Use of MALDI-TOF mass spectrometry for identification of Weissella tructae.

Weissella ceti, the etiological agent of hemorrhagic septicemia in rainbow trout (Oncorhynchus mykiss), was recently reclassified as a new species, Weissella tructae. This taxonomic update underscores the need for diagnostic methods capable of accurately identifying W. tructae and distinguishing it from W. ceti in a rapid, reliable, and cost-effective manner. In this study, we evaluated matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the identification of W. tructae. A total of 60 isolates, including 58 Weissella spp. from rainbow trout, the W. tructae type strain WS08ᵀ, and the W. ceti type strain CECT 7719ᵀ, were identified using MALDI-TOF MS, duplex PCR, and 16S rRNA gene sequencing, and the results were compared. A MALDI-TOF dendrogram and a 16S rRNA phylogenetic tree were constructed, and representative spectra of W. tructae and W. ceti were analyzed using principal component analysis (PCA). MALDI-TOF MS and duplex PCR showed perfect agreement, whereas 16S rRNA gene sequencing failed to discriminate between the two species due to their high sequence similarity. PCA further supported the discriminatory capability of MALDI-TOF MS, revealing species-specific peaks for W. tructae (3700.11, 3720.75, and 7406.47 m/z). Collectively, our findings demonstrate that MALDI-TOF MS is a reliable and rapid method for identifying W. tructae and distinguishing it from W. ceti, representing a valuable diagnostic tool for aquaculture applications.