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1. The changing epidemiology of carbapenemase-producing Klebsiella pneumoniae in Italy: Toward polyclonal evolution with emergence of high-risk lineages

Author

Di Pilato V., Errico G., Monaco M., Giani T., Del Grosso M., Antonelli A., David S., Lindh E., Camilli R., Aanensen D. M., Rossolini G. M., Pantosti A., Manso E., Pedna M. F., Mungiguerra M., Mosca A., Vailati F., Aschbacher R., Imbriani A., Sartore P., Giraldi C., Piana F., Pecile P., de Nittis R., Pini B., Mirri P., Bianchi E., Restelli A., Morelli D., Catania M. R., Barbaro A., Bernaschi P, Parisi G, Gualdi P, Dusi PA, Bona R, D'Andrea M M, Cavallo R, Lanzafame P, Sartor A, Grandesso S, Milano F, Di Pilato, V., Errico, G., Monaco, M., Giani, T., Del Grosso, M., Antonelli, A., David, S., Lindh, E., Camilli, R., Aanensen, D. M., Rossolini, G. M., Pantosti, A., Manso, E., Pedna, M. F., Mungiguerra, M., Mosca, A., Vailati, F., Aschbacher, R., Imbriani, A., Sartore, P., Giraldi, C., Piana, F., Pecile, P., de Nittis, R., Pini, B., Mirri, P., Bianchi, E., Restelli, A., Morelli, D., Catania, M. R., Barbaro, A., Bernaschi, P, Parisi, G, Gualdi, P, Dusi, Pa, Bona, R, D'Andrea, M M, Cavallo, R, Lanzafame, P, Sartor, A, Grandesso, S, and Milano, F

Subjects
Microbiology (medical), Imipenem, Klebsiella pneumoniae, Population, Microbial Sensitivity Tests, Meropenem, beta-Lactamases, Settore MED/07, Antibiotic resistance, Bacterial Proteins, Genotype, medicine, Humans, Pharmacology (medical), education, Pharmacology, Genetics, education.field_of_study, biology, Settore BIO/19, biology.organism_classification, Anti-Bacterial Agents, Klebsiella Infections, Resistome, Infectious Diseases, Italy, Multilocus sequence typing, Multilocus Sequence Typing, medicine.drug
Abstract

BackgroundPrevious studies showed that the epidemic of carbapenem-resistant Klebsiella pneumoniae (CR-KP) observed in Italy since 2010 was sustained mostly by strains of clonal group (CG) 258 producing KPC-type carbapenemases. In the framework of the National Antibiotic-Resistance Surveillance (AR-ISS), a countrywide survey was conducted in 2016 to explore the evolution of the phenotypic and genotypic characteristics of CR-KP isolates.MethodsFrom March to July 2016, hospital laboratories participating in AR-ISS were requested to provide consecutive, non-duplicated CR-KP (meropenem and/or imipenem MIC >1 mg/L) from invasive infections. Antibiotic susceptibility was determined according to EUCAST recommendations. A WGS approach was adopted to characterize the isolates by investigating phylogeny, resistome and virulome.ResultsTwenty-four laboratories provided 157 CR-KP isolates, of which 156 were confirmed as K. pneumoniae sensu stricto by WGS and found to carry at least one carbapenemase-encoding gene, corresponding in most cases (96.1%) to blaKPC. MLST- and SNP-based phylogeny revealed that 87.8% of the isolates clustered in four major lineages: CG258 (47.4%), with ST512 as the most common clone, CG307 (19.9%), ST101 (15.4%) and ST395 (5.1%). A close association was identified between lineages and antibiotic resistance phenotypes and genotypes, virulence traits and capsular types. Colistin resistance, mainly associated with mgrB mutations, was common in all major lineages except ST395.ConclusionsThis WGS-based survey showed that, although CG258 remained the most common CR-KP lineage in Italy, a polyclonal population has emerged with the spread of the new high-risk lineages CG307, ST101 and ST395, while KPC remained the most common carbapenemase.

