First Report of an Enterobacter ludwigii Isolate Coharboring NDM-1 and OXA-48 Carbapenemases

Abstract

b E nterobacter spp. have emerged as pathogens responsible for hospital-acquired infections. Carbapenem resistance is increasingly being reported in this species, which is a matter of concern. Enterobacter spp. can produce abdominal, urinary tract, meningeal, and surgical site infections. In the present study, we report a case of Enterobacter ludwigii infection in a 17-year-old female who underwent surgery for an aneurysmal bone cyst in her left distal femur. She was treated with intravenous cefotaxime and amikacin postoperatively. She developed a purulent discharge from the surgical site after 72 h of surgery. Microbiological culture of the purulent material on two consecutive days grew a Gramnegative, motile bacillus in pure culture, which was identified to the species level as Enterobacter ludwigii by its growth on myoinositol and 3-0-methyl-D-glucopyranose and then phenotypically confirmed by the Vitek-2 test (bioMérieux, Marcy l'Etoile, France). The antibiotic susceptibility test was performed by the disc diffusion method as per Clinical and Laboratory Standards Institute (CLSI) (1) guidelines using commercial antibiotic discs (Hi Media, Mumbai, India). MICs of antibiotics were determined by Vitek-2 per CLSI breakpoints (Table 1). MICs of meropenem and imipenem determined by the Etest were Ͼ32 g/ml for the isolate. The production of metallo-␤-lactamase (MBL) was confirmed by positive results from the double-disc synergy test, modified Hodge test, combined-disc synergy test, and MBL (IP/IPI) Etest method (bioMérieux, Marcy l'Etoile, France). PCR-based screening for beta-lactamase genes (bla CTXM , bla SHV , bla TEM , and bla OXA ) and the Ambler class (B and D) carbapenemase genes bla IMP , bla VIM , bla SPM , bla GIM , bla SIM , bla NDM-1 , bla OXA-23 , bla , and bla OXA48 gave positive results for bla NDM-1 , bla CTX-M-15 , bla OXA-2 , and bla OXA-48 (2-5), confirmed by sequencing. 16S rRNA gene sequence analysis showed a sequence identity value of 99% with GenBank accession number KC139450. PCR primers were procured from Sigma-Aldrich, India. Plasmid analysis using the Kieser technique (6) revealed that E. ludwigii harbored three plasmids, with sizes of 130, 50, and 35 kb. Conjugation using E. coli J53 (azide resistant) as the recipient strain revealed the bla NDM-1 gene to be present on a 130-kb plasmid. Using the PCRbased replicon typing (PBRT) (7) method, we identified an IncA/C type (amplicon size of 465 bp) that was cotransferred with two other plasmids of 35 and 50 kb. The bla CTX-M-15 gene together with bla OXA-2 was identified on the 35-kb plasmid, which belonged to the IncFIB type and had an amplicon size of 702 bp. The bla OXA-48 gene was identified on a 50-kb plasmid that belonged to the IncFIA type and had an amplicon size of 462 bp. Although the coexistence of bla NDM-1 with different carbapenemase genes has been reported, we believe this to be the first report of the cooccurrence of bla NDM-1 and bla OXA-48 in a clinical isolate of E. ludwigii from central India. The growing emergence of ca...

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