community may have a zoonotic origin since the same clone was found in several Mediterranean countries in clinical specimens. These findings need further investigation, since resistance to carbapenems is not commonly tested for in veterinary clinical isolates.