Detection of a NDM-1-producing Klebsiella pneumoniae (ST22) Clinical Isolate at a Pediatric Hospital in Mexico

Abstract

To The Editors: A 12-year-old male patient was hospitalized on April 20, 2013 at the pediatric hospital “Hospital del Niño y del Adolescente Morelense”, in Emiliano Zapata, Morelos, Mexico. The patient had the following diagnoses on admission: infantile cerebralpalsy, secondary malnutrition, secondary thrombocytopenia, pneumonia, dehydration and seizures and with a urinary probe installed. The antimicrobial treatment was ampicillin, cefuroxime, ceftriaxone, clindamycin and amikacin. Candida tropicalis was isolated from a urinary tract infection and fluconazole was given. Subsequently, a Klebsiella pneumoniae isolate (3407-2) was isolated (April 25, 2013) from the urine, and the antimicrobial treatment was changed to ciprofloxacine and amikacin. The patient died after being hospitalized for a period of 1 month and eighteen days. The isolates were identified using the VITEK 2 compact system (BioMérieux, Durham, NC). The K. pneumoniae isolate displayed an unusual multidrug resistance profile and the resistance profile was verified using the disk diffusion method according to Clinical Laboratory Standard Institute.1 MultiLocus sequence typing was carried out2 and it was found that the isolate belonged to an unusual sequence type (ST) 22, which has not been previously related to human infections (www.pasteur.fr/mlst). The minimal inhibitory concentration was determined by broth microdilution following Clinical Laboratory Standard Institute recommendations.1 The currently available susceptibility breakpoints for colistin (2 mg/L) according to the European Committee on Antimicrobial Susceptibility Testing (www.eucast.org) were used. The clinical isolate pointed to a multidrug resistance profile according to minimal inhibitory concentrations obtained to: imipenem (32 mg/L), meropenem (64 mg/L), cefotaxime (>256 mg/L), ceftazidime (>256 mg/L), piperaciline (>256 mg/L), ciprofloxacin (4 mg/L), gentamicin (>64 mg/L) and colistin (8 mg/L). Screening for carbapenemases were carried out by polymerase chain reaction (PCR) using the following generic primers: NDM,3 IMP, VIM, SIM, GIM, and SPM-type carbapenemase families and for plasmid-mediated quinolone resistance (PMQR) qnrA, B, S, C and D, aac(6’)-Ib and qepA genes.4 PCR screening showed the presence of NDM-, qnrB-type and aac(6’)-Ib genes; the nucleotide sequence was obtained on a PerkinElmer/Applied Biosystems 3730 DNA sequencing system (Applied Biosystems, PerkinElmer, Foster City, CA), and it confirmed the presence of MβL NDM-1, qnrB5 gene and the non-PCR-digestion indicated the ciprofloxacin resistance aac(6’)-Ib-cr gene. The plasmid profile was determined according to the Kaiser method;4 it corresponded to 3 plasmids of 150, 7 and 3 kb and the high-molecular-weight plasmid (150 kb) which could self-transfer by mating;4 only the NDM-1 carbapenemase gene was transferred. In this transconjugant (T3407-2), the plasmid incompatibility groups were identified by PCR-replicon typing,4 and corresponded to Inc-FII and Inc-K groups. The minimal inhibitory concentration obtained in the transconjugant was very similar to the clinical isolate: imipenem (8 mg/L), meropenem (8 mg/L), cefotaxime (256 mg/L), ceftazidime (256 mg/L), piperaciline (256 mg/L), gentamicin (16 mg/L) and colistin (4 mg/L), and they showed only susceptibility to ciprofloxacine (0.06 mg/L) and was consistent with the absence of aac(6’)-Ib-cr gene. These findings emphasize the need for continuous molecular epidemiologic surveillance, in pediatric and nonpediatric hospitals in Mexico. Humberto Barrios, PhD Jesus Silva-Sanchez, PhD Fernando Reyna-Flores, Biol Alejandro Sanchez-Perez, Quim Instituto Nacional de Salud Pública (INSP), CISEI, Cuernavaca Domingo Sanchez-Francia, QBP Jesús A. Aguirre-Torres, TLC José Sánchez-Rogel, TLC Hospital del Niño y el Adolescente Morelense Emiliano Zapata, Morelos. México Ulises Garza-Ramos, PhD Instituto Nacional de Salud Pública (INSP), CISEI, Cuernavaca