Development and validation of main spectral profile for rapid identification of Yersinia ruckeri isolated from Atlantic salmon using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Abstract

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows rapid and reliable identification of microorganisms. The accuracy of bacterial identification using MALDI-TOF MS depends on main spectral profiles (MSPs) provided in a quality-assured commercial reference library, which requires ongoing improvement. This study aimed to develop and validate an in-house MALDI-TOF MS MSP to rapidly identify Yersinia ruckeri isolated from Atlantic salmon (Salmo salar). The novel MSP was prepared using an isolate of Y. ruckeri recovered from Atlantic salmon and confirmed by 16S rRNA gene sequencing. Subsequently, a validation set which comprises 29 isolates of Y. ruckeri were examined from three fishes: Atlantic salmon (Salmo salar) (n = 26), American eel (Anguilla rostrata) (n = 1), and Atlantic cod (Gadus morhua) (n = 2). These isolates were randomly selected from the Atlantic Veterinary College, Aquatic Diagnostic Services Bacteriology Laboratory's culture collection to validate the novel MSP. Analytical sensitivity of MALDI-TOF MS using the novel MSP to identify the validation set was 86.2%. Repeatability was assessed by acquiring spectra from 30 different spots of a randomly-selected isolate of Y. ruckeri, and analyzed spectra from each spot were compared against the novel MSP. The coefficient of variation was 3.3%. The novel MSP clustered with Bruker MSPs (n = 3) of Y. ruckeri in the reference library and did not falsely identify any closely related bacteria to Y. ruckeri. This study reports the development of a novel MSP of high analytical sensitivity and specificity for rapid identification of Y. ruckeri using MALDI-TOF MS.

Diagnosis of Renibacterium salmoninarum infection in harvested Atlantic salmon (Salmo salar L.) on the east coast of Canada: Clinical findings, sample collection methods and laboratory diagnostic tests

“…The time‐to‐identification can be decreased when compared to other bacteriologic techniques such as biochemical profiling or immunofluorescent assays, and this is particularly true for slow‐growing organisms like R. salmoninarum . Its application and feasibility in the identification of bacterial species of freshwater and seawater finfish and crustacean species have been extensively reported (Bronzato et al, 2018; Jung‐Schroers et al, 2018; 2019; Moussa et al, 2021; Nhu et al, 2016; Ojasanya et al, 2022; Pečur Kazazić et al, 2019; Rahmani et al, 2021; Yolanda et al, 2019), including for the fastidious salmonid pathogens R. salmoninarum (Jansson et al, 2020), Flavobacterium species (Fernández‐Álvarez et al, 2018; Jansson et al, 2020), Piscirickettsia salmonis (Olate et al, 2016) and Tenacibaculum maritimum (Fernández‐Álvarez et al, 2017).…”

“…One primary limitation that affects the success of MALDI‐TOF MS as a tool for identification depends on the completeness, relatedness and spectral quality of reference library databases (Ausloos et al, 1999; Cuénod et al, 2021; Sánchez‐Romero et al, 2019). It can be challenging to improve the robustness of the library database to differentiate clinical and environmental strains given the high genetic diversity of bacterial species in the aquatic environment (Piamsomboon et al, 2020; Popović et al, 2017); however, commercial reference libraries can be successfully supplemented with new entries (Jansson et al, 2020; Ojasanya et al, 2022). In the current study, the MALDI‐TOF MS reference library only contained an entry for R. salmoninarum that was created and added by the diagnostic laboratory.…”