Objective To investigate the clinical characteristics and bacterial drug resistance of bloodstream infection of gram-negative bacteria, and provide guidance for clinical rational drug use and control of hospital infection. Methods A retrospective analysis was conducted in the patients diagnosed as severe pneumonia with blood culture of gram-negative bacteria from January 2015 to December 2017 in Beijing Anzhen Hospital. Results A total of 60 severe pneumonia patients suffered from bloodstream infection of gram-negative bacteria were recruited including 34 males and 26 females aging from 42 to 89 years and 73.4 years in average. In the 60 patients, 32 cases were infected with Klebsiella pneumonias, 20 cases were infected with Acinetobacter baumanni, and 8 cases were infected with Escherichia coli. The antimicrobial susceptibility testing result of Klebsiella pneumonias showed that the drug susceptibility rate was 100% to tigecycline, and 6.3% to amikacin. Escherichia coli was sensitive to Amikacin, imipenem, ceftazidime and meropenem while resistance to other drugs. The antimicrobial resistance of Acinetobacter baumanni was 28.6% for cefoperazone/sulbactam, and 14.3% for tigecycline. C-reactive protein, procalcitonin and SOFA scores were higher in the patients infected with Acinetobacter baumanni. Neutrophils and blood lactic acid were higher in the patients infected with Klebsiella pneumonias. There were no statistical differences in white blood cell, platelet or motality rate between the patients infected with Acinetobacter baumanni and the patients infected with Klebsiella pneumonias. SOFA scores and blood lactic acid had significantly statistical relevance with prognosis. Conclusion There is a high proportion of drug resistance of Klebsiella pneumoniae and Acinetobacter baumanni in the bloodstream infection of gram-negative bacteria.