1476 © The Author(s) 2021. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com. Short Communication Identifcation and Characterization of Aminopeptidase N 1 Gene of the Indian Malaria Vector Anopheles culicifacies (Diptera: Culicidae) Renu Jakhar 1, and Surendra Kumar Gakhar Centre for Medical Biotechnology, Maharshi Dayanand University, Rohtak-124001, Haryana, India and 1 Corresponding author, e-mail: renujakhar22@gmail.com Subject Editor: Michel Slotman Received 30 June 2020; Editorial decision 4 January 2021 Abstract Aminopeptidase N1 (APN) is one of the important enzymes involved in blood digestion and is up-regulated along with several other enzymes in response to bloodmeal ingestion. APN is a zinc metalloprotease that cleaves one amino acid residue at a time from the amino terminus of the protein. The APN1 gene of the Indian malaria vector Anopheles culicifacies Giles was cloned and characterized. The An. culicifacies APN1 (AcAPN1) gene has an Open Reading Frame of 3084 basepairs which encodes a putative protein of 1027 amino acids. The coding region of the gene shares 81% and 78% similarity to the APN1 genes found in An. stephensi (Diptera: Culicidae) and An. gambiae (Diptera: Culicidae), respectively. The organization of the APN1 gene was studied in available mosquito genomes and a three-dimensional structure of AcAPN1 mod- eled using homology structure modeling. The enzymatic active site was predicted to consist of HEYAH and GAMEN amino acid residues, and a comparison of the protein sequences among different genera revealed the conservation of zinc-binding residues. The expression pattern of AcAPN1 showed that the gene was expressed rapidly in response to the ingestion of the bloodmeal and therefore this gene may be used to exploit its promoter region as an antiparasite candidate molecule. Key words: mosquito, Anopheles culicifacies, Aminopeptidase N1 Malaria continues to impose a huge health burden mostly on popu- lations residing in tropical and semitropical regions of the world. Genetic modifcation of malaria vectors is one promising interven- tion. Accordingly, the genes that permit malarial infection in mos- quito vectors can be identifed and then replaced or altered in terms of their function and thereby disrupt Plasmodium development in the vector. In this way, vector Anopheles populations will become refractory to the parasite, eventually leading to malaria transmis- sion being halted. The path for the creation of such mosquitoes faces many challenges. The expression of a transgene in the mosquito requires the use of an appropriate promoter. Interruption of the parasite developmental process in the mosquito using antiparasite transgenic strategies generally focuses on three different sites within the mosquito: midgut, hemolymph, and salivary glands. The mos- quito midgut probably provides the more fruitful results, because it is the site where parasite syngamy occurs, followed by infection and then asexual reproduction. The Aminopeptidase N (APN) is known to facilitate the malaria parasite invasion into midgut and is perhaps the most important vector/mosquito-based Transmission Blocking Vaccine candidate (Mathias et al. 2012, Wangdi et al. 2016). Antibodies against a midgut aminopeptidase (AgAPN1) reduced P. falciparum oocyst infection intensity in An. gambiae Giles and An. stephensi Liston by 73% and 67%, respectively (Dinglasan et al. 2007, Armistead et al. 2014). In India, An. culicifacies A (Diptera: Culicidae) is an important vector of malaria. It is responsible for 60–70% of malaria cases every year (Goswami et al. 2006); how- ever, the blood inducible APN1 gene has yet to be characterized. This study describes the cloning and expression pattern of the An. culicifacies A APN1 gene, and the in silico analysis of the related protein to APN1 gene. Materials and Methods Mosquitoes Rearing Anopheles culicifacies A (Dhera strain) was used in this study. This strain was obtained from the National Institute of Malaria Research (NIMR), New Delhi. The mosquitoes were reared in an insectary maintained at 28 ± 2°C and 70%–80% relative humidity, with sim- ulated dawn and dusk and 14L: 10D photoperiod (Kumar et al. 2014). Journal of Medical Entomology, 58(3), 2021, 1476–1481 doi: 10.1093/jme/tjab011 Advance Access Publication Date: 4 March 2021 Short Communication Downloaded from https://academic.oup.com/jme/article/58/3/1476/6158877 by guest on 07 April 2022