Discovery of flavivirus-derived endogenous viral elements in two Anopheles

17 The Flavivirus genus encompasses several arboviruses of public health significance 18 such as dengue, yellow fever, and Zika viruses. It also includes insect-specific flaviviruses 19 (ISFs) that are only capable of infecting insect hosts. The vast majority of mosquito-infecting 20 flaviviruses have been associated with mosquito species of the Aedes and Culex genera in the 21 Culicinae subfamily, which also includes most arbovirus vectors. Mosquitoes of the 22 Anophelinae subfamily are not considered significant arbovirus vectors, however flaviviruses 23 have occasionally been detected in field-caught Anopheles specimens. Whether such 24 observations reflect occasional spillover or laboratory contamination or whether Anopheles 25 mosquitoes are natural hosts of flaviviruses is unknown. Here, we provide in silico and in vivo 26 evidence of transcriptionally active, flavivirus-derived endogenous viral elements (EVEs) in 27 the genome of Anopheles minimus and Anopheles sinensis. Such non-retroviral 28 endogenization of RNA viruses is consistent with a shared evolutionary history between 29 flaviviruses and Anopheles mosquitoes. Phylogenetic analyses of the two newly described 30 EVEs support the existence of a distinct clade of Anopheles-associated ISFs. 31 32 . CC-BY-NC-ND 4.0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/064907 doi: bioRxiv preprint first posted online Sep. 19, 2016;