Nosocomial bloodstream infections (BSIs) are an important cause of morbidity and mortality. The current study analysed data from a concurrent surveillance programme to examine the current epidemiological trends for nosocomial BSIs at 22 Japanese university hospitals from 1 April 2008 to 31 March 2012. The number of blood culture sets taken, the rate of multiple blood culture sets and the rates of antibiotic-resistant isolates among six major nosocomial BSI pathogens (Staphylococcus aureus, Enterococcus spp., Escherichia coli, Klebsiella spp., Pseudomonas aeruginosa, and Candida spp.) not including coagulase-negative staphylococci, were evaluated. The clinical characteristics of nosocomial BSIs caused by these pathogens were also collected for 2941 patients. The number of blood culture sets taken per bed increased during the 4-year study period (from 4.07 in 2008 to 5.37 in 2011), and the rates of multiple blood culture sets also increased (from 29.9% in 2008 to 50.0% in 2011). Methicillin resistance was detected in 50.2% of S. aureus isolates. The prevalence rates of extended-spectrum beta-lactamase-producing E. coli and Klebsiella spp. isolates increased annually during the study period, and the average prevalence rates were 12.3% and 5.8%, respectively. The overall crude mortality of nosocomial BSIs due to the six pathogens evaluated was 24.5% (43.2% in ICU settings and 20.5% in non-ICU settings). Thus, our multicentre study evaluated the current epidemiological trends for nosocomial BSIs, and we found that further efforts are needed to increase the use of multiple blood culture sets and improve the prognosis of nosocomial BSIs in Japanese university hospitals.