Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

Abstract

Background: The majority of infectious diseases of cultured fish is caused by bacteria. Rapid identification of bacterial pathogens is necessary for immediate management. The present study developed a custom Main Spectra Profile (MSP) database and validate the method using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for rapid identification of fish bacterial pathogens. Streptococcus agalactiae, Streptococcus iniae, Aeromonas hydrophila, Aeromonas veronii, and Edwardsiella tarda obtained from diseased fish were used as representative bacterial pathogens in this study. Bacterial peptides were extracted to create a Main Spectra Profile (MSP), and the MSPs of each bacterial species was added into the MALDI Biotyper database. Fifteen additional isolates of each bacterial species were tested to validate the utilized technique. Results: The MSPs of all field isolates were clearly distinguishable, and the MSPs of the same species were clustered together. The identification methodology was validated with 75 bacterial isolates. The reliability and specificity of the method were determined with MALDI Biotyper log score values and matching results with 16 s rDNA sequencing. The species identification using the public MALDI Biotyper library (Bruker MALDI Biotyper) showed unreliable results (log score < 2.000) with 42.67% matching result with the reference method. In contrast, accurate identification was obtained when using the custom-made database, giving log score > 2.115, and a 100% matching result. Conclusion: This study demonstrates an effective identification of fish bacterial pathogens when a complete custom-made MSP database is applied. Further applications require a broad, well-established database to accommodate prudent identification of many fish bacterial pathogens by MALDI-TOF MS.

Streptococcus suis serotyping by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

“…S. plurextorum, S. porci and S.porcorum where their species-specific markers corresponded to 6164, 6133 and 4190/8381, respectively [9]. However, some other peptide masses were found to be different from those in previous studies, which can be explained by the different preparation protocols, such as the acid concentration, spotting method, and type of matrices [16][17][18]. In the present study, the principal difference in the preparation protocol was the modified peptides extraction method, that formic acid was not included, gentle dissolve pellet by vortex and vigorously mix peptides with sinapinic matrix, which likely accounted for some of the dissimilar PMFs.…”

Identification of French Guiana anopheline mosquitoes by MALDI-TOF MS profiling using protein signatures from two body parts

“…Moreover, if the cut-off threshold of LSV to consider identification as reliable was raised to 2.0, 100% of the specimens succeeded to reach this cut-off with MS spectra from at least one body part. It is interesting to note that, for bacteria identification, it was assumed that species were assigned for LSVs upper or equal than 2.0, and genera were assigned for scores of upper or equal to 1.7 but lower than 2.0 [49,50]. Then, the combination of the results from both body-parts allowed to enhance ratio of LSVs upper or equal than 2.0 for each specimen compared to the use of only one body-part.…”

Proteomic Strategies to Evaluate the Impact of Farming Conditions on Food Quality and Safety in Aquaculture Products

“…MALDI-TOF MS analyses have been performed to obtain available reference spectral libraries for diverse bacterial strains isolated from seafoods [82]. Recently, the open MALDI Biotyper library (Bruker MALDI Biotyper) allowed for the precise identification of 75 pathogenic bacterial isolates [83].…”