Aquatic enterococci from a Mediterranean lagoon (Bizerte, Tunisia): co-occurrence of antibiotic resistance, heavy metal tolerance, biofilm formation, and ST17 lineage

Coastal lagoons receiving urban, agricultural, and industrial inputs are hotspots for co-selection and dissemination of antimicrobial and heavy metal resistance, yet their role as reservoirs of resistant enterococci remains poorly characterized in North Africa. This study investigated the Lagoon of Bizerte (northern Tunisia) as a reservoir of antimicrobial and heavy metal resistance in Enterococcus spp. A total of 146 isolates from eight sites were analyzed using integrated phenotypic, molecular, and epidemiological approaches. Enterococcus faecium (44.5%) and E. faecalis (26.7%) predominated. Antimicrobial susceptibility testing showed high resistance to tetracycline (44.5%) and erythromycin (43.1%), with multidrug resistance more frequent in E. faecium (46.1%). Fourteen resistance genes were identified, including tet(M/O/L/K), erm(A/B/C), mefA, msrA, aminoglycoside-modifying enzyme genes, and optrA, frequently linked to the Tn916-Tn1545 integrative element. Heavy metal tolerance was most common for copper, lead, zinc, and mercury, with corresponding resistance genes (pcoA, pbrT, zntB, merA, cnrA, silC, nccA, arsA) detected, particularly in E. faecium. Biofilm formation occurred in 81.6% of isolates, strongest in E. faecalis and E. faecium. Multilocus sequence typing of five optrA-positive E. faecium showed predominance of ST17. These findings demonstrate co-selection of resistance traits in lagoon enterococci, underscoring their epidemiological relevance as a public health concern in coastal ecosystems.