Standardized numbering and alignment of the KPC family of β-lactamases

ABSTRACT The KPC family of serine β-lactamases comprises more than 260 members. Some variants are associated with ceftazidime-avibactam resistance in clinical isolates, often linked to substitutions and/or insertions/deletions (i.e., INDELs) in three distinct loops of the KPC sequence: (i) the 164–179 loop (i.e., the Ω loop); (ii) the 237–243 loop; and (iii) the 267–275 loop. Inconsistencies in residue numbering across published reports, however, complicate the accurate annotation of KPC variants. We retrieved 267 KPC variant sequences from the Beta-Lactamase Database (BLDB) in September 2025 and analyzed sequence differences between variants using combined nucleotide and structure-guided alignment algorithms, supported by AlphaFold3 modeling in ambiguous cases. Variants were classified into four groups to comprehensively review sequence changes across the KPC family: substitutions only ( n = 126), deletions only ( n = 17), insertions only ( n = 66), and variants with two or more types of amino acid changes ( n = 57). Comparisons with previous reports indicate that many annotation errors stem from overlooking the absent residues 58 and 253 in the KPC consensus sequence, and that most inconsistencies in annotation and residue assignment occur in INDEL variants. To address these issues, we propose a standardized annotation scheme for substitutions, deletions, and insertions for the KPC family, based on the Ambler numbering system, and supported by structural information. This systematic scheme will help to standardize the description of newly emerging KPC variants and prevent discrepancies in future reports in the context of antimicrobial resistance.