Total RNA sequencing of Phlebotomus chinensis , a neglected vector in China, simultaneously revealed viral, bacterial, and eukaryotic microbes that are potentially pathogenic to humans.

sandfly is a neglected insect vector in China which is well-known for carrying . Recent studies have expanded its pathogen repertoire with two novel arthropod-borne phleboviruses capable of infecting human and animals. Despite these discoveries, our knowledge on the general pathogen diversity and overall microbiome composition of this vector species are still very limited. Here we carried out a meta-transcriptomics analysis which simultaneously revealed the actively replicating/transcribing RNA viruses, DNA viruses, bacteria and eukaryotic microbes, namely, "total microbiome", of several sandfly populations in China. Strikingly, "microbiome" made up 1.8% of total non-ribosomal RNA and were comprised of more than 87 species, among which 70 were novel, including divergent members of the genera and of the family Trypanosomatidae. Importantly, among these microbes we were able to reveal four distinguished types of human and/or mammalian pathogens, including two phleboviruses (hedi and wuxiang viruses), one novel Spotted fever group rickettsia, as well as a member of complex, among which hedi virus and each had > 50% pool prevalence rate and relatively high abundance levels. Our study also showed the ubiquitous presence of an endosymbiont, namely , although no anti-viral or anti-pathogen effect were detected based on our data. In summary, our results uncovered the much un-explored diversity of microbes harbored by sandflies in China and demonstrated that high pathogen diversity and abundance is currently present in multiple populations, implying disease potential for exposed local human population or domestic animals.