Escherichia coli are one of the commonest bacteria causing bloodstream infection (BSI). The aim of the research was to identify the serotypes, MLST (Multi Locus Sequence Type), virulence genes, and antimicrobial resistance of E. coli isolated from bloodstream infection hospitalized patients in Cipto Mangunkusumo National Hospital Jakarta. We used whole genome sequencing methods rather than the conventional one, to characterized the serotypes, MLST (Multi Locus Sequence Type), virulence genes, and antimicrobial resistance (AMR) of E. coli. The composition of E. coli sequence types (ST) was as follows: ST131 (n=5), ST38 (n=3), ST405 (n=3), ST69 (n=3), and other STs (ST1057, ST127, ST167, ST3033, ST349, ST40, ST58, ST6630). Enteroaggregative E. coli (EAEC) and Extra-intestinal pathogenic E. coli (ExPEC) groups were found dominant in our samples. Twenty isolates carried virulence genes for host cells adherence and 15 for genes that encourage E. coli immune evasion by enhancing survival in serum. ESBL-genes were present in 17 E. coli isolates. Other AMR genes also encoded resistance against aminoglycosides, quinolones, chloramphenicol, macrolides and trimethoprim. The phylogeny analysis showed that phylogroup D is dominated and followed by phylogroup B2. The E. coli isolated from 22 patients in Cipto Mangunkusumo National Hospital Jakarta showed high diversity in serotypes, sequence types, virulence genes, and AMR genes. Based on this finding, routinely screening all bacterial isolates in health care facilities can improve clinical significance. By using Whole Genome Sequencing for laboratory-based surveillance can be a valuable early warning system for emerging pathogens and resistance mechanisms. © 2020 Paramita et al.