Anopheles gambiae is the predominant vector of malaria in Africa, which is caused by an apicomplexan belonging to the genus Plasmodium. In 2018, 214 million cases of malaria were reported world-wide, of which 88% of those cases were in Africa, predominately sub-Saharan Africa where malaria was the fourth largest cause of infant mortality [1]. Anopheles gambiae also vectors O’nyong-nyong arbovirus and other viruses that pose emerging threats to human health.
The vectorial capacity of mosquitoes to transmit infectious pathogens depends on many pathogen-host interactions such as pathogen entry and development in the host, each of which is countered by the innate immune response of the host [2-5]. Environmental factors provide challenges for studying mosquito-infectious microbe interactions at the whole animal level. The handling of mosquitoes and infectious pathogens requires a range of skills and facilities including the ability to raise large numbers of mosquitoes, facilities for handling both the insect vector and the pathogen and expertise in vector and pathogen biology. Mosquito cell lines have been used as models to investigate mosquito pathogen interactions [6-8].
Mosquito cell lines have been used to investigate mosquito-virus interactions [6, 7, 9], as models for deducing the host range of arboviruses [10, 11], for isolating and characterizing mosquito-specific flaviviruses [12, 13], for the potential in vitro development of Plasmodium ookinetes [14, 15], and for screening of insecticides [16]. High throughput RNAi screens have made cell lines useful models for reverse genetic studies [17-19]. Furthermore, experiments with Drosophila melanogaster cell lines have been shown to correlate well with experiments using whole flies for genetic and developmental studies [20-23]. A. gambiae cells lines have been used as models to study mosquito immune responses [24-27]. Further, these cells have been shown to express immune factors upon microbial challenge and perform complex immune tasks such as phagocytosis of beads and bacteria [24, 28]. The discovery of A. gambiae densonucleovirus (AgDNV) is important as this virus replicates in adult mosquitoes and in cultured An. gambiae MOS55 cells with minimal physiological effects on the host. It is likely that AgDNV can be engineered as a transducing virus.
Cell lines have been derived from A. gambiae as it is the primary vector of malaria in sub-Saharan Africa. The Ag55 [29], Sua1B and Sua4a-3B [25] cell lines are derived from neonate first instar larvae. Further, a Sua5B cell line was derived by splitting Sua1 cell line [30]. The Sua1B, Sua4a-3B, and Sua5B cell lines are considered to have hemocyte-like properties [25, 30].
Anopheles Ag55 cells were tested as a model to study Plasmodium ookinete interaction [8], and Lysinibacillus sphaericus Bin toxin mode of action [31]. Ag55 cells are amenable to RNA inhibition-based knock-down of targeted mosquito genes [32, 33]. We used transcriptomic analyses, confocal microscopy and flow cytometry towards the goal of enhancing the utility of Ag55 cells for pathogen interaction, genetic and immune response studies. KEGG analysis of genes with transcripts level ≥200 FPKM identified phagosome term enriched. Confocal microscopy images and flow cytometry data showed internalization of FITC-E.coli bioparticles. The presence of phagocytic receptors, hemocyte markers and anti-microbial peptide transcripts suggest Ag55 cells have hemocyte like properties.