The whole research work was conducted in the Microbiology Laboratory, Department of Veterinary and Animal Sciences, University of Rajshahi, Rajshahi-6205, Bangladesh. A total of 180 chicken egg samples were aseptically collected from randomly selected commercial layer farms, whole-sellers, and retail outlets. Out of 180 egg samples, 60 eggs were collected from commercial layer farms, 60 from whole sellers, and 60 from retailers. Among these, 60 were brown the opened, 60 were white the opened, and 60 were indigenous chicken eggs. For egg peels, a pool of five eggs was used and one sterile swab was moistened with sterile saline and was applied on the surface of each egg. Each swab applied on five egg peels was submerged in 6 ml of sterile saline as ‘peel wash’. The peel wash was mixed with a mini vortex and inoculated into different culture media. After that, the egg peels were sterilized by swabbed with 70% ethyl alcohol, flamed, and broken with a sterile forceps from the broad ends, and the yolk and albumen of five eggs were pooled. The contents were thoroughly mixed for nearly one minute using a blender and a mixer was used to inoculate into different culture media. After enrichment in nutrient broth, a loopful of enriched culture was taken on a sterile glass slide and Gram’s staining was performed for morphology study. Then the broth culture of bacteria was inoculated on nutrient agar by streak plate technique and inoculated at 37°C for 24 hours for the development of colonies. The colony on primary culture was repeatedly sub-cultured on different selective culture media by the streak plate technique until the pure culture with homogenous colonies was obtained. Identification of E. coli was done through a series of biochemical tests. The colony morphology of the isolated E. coli was studied as mentioned by Merchant and Packer. Colony morphology such as shape, size, surface texture, edge and elevation, color, and opacity developed after 24 hour of incubation were carefully studied and recorded. Gram’s staining was performed according to the method described by Cheesbrough. In brief, a small colony was picked up with a bacteriological loop, smeared on separate glass slides, and fixed by gently heating. Crystal violet was applied on each smear to stain for two minutes and washed with running tap water. A few drops of Gram’s iodine were added as mordent for one minute and again washed with running tap water. Acetone alcohol was added for a few seconds as a decoloring agent. After washing with water, safranin was added as a counterstain and allowed to stain for two minutes. The slides were washed with water, blotted and dried in the air, and examined under a light microscope with a high power objective (100X) using immersion oil. Pure culture of E. coli was subjected to different biochemical tests like sugar fermentation test, catalase test, indole test, MR test, VP test, and TSI agar slant reaction. Standard methods were followed to conduct these tests and interpretation. The disk diffusion method was used to test the susceptibility of the E. coli isolates. In brief, pure colonies of the E. coli isolates were separately inoculated in nutrient broth and incubated at 37°C for overnight. Then 100 µl of broth culture was taken and placed on Mueller Hinton agar plate and spread evenly with a sterile glass rod spreader. The antibiotic disks were placed on the surface of the plates keeping about 1cm apart. After 18 to 20 hours of incubation at 37°C, each plate was examined. The susceptibility test of E. coli was done against ten antibiotic disks namely, gentamycin, ciprofloxacin, erythromycin, doxycycline, chloramphenicol, tetracycline, amoxicillin, ampicillin, ceftriaxone, and meropenem. The susceptibility zones were measured and interpreted according to the Clinical and Laboratory Standards Institute.