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In the past decade, there has been increasing interest in mosquito microbiome research, leading to large amounts of data on different mosquito species, with various underlying physiological characteristics, and from diverse geographical locations. However, guidelines and standardized methods for conducting mosquito microbiome research are lacking. To streamline methods in mosquito microbiome research and optimize data quality, reproducibility, and comparability, as well as facilitate data curation in a centralized location, we are establishing the Mosquito Microbiome Consortium, a collaborative initiative for the advancement of mosquito microbiome research. Our overall goal is to collectively work on unraveling the role of the mosquito microbiome in mosquito biology, while critically evaluating its potential for mosquito-borne disease control. This perspective serves to introduce the consortium and invite broader participation. It highlights the issues we view as most pressing to the community and proposes guidelines for conducting mosquito microbiome research. We focus on four broad areas in this piece: (1) sampling/experimental design for field, semi-field, or laboratory studies; (2) metadata collection; (3) sample processing, sequencing, and use of appropriate controls; and (4) data handling and analysis. We finally summarize current challenges and highlight future directions in mosquito microbiome research. We hope that this piece will spark discussions around this area of disease vector biology, as well as encourage careful considerations in the design and implementation of mosquito microbiome research. Video Abstract.

Many blood-feeding arthropods are known vectors of viruses that are a source of unprecedented global health concern. Mosquitoes are an integral part of these arthropod vectors. Advancements in next-generation sequencing and bioinformatics has expanded our knowledge on the richness of viruses harbored by arthropods. In the present study, we applied a metagenomic approach to determine the intercontinental virome diversity of Culex quinquefasciatus and Culex tritaeniorhynchus in Kwale, Kenya and provinces of Hubei and Yunnan in China. Our results showed that viromes from the three locations were strikingly diverse and comprised 30 virus families specific to vertebrates, invertebrates, plants, and protozoa as well as unclassified group of viruses. Though sampled at different times, both Kwale and Hubei mosquito viromes were dominated by vertebrate viruses, in contrast to the Yunnan mosquito virome, which was dominated by insect-specific viruses. However, each virome was unique in terms of virus proportions partly influenced by type of ingested meals (blood, nectar, plant sap, environment substrates). The dominant vertebrate virus family in the Kwale virome was Papillomaviridae (57%) while in Hubei it was Herpesviridae (30%) and the Yunnan virome was dominated by an unclassified viruses group (27%). Given that insect-specific viruses occur naturally in their hosts, they should be the basis for defining the viromes. Hence, the dominant insect-specific viruses in Kwale, Hubei, and Yunnan were Baculoviridae , Nimaviridae and Iflaviridae , respectively. Our study is preliminary but contributes to growing and much needed knowledge, as mosquito viromes could be manipulated to prevent and control pathogenic arboviruses., Competing Interests: The authors declare no conflict of interest.