Published
2021

2. Molecular epidemiology of KPC-producing Klebsiella pneumoniae from invasive infections in Italy: Increasing diversity with predominance of the ST512 clade II sublineage

Author

Conte, Viola, Monaco, Monica, Giani, Tommaso, D'Ancona, Fortunato, Moro, Maria Luisa, Arena, Fabio, D'Andrea, Marco Maria, Rossolini, Gian Maria, Pantosti, Annalisa, Bianchi, E., Catania, M. R., Cavalcanti, P., De Nittis, R., Dusi, P. A., Grandesso, S., Gualdi, P., Imbriani, A., Pini, B., Vincenzi, C., Meledandri, M., Agrappi, C., Parisi, G., Pollini, S., Restelli, A., Rocchetti, A., Vailati, F., Aschbacher, R., Barbaro, A., Bona, R., Chirillo, M., Corradini, S., Cuccurullo, S., De Bernochi, A., Dodi, C., Giammanco, A., Mencacci, Antonella, Milano, F., Miragliotta, G., Mungiguerra, M., Pedna, M. F., Piana, Francesca, Porcheddu, G. M., Rossi, M. R., Santino, I., Sartor, A., Sartore, P., Conte V., Monaco M., Giani T., D'Ancona F., Moro M.L., Arena F., D'Andrea M.M., Rossolini G.M., Pantosti A., Bianchi E., Catania M.R., Cavalcanti P., De Nittis R., Dusi P.A., Grandesso S., Gualdi P., Imbriani A., Pini B., Vincenzi C., Meledandri M., Agrappi C., Parisi G., Pollini S., Restelli A., Rocchetti A., Vailati F., Aschbacher R., Barbaro A., Bona R., Chirillo M., Corradini S., Cuccurullo S., De Bernochi A., Dodi C., Giammanco A., Mencacci A., Milano F., Miragliotta G., Mungiguerra M., Pedna M.F., Piana F., Porcheddu G.M., Rossi M.R., Santino I., Sartor A., Sartore P., Conte, V, Monaco, M, Giani, T, D'Ancona, F, Moro, Ml, Arena, F, D'Andrea, Mm, Rossolini, Gm, Pantosti, A, AR-ISS Study Group on Carbapenemase-Producing K., pneumoniae: Bianchi E, Catania, Mr, Cavalcanti, P, De Nittis, R, Dusi, Pa, Grandesso, S, Gualdi, P, Imbriani, A, Pini, B, Vincenzi, C, Meledandri, Alessio, Agrappi, C, Parisi, G, Pollini, S, Restelli, Ugo, Rocchetti, Romualdo, Vailati, F, Aschbacher, R, Barbaro, A, Bona, R, Chirillo, M, Corradini, S, Cuccurullo, S, De Bernochi, A, Dodi, C, Giammanco, Anna, Mencacci, A, Milano, F, Miragliotta, Giuseppe, Mungiguerra, M, Pedna, Mf, Piana, Daniela, Porcheddu, Gm, Rossi, Mr, Santino, I, Sartor, A, and Sartore, P.

Subjects
0301 basic medicine, Microbiology (medical), gel, Klebsiella pneumoniae, genotype, 030106 microbiology, Population, cross-sectional studies, pulsed-field, multilocus sequence typing, electrophoresis, gel, pulsed-field, humans, italy, klebsiella infections, klebsiella pneumoniae, molecular epidemiology, serogroup, beta-lactamases, genetic variation, Settore BIO/19 - Microbiologia Generale, Cross-Sectional Studies, Electrophoresis, Gel, Pulsed-Field, Humans, Italy, Klebsiella Infections, Molecular Epidemiology, Multilocus Sequence Typing, Serogroup, beta-Lactamases, Genetic Variation, Genotype, Pharmacology, Pharmacology (medical), Infectious Diseases, law.invention, Microbiology, beta-Lactamase, 03 medical and health sciences, law, Pulsed-field gel electrophoresis, Typing, education, Polymerase chain reaction, Cross-Sectional Studie, education.field_of_study, biology, Molecular epidemiology, biochemical phenomena, metabolism, and nutrition, biology.organism_classification, Settore MED/07 - Microbiologia e Microbiologia Clinica, bacterial infections and mycoses, Virology, electrophoresis, Multilocus sequence typing, Human, Klebsiella Infection
Abstract

