Objective To explore the sample conditions for the matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF MS) method used for mosquito species identification, so as to provide a basis for establishing a MALDI-TOF MS method for rapid and accurate mosquito species identification. Methods Four laboratory mosquito strains, namely Aedes albopictus, Culex pipiens pallens, Cx. tritaeniorhynchus, and Anopheles sinensis, were selected as the study subjects. The MALDI-TOF MS method was used to analyze the samples from these mosquitoes for collecting peptide mass fingerprint and establishing a fingerprint database for mosquito species identification. The samples varied by body part (whole body, head, thorax, and foot), preservation method (cryopreservation at -20°C or dry preservation in silica gel), and blood-sucking status (non-blood-sucked and blood-sucked). The accuracy of this database was evaluated through internal cross-validation with 120 samples and identification scores using the software's standard parameters. Results Except for the abdomen, the fingerprints of the thorax, foot, and head of each mosquito species were clustered into one group, and different preservation method and blood-feeding status of those mosquitoes did not have any impact on the clustering results of mosquito species identification. At the same time, 120 samples were cross validated within the database, and the results were consistent with the morphological identification results, with identification scores >2.00. Only 1 μl of protein extract was allowed for the identification of mosquito species. Conclusions The head, thorax, and legs of mosquitoes can all be used for mosquito species identification by the MALDI-TOF MS method, and the blood-feeding status of mosquitoes and sample preservation methods do not affect mosquito identification results.The MALDI-TOF MS method can be used for mosquito species identification and offers the advantages of rapidity, accuracy, and sensitivity. These features make it highly useful for mosquito species identification.
GE Wen-jun, XIAO Di, ZHANG Hui-fang, ZHAO Chun-chun, LUN Xin-chang, SONG Xiu-ping, GUO Yu-hong, LIANG Ying, LIU Qi-yong, MENG Feng-xia
. A preliminary study on the identification of mosquito species based on MALDI-TOF MS[J]. Chinese Journal of Vector Biology and Control, 2025
, 36(2)
: 238
-245
.
DOI: 10.11853/j.issn.1003.8280.2025.02.016
[1] Nebbak A，Almeras L，Parola P，et al. Mosquito vectors （Diptera：Culicidae） and mosquito-borne diseases in north Africa[J]. Insects，2022，13（10）：962. Doi：10.3390/insects13100962.
[2] The Lancet Global Health. Vector control：Time for a planetary health approach[J]. Lancet Glob Health，2017，5（6）：e556. DOI：10.1016/S2214-109X（17）30185-7.
[3] World Health Organization. Ten threats to global health in 2019[EB/OL]. （2019-01-10）[2024-11-06]. https：//www.who.int/news-room/spotlight/ten-threats-to-global-health-in-2019/.
[4] Harbach RE. Family Culicidae Meigen，1818[EB/OL]. （2024-10-18）[2024-11-06]. https：//mosquito-taxonomic-inventory.myspecies.info/node/13679.
[5] Liu QY. Reported cases of vector-borne diseases in China，2005-2020：Epidemic trend，challenges in prevention and control，and related coping strategies[J]. Chin J Vector Biol Control，2022，33（1）：1-7. DOI：10.11853/j.issn.1003.8280.2022.01.001.（in Chinese） 刘起勇. 2005－2020年我国媒介生物传染病报告病例：流行趋势、防控挑战及应对策略[J]. 中国媒介生物学及控制杂志，2022，33（1）：1-7. DOI：10.11853/j.issn.1003.8280.2022.01.001.
[6] Atoni E，Zhao L，Hu C，et al. A dataset of distribution and diversity of mosquito-associated viruses and their mosquito vectors in China[J]. Sci Data，2020，7（1）：342. DOI：10.1038/s41597-020-00687-9.
[7] Fu WB，Chen B. Taxonomy and fauna of Culicidae：History and current knowledge[J]. Acta Entomol Sin，2018，61（1）：122-138. DOI：10.16380/j.kcxb.2018.01.014.（in Chinese） 付文博，陈斌. 蚊科昆虫分类及区系研究历史和现状概述[J]. 昆虫学报，2018，61（1）：122-138. DOI：10.16380/j.kcxb.2018. 01.014.
[8] Jourdain F，Picard M，Sulesco T，et al. Identification of mosquitoes （Diptera：Culicidae）：An external quality assessment of medical entomology laboratories in the MediLabSecure Network[J]. Parasit Vectors，2018，11（1）：553. DOI：10.1186/s13071-018-3127-7.
[9] Beebe NW. DNA barcoding mosquitoes：Advice for potential prospectors[J]. Parasitology，2018，145（5）：622-633. DOI：10.1017/S0031182018000343.
