Large-Scale Cytochrome C Oxidase Subunit I Gene Data Analysis for the Development of a Multiplex Polymerase Chain Reaction Test Capable of Identifying Biting Midge Vector Species and Haplotypes (Diptera: Ceratopogonidae) of the Culicoides Subgenus Avaritia Fox, 1955

Author:

Dähn Oliver1,Werner Doreen2,Mathieu Bruno3ORCID,Kampen Helge1

Affiliation:

1. Friedrich-Loeffler-Institut (FLI), Federal Research Institute for Animal Health, Südufer 10, 17493 Greifswald, Germany

2. Leibniz Centre for Agricultural Landscape Research (ZALF), Eberswalder Str. 84, 15374 Müncheberg, Germany

3. Institutes of Bacteriology and Parasitology, Medical Faculty, University of Strasbourg, UR 3073 PHAVI, 67000 Strasbourg, France

Abstract

The emergence of culicoid-transmitted bluetongue and Schmallenberg viruses in several European countries demonstrated the ability of indigenous biting midge species to transmit pathogens. Entomologic research programs identified members of the Obsoletus Group (Culicoides subgenus Avaritia) as keyplayers in disease epidemiology in Europe. However, morphological identification of potential vectors is challenging due to the recent discovery of new genetic variants (haplotypes) of C. obsoletus sensu stricto (s.s.), forming distinct clades. In this study, 4422 GenBank entries of the mitochondrial cytochrome c oxidase subunit I (COI) gene of subgenus Avaritia members of the genus Culicoides were analyzed to develop a conventional multiplex PCR, capable of detecting all vector species and clades of the Western Palearctic in this subgenus. Numerous GenBank entries incorrectly assigned to a species were identified, analyzed and reassigned. The results suggest that the three C. obsoletus clades represent independent species, whereas C. montanus should rather be regarded as a genetic variant of C. obsoletus s.s. Based on these findings, specific primers were designed and validated with DNA material from field-caught biting midges which achieved very high diagnostic sensitivity (100%) when compared to an established reference PCR (82.6%).

Funder

German Federal Ministry of Food and Agriculture

Publisher

MDPI AG

Reference101 articles.

1. Culicoides biting midges: Their role as arbovirus vectors;Mellor;Annu. Rev. Entomol.,2000

2. The taxonomy of Culicoides vector complexes—Unfinished business;Meiswinkel;Vet. Ital.,2004

3. Marquardt, W. (2005). Biology of Disease Vectors, Elsevier. [2nd ed.].

4. Observations on breeding sites and light-trap collections of Culicoides during an outbreak of bluetongue in Cyprus;Mellor;Bull. Entomol. Res.,1979

5. Isolations of African horse sickness virus from vector insects made during the 1988 epizootic in Spain;Mellor;Epidemiol. Infect.,1990

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