Microscan walkaway 96 plus system

Blood cultures were incubated on the BacT/Alert system (bio Mérieux, Marcy l’Etoile, France) for five days. When growth was detected, the sample was subcultured, and an isolated colony was used in the subsequent processes. Identification of clinical isolates was performed as follows: from January 2005 to March 2010, manual techniques; a test for clumping factor (PS LATEX kit, Eiken, Tokyo, Japan) and biochemical properties (ID test SP-18, Nissui Pharmaceutical Co. Ltd, Japan); from March 2010 to December 2016, pos combo 3.1 J panels in the automated MicroScan WalkAway 96 plus system (Siemens, Berlin, Germany); and from January 2017, matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). Antibiotic susceptibilities were determined using the MicroScan WalkAway 96 plus system. Susceptibility of isolates was interpreted according to the Clinical and Laboratory Standards Institute (CLSI) M100-S31 [17 ]. Oxacillin or cefoxitin susceptibilities were used to detect methicillin resistance according to the CLSI documents. Identification of S. lugdunensis isolates prior to January 2017 was reconfirmed by MALDI-TOF MS.

Peptides were purchased from GL Biochemistry Inc. (Shanghai, China). The purity of peptides was ≥5%. Amino acid sequences and physical characteristics for peptides are shown in Supplementary Table S1. TRIzol reagent and Super Script^TM III kits were purchased from TaKaRa Inc. (Dalian, China). Ceftazidime, piperacillin and levofloxacin were purchased from Solarbio Inc. (Beijing, China). C4-HSL standards is purchased from Sigma (Sigma, China). All other reagents were of analytical grade. The MDR P. aeruginosa (MRPA) strain 0108 (MRPA0108) was obtained from the Clinical Laboratory Department, the First Affiliated Hospital of Dalian Medical University. MRPA0108 exhibits resistance to gentamicin, kanamycin, ampicillin, cefepime, streptomycin, tetracycline, amoxicillin, and ciprofloxacin. Identification and antimicrobial susceptibility testing (AST) of MRPA0108 were performed by an automated MicroScan WalkAway 96 Plus system (Siemens Ltd., Germany) at the First Affiliated Hospital of Dalian Medical University.

Patient bloodstream samples at three time points (Table 1) were collected and cultured in the Emory Healthcare microbiology laboratory (under Institutional Review Board approval 50685). For clinical care of the patient, the S. aureus isolates were subjected to automated susceptibility testing using the MicroScan WalkAway 96 Plus system (Siemens Healthcare Diagnostics Inc., Tarrytown, NY), and vancomycin susceptibility was confirmed by Etest (bioMérieux, Inc., Durham, NC). The initial subculture plate was entirely swept using a plastic loop (after at least 48 h of incubation) and frozen at −80°C for DNA isolation in order to ensure the representation of isolate subpopulations for subsequent deep sequencing and analysis.

Bacterial identification and antimicrobial susceptibility testing were performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (Bruker Daltonics, Bremen, Germany) and a MicroScan WalkAway 96 plus System (Siemens Healthcare Diagnostics, Berkeley, CA, USA), respectively. The susceptibility results were interpreted based on the standard criteria defined by the Clinical and Laboratory Standards Institute [32 ].