Using MALDI-TOF-MS to test Staphylococcus aureus–infected vitreous

PURPOSE This study aimed to establish a method for testing Staphylococcus aureus in the vitreous of endophthalmitis with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS), which is simple, fast, and sensitive. METHODS S. aureus at different numbers was either mixed with homogenized vitreous or inoculated in porcine eyes for culturing, followed by homogenization. The homogenized vitreous samples, with or without centrifugation, were stained with Gram and Coomassie Blue (CBB) dyes and cultured with blood agar. The pellet of the vitreous mixture was analyzed with MALDI-TOF-MS. RESULTS The minimum detectable levels of S. aureus in H2O and in the pellet of homogenized vitreous were 9.0 × 103 (positive rate, 22.2%) and 1.0 × 104 CFU/μl (positive rate, 11.1%), respectively. In the vitreous samples inoculated with S. aureus and cultured for 12 h, the number of S. aureus increased in a dose-dependent manner to the number of bacteria in the inoculate. In the supernatant of the homogenized vitreous, there were traces of bacteria identified with Gram staining. On the blood agar plates, the supernatant grew a few colonies, while the pellet grew intensive colonies. The vitreous fragments that were stained with CBB were displayed in the supernatants, in small numbers, and in the pellets. When the inoculated number was 1.0 × 104 CFU/μl or higher, the bacteria in the vitreous pellets could be identified in all samples (100%, n = 9). However, bacteria could be detected in only two out of nine spots of pellets (22.2%) if the number of inoculated S. aureus was 1.0 × 103 CFU/μl. CONCLUSIONS A method for testing S. aureus directly from vitreous samples of endophthalmitis by the combination of easy extraction methods and a MALDI-TOF- MS assay was provided. This rapid identification method is easily adaptable for use in clinical routine and can help reduce the delay in diagnosis, allowing for earlier therapeutic intervention in patients.