S. aureus analyzed in this study were obtained from positive blood cultures after incubation in Bactec Plus Aerobic/F and Bactec Lytic/10Anaerobic/F blood culture bottles (Becton Dickinson, Sparks, MD, USA). The blood samples were collected from patients treated at different public hospitals in São Luis, Northeast Brazil, during a period of seven months. The isolation from positive blood cultures was performed using blood agar plates (BioMérieux, Marcy l’Etoile, France). Subsequently, the isolates were identified by MALDI-TOF (Matrix Assisted Laser-Desorption Ionization-Time of Flight) Mass Spectrometry (MS). The mass spectra acquired for each bacterial strain were compared to the mass spectra contained in the database using Biotyper 3.0 software (Bruker, Billerica, MA, USA). All isolates were kept in Luria-Bertani broth supplemented with 15% glycerol at −80 °C.
Bactec lytic 10 anaerobic f
Manufactured by BD
Sourced in United States
The Bactec Lytic/10 Anaerobic/F is a laboratory equipment product designed for the detection of anaerobic bacteria in clinical samples. It provides a reliable and efficient method for the identification and growth of anaerobic microorganisms.
Automatically generated - may contain errors
Lab products found in correlation
Bactec plus aerobic f, by BD (6 mentions)
Blood agar plate, by bioMérieux (1 mentions)
Biotyper 3, by Bruker (1 mentions)
Bhi media, by Thermo Fisher Scientific (1 mentions)
Iif kit, by EUROIMMUN (1 mentions)
Bactec peds plus f, by BD (1 mentions)
Chocolate agar, by BD (1 mentions)
Macconkey agar, by BD (1 mentions)
Columbia agar, by BD (1 mentions)
Bactec peds plus, by BD (1 mentions)
Bactec fx, by BD (1 mentions)
Bactec peds plus f culture vial, by BD (1 mentions)
Bactec 9240 system, by BD (1 mentions)
Rapid id 32 strep, by bioMérieux (1 mentions)
Api staph, by bioMérieux (1 mentions)
Vitek 2 automated system, by bioMérieux (1 mentions)
Api20, by bioMérieux (1 mentions)
Bactec fx blood culture system, by BD (1 mentions)
Microscan walkaway plus, by Beckman Coulter (1 mentions)
Vitek 2, by bioMérieux (1 mentions)
8 protocols using bactec lytic 10 anaerobic f
1
Isolation and Identification of S. aureus from Blood Cultures
2
MALDI-TOF MS Bacterial Identification
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MALDI-TOF MS reagents were purchased from Bruker Daltonics, Coventry, UK. All HPLC-grade chemicals were purchased from Fluka (Honeywell). Agar and BHI media were purchased from Oxoid (Thermofisher, UK). Bactec Plus Aerobic/F (paediatric) and Bactec Lytic/10 Anaerobic/F blood culture bottles were purchased from Becton Dickinson, UK. FcMBL magnetic beads were produced using biotinylated FcMBL and one micron Streptavidin beads (Thermofisher) as previously described [14 ] and were provided by the Wyss Institute and BOA Biomedical.
3
Comprehensive CSF and Blood Analysis in AE
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The CSF of all patients was collected when virus infection-like symptoms (T >38°C, headache, etc) were present. DNA sequences were extracted from samples and amplified by Applied Biosystems TM 7500 RT-qPCR instrument (Thermo Fisher Scientific, USA). The CSF of all patients was collected when bacterial infection-like symptoms (T >38°C, neck stiffness, etc) were present, and transferred to different culture bottles (Bactec Plus Aerobic/F, Bactec Lytic/10 Anaerobic/F, Bactec Peds Plus/F) (Becton, Dickinson and Company Sparks, MD 21152 USA) immediately, and underwent etiology inspection by BacrecTM FX automated blood culture system (Becton, Dickinson and Company, USA) in microbiological laboratory. All serum and CSF samples from patients with AE were evaluated for NSAbs by IIF kits (Euroimmun, Lübeck, Germany) and CBA kits (Euroimmun, Lübeck, Germany) according to the manufacturer’s instructions. All patients underwent serum and CSF laboratory tests, including all of the following: CSF examinations included: Pandy test, intracranial pressure, WBC counts, protein levels, chloride, glucose, lactate dehydrogenase (LDH), lactic acid (Lac), adenosine deaminase (ADA) and C-reactive protein (CRP). Blood examinations included: leukocyte, erythrocyte and platelet counts, and the levels of sodium, potassium, calcium, and chlorine.
