KPC-2 allelic variants in
Klebsiella pneumoniae
isolates resistant to ceftazidime-avibactam from Argentina:
bla
KPC-80
,
bla
KPC-81
,
bla
KPC-96
and
bla
KPC-97
Authors: Sanz María Belén , Pasteran Fernando , de Mendieta Juan Manuel , Brunetti Florencia , Albornoz Ezequiel , Rapoport Melina , Lucero Celeste , Errecalde Laura , Nuñez Maria Rosa , Monge Renata , Pennini Magdalena , Power Pablo , Corso Alejandra , Gomez Sonia A. 
Published: February 2024

Ceftazidime-avibactam (CZA) therapy has significantly improved survival rates for patients infected by carbapenem-resistant bacteria, including KPC producers. However, resistance to CZA is a growing concern, attributed to multiple mechanisms. In this study, we characterized four clinical CZA-resistant
Klebsiella pneumoniae
isolates obtained between July 2019 and December 2020. These isolates expressed novel allelic variants of
bla
KPC-2
resulting from changes in hotspots of the mature protein, particularly in loops surrounding the active site of KPC. Notably, KPC-80 had an K269_D270insPNK mutation near the Lys270-loop, KPC-81 had a del_I173 mutation within the Ω-loop, KPC-96 showed a Y241N substitution within the Val240-loop and KPC-97 had an V277_I278insNSEAV mutation within the Lys270-loop. Three of the four isolates exhibited low-level resistance to imipenem (4 µg/mL), while all remained susceptible to meropenem. Avibactam and relebactam effectively restored carbapenem susceptibility in resistant isolates. Cloning mutant
bla
KPC
genes into pMBLe increased imipenem MICs in recipient
Escherichia coli
TOP10 for
bla
KPC-80
,
bla
KPC-96
, and
bla
KPC-97
by two dilutions; again, these MICs were restored by avibactam and relebactam. Frameshift mutations disrupted
omp
K35 in three isolates. Additional resistance genes, including
bla
TEM-1
,
bla
OXA-18
and
bla
OXA-1
, were also identified. Interestingly, three isolates belonged to clonal complex 11 (ST258 and ST11) and one to ST629. This study highlights the emergence of CZA resistance including unique allelic variants of
bla
KPC-2
and impermeability. Comprehensive epidemiological surveillance and in-depth molecular studies are imperative for understanding and monitoring these complex resistance mechanisms, crucial for effective antimicrobial treatment strategies.

IMPORTANCE

The emergence of ceftazidime-avibactam (CZA) resistance poses a significant threat to the efficacy of this life-saving therapy against carbapenem-resistant bacteria, particularly
Klebsiella pneumoniae
-producing KPC enzymes. This study investigates four clinical isolates exhibiting resistance to CZA, revealing novel allelic variants of the key resistance gene,
bla
KPC-2
. The mutations identified in hotspots surrounding the active site of KPC, such as K269_D270insPNK, del_I173, Y241N and V277_I278insNSEAV, prove the adaptability of these pathogens. Intriguingly, low-level resistance to imipenem and disruptions in porin genes were observed, emphasizing the complexity of the resistance mechanisms. Interestingly, three of four isolates belonged to clonal complex 11. This research not only sheds light on the clinical significance of CZA resistance but also shows the urgency for comprehensive surveillance and molecular studies to inform effective antimicrobial treatment strategies in the face of evolving bacterial resistance.