Detecting KPC-2 and NDM-1 Coexpression in Klebsiella pneumoniae Complex from Human and Animal Hosts in South America.
Felipe Vásquez-Ponce, Karine Dantas, Johana Becerra, Gregory Melocco, Fernanda Esposito, Brenda Cardoso, Larissa Rodrigues, Keila Lima, Aline V de Lima, Fábio P Sellera, Renata Mattos, Lucas Trevisoli, Marco A Vianello, Thais Sincero, Jose Di Conza, Eliana Vespero, Gabriel Gutkind, Jorge Sampaio, Nilton Lincopan
Author Information
Felipe Vásquez-Ponce: Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil. ORCID
Karine Dantas: Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil.
Johana Becerra: Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil.
Gregory Melocco: Department of Clinical Analysis, School of Pharmacy, Universidade de São Paulo, São Paulo, Brazil.
Fernanda Esposito: Department of Clinical Analysis, School of Pharmacy, Universidade de São Paulo, São Paulo, Brazil.
Brenda Cardoso: Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil.
Larissa Rodrigues: Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil.
Keila Lima: Department of Clinical Analysis, School of Pharmacy, Universidade de São Paulo, São Paulo, Brazil.
Aline V de Lima: Department of Clinical Analysis, School of Pharmacy, Universidade de São Paulo, São Paulo, Brazil.
Fábio P Sellera: Department of Internal Medicine, School of Veterinary Medicine and Animal Science, Universidade de São Paulo, São Paulo, Brazil.
Renata Mattos: Laborclin, Pinhais, Brazil.
Lucas Trevisoli: Laborclin, Pinhais, Brazil.
Marco A Vianello: Natal Garrison Hospital, Brazilian Army, Natal, Brazil.
Thais Sincero: Department of Clinical Analysis, Health Sciences Center, Federal University of Santa Catarina, Florianópolis, Brazil.
Jose Di Conza: Facultad de Farmacia y Bioquímica, Instituto de Investigaciones en Bacteriologia y Virología Molecular, Universidad de Buenos Aires, Buenos Aires, Argentina.
Eliana Vespero: Department of Pathology, Clinical and Toxicological Analysis, Health Sciences Center, University Hospital of Londrina, Paraná, Brazil.
Gabriel Gutkind: Facultad de Farmacia y Bioquímica, Instituto de Investigaciones en Bacteriologia y Virología Molecular, Universidad de Buenos Aires, Buenos Aires, Argentina. ORCID
Jorge Sampaio: Department of Clinical Analysis, School of Pharmacy, Universidade de São Paulo, São Paulo, Brazil.
Nilton Lincopan: Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil. ORCID
Reports of Gram-negative bacteria harboring multiple carbapenemase genes have increased in South America, leading to an urgent need for appropriate microbiological diagnosis. We evaluated phenotypic methods for detecting Klebsiella pneumoniae carbapenemase 2 (KPC-2) and New Delhi metallo-β-lactamase-1 (NDM-1) coexpression in members of the K. pneumoniae complex (i.e., K. pneumoniae, K. quasipneumoniae, and K. variicola) isolated from human and animal hosts, based on inhibition of ceftazidime-avibactam (CZA) and aztreonam (ATM) by dipicolinic acid (DPA), EDTA, or avibactam (AVI). While the presence of and genes was confirmed by whole-genome sequencing, PCR, and/or GeneXpert, coexpression was successfully detected based on the following: (i) a ≥5-mm increase in the zone diameter of ATM (30 µg) disks plus AVI (4 or 20 µg) and ≥4-mm and ≥10-mm increases in the zone diameters for "CZA 50" (30 µg ceftazidime [CAZ] and 20 µg AVI) and "CZA 14" (10 µg CAZ and 4 µg AVI) disks, respectively, when we added DPA (1 mg/disk) or EDTA (5 mM) in a combined disk test (CDT); (ii) a positive ghost zone (synergism) between ATM (30 µg) and CZA 50 disks and between CZA 50 and DPA (1 mg) disks, using the double-disk synergy test (DDST) at a disk-disk distance of 2.5 cm; (iii) ≥3-fold MIC reductions of ATM and CZA in the presence of AVI (4 µg/mL), DPA (500 µg/mL), or EDTA (320 µg/mL); and (iv) immunochromatography. Although our results demonstrated that inhibition by AVI, DPA, and EDTA may provide simple and inexpensive methods for the presumptive detection of coexpression of KPC-2 and NDM-1 in members of the K. pneumoniae complex, additional studies are necessary to confirm the accuracy of these methodologies by testing other Gram-negative bacterial species and other KPC and NDM variants coexpressed by WHO critical priority pathogens detected worldwide. Alerts regarding the emergence and increase of combinations of carbapenemases in in Latin America and the Caribbean have recently been issued by PAHO and WHO, emphasizing the importance of appropriate microbiological diagnosis and the effective and articulated implementation of infection prevention and control programs. In this study, we evaluated methods based on inhibition of ceftazidime (CAZ), ceftazidime-avibactam (CZA), and aztreonam (ATM) by dipicolinic acid (DPA), EDTA, and avibactam (AVI) inhibitors for the identification of KPC-2- and NDM-1-coexpression in members of the K. pneumoniae complex recovered from human and animal hosts. Our results demonstrate that inhibition by AVI, DPA, and EDTA may provide simple and inexpensive methods for the presumptive detection of coexpression of KPC-2 and NDM-1 in members of the K. pneumoniae complex.
