Mine Altinli

The inadequacy of standard mosquito control strategies calls for ecologically safe novel approaches, for example the use of biological agents such as the endosymbiotic α-proteobacteria Wolbachia or insect-specific viruses (ISVs). Understanding the ecological interactions between these “biocontrol endosymbionts” is thus a fundamental step. Wolbachia are transmitted vertically from mother to offspring and modify their hosts’ phenotypes, including reproduction (e.g., cytoplasmic incompatibility) and survival (e.g., viral interference). In nature, Culex pipiens (sensu lato) mosquitoes are always found infected with genetically diverse Wolbachia called wPip that belong to five phylogenetic groups. In recent years, ISVs have also been discovered in these mosquito species, although their interactions with Wolbachia in nature are unknown. Here, we studied the interactions between a widely prevalent ISV, the Culex pipiens densovirus (CpDV, Densovirinae), and Wolbachia in northern Tunisian C. pipiens populations. We showed an influence of different Wolbachia groups on CpDV prevalence and a general positive correlation between Wolbachia and CpDV loads. By investigating the putative relationship between CpDV diversification and wPip groups in the different sites, we detected a signal linked to wPip groups in CpDV phylogeny in sites where all larvae were infected by the same wPip group. However, no such signal was detected where the wPip groups coexisted, suggesting CpDV horizontal transfer between hosts. Overall, our results provide good evidence for an ecological influence of Wolbachia on an ISV, CpDV, in natural populations and highlight the importance of integrating Wolbachia in our understanding of ISV ecology in nature.

Mosquitoes not only transmit human and veterinary pathogens called arboviruses (arthropod-borne viruses) but also harbor mosquito-associated insect-specific viruses (mosquito viruses) that cannot infect vertebrates. In the past, studies investigating mosquito viruses mainly focused on highly pathogenic interactions that were easier to detect than those without visible symptoms. However, the recent advances in viral metagenomics have highlighted the abundance and diversity of viruses which do not generate mass mortality in host populations. Over the last decade, this has facilitated the rapid growth of virus discovery in mosquitoes. The circumstances around the discovery of mosquito viruses greatly affected how they have been studied so far. While earlier research mainly focused on the pathogenesis caused by DNA and some doublestranded RNA viruses during larval stages, more recently discovered single-stranded RNA mosquito viruses were heavily studied for their putative interference with arboviruses in female adults. Thus, many aspects of mosquito virus interactions with their hosts and host-microbiota are still unknown. In this context, considering mosquito viruses as endosymbionts can help to identify novel research areas, in particular in relation to their long-term interactions with their hosts (e.g. relationships during all life stages, the stability of the associations at evolutionary scales, transmission routes and virulence evolution) and the possible context-dependent range of interactions (i.e. beneficial to antagonistic). Here, we review the symbiotic interactions of mosquito viruses considering different aspects of their ecology, such as transmission, host specificity, host immune system and interactions with other symbionts within the host cellular arena. Finally, we highlight related research gaps in mosquito virus research.

Arboviruses transmitted by mosquitoes are responsible for the death of millions of people each year. In addition to arboviruses, many insect-specific viruses (ISVs) have been discovered in mosquitoes in the last decade. ISVs, in contrast to arboviruses transmitted by mosquitoes to vertebrates, cannot replicate in vertebrate cells even when they are evolutionarily closely related to arboviruses. The alphavirus genus includes many arboviruses, although only a few ISVs have been discovered from this genus so far. Here, we investigate the interactions of a recently isolated insect-specific alphavirus, Agua Salud alphavirus (ASALV), with its mosquito host. RNA interference (RNAi) is one of the essential antiviral responses against arboviruses, although there is little knowledge on the interactions of RNAi with ISVs. Through the knockdown of transcripts of the different key RNAi pathway (small interfering RNA [siRNA], microRNA [miRNA], and P-element-induced wimpy testis [PIWI]-interacting RNA [piRNA]) proteins, we show the antiviral role of Ago2 (siRNA), Ago1 (miRNA), and Piwi4 proteins against ASALV in Aedes aegypti-derived cells. ASALV replication was increased in Dicer2 and Ago2 knockout cells, confirming the antiviral role of the siRNA pathway. In infected cells, mainly ASALV-specific siRNAs are produced, while piRNA-like small RNAs, with the characteristic nucleotide bias resulting from ping-pong amplification, are produced only in Dicer2 knockout cells. Taken together, ASALV interactions with the mosquito RNAi response differ from those of arthropod-borne alphaviruses in some aspects, although they also share some commonalities. Further research is needed to understand whether the identified differences can be generalized to other insect-specific alphaviruses.

Mosquitoes are competent vectors for many important arthropod-borne viruses (arboviruses). In addition to arboviruses, insect-specific viruses (ISV) have also been discovered in mosquitoes. ISVs are viruses that replicate in insect hosts but are unable to infect and replicate in vertebrates. They have been shown to interfere with arbovirus replication in some cases. Despite the increase in studies on ISV–arbovirus interactions, ISV interactions with their hosts and how they are maintained in nature are still not well understood. In the present study, we investigated the infection and dissemination of the Agua Salud alphavirus (ASALV) in the important mosquito vector Aedes aegypti through different infection routes (per oral infection, intrathoracic injection) and its transmission. We show here that ASALV infects the female Ae. aegypti and replicates when mosquitoes are infected intrathoracically or orally. ASALV disseminated to different tissues, including the midgut, salivary glands and ovaries. However, we observed a higher virus load in the brain than in the salivary glands and carcasses, suggesting a tropism towards brain tissues. Our results show that ASALV is transmitted horizontally during adult and larval stages, although we did not observe vertical transmission. Understanding ISV infection and dissemination dynamics in Ae. aegypti and their transmission routes could help the use of ISVs as an arbovirus control strategy in the future.