Location: Arthropod-borne Animal Diseases ResearchTitle: Gene discovery and differential expression analysis of humoral immune response elements in female Culicoides sonorensis (Diptera: Ceratopogonidae) Author
Submitted to: Medical and Veterinary Entomology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/26/2014
Publication Date: N/A
Citation: N/A Interpretive Summary: Female Culicoides sonorensis midges are important in the transmission of pathogenic viruses such as bluetongue virus and epizootic hemorrhagic disease virus, which have morbidity, mortality, fitness and economic impacts on US livestock and wildlife. We recently completed and published the transcriptome, which is a catalog of expressed genes, for this vector and also looked at how the expression of genes changes with diet (unfed vs. blood fed vs. sugar fed). Subsequently, those transcriptomes were analyzed in order to identify components of the insect innate immune system. Because the insect immune response is a key player in important processes including vector competence, or the ability for the midge to transmit pathogens, we also examined immune gene expression profiles in response to diet across time, on both a transcriptome-wide, whole-midge level and more specifically by examining antimicrobial peptide (AMP) gene expression in the digestive tract, which serves as a primary barrier to infection. We identified functional components of the three main insect humoral immune pathways including: immune deficiency (Imd), Toll and JAK/STAT. We found complete or nearly-complete sequences for all elements, including humoral factors, transmembrane receptors, signaling components, gene transcription factors/regulators and antimicrobial effectors (e.g. AMPs, thioester-containing proteins, and prophenoloxidase). Midges that were fed on either sugar or blood had altered expression of receptors, regulators, effectors (AMPs, PPO and TEPs). Further, blood had a greater effect than sucrose on the expression profiles of most innate immune components. All five AMPs were significantly upregulated in the alimentary canal after blood feeding, possibly in response to proliferating populations of gut bacteria. Functional insight into C. sonorensis humoral/innate immune components updates our knowledge of the molecular functioning of this important vector. Future studies involving the manipulation of immune gene expression, such as gene silencing, will help in uncovering genetic components of vector competence. The results of such studies would serve as a platform for designing novel transmission-blocking strategies.
Technical Abstract: Background: Female Culicoides sonorensis midges (Diptera: Ceratopogonidae) are vectors of pathogens that impact livestock and wildlife in the United States. Little is known about their molecular functioning, including components of their immune system. Because the insect immune response is involved with important processes like gut microbial homeostasis and vector competence, our aims were to identify components of the midge innate immune system and examine their expression profiles in response to diet across time. Methods: In our previous work, we de novo sequenced and analyzed the transcriptional landscape of female midges under several feeding states including teneral (unfed) and early and late time points after blood and sucrose. Here, those transcriptomes were further analyzed to identify insect innate immune orthologs, particularly humoral immune response elements. Additionally, we examined immune gene expression profiles in response to diet across time, on both a transcriptome-wide, whole-midge level and more specifically via qRTPCR analysis antimicrobial peptide (AMP) expression in the alimentary canal. Results: We identified functional units comprising complete, or nearly complete, immune deficiency (Imd), Toll and JAK/STAT pathways, including humoral factors, transmembrane receptors, signaling components, transcription factors/regulators and effectors such as AMPs. Feeding altered the expression of receptors, regulators, AMPs, prophenoloxidase and thioester-containing proteins, where blood had a greater effect than sucrose on the expression profiles of most innate immune components. qRTPCR of AMP genes showed that all five were significantly upregulated in the alimentary canal after blood feeding, possibly in response to proliferating populations of gut bacteria.