Eur. J. Entomol. 122: 88-98, 2025 | DOI: 10.14411/eje.2025.011

COI barcodes for the identification of anthropophilic Biting Midges (Diptera: Ceratopogonidae: Culicoides) from the Brazilian AmazonOriginal article

Emanuelle DE SOUSA FARIAS1, 2, Moises Thiago DE SOUZA FREITAS3, Sanmya Silva DOS SANTOS1, 4, Jokebede Melynda DOS SANTOS PAULINO-ROSA1, 5, Luiz DE SOUZA COELHO6, Jordam William PEREIRA-SILVA1, James Lee CRAINEY1, *, Claudia María RÍOS-VELÁSQUEZ1, Felipe Arley COSTA PESSOA1
1 Instituto Leônidas e Maria Deane - FIOCRUZ, Laboratório de Ecologia e Doenças Transmissíveis na Amazônia, Manaus, Amazonas, Brazil; e-mails: emanuellefarias82@gmail.com, sanmyasantos23@gmail.com, James.Lee@fiocruz.br, JokebedeMelynda@gmail.com, jordamwilliam@gmail.com, criosvelasquez@gmail.com, facpessoa@gmail.com
2 Instituto Oswaldo Cruz-IOC/FIOCRUZ, Programa de Pós-Graduação em Biodiversidade e Saúde, Rio de Janeiro, Brazil
3 Universidade Federal de Pernambuco, Departamento de Genética, Pernambuco, Brazil; e-mail: moisesfrts@gmail.com
4 Instituto Leônidas e Maria Deane - FIOCRUZ, Programa de Bolsa de Iniciação Científica - PIBIC/CNPq, Manaus, Amazonas, Brazil
5 Instituto Leônidas e Maria Deane - FIOCRUZ, Programa de Pós-Graduação em Biologia da Interação Patógeno Hospedeiro, Manaus, Amazonas, Brazil
6 Instituto Nacional de Pesquisa na Amazônia - INPA, Laboratório de Inventário Florístico e Botânica Econômica, Manaus, Amazonas, Brazil; e-mail: lscoelhos@gmail.com

The genus Culicoides is the best known of the family Ceratopogonidae. Hematophagous females of the genus typically feed on the blood of vertebrate animals and in the Brazilian Amazon often on the blood of human beings. Amazon region anthropophilic Culicoides bites can provoke allergic reactions and transmit Mansonella ozzardi as well as the Oropouche virus. Past integrated taxonomy studies, combining morphometric and molecular analyses, have revealed hidden disease vector biodiversity and cryptic species with epidemiological and disease control relevance and have provided new tools to assist with vector identification. For this study we used light traps set in 12 distinct sites from three different Amazon states: Rondonia (1 site), Amazonas (3 sites) and Para (8 sites). We captured 12 different species of Culicoides representing seven different subgenera: C. foxi, C. fusipalpis, C. hylas, C. insignis, C. plaumanni, C. pseudodiabolicus, C. ruizi, C. debilipalpis, C. glabrior, C. jurutiensis, C. paraensis, C. pauci­enfuscatus. Between two and nine specimens were barcoded of each species. Neighbor joining and Maximum likelihood phylogenetic analysis with these COI barcodes showed the utility of these barcode sequences for species identification by clustering the barcode sequences into bootstrap-supported, species-specific monophyletic groups. Although this barcoding analysis did not resolve relationships between the species studied, it did reveal cryptic diversity within C. paucienfuscatus, C. glabrior, C. plaumanni, C. insignis and C. pseudodiabolicus. Two-dimensional geometric morphometrics, using eight wing-vein landmarks, robustly separated the analyzed species and raised questions about the validity of the subgenus Haematomyidium. Importantly, our GM wing landmark analysis separated C. paraensis from all the other analyzed species suggesting this type of analysis could be harnessed for epidemiological monitoring of this key Amazon-region vector species.

