Bbl crystal identification system

For patients with suspected sepsis, local guidelines recommended the inoculation of 1–3 mL (for paediatric patients; however, for older children, larger blood inoculums of 10 mL were encouraged) and 8–10 mL (for adults) directly into Bactec^® culture vials (Becton–Dickinson, USA). Routinely at the laboratory, cultures were processed with the BACTEC 9240 blood culture system (Becton–Dickinson, NJ, USA) according to manufacturer’s instructions. Where bacterial growth was detected in vials, Gram-stains were performed; and subcultures were typically made onto appropriate media based on Gram-stain results. Bacterial isolates were identified using routine biochemical methods. Bacteria speciation was done with the BBL^® Crystal identification system (Becton–Dickinson, NJ, USA). For positive fungal blood cultures, organisms were identified on the basis of morphology. As part of regular practice at the laboratory, consultant microbiologists evaluated all positive blood cultures to categorize isolates as contaminants or true pathogens. Susceptibility testing for bacterial pathogens were conducted using the Kirby-Bauer disc diffusion method with antibiotic discs; and these tests were interpreted according to guidelines by the Clinical and Laboratory Standards Institute (CLSI) [13 ].

Gut flora was determined by the development of bacteria (Lactobacillus spp., Escherichia coli, fecal coliforms, and Clostridium spp.) and Mycétes in stools, as previously described [29 ]. Stool samples were collected from intestine after euthanasia with strikers, and inserted into hermetic vials using a specific medium. Subsequently, the microbiota was measured after 48 h of incubation under proper conditions using a selective agar. Further proof of isolation was performed by using bacterial metabolic tests on isolated organisms through the BBL Crystal Identification System (Becton Dickinson, Franklin Lakes, NJ, USA). The results are expressed in colony-forming units (c.f.u.) per milliliter of stool. Test was performed by Functional Point (Bergamo, Italy), a clinical and virology laboratory that adheres to international quality control standards and is accredited as an official laboratory within the National Health System. The test coefficient of variation was <9%.

For each patient, 1–3 mL (for paediatric patients) and 8–10 mL (for adults) of blood cultures injected directly into Bactec^® culture vials (Becton–Dickinson, USA) were submitted for laboratory diagnosis. Blood cultures were processed with Bactec^® 9240 blood culture system (Becton–Dickinson, NJ, USA) according to manufacturer’s instructions. The presence of BSI was defined by at least one set of positive blood culture for bacteria or fungi in patients. When blood cultures yielded positive results, they were evaluated to determine whether it represented true bacteraemia, fungaemia, or simply contamination. For potential skin contaminants (e.g., micrococci, Staphylococcus epidermidis), at least two separate positive blood cultures obtained from different sites had to be isolated for the results to be considered as true infection. A total of 111 blood cultures from cancer patients were positive for various organisms. Aerobic subcultures were made from positive culture vials onto blood, chocolate, MacConkey and Saboraund agar. Identification of bacteria were carried out by Gram-stain microscopy and by conventional biochemical methods. Species identity were confirmed by the BBL crystal identification system (Becton–Dickinson, NJ, USA). Positive blood cultures for yeast and non-yeast fungi were identified on the basis of morphology.