Isolation of a novel rhabdovirus and detection of multiple novel viral sequences in Culex species mosquitoes in the United States

To increase our understanding of the diversity of the mosquito virome, 6956 mosquitoes of five species ( Culex erraticus , Culex pipiens , Culex restuans , Culex tarsalis , and Culex territans ) collected in Iowa in the United States in 2017 and 2020 were assayed for novel viruses by performing polyethylene glycol precipitation, virus isolation in cell culture, and unbiased high-throughput sequencing. A novel virus, provisionally named “Walnut Creek virus”, was isolated from Cx. tarsalis , and its genomic sequence and organization are characteristic of viruses in the genus Hapavirus (family Rhabdoviridae ). Replication of Walnut Creek virus occurred in avian, mammalian, and mosquito, but not tick, cell lines. A novel virus was also isolated from Cx. restuans , and partial genome sequencing revealed that it is distantly related to an unclassified virus of the genus Phytoreovirus (family Sedoreoviridae ). Two recognized viruses were also isolated: Culex Y virus (family Birnaviridae ) and Houston virus (family Mesoniviridae ). We also identified sequences of eight novel viruses from six families ( Amalgaviridae , Birnaviridae , Partitiviridae, Sedoreoviridae , Tombusviridae , and Totiviridae ), two viruses that do not belong to any established families, and many previously recognized viruses. In summary, we provide evidence of multiple novel and recognized viruses in Culex spp. mosquitoes in the United States.