Phenotypic and molecular characterisation of multidrug-resistant Enterobacter cloacae isolated from two cyprinid carp species and evaluation of the antibacterial activity of Curcuma longa aqueous extract

The emergence of multidrug-resistant (MDR) bacterial pathogens poses a major challenge to freshwater aquaculture by compromising fish health, production sustainability and antimicrobial stewardship. This study investigated the phenotypic and molecular characteristics of multidrug-resistant members of the Enterobacter cloacae complex isolated from naturally diseased fish of two cyprinid carps (Labeo rohita and Catla catla) and evaluated the in vitro antibacterial activity of aqueous Curcuma longa extract. Eight naturally diseased fish belonging to two cyprinid carp species were collected from three independent disease outbreaks and subjected to bacteriological examination. Morphological and biochemical analyses, together with 16S rRNA gene sequencing, identified all isolates as members of the E. cloacae complex with 99.86–100% 16S rRNA gene sequence similarity to reference sequences in the NCBI GenBank database. Antimicrobial susceptibility testing against 12 antibiotics demonstrated that all isolates were multidrug resistant, with multiple antibiotic resistance (MAR) indices ranging from 0.25 to 0.33. Universal resistance to tetracycline was observed, whereas all isolates remained susceptible to chloramphenicol, levofloxacin and ofloxacin. PCR analysis detected multiple antimicrobial resistance genes, while phenotypic characterisation revealed isolate-dependent variation in biofilm formation, protease and lipase activities. The aqueous C. longa extract exhibited significant concentration-dependent antibacterial activity, producing inhibition zones of 18.0–20.0 mm at 100 mg mL⁻¹ and minimum inhibitory concentration values of 12.5–25 mg mL⁻¹. These findings identify MDR members of the E. cloacae complex as emerging opportunistic pathogens of two cyprinid carps and highlight the potential of aqueous C. longa extract as a sustainable plant-derived antibacterial agent for freshwater aquaculture.