Keywords: Taxonomy, hematophagy, disease vector, wing morphometrics, landmark analysis

Received: September 11, 2024; Revised: April 4, 2025; Accepted: April 4, 2025; Published online: May 7, 2025  Show citation

ACS AIP APA ASA Harvard Chicago Chicago Notes IEEE ISO690 MLA NLM Turabian Vancouver
DE SOUSA FARIAS, E., DE SOUZA FREITAS, M.T., DOS SANTOS, S.S., DOS SANTOS PAULINO-ROSA, J.M., DE SOUZA COELHO, L., PEREIRA-SILVA, J.W., ... COSTA PESSOA, F.A. (2025). COI barcodes for the identification of anthropophilic Biting Midges (Diptera: Ceratopogonidae: Culicoides) from the Brazilian Amazon. EJE, 122, Article 88-98. https://doi.org/10.14411/eje.2025.011
Share...
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)
    • Open full article

    References

    1. Adler P.H., Cheke R.A. & Post R.J. 2010: Evolution, epidemiology, and population genetics of black flies (Diptera: Simuliidae).­­­­ - Infect. Genet. Evol. 10: 846-865. Go to original source...
    2. Ander M., Troell K. & Chirico J. 2013: Barcoding of biting midges in the genus Culicoides: a tool for species determination. - Med. Vet. Entomol. 27: 323-331. Go to original source...
    3. Augot D., Sauvage F., Jouet D., Simphal E., Veuille M., Couloux A., Kaltenbach M.L. & Depaquit J. 2010: Discrimination of Culicoides obsoletus and C. scoticus, potential bluetongue vectors, by morphometrical and mitochondrial cytochrome oxidase subunit I analysis. - Infect. Genet. Evol. 10: 629-637. Go to original source...
    4. Bellis G.A., Dyce A.L., Gopurenko D. & Mitchell A. 2013: Revision of the Immaculatus group of Culicoides Latreille (Diptera: Ceratopogonidae) from the Australasian Region with description of two new species. - Zootaxa 3680: 15-37. Go to original source...
    5. Borkent A. 2016a: The Subgeneric Classification of Species of Culicoides - Thoughts and a Warning. URL: https://wwv.inhs.illinois.edu/files/5014/6532/8290/CulicoidesSubgenera.pdf
    6. Borkent A. 2016b: Numbers of Extant and Fossil Species of Ceratopogonidae. URL: https://wwv.inhs.illinois.edu/files/4014/6785/5847/WorldCatalogtaxa.pdf
    7. Borkent A. & Dominiak P. 2020: Catalog of the Biting Midges of the World (Diptera: Ceratopogonidae). - Zootaxa 4787: 1-377. Go to original source...
    8. Calle D.A., Quiñones M.L., Erazo H.J. & Jaramillo N. 2008: Discriminación por morfometría geométrica de once especies de Anopheles (Nyssorhynchus) presentes en Colombia. - Biomedica 28: 371-385. Go to original source...
    9. Cannet A., Simon-Chane C., Histace A., Akhoundi M., Romain O., Souchaud M., Jacob P., Sereno D., Gouagna L.C., Bousses P., Mathieu-Daude F. & Sereno D. 2023a: Wing Interferential Patterns (WIPs) and machine learning for the classification of some Aedes species of medical interest. - Sci. Rep. 13: 17628, 12 pp. Go to original source...
    10. Cannet A., Simon-Chane C., Histace A., Akhoundi M., Romain O., Souchaud M., Jacob P., Sereno D., Volf P., Dvorak V. & Sereno D. 2023b: Species identification of phlebotomine sandflies using deep learning and wing interferential pattern (WIP). - Sci. Rep. 13: 21389, 9 pp. Go to original source...
    11. Cannet A., Simon-Chane C., Akhoundi M., Histace A., Romain O., Souchaud M., Jacob P., Sereno D., Mouline K., Barnabe C., Lardeux F., Boussès P. & Sereno D. 2023c: Deep learning and wing interferential patterns identify Anopheles species and discriminate amongst Gambiae complex species. - Sci. Rep. 13: 13895, 13 pp. Go to original source...
