Bactec 9050

CISM has been carrying out high-quality surveillance of pediatric invasive bacterial infections since 2001 in jointly operation with the Manhiça District Hospital. For that, blood culture is routinely performed upon hospital admission in all children aged less than 2 years and for older children (up to 15 years) with axillary temperature ≥39°C, while culture of cerebrospinal fluid (CSF) is routinely performed for all children with suspected meningitis (Sigaúque et al., 2009 (link)).
For blood culture, 1–3 ml of whole blood from venous puncture were inoculated into a pediatric bottle (Pedibact®, Becton-Dickinson, Franklin Lakes, NJ, USA) and incubated into automatic system Bactec9050 (Becton-Dickinson, Franklin Lakes, NJ, USA), for 5 days. All positive cultures with a Gram stain compatible to S. aureus were sub-cultured into blood agar plates and incubated overnight at 37°C in a 5% CO₂ atmosphere. Presumptive identification of Staphylococci was performed on the basis of colony morphology and β-hemolysis test. Colonies compatible with S. aureus were confirmed by catalase and third generation Pastorex coagulase (Hercules California, USA) test. Due to financial limitations associated to the cost of DNA microarray assay, we randomly selected 84 isolates (±20% of the total of positives) from 2001 to 2009 for molecular characterization and assessment of antimicrobial susceptibility.

A blood sample was taken from every child by venepuncture for malaria diagnosis and bacterial blood culture. One to three millilitres of blood were injected into vials for paediatric blood cultures (Becton Dickinson, NJ 07417, USA) and incubated in an automated BACTEC 9050 culturing instrument (Becton Dickinson). Broth from positive bottles was examined microscopically (Gram stain) and was further cultured on standard media (chocolate agar, MacConkey agar, and Columbia agar with 5% sheep blood). For malaria diagnosis Giemsa-stained thin and thick blood slides were prepared. Two independent readers examined the slides and a third reading was performed in case of discrepancies. In 39 (1.7%) patients malaria results were not available, and these patients were excluded from the respective analyses.