Transcriptome analyses of blood and sugar digestive processes in female Culicoides sonorensis midges (Diptera: Ceratopogonidae)

Female Culicoides sonorensis Wirth & Jones (Diptera:Ceratopogonidae) midges vector numerous diseases impacting livestock and humans. The molecular physiology of this midge has been under-studied, so our approach was to gain an understanding of basic processes of blood and sucrose digestion using transcriptomics. Two day-old female midges remained teneral or were fed a meal either of sterile sheep blood or 10% sucrose and maintained for 36 h for subsequent RNA extraction; two biological replicates were performed for each. Total RNA was extracted and used for TruSeq library construction. 100 bp paired-end (PE) HiSeq2000 (Illumina) sequencing was performed. In the absence of a reference genome, trimmed PE sequences were subjected to de novo clustering and assembled into a set of contigs, and singletons and unigenes were generated. The unigene assembly was functionally annotated using Blast2GO. Enzyme codes were obtained by mapping Gene Ontology terms, functional proteins were determined using InterProScan, and enzyme codes were highlighted on KEGG biochemical pathways. A comparative transcriptomics approach was used to elucidate genes and gene networks that were upregulated in response to sugar and blood exposure when compared to teneral midges, and statistical analyses were performed using Gossip. In addition, with the use of biological replicates and deep sequencing, we assessed the level of differential transcript expression quantitatively (upregulated, downregulated, or no change compared to teneral conditions) using the edgeR package. This type of RNA catalogue is the first of its kind for midges and serves an invaluable resource for exploiting the functional elements of the midge.