Integrative taxonomy to investigate species boundaries within Culicoides (Diptera: Ceratopogonidae): a case study using subgenus Avaritia from Australasia and Eastern Asia.

In this study, species boundaries were examined for 15 described and 2 undescribed species within the economically important Culicoides subg. Avaritia Fox from Australasia and Eastern Asia. We used an integrative taxonomic approach incorporating DNA barcoding, nuclear gene sequencing, and retrospective morphological analyses. Some arbovirus vector species such as Culicoides fulvus Sen and Das Gupta and Culicoides wadai Kitaoka were genetically and morphologically uniform across sampled distributions, but others including Culicoides actoni Smith and Culicoides brevipalpis Delfinado contained 2 or more genetically independent populations of 'cryptic species' that in some cases were sympatric. Some of these 'cryptic species' exhibited consistent morphological differences, while differences are yet to be found for others species. Additionally, an undescribed species, C. Avaritia sp. No. 3, was found to be synonymous with C. fulvus. These results refine our understanding of the distribution of individual species of C. subg. Avaritia and demonstrate that species descriptions and distribution records need revision for part of the Culicoides fauna. Furthermore, because vector competence studies for most of these species are based entirely on Australian populations, the competence of the putative cryptic species identified elsewhere will require independent assessment. Finally, integrative taxonomic assessment requires genetic and morphological assessment of material from the type localities in order to clarify the status and distribution of species, especially for clades containing cryptic species. International collaboration is needed to facilitate this research.