Culture conditions and sample preparation methods affect spectrum quality and reproducibility during profiling of Staphylococcus aureus with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Abstract

Significance and Impact of the Study: Two culture conditions (agar or broth) and two sample preparation methods (intact cell or protein extraction) were evaluated for their effects on profiling of Staphylococcus aureus using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results indicated that MALDI-enabled profiling of S. aureus is most effective when cultures are grown in broth and processed using a protein extraction-based approach. These findings should enhance future efforts to maximize the performance of this approach to characterize strains of S. aureus. Abstract Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has emerged as a promising tool to rapidly characterize Staphylococcus aureus. Different protocols have been employed, but effects of experimental factors, such as culture condition and sample preparation, on spectrum quality and reproducibility have not been rigorously examined. We applied MALDI-TOF MS to characterize a model system consisting of five methicillin-sensitive (MSSA) and five methicillin-resistant S. aureus isolates (MRSA) under two culture conditions (agar and broth) and using two sample preparation methods [intact cell method and protein extraction method (PEM)]. The effects of these treatments on spectrum quality and reproducibility were quantified. PEM facilitated increases in the number of peaks and mass range width. Broth cultures further improved spectrum quality in terms of increasing the number of peaks. In addition, PEM increased reproducibility in samples prepared using identical culture conditions. MALDI imaging data suggested that the improvement in reproducibility may result from a more homogeneous distribution of sample associated with the broth/ PEM treatment. Broth/PEM treatment also yielded the highest rate (96%) of correct classification for MRSA. Taken together, these results suggest that broth/PEM maximizes the performance of MALDI-TOF MS to characterize S. aureus.

Identification of Highly Pathogenic Microorganisms by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry: Results of an Interlaboratory Ring Trial

“…Although the TFA sample preparation protocol has been specifically developed as a MALDI-TOF MScompatible method for microbial inactivation of HPB, it was decided not to employ this protocol. It is well-known that spectra produced by acid-based methods exhibit systematic changes compared to spectra created by the direct transfer method (41). Differences between spectra obtained by the ethanol-FA and the TFA method, however, are much smaller, since both techniques are ultimately based on acid extraction.…”

Rapid Detection of Vancomycin-Intermediate Staphylococcus aureus by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

“…Instead, we extracted all of our isolates, which modestly increases the time and expense of the assay. However, extraction is known to improve spectrum reproducibility and increase the number of peaks and mass range width for S. aureus isolates (48). Finally, we did not fully evaluate whether the use of a single spectrum per isolate tested could achieve similar results.…”

Rapid Identification of Methicillin-Resistant Staphylococcus aureus (MRSA) by the Vitek MS Saramis system

“…J.E. Goldstein et al [4] reported that culture conditions and sample preparation prior to spectral acquisition would affect the spectrum quality of S. aureus spectra for MALDI-TOF MS identification. Therefore, in this study, the conditions including the extraction method and experimental environment were strictly controlled.…”