| Summary: | In this paper, we aimed to characterize the fecal microbiome and its resistomes of healthy and diseased subjects infected with multidrug-resistant <i>Escherichia coli</i> using next-generation sequencing (NGS). After initial screening, 26 stools samples belonging to healthy (<i>n</i> = 13) and diseased subjects (<i>n</i> = 13) were selected and subjected to NGS. A total of 23 and 42 antibiotic-resistant genes (ARGs) conferring resistance to 6 and 9 classes of antibiotics were identified in the resistomes of healthy and diseased subjects, respectively. <i>Bacteroidetes</i> were found to be the major phylum in both healthy and diseased subjects; however, <i>Proteobacteria</i> was predominantly present in the diseased subjects only. Microbial dysbiosis and predominance of various ARGs in the resistome of diseased subjects reflect the excessive usage of antibiotics in Pakistan and warrants immediate attention to regulate the use of various antimicrobials.
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