Nhumirim virus (NHUV) is an insect-specific virus that phylogenetically affiliates with dual-host mosquito-borne flaviviruses. Previous in vitro co-infection experiments demonstrated prior or concurrent infection of Aedes albopictus C6/36 mosquito cells with NHUV resulted in a 10,000-fold reduction in viral production of West Nile virus (WNV). This interference between WNV and NHUV was observed herein in an additional Ae. albopictus mosquito cell line, C7-10. A WNV 2K peptide (V9M) mutant capable of superinfection with a pre-established WNV infection demonstrated a comparable level of interference from NHUV as the parental WNV strain in C6/36 and C7-10 cells. Culex quinquefasciatus and Culex pipiens mosquitoes intrathoracically inoculated with NHUVandWNV, or solely withWNVas a control, were allowed to extrinsically incubate the viruses up to nine and 14 days, respectively, and transmissibility and replication of WNV was determined. The proportion of Cx. quinquefasciatus mosquitoes capable of transmitting WNV was significantly lower for the WNV/NHUV group than the WNV control at seven and nine days post inoculation (dpi), while no differences were observed in the Cx. pipiens inoculation group. By dpi nine, a 40% reduction in transmissibility in mosquitoes from the dual inoculation group was observed compared to the WNV-only control. These data indicate the potential that infection of some Culex spp. vectors with NHUV could serve as a barrier for efficient transmissibility of flaviviruses associated with human disease.

The importance of Zika virus (ZIKV) has increased noticeably since the outbreak in the Americas in 2015, when the illness was associated with congenital disorders. Although there is evidence of sexual transmission of the virus, the mosquito Aedes aegypti is believed to be the main vector for transmission to humans. This species of mosquito has not only been found naturally infected with ZIKV, but also has been the subject of study in many vector competence assays that employ different strains of ZIKV around the world. In Argentina, the first case was reported in February 2016 and a total of 278 autochthonous cases have since been confirmed, however, ZIKV virus has not been isolated from any mosquito species yet in Argentina. In order to elucidate if Argentinian Ae. aegypti populations could be a possible vector of ZIKV, we conducted vector competence studies that involved a local strain of ZIKV from Chaco province, and a Venezuelan strain obtained from an imported case. For this purpose, Ae. aegypti adults from the temperate area of Argentina (Buenos Aires province) were fed with infected blood. Body, legs and saliva were harvested and tested by plaque titration on plates of Vero cells for ZIKV at 7, 11 and 14 days post infection (DPI) in order to calculate infection, transmission, and dissemination rates, respectively. Both strains were able to infect mosquitoes at all DPIs, whereas dissemination and transmission were observed at all DPIs for the Argentinian strain but only at 14 DPI for the Venezuelan strain. This study proves the ability of Ae. aegypti mosquitoes from Argentina to become infected with two different strains of ZIKV, both belonging to the Asian lineage, and that the virus can disseminate to the legs and salivary glands. Zika virus is a flavivirus transmitted by mosquitoes, isolated for the first time in the Ziika Forest in Uganda in 1947 from a rhesus macaque monkey. The disease is usually asymptomatic, but sometimes it causes a mild illness that comes with fever, rash, joint pain, and conjunctivitis. The World Health Organization focused the attention on this virus after the outbreak in the Americas, when the virus was linked to microcephaly and serious neurological diseases, including Guillain-Barré syndrome. Aedes aegypti was incriminated as the main vector of the virus as it was found both naturally and experimentally infected. This mosquito species was declared eradicated in Argentina by 1970 but re-emerged in 1989. Recent studies found a peculiarity in the genetics of Argentinian Ae. aegypti populations that consists in a combination between both subspecies: Ae. aegypti formosus and Ae. aegypti aegypti. Our study tries to elucidate if Ae. aegypti from Argentina are able to transmit the virus in order to add these mosquitoes to the list of possible vectors of ZIKV and, in future prospect, orient to fight the virus by controlling the vector.