Identification of Tsetse (Glossina spp.) Using Matrix-Assisted Laser Desorption/Ionisation Time of Flight Mass Spectrometry

Abstract

Glossina (G.) spp. (Diptera: Glossinidae), known as tsetse flies, are vectors of African trypanosomes that cause sleeping sickness in humans and nagana in domestic livestock. Knowledge on tsetse distribution and accurate species identification help identify potential vector intervention sites. Morphological species identification of tsetse is challenging and sometimes not accurate. The matrix-assisted laser desorption/ionisation time of flight mass spectrometry (MALDI TOF MS) technique, already standardised for microbial identification, could become a standard method for tsetse fly diagnostics. Therefore, a unique spectra reference database was created for five lab-reared species of riverine-, savannah- and forest- type tsetse flies and incorporated with the commercial Biotyper 3.0 database. The standard formic acid/acetonitrile extraction of male and female whole insects and their body parts (head, thorax, abdomen, wings and legs) was used to obtain the flies' proteins. The computed composite correlation index and cluster analysis revealed the suitability of any tsetse body part for a rapid taxonomical identification. Phyloproteomic analysis revealed that the peak patterns of G. brevipalpis differed greatly from the other tsetse. This outcome was comparable to previous theories that they might be considered as a sister group to other tsetse spp. Freshly extracted samples were found to be matched at the species level. However, sex differentiation proved to be less reliable. Similarly processed samples of the common house fly Musca domestica (Diptera: Muscidae; strain: Lei) did not yield any match with the tsetse reference database. The inclusion of additional strains of morphologically defined wild caught flies of known origin and the availability of large-scale mass spectrometry data could facilitate rapid tsetse species identification in the future.

Concurrent Proteomic Fingerprinting and Molecular Analysis of Cyathostomins

“…Submission of one part for MALDI‐TOF MS and use of remaining parts for PCR is a straightforward strategy . However, this approach bears potential pitfalls as for example different protein patterns in different body parts were observed in tsetse flies . This approach is not applicable in strongylid nematodes, since the body of nematodes cannot be separated into representative tagmata.…”

“…A promising method for species‐identification is proteomic fingerprinting using MALDI‐TOF MS . It is routinely used for bacteria and has also been applied to ectoparasites such as insects and ticks . The few reports previously published on the application of MALDI‐TOF MS to parasitic nematodes used well established single species nematode isolates to obtain master spectra libraries .…”

Comparative analysis of storage conditions and homogenization methods for tick and flea species for identification by MALDI‐TOF MS

“…A Composite Correlation Index (CCI) tool from MALDI-Biotyper v3.0. Software (Bruker Daltonics), was also used, to assess the spectral variations within and between each sample group, according to the storage conditions tested, as previously described (Hoppenheit et al, 2013;Nebbak et al, 2016). The CCI was computed using the standard settings for the mass range 4000-16 000 Da, resolution four, eight intervals, and autocorrelation off.…”

Accurate identification of Culicidae at aquatic developmental stages by MALDI-TOF MS profiling

“…Based on the introduction of the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) as an economic, rapid, and highly informative tool for bacterial identification and classification [14], this proteomic approach was explored as a taxonomic tool for insects. Since a pioneering study that evaluated MALDI-TOF MS for the discrimination of fruit fly species a decade ago [15], this technique has proven to be applicable for the identification of different arthropod groups, including Drosophila [16], Culicoides [17][18][19][20], Ixodidae [21,22], Glossina [23,24], Phlebotominae [25] and Siphonaptera [26]. MALDI-TOF MS was also successfully applied for the identification of adult mosquitoes allowing the discrimination of cryptic species such as the An.…”