Morphological, molecular and MALDI-TOF mass spectrometry identification of ixodid tick species collected in Oromia, Ethiopia.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) technology has recently been reported as a promising method for arthropods identification. More recently, our laboratory reported the correct identification of tick species via the MALDI-TOF MS protein spectra profiling of legs from fresh specimens. The aim of the present study was to assess the use of MALDI-TOF MS for correct identification of ixodid tick species preserved in 70 % ethanol during field collection in Ethiopia. Following morphological identification of 12 tick species, the legs from 85 tick specimens were subjected to MALDI-TOF MS. Spectral analysis revealed an intra-species reproducibility and inter-species specificity that were consistent with the morphological classification. To support the results of the MALDI-TOF MS tick species identification, 41 tick specimens comprising 3 to 5 specimens per tick species were used to create a reference spectra database, which was evaluated using the spectra of the 44 remaining tick specimens. The blind tests revealed that 100 % of the tick specimens studied by MALDI-TOF MS were correctly identified. A relevant Log score value (LSV) of >1.8 was recorded for all of the tick species studied by MALDI-TOF MS, except for Rhipicephalus praetextatus. The morphological and MALDI-TOF MS identifications were confirmed by sequencing the 12S ribosomal RNA (rRNA) gene of 40 tick specimens belonging to 11 ixodid species. Taken together, the results of the present study indicate that MALDI-TOF MS is a reliable tool for tick species identification, even after preservation in ethanol, provided that a reference spectra database is built from specimens that represent the respective species stored under the same conditions.