Comparative Genomics of the Enterobacter cloacae Mobilome: Prophage Diversity and Anti-Phage Defense Systems in Phage Satellites.

Enterobacter cloacae (E. cloacae) is an important healthcare-associated pathogen with increasing antimicrobial resistance. Prophages and phage-inducible chromosomal islands (PICIs) are integral components of the E. cloacae genome, yet their contribution to resistance, virulence, and anti-phage defense remains poorly understood. To address this gap, we analyzed 35 complete E. cloacae genomes, identifying 139 intact prophages (10.8-85.7 kb) with an average GC content (~52%) close to that of the host (54-55%). Most prophages (86.6%) were assigned to the families Peduoviridae and Drexlerviridae within the class Caudoviricetes, and proteome-based clustering showed strong relatedness to phages infecting Enterobacteriaceae. The E. cloacae genomes harbored multiple antimicrobial resistance genes, notably fosA (97%), oqxA/oqxB (89%) and blaCMH-3 (77%). However, on genomic-context verification none were genuinely prophage-encoded; they were instead located on the chromosome or plasmids. Virulence profiling revealed universal presence of ompA and high prevalence of csgG (94%), while the only confirmed prophage-encoded virulence determinants were the Shiga toxin genes stxA and stx1B. Additionally, 16 phage satellites were identified, containing conserved integration, replication, and packaging modules together with diverse accessory and anti-phage defense systems. Overall, prophages and phage satellites drive genome diversification in E. cloacae, with satellites notably serving as reservoirs of anti-phage defense, whereas resistance and virulence genes are located outside prophage and satellite regions.