Introduction Bloodstream infections (BSI) are a leading source of fatalities and morbidity in hospitals. However, the clinical spectrum and antimicrobial resistance differ globally. Identifying the pathogenic spectrum and variations in antibiotic resistance is crucial for controlling BSI and preventing inappropriate antibiotic use. Material and methods This retrospective observational study was conducted at the Dr. Ram Manohar Lohia Institute of Medical Sciences, Lucknow, UP, India, for one year between June 2022 and June 2023. A total of 669 adult patients' blood cultures were obtained from ICUs. Blood culture was done using a BacT/Alert 3D (BioMérieux SA,Marcy-l'Étoile,France) automated system. Identification of the bacterial as well as fungal isolates was done using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS), and the antimicrobial susceptibility profile was analyzed using the VITEK 2 Compact system (BioMérieux SA). Results Of the 669 blood culture samples, 213 (31.8%) showed bacterial or fungal growth. Of these 213 isolates, the most common isolate was coagulase-negativeStaphylococci(21.6%), followed byKlebsiella pneumoniae(19.3%)and Acinetobacterspp.(17.8%). The majority of gram-negative bacteria were resistant to most drugs, and vancomycin and linezolid were both effective against the majority of gram-positive bacteria. Conclusion The current study found that septicemia was more frequently caused by gram-negative bacteria than by gram-positive bacteria. Blood cultures are always necessary in cases of suspected septicemia, and once the antimicrobial susceptibility profile of the pathogen causing septicemia has been determined, suitable antimicrobials should be prescribed and used to lower the antimicrobial resistance burden.