Repeatability and reproducibility of the wzi high resolution melting-based clustering analysis for Klebsiella pneumoniae typing
Ajay Ratan Pasala,
Matteo Perini,
Aurora Piazza,
Simona Panelli,
Domenico Di Carlo,
Cristian Loretelli,
Alessandra Cafiso,
Sonia Inglese,
Floriana Gona,
Daniela Maria Cirillo,
Gian Vincenzo Zuccotti,
Francesco Comandatore
Affiliations
Ajay Ratan Pasala
Department of Biomedical and Clinical Sciences “L. Sacco”, Università di Milano, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”
Matteo Perini
Department of Biomedical and Clinical Sciences “L. Sacco”, Università di Milano, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”
Aurora Piazza
Department of Clinical Surgical Diagnostic and Pediatric Sciences, Microbiology and Clinical Microbiology Unit, University of Pavia
Simona Panelli
Department of Biomedical and Clinical Sciences “L. Sacco”, Università di Milano, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”
Domenico Di Carlo
Department of Biomedical and Clinical Sciences “L. Sacco”, Università di Milano, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”
Cristian Loretelli
International Center for T1D, Pediatric Clinical Research Center “Romeo Ed Enrica Invernizzi”, Department of Biomedical and Clinical Science L. Sacco, Università di Milano
Alessandra Cafiso
Department of Veterinary Medicine, Università di Milano
Sonia Inglese
Microbiology and Virology Unit, Fondazione IRCCS Policlinico San Matteo
Floriana Gona
Laboratorio Microbiologia e Virologia-Ospedale San Raffaele Dibit
Daniela Maria Cirillo
Emerging Bacterial Pathogens Unit, Division of Immunology, Transplantation and Infectious Diseases, IRCCS San Raffaele Scientific Institute
Gian Vincenzo Zuccotti
Department of Biomedical and Clinical Sciences “L. Sacco”, Università di Milano, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”
Francesco Comandatore
Department of Biomedical and Clinical Sciences “L. Sacco”, Università di Milano, Pediatric Clinical Research Center “Romeo and Enrica Invernizzi”
Abstract High resolution melting (HRM) is a fast closed-tube method for nucleotide variant scanning applicable for bacterial species identification or molecular typing. Recently a novel HRM-based method for Klebsiella pneumoniae typing has been proposed: it consists of an HRM protocol designed on the capsular wzi gene and an HRM-based algorithm of strains clustering. In this study, we evaluated the repeatability and reproducibility of this method by performing the HRM typing of a set of K. pneumoniae strains, on three different instruments and by two different operators. The results showed that operators do not affect melting temperatures while different instruments can. Despite this, we found that strain clustering analysis, performed using MeltingPlot separately on the data from the three instruments, remains almost perfectly consistent. The HRM method under study resulted highly repeatable and thus reliable for large studies, even when several operators are involved. Furthermore, the HRM clusters obtained from the three different instruments were highly conserved, suggesting that this method could be applied in multicenter studies, even if different instruments are used.