[10] Tang X，Xiao D，Zhang HF，et al. Establishment and evaluation of identification of Brucella by MALDI-TOF-MS[J]. Chin J Zoonoses，2016，32（9）：772-778. DOI：10.3969/j.issn.1002-2694.2016.09.002.（in Chinese） 汤旭，肖迪，张慧芳，等. MALDI-TOF-MS鉴定布鲁氏菌方法建立和评价[J]. 中国人兽共患病学报，2016，32（9）：772-778. DOI：10.3969/j.issn.1002-2694.2016.09.002.
[11] Yssouf A，Flaudrops C，Drali R，et al. Matrix-assisted laser desorption ionization-time of flight mass spectrometry for rapid identification of tick vectors[J]. J Clin Microbiol，2013，51（2）：522-528. DOI：10.1128/JCM.02665-12.
[12] Chavy A，Nabet C，Normand AC，et al. Identification of French Guiana sand flies using MALDI-TOF mass spectrometry with a new mass spectra library[J]. PLoS Negl Trop Dis，2019，13（2）：e0007031. DOI：10.1371/journal.pntd.0007031.
[13] Sánchez-Juanes F，Calvo Sánchez N，Belhassen García M，et al. Applications of MALDI-TOF mass spectrometry to the identification of parasites and arthropod vectors of human diseases[J]. Microorganisms，2022，10（11）：2300. DOI：10.3390/microorganisms10112300.
[14] Lu BL. Fauna Sinica. Insecta，vol. 8. Diptera：Culicidae Ⅰ[M]. Beijing：Science Press，1997：.（in Chinese） 陆宝麟. 中国动物志. 昆虫纲. 第8卷. 双翅目. 蚊科（上）[M]. 北京：科学出版社，1997：.
[15] Lu BL. Fauna Sinica. Insecta，vol. 9. Diptera：Culicidae Ⅱ[M]. Beijing：Science Press，1997：31-34.（in Chinese） 陆宝麟. 中国动物志. 昆虫纲. 第9卷. 双翅目. 蚊科（下）[M]. 北京：科学出版社，1997：31-34.
[16] Yssouf A，Almeras L，Raoult D，et al. Emerging tools for identification of arthropod vectors[J]. Future Microbiol，2016，11（4）：549-566. DOI：10.2217/fmb.16.5.
[17] Bamou R，Costa MM，Diarra AZ，et al. Enhanced procedures for mosquito identification by MALDI-TOF MS[J]. Parasit Vectors，2022，15（1）：240. DOI：10.1186/s13071-022-05361-0.
[18] Batovska J，Blacket MJ，Brown K，et al. Molecular identification of mosquitoes （Diptera：Culicidae） in southeastern Australia[J]. Ecol Evol，2016，6（9）：3001-3011. DOI：10.1002/ece3.2095.
[19] Guo XX，Cheng P，Liu LJ，et al. Analysis of population genetic diversity of mosquitoes from Shandong Province based on mitochondrial DNA cytochrome oxidase subunit Ⅰgene fragment[J]. Chin J Schisto Control，2018，30（1）：37-41. DOI：10.16250/j.32.1374.2017091.（in Chinese） 郭秀霞，程鹏，刘丽娟，等. 基于mtDNA-COⅠ基因的山东省蚊种群遗传多样性分析[J]. 中国血吸虫病防治杂志，2018，30（1）：37-41. DOI：10.16250/j.32.1374.2017091.
[20] Sevestre J，Diarra AZ，Laroche M，et al. Matrix-assisted laser desorption/ionization time-of flight mass spectrometry：an emerging tool for studying the vectors of human infectious diseases[J]. Future Microbiol，2021，16：323-340. DOI：10.2217/fmb-2020-0145.
[21] Vega-Rúa A，Pagès N，Fontaine A，et al. Improvement of mosquito identification by MALDI-TOF MS biotyping using protein signatures from two body parts[J]. Parasit Vectors，2018，11（1）：574. DOI：10.1186/s13071-018-3157-1.
[22] Niare S，Berenger JM，Dieme C，et al. Identification of blood meal sources in the main African malaria mosquito vector by MALDI-TOF MS[J]. Malar J，2016，15：87. DOI：10.1186/s12936-016-1152-6.
[23] Tahir D，Almeras L，Varloud M，et al. Assessment of MALDI-TOF mass spectrometry for filariae detection in Aedes aegypti mosquitoes[J]. PLoS Negl Trop Dis，2017，11（12）：e0006093. DOI：10.1371/journal.pntd.0006093.
[24] Müller P，Pflüger V，Wittwer M，et al. Identification of cryptic Anopheles mosquito species by molecular protein profiling[J]. PLoS One，2013，8（2）：e57486. DOI：10.1371/journal.pone.0057486.
[25] Raharimalala FN，Andrianinarivomanana TM，Rakotondrasoa A，et al. Usefulness and accuracy of MALDI-TOF mass spectrometry as a supplementary tool to identify mosquito vector species and to invest in development of international database[J]. Med Vet Entomol，2017，31（3）：289-298. DOI：10.1111/mve.12230.
