Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been reported as a promising and reliable tool for arthropod identification, including the identification of alcohol-preserved ticks based on extracted leg protein spectra. In this study, the legs of 361 ticks collected in Vietnam, including 251 Rhiphicephalus sanguineus s.l, 99 Rhipicephalus (Boophilus) microplus, two Amblyomma varanensis, seven Dermacentor auratus, one Dermacentor compactus, and one Amblyomma sp. were submitted for MALDI-TOF MS analyses. Spectral analysis showed intra-species reproducibility and inter-species specificity and the spectra of 329 (91%) specimens were of excellent quality. The blind test of 310 spectra remaining after updating the database with 19 spectra revealed that all were correctly identified with log score values (LSV) ranging from 1.7 to 2.396 with a mean of 1.982 ± 0.142 and a median of 1.971. The DNA of several microorganisms including Anaplasma platys, Anaplasma phagocytophilum, Anaplasma marginale, Ehrlichia rustica, Babesia vogeli, Theileria sinensis, and Theileria orientalis were detected in 25 ticks. Co-infection by A. phagocytophilum and T. sinensis was found in one Rh. (B) microplus.
Background
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a tool that has revolutionised clinical microbiology and has recently been described as an innovative and effective approach to arthropod identification.
Methods
In this study, mosquitoes were captured in Vietnam using four different methods (human landing catch, CDC light traps, BG-Sentinel traps, animal-baited net traps). A total of 4215 mosquitoes were captured and morphologically identified as belonging to three genera: Aedes, Anopheles and Culex. We randomly selected 1253 mosquitoes, including 662 specimens of 14 Anopheles species, 200 specimens of two Aedes species and 391 morphologically unidentified Culex specimens, for molecular and MALDI-TOF MS analysis. The DNA from 98 mosquitoes (69 Anopheles specimens, 23 Culex specimens and six Aedes sp. specimens) was subjected to molecular analysis, either to confirm our morphological identification or the MALDI-TOF MS results, as well as to identify the Culex species that were morphologically identified at the genus level and to resolve the discrepancies between the morphological identification and the MALDI-TOF MS identification.
Results
High-quality MS spectra were obtained for 1058 of the 1253 specimens (84%), including 192/200 for Aedes, 589/662 for Anopheles and 277/391 for Culex. The blind test showed that 986/997 (99%) of the specimens were correctly identified by MALDI-TOF MS, with log score values ranging from 1.708 to 2.843. Eleven specimens of Culex could not be identified based on morphological features, MALDI-TOF MS or molecular analysis.
Conclusions
This study enabled us to identify several species of mosquitoes from Vietnam using MALDI-TOF MS, and to enrich our database of MALDI-TOF MS reference spectra.
Graphical Abstract
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