    12. Cariou M., Duret L. & Charlat S. 2017: The global impact of Wolbachia on mitochondrial diversity and evolution. - J. Evol. Biol. 30: 2204-2210. Go to original source...
    13. Carvalho L.P.C., Costa Silva G., Pereira Júnior A.M., Paulo P.F.M., Silva G.S., Carioca A.P.M., Rodrigues B.L., Pessoa F.A.C. & Medeiros J.F. 2022: DNA barcoding of genus Culicoides biting midges (Diptera: Ceratopogonidae) in the Brazilian Amazon. - Acta. Trop. 235: 106619, 8 pp. Go to original source...
    14. Cêtre-Sossah C., Baldet T., Delécolle J.C., Mathieu B., Perrin A., Grillet C. & Albina E. 2004: Molecular detection of Culicoides spp. and Culicoides imicola, the principal vector of African horse sickness (AHS) in Africa and Europe. - Vet. Res. 35: 325-337. Go to original source...
    15. Conceição P.A., Crainey J.L., Almeida T.P., Shelley A.J. & Luz S.L. 2013: New molecular identifiers for Simulium limbatum and Simulium incrustatum sl and the detection of genetic substructure with potential implications for onchocerciasis epidemiology in the Amazonia focus of Brazil. - Acta. Trop. 127: 118-125. Go to original source...
    16. Crainey J.L., Marín M.A., Silva T.R.R.D., de Medeiros J.F., Pessoa F.A.C., Santos Y.V., Vicente A.C.P. & Luz S.L.B. 2018: Mansonella ozzardi mitogenome and pseudogene characterisation provides new perspectives on filarial parasite systematics and CO-1 barcoding. - Sci. Rep. 8: 6158, 11 pp. Go to original source...
    17. Delgado-Noguera L.A., Hernández-Pereira C.E., Ramírez J.D., Hernández C., Velasquez-Ortíz N., Clavijo J., Ayala J.M., Forero-Peña D., Marquez M., Suarez M.J. et al. 2022: Tele-entomology and tele-parasitology: A citizen science-based approach for surveillance and control of Chagas disease in Venezuela. - Parasite Epidemiol. Control 19: e00273, 9 pp. Go to original source...
    18. de Lima V.R., de Morais M.C.C. & Kirchgatter K. 2023: Integrating artificial intelligence and wing geometric morphometry to automate mosquito classification. - Acta. Trop. 249: 107089, 7 pp. Go to original source...
    19. Demari-Silva B., Suesdek L., Sallum M.A.M. & Marrelli M.T. 2014: Wing geometry of Culex coronator (Diptera: Culicidae) from South and Southeast Brazil. - Parasit Vectors 7: 174, 9 pp. Go to original source...
    20. Deng J., Assandri G., Chauhan P., Futahashi R., Galimberti A., Hansson B., Lancaster L.T., Takahashi Y., Svensson E.I. & Duplouy A. 2021: Wolbachia-driven selective sweep in a range expanding insect species. - BMC Ecol. Evol. 21: 1-17. Go to original source...
    21. Farias E.S., Almeida J.F., Pereira-Silva J.W., Coelho L.S., Ríos-Velásquez C.M., Luz S.L.B. & Pessoa F.A.C. 2020: Diversity of biting midges Culicoides (Diptera: Ceratopogonidae), potential vectors of disease, in different environments in an Amazonian rural settlement, Brazil. - Rev. Soc. Bras. Med. Trop. 53: e20200067, 8 pp. Go to original source...
    22. Feitoza L.H.M., de Carvalho L.P.C., da Silva L.R., Meireles A.C.A., Rios F.G.F., Silva G.S., Medeiros de Paulo P.F., Pessoa F.A.C. & Julião G.R. 2023: Influence of meteorological and seasonal parameters on the activity of Culicoides paraensis (Diptera: Ceratopogonidae), an annoying anthropophilic biting midge and putative vector of Oropouche virus in Rondônia, Brazilian Amazon. - Acta. Trop. 243: 106928, 8 pp. Go to original source...
