TEM- and SHV-derived extended-spectrum β-lactamases (ESBLs) arose by point mutations in genes encoding broad-spectrum variants that were extensively disseminated into mobile elements, while CTX-M β-lactamases are derived from natural (chromosomal) counterparts that already have oxyimino-cephalosporinase activity. To date, CTX-M/KLU β-lactamases account for nearly 110 representatives clustered in five main groups: CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, and CTX-M-25, with a hypothetical chromosomal counterpart for the CTX-M-25 subfamily still missing. Kluyvera georgiana 14751 was isolated from a bloodstream infection in a 62-year-old male patient, and its identity was resolved by 16S rRNA gene sequencing (99.7% identity with K. georgiana ATCC 51603). Partially EcoRI-digested chromosomal DNA from K. georgiana 14751 was cloned into pK19 vector and transformed into E. coli Top10F⁻, yielding recombinant plasmid pTKE-1KA2 with an 8-kb insert harboring a 876-bp bla gene that encodes a novel CTX-M-78 (EMBL accession no. AM982522). CTX-M-78 is closely related to CTX-M-39 (96.2% amino acid identity) and clustered in the CTX-M-25 subgroup. Upstream of blaCTX-M-78, there is a 1,227-bp att-like gene encoding a putative aspartate aminotransferase (≥96% identity with other Kluyvera entries), and downstream are an orf3-like gene (similar to a putative ttrR response regulator from K. georgiana) and an orf4-like gene (encoding a putative autotransporter in K. ascorbata). CTX-M-78 possesses high similarity with members of the CTX-M-25 subgroup, making it the closest representative to be considered one of the probable progenitors of the subfamily.