Bact alert fa plus

Nurses and other medical staff at our institution are not permitted to collect blood culture samples. Only physicians, especially first- or second-year interns, are permitted to collect blood samples in the ED.
Blood (14–20 mL) from peripheral veins or arteries was sampled for aerobic and anaerobic culture (7–10 mL each) in BacT/Alert FA Plus and FN Plus resin bottles (bioMérieux Inc., Durham, NC, USA). Physicians selected the topical disinfectant such as ACHX, PVI, alcohol and others available in the ED, according to their personal preferences. A blood culture was considered contaminated if one or more of the following organisms were identified in one of two blood cultures: coagulase-negative Staphylococci (CoNS), Propionibacterium acnes, Micrococci, Corynebacteria, Bacillus species other than Bacillus anthracis, or Clostridium perfringens^{10 (link),15 (link),16 (link)}. Viridans streptococci are regarded as contaminants based on the described criteria^{10 (link),15 (link)}, but they are not considered as contaminants at our institute because they were common causative agents of infective endocarditis. Polymicrobial cultures with a mixture of contaminant and true pathogens were regarded as contaminated^{14 (link)}. A culture was defined as “negative” when bacterial growth was absent or when a bacterium was regarded by the attending microbiologist as having low pathogenicity.

Colonies of S. aureus (VSSA and hVISA) were suspended in PBS to 1.0 McFarland standard. 400 μl of the suspension were mixed with 10 mL defibrinated horse blood (sourced locally) and then added to a BacT/Alert® FA Plus aerobic blood culture bottle (Biomérieux, Craponne, France) containing 30 mL growth medium. After 30 min of incubation at 37°C, 2 mL of the culture was centrifuged for 5 min at room temperature at 170 RCF (relative centrifugal force, Universal 320 Benchtop Centrifuge, rotor 1324, Hettich Lab Technology, Tuttlingen, Germany) to separate bacteria from blood cells. A 100 μL sample was plated on MHII agar before and after centrifugation for colony counting to estimate the loss of bacteria. The supernatant was mixed 1:1 with 0.5% w/v Top Vision low melting point agarose and used in CellDirector 3D. All experiments were done in triplicate.

Clinical blood culture samples collected for 13 weeks between November 2019 and March 2020 by standard protocols at Karolinska University Hospital (Huddinge, Sweden) in BacT/Alert-FA Plus and BacT/Alert-FN Plus blood culture bottles (bioMérieux, Marcy-l'Étoile, France) were used for the study. All blood culture bottles that were received at the Clinical Microbiology Department, Karolinska University Hospital (Huddinge, Sweden), were incubated in the automated BacT/Alert 3D blood culture system (bioMérieux, Marcy-l'Étoile, France) until positivity, or for a maximum of 5 days.

12 ml of the venous blood sample was obtained aseptically from each patient via venepuncture; according to a standard technique after skin disinfection.10 ml of collected blood was used for blood bottle inoculation. Inoculated blood bottles were incubated at 35 ± 2°C in the BacT/ALERT blood culture System (BacT/Alert FA Plus, bioMerieux SA, France). The BACT/Alert bottles that showed a sign of growth were alarmed. All samples were sent to Payvand's clinical and special laboratory, immediately and were incubated at 35 ± 2°C in Bact/Alert blood culture system. 2 ml of remained collected blood was added to CBC container including EDTA and was sent to the hospital lab.

Blood cultures were sampled, according to predefined indications, in BacT/ALERT FA Plus and PF Plus (bioMérieux, Marcy-l’Etoile, France) blood culture bottles. Isolates retrieved from the blood culture bottles were processed locally and later shipped to Huashan Hospital for reference identification [matrix-assisted laser desorption/ionization time-of-flight spectrometry (MALDI-TOF), bioMérieux)] and antimicrobial susceptibility testing (broth microdilution method). All microbiological methods were consistent with current Clinical and Laboratory Standards Institute (CLSI) recommendations. The isolates were stored at −70°C for further tests. The results were interpreted according to the breakpoints recommended by CLSI in 2018 (CLSI, 2018b ; CLSI, 2018b ).

The Public Health Laboratory, Birmingham, UK provides a diagnostic microbiology service to the Heart of England NHS Foundation Trust in Birmingham from whom it receives about 27,000 blood culture samples per year. All blood culture bottles are incubated and microbial growth is detected using the BacT/ALERT^® system (bioMérieux). Blood culture media used are BacT/ALERT^® SA Standard Aerobic, BacT/ALERT^® SN Standard Anaerobic and BacT/ALERT^® FA Plus (antimicrobial neutralization) media (bioMérieux). No charcoal containing bottles are used. When blood culture bottles flag positive on the system (indicating microbial growth), aliquots of the blood culture medium are used to prepare Gram stains and agar plate subcultures for identification of cultured organisms. The laboratory routinely uses MALDI-TOF (Bruker MALDI Biotyper system) for identification of bacterial isolates from agar plates. Some bacterial species are confirmed with biochemical tests in addition to MALDI-TOF according to ISO 15189:2012 United Kingdom Accreditation Service accredited methods used for routine clinical diagnostics. Routine clinical diagnostic results reported by the laboratory were considered the definitive result with which direct MALDI-TOF results (see below) were compared.