    23. Fontaine M.C., Pease J.B., Steele A., Waterhouse R.M., Neafsey D.E., Sharakhov I.V., Hall A.B., Evdoxia Kakani F.C.E., Mitchell S.N., Wu Y.C., Smith H.A., Love R.R., Lawniczak M.K., Slotman M.A., Emrich S.J., Hahn M.W. & Besansky N.J. 2015: Extensive introgression in a malaria vector species complex revealed by phylogenomics. - Science 347: 1258524, 20 pp. Go to original source...
    24. Freitas M.T.S., Ríos-Velasquez C.M., Costa C.R. Jr., Figueiredo C.A., Aragão N.C., Balbino T.C.L., Pessoa F.A.C. & Balbino V.Q. 2015: Phenotypic and genotypic variations among three allopatric populations of Lutzomyia umbratilis, main vector of Leishmania guyanensis. - Parasit Vectors 8: 448, 10 pp. Go to original source...
    25. Georgiadou S.P., Makaritsis K.P. & Dalekos G.N. 2015: Leishmaniasis revisited: current aspects on epidemiology, diagnosis and treatment. - J. Transl. Int. Med. 3: 43-50. Go to original source...
    26. Giordani B.F., Andrade A.J., Galati E.A.B. & Gurgel-Gonçalves R. 2017: The role of wing geometric morphometrics in the identification of sandflies within the subgenus Lutzomyia. - Med. Vet. Entomol. 31: 373-380. Go to original source...
    27. Godoy R.E., Galati E.A.B., Estrela P.C., Souza N.A., Santos T.V., Sousa L.G. & Rangel E. 2014: Comparative study of the phlebotomine sand fly species (Diptera: Psychodidae: Phlebotominae) of the genera Nyssomyia Barretto, 1962, Bichromomyia Artemiev, 1991 and Migonemyia Galati, 1995 vectors of American cutaneous leishmaniasis in Brazil. - Zootaxa 3838: 501-517. Go to original source...
    28. Godoy R.E., Shimabukuro P.H., Santos T.V., Pessoa F.A.C., Cunha A.E.F.L., Santos F.K.M., Vilela M.L., Rangel A.F. & Galati E.A.B. 2018: Geometric morphometry of the head in sand flies (Diptera: Psychodidae: Phlebotominae), an alternative approach to taxonomy studies. - Zootaxa 4504: 566-576. Go to original source...
    29. Guindon S., Dufayard J.F., Lefort V., Anisimova M., Hordijk W. & Gascuel O. 2010: New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. - Syst. Biol. 59: 307-321. Go to original source...
    30. Harbach R.E. 2011: Classification within the cosmopolitan genus Culex (Diptera: Culicidae): The foundation for molecular systematics and phylogenetic research. - Acta. Trop. 120: 1-14. Go to original source...
    31. Henni H.L., Sauvage F., Ninio C., Depaquit J. & Augot D. 2014: Wing geometry as a tool for discrimination of Obsoletus group (Diptera: Ceratopogonidae: Culicoides) in France. - Infect. Genet. Evol. 21: 110-117. Go to original source...
    32. Hernandez-Triana L.M., Crainey J.L., Hall A., Fatih F., Mackenzie-Dodds J., Shelley A.J., Zhou X., Post R.J., Gregory T.R. & Hebert P.D.N. 2012: DNA barcodes reveal cryptic genetic diversity within the blackfly subgenus Trichodagmia Enderlein (Diptera: Simuliidae: Simulium) and related taxa in the New World. - Zootaxa 3514: 43-69. Go to original source...
    33. Jiggins F.M. 2003: Male-killing Wolbachia and mitochondrial DNA: selective sweeps, hybrid introgression and parasite population dynamics. - Genetics 164: 5-12. Go to original source...
