Development of an improved rapid BACpro® protocol and a method for direct identification from blood-culture-positive bottles using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been incorporated into pathogenic bacterial identification methods and has improved their rapidity. Various methods have been reported to directly identify bacteria with MALDI-TOF MS by pretreating culture medium in blood culture bottles. Rapid BACpro® (Nittobo Medical Co., Ltd.) is a pretreatment kit for effective collection of bacteria with cationic copolymers. However, the Rapid BACpro® pretreatment kit is adapted only for MALDI Biotyper (Bruker Daltonics K.K.), and there has been a desire to expand its use to VITEK MS (VMS; bioMerieux SA). We improved the protocol and made it possible to analyze with VMS. The culture medium bacteria collection method was changed to a method with centrifugation after hemolysis using saponin; the cationic copolymer concentration was changed to 30% of the original concentration; the sequence with which reagents were added was changed; and a change was made to an ethanol/formic acid extraction method. The improved protocol enhanced the identification performance. When VMS was used, the identification rate was 100% with control samples. With clinical samples, the identification agreement rate with the cell smear method was 96.3%. The improved protocol is effective in blood culture rapid identification, being both simpler and having an improved identification performance compared with the original.