In vivo
evolution to hypermucoviscosity and ceftazidime/avibactam resistance in a liver abscess caused by
Klebsiella pneumoniae
sequence type 512
Authors: Capitani Valerio , Arcari Gabriele , Ambrosi Cecilia , Scribano Daniela , Ceparano Mariateresa , Polani Riccardo , De Francesco Alice , Raponi Giammarco , Ceccarelli Giancarlo , Villari Paolo , Palamara Anna Teresa , Marzuillo Carolina , Carattoli Alessandra 
Published: August 2024

ABSTRACT

Carbapenemase-producing
Klebsiella pneumoniae
represents a major public health issue globally. Isolates with resistance to the newest drugs, like ceftazidime/avibactam (CZA), are increasingly reported. In this study, we analyzed the evolution of KPC-3-producing sequence type (ST) 512
K
.
pneumoniae
strains isolated at three different times (hospitalization days 45, 56, and 78) from the same patient, two of which were observed in a pericholecystic liver abscess. The three
K. pneumoniae
isolates (295Kp, 304Kp, and hmv-318Kp) from the same patient were subjected to antimicrobial susceptibility testing, whole-genome sequencing, sedimentation assay, biofilm measurement, serum resistance assay, macrophage phagocytosis, and adhesion assays. KPC-producing isolate hmv-318Kp exhibited carbapenem susceptibility, hypermucoviscous (hmv) colony phenotype and CZA resistance. Virulence markers of hypervirulent
Klebsiella
were absent. Two non-synonymous mutations were identified in the hmv-318Kp genome comparing with isogenic strains: a single-nucleotide polymorphism (SNP) occurred in the pKpQIL plasmid, changing
bla
KPC-3
in the
bla
KPC-31
gene variant, conferring CZA resistance; and a second SNP occurred in the
wzc
gene of the capsular biosynthesis cluster, encoding a tyrosine kinase, resulting in the F557S Wzc protein mutation. The
Klebsiella pneumoniae
strain exhibiting an hmv phenotype (hmv-Kp) phenotype has been previously associated with amino acid substitutions occurring in the Wzc tyrosin kinase protein. We observed
in vivo
evolution of the ST512 strain to CZA resistance and acquisition of hypermucoviscosity. The pathogenetic role of the detected Wzc substitution is not fully elucidated, but other Wzc mutations were previously reported in hmv
K. pneumoniae
. Wzc mutants may be more frequent than expected and an underreported cause of hypermucoviscosity in
K. pneumoniae
clinical isolates.

IMPORTANCE

Here we describe the evolution of KPC-3-producing ST512
K. pneumoniae
isolated at three different times from the same patient of which the last one, from a biliary abscess, showed CZA resistance by KPC-31 production and manifested hmv colony phenotype. Hypervirulent
Klebsiella pneumoniae
(hv-Kp) isolates are increasingly reported worldwide. Their hypervirulent traits are associated with the presence of rmpA/A2 genes and an hmv. In this study, we identified an hmv-Kp that lacked the rmpA-D cluster but showed an amino acid substitution in the Wzc tyrosin kinase protein, involved in the capsular biosynthesis. This hmv-Kp strain emerged
in vivo
and evolved resistance to ceftazidime/avibactam resistance in a liver abscess of a patient. Our findings suggest that wzc mutations may be underreported, making it challenging to distinguish hv-Kp from “classic”
K. pneumoniae
with an hmv phenotype.