Taxonomic Study On Culicoides Latreille (Diptera:Ceratopogonidae) From China

Objective: Through the investigation and sorting of the species of Culicoides in China,the data and morphological descriptions of 12 species of Avaritia were revised and new species was discovered,which enriched the species diversity of Culicoides in China and provided the basic data for the study of the diseases from Culicoides.To identify and comparatively analyze the species of Culicoides more objectively and accurately by using geometric morphology method and digitizing the morphological characteristics of the traditional classification,which achieve the purpose of rapid and accurate identification of blood-sucking midges.Scanning electron microscope(SEM)is used to observe the ultrastructure of blood-sucking midges among subgenus,species or one species,which shows its morphological characteristics or differences more clearly and intuitively,making it possible to distinguish the species of blood-sucking midges accurately,especially to similar species.Methods: 1.Species description: examine the specimens of the Culicoides to observe their morphological characteristics and record the detailed characteristics of species,such as whether there is pubescence,whether the compound eyes are connected,the number of bristles of clypeus,mandibles and maxil,the distribution of antennal flagellomeres,the presence or absence of sensory pits,the distribution of wing spots,the number of tibiae bristles,spermatheca and the characteristics of hypopygium,the PR and AR values and the wing length and width.We also gathered the new species of subgenus Avaritia since 2005 to 2018 and investigate in the wild to enrich the catalog of subgenus Avaritia.2.Geometric morphology study: The specimens of the adult species of Culicoides were selected in the laboratory,which were preliminary classified,to be made into permanent slides and identified under the microscope according to their morphological characteristics.All of them were photographed under a microscope.All of the wing images were numbered to obtain wing morphological data and analyzed by comparing the Principal component analysis(PCA)and Canonical variate analysis(CVA).3.Scanning electron microscope section: We selected some species of Culicoides and dehydrated them step by step,then dried them in Leica EM CPD300 CO2 automated critical point dryer.The dried specimens were dissected under the microscope and the structure which would be observed was adhered to the sample stage with a double-sided conductive adhesive under a dissecting microscope,then it was sprayed gold by a Supper cool sputter coater(Leica EM SCD050).Finally,the ultrastructure of species of Culicoides was observed by Environmental scanning electron microscopy(FEI Quanta 450).Results: 1.In the description of the morphological characteristics of Culicoides,this study reorganized and re-examined 12 species of subgenus Avaritia in China,the original description and some of the data were revised.Meanwhile,a new species of Avaritia was obtained in the field,which was named for C.(A.)fenggangensis sp.n.and described in details.2.Geometric morphology study of genus Culicoides,the results of CVA analysis of species among the subgenus,species and similar species showed that there was quite different in the wing shape between subgenus Trithecoides and Monoculicoides,and slight different between subgenus Sinocoides and Trithecoides.The P values of Mahalanobis distance were P <0.05 between every two subgenus,showing that there are significant differences in the wing shape;The differences of wing shape among species showed that C.gemellus(DS)and C.xinghaiensis(XH)had quite difference,C.festivipennis(XJ)and C.pictimargo(BY)had slight differences.The P values of Mahalanobis distance in addition to the three groups of C.festivipennis(XJ)-C.pictimargo(BY),C.mcdonaldi(MS)-C.punctatus(CS),C.elbeli(TB)-C.humeralis(JH)were P>0.05,showing that there was no significant difference between the wings,the other groups were all had significant different in wing shape;The results of similar species study showed that there were all significantly different among C.punctatus,C.pulicaris,C.newsteadi.Cluster analysis showed that subgenus Culicoides and Beltranmyia were located on the same branch,revealing that the average shape of wings was the closest among the subgenus;the subgenus Haemophoructus and the other subgenus were quite different;the shape of wing between C.pictimargo and C.festivipennis were the closest;the shape of wing of C.oxystoma and C.gemellus with had a quite difference compared with the others.The result of three similar species showed that the shape of wings of C.punctatus and C.pulicariswing were the closest and C.newsteadi had a significant different with the other.3.Scanning electron microscopy observations of 9 species of 7 subgenus of Culicoides showed that the main morphological characteristics among the subgenus were significance different;the observation of the microstructure of three similar species of subgenus Culicoides indicated the aedeagus and paramere with a certain difference in the process,especially in the distal branches of paramere.The detailed observation and description of the adult male and female of C.Arakawa reservoir complemented the distribution of the sensilla of the antenna and described the maxil,spur and cerci,which have not been described by the predecessors.Conclusion: 1.Morphological description of 12 species of the subgenus Avaritia in our country has been revised and a new species has been found and described,which improved to enrich the species diversity of Culicoides and provided the basic data for the occurrence and control of the diseases from Culicoides and field pest control in China.2.The results of geometric morphology study of the genus Culicoides showed that the shape of the wing can be used as a basis to distinguish subgenus or species and geometric morphology can effectively distinguish the Culicoides rapidly and accurately,which provided a new method and new idea for the study of Ceratopogonidae insects in the future.3.Scanning electron microscopy studies shows the microstructures and the differences of morphological characteristics among subgenus or species of Culicoides intuitively and accurately.The detailed morphological observations and descriptions not only corrected the existing characterization data of Culicoides,but also find and describe the new features,which makes up for the deficiencies of the morphological description under the light microscope and provides a reference basis for the classification and further study of the Culicoides in China.