Identification of French Guiana sand flies using MALDI-TOF mass spectrometry with a new mass spectra library

Abstract

Phlebotomine sand flies are insects that are highly relevant in medicine, particularly as the sole proven vectors of leishmaniasis. Accurate identification of sand fly species is an essential prerequisite for eco-epidemiological studies aiming to better understand the disease. Traditional morphological identification is painstaking and time-consuming, and molecular methods for extensive screening remain expensive. Recent studies have shown that matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a promising tool for rapid and cost-effective identification of arthropod vectors, including sand flies. The aim of this study was to validate the use of MALDI-TOF MS for the identification of Northern Amazonian sand flies. We constituted a MALDI-TOF MS reference database comprising 29 species of sand flies that were field-collected in French Guiana, which are expected to cover many of the more common species of the Northern Amazonian region, including known vectors of leishmaniasis. Carrying out a blind test, all the sand flies tested ( n = 157) with a log (score) threshold greater than 1.7 were correctly identified at the species level. We confirmed that MALDI-TOF MS protein profiling is a useful tool for the study of sand flies, including neotropical species, known for their great diversity. An application that includes the spectra generated here will be available to the scientific community in the near future via an online platform.

New assessment of Anopheles vector species identification using MALDI-TOF MS

“…As all the previous studies have included in the databases specimens of the same origin as the specimens to be tested, they did not reveal as much the importance of this methodology. However, heterogeneity of mass spectra was also reported when comparing mosquito species [ 10 , 11 , 16 ] or sand fly species [ 22 , 32 , 33 ] from various geographical origins and between reared and field mosquito spectra [ 10 , 15 ]. For a same species, the observation of biomarkers specific to colony specimens and to field specimens [ 10 , 15 ] suggests a great variability in protein content due to phenotypic distinctness in relation to the genetic diversity of Anopheles, influenced by environmental settings, evolutionary history adaptation, demographic history or genetic drift.…”

“…Once dried, the deposits were covered with a 1-µL alpha-cyano-4-hydroxycinnamic acid (HCCA) matrix prepared in 50% acetonitrile and 2.5% trifluoroacetic acid and 47.5% of HPLC grade water (final concentration of 10 mg/mL). To ensure the reproducibility of the results, a total of ten replicates were spotted for each specimen to be included in the database, and a total of four replicates were spotted for each specimen of the panel to be tested, as previously published [ 22 , 23 ].…”

“…During crosswise comparison, the first hits corresponding to cross-identification with any spectrum of the same specimen were eliminated, and the second or third hits of cross-identification were selected for the analysis. The LSV threshold for an interpretable identification result was defined at 1.7 because it is a commonly used threshold for arthropod vector species identification using MALDI-TOF [ 13 , 22 ]. The impact of various LSV thresholds from 1.7 to 2 was assessed, and modifying the threshold did not impact the main results presented in the study (Additional file 2 : Fig.…”

“…Each of the four raw spectra of the panels obtained from each anatomic part was analysed against databases 1 (panels A and B), 2, 3 and 4 (panel A). As previously published [ 22 , 23 ], only the replicate with the highest LSV was selected, and the identification corresponded to the first hit obtained for this replicate. MALDI-TOF MS identifications were compared to molecular identifications for every specimen.…”

An integrative approach to identify sand fly vectors of leishmaniasis in Ethiopia by morphological and molecular techniques

“…Recently, an alternative molecular method, protein profiling by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), has been introduced for species identification in different arthropod families, including sand flies [20,[23][24][25][26][27][28]. This approach provides unique protein profiles that allow unambiguous species identification and sample processing is rapid, simple and cost-effective.…”