    34. Jirakanjanakit N. & Dujardin J.P. 2005: Discrimination of Aedes aegypti (Diptera: Culicidae) laboratory lines based on wing geometry. - Southeast Asian J. Trop. Med. Public Health 36: 858-861.
    35. Lane R.P. & Crosskey R.W. 1993: Medical Insects and Arachnids. Chapman & Hall, London, xv + 723 pp. Go to original source...
    36. Linton Y.M., Mordue A.J., Cruickshank R.H., Meiswinkel R., Mellor P.S. & Dallas J.F. 2002: Phylogenetic analysis of the mitochondrial cytochrome oxidase subunit I gene of five species of the Culicoides imicola species complex. - Med. Vet. Entomol. 16: 139-146. Go to original source...
    37. Lorenz C., Marques T.C., Sallum M.A.M. & Suesdek L. 2012: Morphometrical diagnosis of the malaria vectors Anopheles cruzii, An. homunculus and An. bellator. - Parasit Vectors 5: 257, 7 pp. Go to original source...
    38. Lowe R., Lee S., Lana R.M., Codeço C.T., Castro M.C. & Pascual M. 2020: Emerging arboviruses in the urbanized Amazon rainforest. - B.M.J. 371: m4385, 5 pp. Go to original source...
    39. McAlpine J.F. 1981: Morphology and terminology - adults. In McAlpine J.F., Peterson B.V., Shewell G.E., Teskey H.J., Vockeroth J.R. & Wood D.M. (eds): Manual of Nearctic Diptera, Vol. 1. Research Branch, Agriculture Canada (Monograph No. 27), Ottawa, pp. 9-63.
    40. Mohapatra R.K., Mishr S., Satapathy P., Kandi V. & Tuglo L.S. 2024: Surging Oropouche Virus (OROV) cases in the Americas: A public health challenge. - New Microbes New Infect. 59: 101243, 2 pp. Go to original source...
    41. Muñoz-Muñoz F., Talavera S. & Pagès N. 2011: Geometric morphometrics of the wing in the subgenus Culicoides (Di­ptera: Ceratopogonidae): from practical implications to evolutionary interpretations. - J. Med. Entomol. 48: 129-139. Go to original source...
    42. Pagès N. & Sarto i Monteys I. 2005: Differentiation of Culicoides obsoletus and Culicoides scoticus (Diptera: Ceratopogonidae) based on mitochondrial cytochrome oxidase subunit I. - J. Med. Entomol. 42: 1026-1034. Go to original source...
    43. Pagès N., Muñoz-Muñoz F., Talavera S., Sarton V., Lorca C. & Núñez J.I. 2009: Identification of cryptic species of Culicoides (Diptera: Ceratopogonidae) in the subgenus Culicoides and development of species-specific PCR assays based on Barcode regions. - Vet. Parasitol. 165: 298-310. Go to original source...
    44. Pataki B.A., Garriga J., Eritja R., Palmer J.R., Bartumeus F. & Csabai I. 2021: Deep learning identification for citizen science surveillance of tiger mosquitoes. - Sci. Rep. 11: 4718, 12 pp. Go to original source...
    45. Peng N.Y., Hall R.N., Huang N., West P., Cox T.E., Mahar J.E., Mason H., Campbell S., O'Connor T., Read A.J. et al. 2023: Utilizing molecular epidemiology and citizen science for the surveillance of lagoviruses in Australia. - Viruses 15: 2348, 17 pp. Go to original source...
    46. Portela C.S., de Araújo C.P.M., Sousa P.M., Simão C.L.G., de Oliveira J.C.S. & Crainey J.L. 2024: Filarial disease in the brazilian amazon and emerging opportunities for treatment and control. - Curr. Res. Parasitol. Vector Borne Dis. 5: 100168, 10 pp. Go to original source...
