Background: The identification of mosquito vectors is typically based on morphological characteristics using morphological keys of determination, which requires entomological expertise and training. The use of protein profiling by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), which is increasingly being used for the routine identification of bacteria, has recently emerged for arthropod identification.

Methods: To investigate the usefulness of MALDI-TOF-MS as a mosquito identification tool, we tested protein extracts made from mosquito legs to create a database of reference spectra. The database included a total of 129 laboratory-reared and field-caught mosquito specimens consisting of 20 species, including 4 Aedes spp., 9 Anopheles spp., 4 Culex spp., Lutzia tigripes, Orthopodomyia reunionensis and Mansonia uniformis. For the validation study, blind tests were performed with 76 specimens consisting of 1 to 4 individuals per species. A cluster analysis was carried out using the MALDI-Biotyper and some spectra from all mosquito species tested.

Results: Biomarker mass sets containing 22 and 43 masses have been detected from 100 specimens of the Anopheles, Aedes and Culex species. By carrying out 3 blind tests, we achieved the identification of mosquito vectors at the species level, including the differentiation of An. gambiae complex, which is possible using MALDI-TOF-MS with 1.8 as the cut-off identification score. A cluster analysis performed with all available mosquito species showed that MALDI-Biotyper can distinguish between specimens at the subspecies level, as demonstrated for An gambiae M and S, but this method cannot yet be considered a reliable tool for the phylogenetic study of mosquito species.

Conclusions: We confirmed that even without any specific expertise, MALDI-TOF-MS profiling of mosquito leg protein extracts can be used for the rapid identification of mosquito vectors. Therefore, MALDI-TOF-MS is an alternative, efficient and inexpensive tool that can accurately identify mosquitoes collected in the field during entomological surveys.

Competing Interests: The authors have declared that no competing interests exist.

Figure 1

Figure 1. Study design to evaluate the...

Figure 1. Study design to evaluate the potential of MALDI-TOF-MS for identifying mosquito vectors.

Figure 2

Figure 2. MALDI-TOF-MS spectra obtained from leg...

Figure 2. MALDI-TOF-MS spectra obtained from leg protein extraction of four An. gambiae form...

Figure 3

Figure 3. Matrix-assisted laser desorption time of...

Figure 3. Matrix-assisted laser desorption time of flight mass spectra profiles of legs only from ...

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Figure 4. Dendrogram obtained by a cluster...

Figure 4. Dendrogram obtained by a cluster analysis of the spectra obtained from laboratory-reared and...