Background and Aim:Coagulase-negative staphylococci (CoNS) are considered to be one of the emerging pathogens in human and animals in recent times. Staphylococcus pettenkoferi, a novel pathogen under CoNS, is discovered in 2002 in humans with multiple clinical manifestations in various patients. To date, the pathogens have not yet been reported from any animals. The present study reported the first ever isolation, identification, and characterization of multidrug-resistant S. pettenkoferi from a cat with peritonitis in India.Materials and Methods:Peritoneal fluid was collected aseptically from 3 years old cat processed for bacteriological culture by standard techniques. Isolates were confirmed by BD Phoenix™ automated bacterial identification system and were subjected to plate and tube coagulase tests. All the isolates were tested for antimicrobial sensitivity profile by disc diffusion assay, extended-spectrum β-lactamase production by double disc diffusion assay, in vitro biofilm production ability by microtiter plate assay, and detection of virulence genes and mecA gene by polymerase chain reaction assay.Results:A total of five clonally expanded isolates of S. pettenkoferi were isolated from peritoneal fluid of the affected cat. All the isolates were resistant against 36 antimicrobial agents and were also methicillin-resistant staphylococci. Phenotypically, all the isolates were negative for biofilm production but were carrying multiple biofilm-producing genes (icaA, IS257, nuc, and mecA).Conclusion:Although S. pettenkoferi was previously reported once from animal (cat) environment, this is probably the first ever report of isolation of the organism directly from any animals. This is also probably the first report from any species in India.