Escherichia coli (E. coli) strains placed in predominant nosocomial bacteria in intensive care units (ICUs), resulting in severe drug-resistant infections. Non-susceptibility to β-lactams and last-line drugs such as Fosfomycin and Colistin cause limited availability of infections eradication. The objective of this study included the determination of genes encoding extended-spectrum β-lactamase (ESBL), Carbapenemase, Colistin, and Fosfomycin resistance in clinical isolates of E. coli in ICUs. A total of 200 E. coli isolates were identified from ICU settings. The CTXM-1, SHV, IMP and OXA-48 genes were detected for β-lactamases using the polymerase chain reaction (PCR) technique. The fosA3 and mcr-1 and mcr-2 genes were also detected for resistance against Fosfomycin and Colistin. The CTX-M1, SHV, IMP and OXA-48 genes were detected in 60 (30%), 56 (28%), 28 (14%) and 4 (8%) of isolates. none of the E. coli isolates had the mcr-2 and fosA3 genes. Despite the existence of resistance genes to the third-generation antibiotics and Carbapenemase s, any isolates had genes for resistance to Fosfomycin and Colistin. More studies are needed to follow the resistance genes against last-resort antibiotics.