    47. Saitou N. & Nei M.1987: The neighbor-joining method: a new method for reconstructing phylogenetic trees. - Mol. Biol. Evol. 4: 406-425.
    48. Sakkas H., Bozidis P., Franks A. & Papadopoulou C. 2018: Oropouche fever: a review. - Viruses 10: 175, 16 pp. Go to original source...
    49. Santarém M.C. & Felippe-Bauer M.L. 2022: Brazilian species of biting midges - espécies de maruins do Brasil (Diptera: Ceratopogonidae). URL: https://portal.fiocruz.br/sites/portal.fiocruz.br/files/documentos/brazilian_species_of_biting_midges_2022_0.pdf
    50. Santarém M.C.A., Farias E.S. & Felippe-Bauer M.L. 2015: Culicoides castelloni sp. nov. from the Brazilian Amazon Region with a revision of the reticulatus species group (Diptera, 63 Ceratopogonidae). - An. Acad. Bras. Cienc. 87: 955-972. Go to original source...
    51. Sick F., Beer M., Kampen H. & Wernike K. 2019: Culicoides biting midges - underestimated vectors for arboviruses of public health and veterinary importance. - Viruses 11: 376, 19 pp. Go to original source...
    52. Slama D., Haouas N., Remadi L., Mezhoud H., Babba H. & Chaker E. 2014: First detection of Leishmania infantum (Kinetoplastida: Trypanosomatidae) in Culicoides spp. (Diptera: Ceratopogonidae). - Parasit. Vectors 7: 51, 3 pp. Go to original source...
    53. Song H., Buhay J.E., Whiting M.F. & Crandall K.A. 2008: Many species in one: DNA barcoding overestimates the number of species when nuclear mitochondrial pseudogenes are coamplified. - Proc. Natl. Acad. Sci. U.S.A. 105: 13486-13491. Go to original source...
    54. Spinelli G.R. & Wirth W.W. 1993: Los Ceratopogonidae de la Argentina (Insecta: Diptera). In Castellanos Z.A. (ed.): Fauna de Agua Dulce de la Republica Argentina, Vol. 38(3). Profadu (Conicet), Buenos Aires, 124 pp.
    55. Ta-Tang T.H., Crainey J.L., Post R.J., Luz S.L. & Rubio J.M. 2018: Mansonellosis: current perspectives. - Res. Rep. Trop. Med. 9: 9-24. Go to original source...
    56. Ta-Tang T.H., Luz S.L., Crainey J.L. & Rubio J.M. 2021: An overview of the management of mansonellosis - Res. Rep. Trop. Med 12: 93-105. Go to original source...
    57. Tamura K., Peterson D., Peterson N., Stecher G., Nei M. & Kumar S. 2011: MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. - Mol. Biol. Evol. 28: 2731-2739. Go to original source...
    58. Vidal P.O., Carvalho E. & Suesdek L. 2012: Temporal variation of wing geometry in Aedes albopictus. - Mem. Inst. Oswaldo Cruz 107: 1030-1034. Go to original source...
    59. Wirth W.W. & Blanton F.S. 1959: Biting midges of the genus Culicoides from Panama (Diptera: Heleidae). - Proc. U.S. Natl. Mus. 109: 237-482. Go to original source...
    60. Wirth W.W. & Marston N. 1968: Method for mounting small insects on microscope slides in Canada balsam. - Ann. Entomol. Soc. Am. 61: 783-784. Go to original source...
    61. Yildirim A., Dik B., Duzlu O., Onder Z., Ciloglu A., Yetismis G. & Inci A. 2019: Genetic diversity of Culicoides species within the Pulicaris complex (Diptera: Ceratopogonidae) in Turkey inferred from mitochondrial COI gene sequences. - Acta. Trop. 190: 380-388. Go to original source...

    This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International License (CC BY 4.0), which permits use, distribution, and reproduction in any medium, provided the original publication is properly cited. No use, distribution or reproduction is permitted which does not comply with these terms.


    Return to the content