Molecular evolution of opsin visual receptors in the mosquitoes Ae. aegypti, An. gambiae and Cx. quinquefasciatus

Gloria Isabel Giraldo-Calderon, Purdue University

Abstract

Female mosquitoes (Diptera: Culicidae) are hematophagous insects and many are obligatory vectors of some of the most significant vector-borne diseases impacting human health worldwide. Three of the most important species are the yellow fever mosquito, Aedes aegypti, the malaria mosquito, Anopheles gambiae, and the southern house mosquito, Culex quinquefasciatus. Understanding the role of mosquito vision in mating, host detection and oviposition may help to improve or develop new control strategies to reduce the incidence of vector-borne diseases. Opsins are receptors that interact with photons to initiate visual processes. Typically, insects have three classes of opsins that are stimulated by ultraviolet, short, and long (LW) wavelengths. Here we report the first molecular analysis of light receptors (opsins) from Ae. aegypti, An. gambiae , and Cx. quinquefasciatus. We used expression data to improve the 10 Ae. aegypti and 11 An. gambiae published opsin gene models, and we report the first manual annotation of 13 opsin genes from Cx. quinquefasciatus . Opsin transcripts were confirmed using Reverse Transcriptase-PCR and published expression data. Phylogenetic analyses were used to predict the wavelength sensitivity of the mosquito opsins and to determine evolutionary relationships of mosquito opsins relative to other animal opsins. Results predicted six putative LW opsins in Ae. aegypti, six in An. gambiae and eight in Cx. quinquefasciatus, suggesting an expansion of these genes in the Culicidae relative to other insect taxa. Time of divergence estimates suggest the mosquito LW opsins originated from 18 or 19 duplication events between 189.87 to 0.85 million years ago (MYA), and that all except the first and possibly the second likely occured after the presumed divergence of the Anophelinae and Culicinae lineages. The transmembrane (TM) domains of these opsins share between 60 to 100% amino acid identity and this raises intriguing questions regarding the retention of these genes in the three mosquito genomes. Molecular evolutionary studies were conducted to evaluate mechanisms responsible for retention of the LW opsins in mosquito genomes. Fifteen LW opsins contained residues predicted to be under positive selection. Eight amino acids were identified under positive selection in the N and C termini, and one in a third transmembrane domain suggestive of opsin positive selection and spectral tuning. Eleven LW opsins were predicted to share putative transcription factor binding sites. Conserved nucleotide sequence was identified in 5' non coding regions, between 7 of the 189 possible mosquito opsin gene pairs and is indicative of coordinated gene regulation in 11 genes. Ten LW opsins had evidence of both positive selection and coordinated regulation and four LW opsin genes had no evidence of either positive selection or coordinated regulation. We discuss potential mechanisms, including positive selection and differential gene regulation for the conservation of LW opsins in these mosquitoes. To study the behavior of mosquito larvae and to develop a tool to evaluate the role of genes in the phototransduction pathway and their impact on mosquito vision, a behavioral assay using Ae. aegypti late third (L3) and early fourth (L4) larvae was developed based on that of Kassap (1977). Experiments using either individual larvae or groups of 50 larvae showed that mosquitoes placed in a 15 cm diameter glass Petri dish, the sides and bottom of which were covered with either black or white foam, were randomly distributed on an all white or all black background but show a significant preference for a black over white background in a choice test. Results suggest that the test can be used with individual larva or groups of 50 larvae at 1 minute, where larva are randomly distributed in the negative control and prefer the black side of the treatments (>80-90%).

Degree

Ph.D.

Advisors

Hill, Purdue University.

Subject Area

Entomology|Evolution and Development|Bioinformatics

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