Nosocomial bloodstream infections (BSIs) are important causes of morbidity and mortality in the United States. Data from a nationwide, concurrent surveillance study (Surveillance and Control of Pathogens of Epidemiological Importance [SCOPE]) were used to examine the secular trends in the epidemiology and microbiology of nosocomial BSIs. Our study detected 24,179 cases of nosocomial BSI in 49 US hospitals over a 7-year period from March 1995 through September 2002 (60 cases per 10,000 hospital admissions). Eighty-seven percent of BSIs were monomicrobial. Gram-positive organisms caused 65% of these BSIs, gram-negative organisms caused 25%, and fungi caused 9.5%. The crude mortality rate was 27%. The most-common organisms causing BSIs were coagulase-negative staphylococci (CoNS) (31% of isolates), Staphylococcus aureus (20%), enterococci (9%), and Candida species (9%). The mean interval between admission and infection was 13 days for infection with Escherichia coli, 16 days for S. aureus, 22 days for Candida species and Klebsiella species, 23 days for enterococci, and 26 days for Acinetobacter species. CoNS, Pseudomonas species, Enterobacter species, Serratia species, and Acinetobacter species were more likely to cause infections in patients in intensive care units (P<.001). In neutropenic patients, infections with Candida species, enterococci, and viridans group streptococci were significantly more common. The proportion of S. aureus isolates with methicillin resistance increased from 22% in 1995 to 57% in 2001 (P<.001, trend analysis). Vancomycin resistance was seen in 2% of Enterococcus faecalis isolates and in 60% of Enterococcus faecium isolates. In this study, one of the largest multicenter studies performed to date, we found that the proportion of nosocomial BSIs due to antibiotic-resistant organisms is increasing in US hospitals.