4
Anaerobic Cultivation of Stool Samples
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The stool specimen was incubated at 37°C into an anaerobic blood bottle Bactec Lytic/10 Anaerobic/F (Becton Dickinson, Le Pont de Claix, France) supplemented with 5% sheep's blood, after a thermal shock of 20 minutes at 80°C. Then dilution cultures were performed, and characterization of growth conditions was tested as previously described [10] (link). Finally, sporulation, different pH levels and NaCl concentrations were tested in the agar plate under the best culture conditions [11] (link).
5
Blood Culture and Bacterial Identification
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All blood specimens of at least 8 mL were injected into aerobic and anaerobic blood culture bottles (BACTEC Lytic/10/Anaerobic/F and BACTEC Peds Plus, Becton Dickinson, Poland). Samples were taken in duplicate (right and left hand) from approximately 70% of the patients. The remaining 30% of patients were sampled in one replicate because, for various reasons (including the patient’s condition), it was not possible to collect more samples. Then, material was cultured on MacConkey agar, Columbia agar (at 37 °C, each for 24 h) and chocolate agar (at 37 °C, 48 h) (all from Becton Dickinson, Warszawa, Poland). The identification of the bacterial strains was performed by the MALDI-TOF automated identification system (Biotyper Bruker Daltonics, Leiderdorp, The Netherlands).
6
Blood Culture Collection Protocol
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Blood was taken with aseptic technique, directly inoculated into aerobic (BD BACTEC Plus Aerobic/F) and anaerobic culture vials (BD BACTEC Lytic/10 Anaerobic/F), or peds culture vials (BD BACTEC Peds Plus/F culture vials) (Becton Dickinson & Co, BD, Shanghai, China). Each adult patient needed an Aerobic and an anaerobic culture vials, the Peds vials were used only for pediatric patients. All blood culture bottles were loaded onto the BD BACTEC FX instrument (Becton Dickinson, Franklin Lakes, NJ, USA).
7
Blood Culture Identification and Antibiotic Susceptibility
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Two peripheral blood samples were obtained from at least two different venipuncture sites in each patient, as recommended by the Infection Control Division of the hospital. When a long-term catheter-associated infection was suspected, an additional blood sample was collected from the suspected vascular device at the same time the peripheral blood sample was drawn. Each blood sample was placed into a separate culture bottle (BD BACTEC Lytic/10 Anaerobic/F, BACTEC plus Aerobic/F and BACTEC MYCO/F Lytic; Becton, Dickinson and Company; Maryland USA). Microorganism growth was detected by the BACTEC® 9240 system (Becton Dickinson). The identification of different species was performed by the Vitek2® automated system (BioMérieux), API 20, API Staph and rapid ID 32 Strep (BioMérieux®). Testing for antibiotic susceptibility was performed with Vitek2® and antibiotic gradient tests (BioMérieux) and interpreted in accordance with the Clinical and Laboratory Standards Institute (CLSI) recommendations, applying the versions updated for the year in which the blood culture was performed, from 2012 to 2017. The resistance to cefoxitin is considered a phenotypic marker of the presence of chromosomal AmpC.
8
Identification and Antimicrobial Susceptibility of Bloodborne Pathogens
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Blood specimens were inoculated in bottles containing liquid medium (BD Bactec Plus aerobic/F and BD Bactec Lytic/10 anaerobic/F, Becton Dickinson) and incubated for a maximum of 5 days in a Bactec™ FX blood culture system (Becton Dickinson). In case of positive signal, the blood cultures were further seeded on CHRO-MagarTM Candida (VacuTest Kima) and Sabouraud Dextrose Agar (VacuTest Kima) plates in order to isolate Candida spp. and on Blood Agar Base with Horse Blood (VacuTest Kima) plates, both in aerobic and anaerobic conditions, in order to isolate bacterial cells. The plates were incubated at 35e37 C for 16 h. Antimicrobial susceptibility testing was performed using both MicroScan Walkaway plus (Beckman Coulter) and VITEK 2 (Biomerieux) automated systems [19] .
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