Microflex lt spectrometer

The identification of isolated colonies was performed using a MICROFLEX LT spectrometer (Bruker Daltonics) and an MSP 96 MALDI-TOF target plate (Bruker Daltonics, Brenen, Germany), as previously described^{44 (link)}. The obtained spectra were imported into MALDI Biotyper 3.0 software (Bruker) and compared with databases from Bruker and the base-specific laboratory at the La Timone hospital in Marseille, France. The resulting score permitted the identification (or not) of tested species as follows: species were labelled as correctly identified at the species level with a score ≥2.0 and at the genus level with a score ≥1.7 and <2.0, while no identification was made with a score <1.7.

Bacterial species identification was performed using the automated system Vitek2 (bioMérieux, Hazelwood, MO) and confirmed with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), using a Microflex LT spectrometer (BrukerDaltonics, MA, USA), as previously described[12 (link)]. The minimal inhibitory concentrations (MICs) of antimicrobials were determined using Vitek2 for the following drugs:ampicillin/sulbactam,piperacillin/tazobactam,ceftazidime, ceftriaxone, cefepime, imipenem, meropenem, gentamicin, ciprofloxacin, colistin, and tigecycline. The tigecycline MICs were confirmed using Etest strips (bioMerieux Marcy l’Étoile, France) according to the manufacturer’s recommendations. Susceptibility results were interpreted according to the breakpoints recommended by the Clinical and Laboratory Standards Institute (CLSI)[13 ], except for tigecycline for which there is no breakpoint available for A. baumannii[14 ]. Preliminary screening for the production of carbapenemases was performed usingan ertapenem hydrolysis assay (2 and 4 hours) using MALDI-TOF MS, as previously described[15 (link)]. Carbapenem hydrolysis was considered positive if the ertapenem intact-molecule mass peak (475 m/z), and that of its monosodium salt (497 m/z), disappeared completely[15 (link),16 (link)].

Culture was carried out by enriching 10 μl of breast milk samples in Fraser broth (Fisher Scientific, Illkirch, France) which were incubated for 48 hours at 37°C. 50 μL of the culture was then seeded in PALCALM agar (Fisher Scientific, Illkirch, France), incubated for between 24 and 48 hours at 37°C before colony growth. The identification of these colonies was achieved using Matrix Assisted Laser Desorption Ionisation—Time of Flight mass spectrometry (MALDI-TOF MS) on a Microflex LT spectrometer (Bruker Daltonics, Heidelberg, Germany).