Abstract Successful treatment against infectious agents depends on rapid and accurate detection of the causative organisms. Misdiagnosis can hamper such success while leading to improper advising of antibiotics. In Bangladesh, the majority of the diagnostic centers detect and identify pathogens through culture and biochemical test-based methods and suggest antibiotics based solely on the results of disk-diffusion methods. This pilot study tried to validate the identity of the isolates characterized by diagnostic facilities near Dhaka. One hundred and twenty pre-characterized clinical isolates were analyzed biochemically and genotypically. Random Amplification of Polymorphic DNA-PCR, rcsA, and phoA genes-based PCR, and Loop-Mediated Isothermal Amplification (LAMP)-based identification of Klebsiella pneumoniae and Escherichia coli, respectively, followed by 16S rRNA sequencing confirmed misidentification of some clinical pathogens of other genera as Klebsiella spp. and E. coli. According to the antibiotic susceptibility testing guidelines, antibiotic choice, sensitivity pattern, and breakpoint measurement are different for each group of organisms. The lack of adherence to proper standards results in misdiagnosis and may facilitate the development of antibiotic resistance. The pilot study observers misidentification of clinical pathogens identified by the diagnostic centers. Well-characterized rapid molecular techniques like LAMP are suggested in clinical diagnosis to avoid misdiagnosis and subsequently circumvent antibiotic resistance development.