Objectives The spread of carbapenem-resistant Enterobacteriaceae (CRE) represents one of the most worrisome problems for clinical medicine worldwide. In Italy, the Antibiotic-Resistance-Istituto Superiore di Sanita surveillance network, in collaboration with the Committee for Antimicrobial Agents of the Italian Society of Clinical Microbiologists, promoted a study to investigate the carbapenem-resistance mechanisms, clonal relatedness and capsular typing of a recent collection of carbapenem-resistant Klebsiella pneumoniae (CR-KP). Methods A total of 17 laboratories distributed across Italy collected all consecutive non-replicate CR-KP isolated from invasive infections during two different study periods (2011-12 and 2013). Carbapenemase genes were searched for by filter hybridization and confirmed by PCR and sequencing. KPC-producing K. pneumoniae (KPC-KP) were typed by PFGE and MLST. Capsular types were identified by wzi gene typing. Results Of the collected K. pneumoniae isolates (n = 461), the overall proportion of CR-KP was 36.2% (n = 167). The majority (97%) of the CR-KP were positive for the blaKPC gene. Among the KPC-KP population, nine different STs were detected with the majority of isolates (94%) belonging to the clonal group (CG) 258. A subpopulation that belonged to ST512 and showed an identical PFGE profile represented the majority (57%) of KPC-KP strains, with a countrywide distribution. Capsular characterization showed the predominance of the wzi154, cps-2 capsular type (88.8% of all CG258 strains). ST258 strains were associated with both cps-1 and cps-2 capsular types, while ST512 was associated with cps-2 only. Conclusions Although a trend to a polyclonal evolution of the Italian KPC-KP was noted, this study showed that the KPC-KP population remained largely oligoclonal with the wide diffusion of an ST512 lineage carrying cps-2 capsular type and producing the KPC-3 enzyme.

Published
2016

3. Comparative Analysis of Five Multiplex RT-PCR Assays in the Screening of SARS-CoV-2 Variants.

Author

De Pace V, Bruzzone B, Orsi A, Ricucci V, Domnich A, Guarona G, Randazzo N, Stefanelli F, Battolla E, Dusi PA, Lillo F, and Icardi G

Abstract

The rapid and presumptive detection of SARS-CoV-2 variants may be performed using multiplex RT-PCR assays. The aim of this study was to evaluate the diagnostic performance of five qualitative RT-PCR tests as compared with next-generation sequencing (NGS). We retrospectively examined a multi-variant panel ( n = 72) of SARS-CoV-2-positive nasopharyngeal swabs categorized as variants of concern (Alpha, Beta, Gamma and Delta), variants under monitoring (Iota and Kappa) and wild-type strains circulating in Liguria (Italy) from January to August 2021. First, NGS libraries of study samples were prepared and mapped to the reference genome. Then, specimens were screened for the detection of L452R, W152C, K417T, K417N, E484Q, E484K and N501Y mutations using the SARS-CoV-2 Variants II Assay Allplex, UltraGene Assay SARS-CoV-2 452R & 484K & 484Q Mutations V1, COVID-19 Ultra Variant Catcher, SARS-CoV-2 Extended ELITe MGB and Simplexa SARS-CoV-2 Variants Direct. The overall accuracy of these assays ranged from 96.9% to 100%. Specificity and sensitivity were 100% and 96-100%, respectively. We highly recommend the use of these assays as second-level tests in the routine workflow of SARS-CoV-2 laboratory diagnostics, as they are accurate, user friendly, low cost, may identify specific mutations in about 2-3 h and, therefore, optimize the surveillance of SARS-CoV-2 variants.

Published
2022
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