Species-level microbiota of ticks and fleas from in the Qinghai-Tibet Plateau.

Introduction: Ticks and fleas, as blood-sucking arthropods, carry and transmit various zoonotic diseases. In the natural plague foci of China, monitoring of has been continuously conducted in and other host animals, whereas other pathogens carried by vectors are rarely concerned in the Qinghai-Tibet Plateau.

Methods: In this study, we investigated the microbiota of ticks and fleas sampling from in the Plateau, China by metataxonomics combined with metagenomic methods.

Results: By metataxonomic approach based on full-length 16S rDNA amplicon sequencing and operational phylogenetic unit (OPU) analyses, we described the microbiota community of ticks and fleas at the species level, annotated 1,250 OPUs in ticks, including 556 known species and 492 potentially new species, accounting for 48.50% and 41.71% of the total reads in ticks, respectively. A total of 689 OPUs were detected in fleas, consisting of 277 known species (40.62% of the total reads in fleas) and 294 potentially new species (56.88%). At the dominant species categories, we detected the (OPU 421) and potentially pathogenic new species of , and . Using shotgun sequencing, we obtained 10 metagenomic assembled genomes (MAGs) from vector samples, including a known species ( DFT2), and six new species affliated to four known genera, i.e., , and . By the phylogenetic analyses based on full-length 16S rRNA genes and core genes, we identified that ticks harbored pathogenic . Moreover, these potentially pathogenic novel species were more closely related to subsp. , and , respectively. The OPU 422 Ehrlichia sp1 was most related to and . The OPU 230 sp1 and spp. (DTF8 and DTF9) was clustered with . The OPU 427 sp1 was clustered with .

Discussion: The findings of the study have advanced our understanding of the potential pathogen groups of vectors in marmot () in the Qinghai-Tibet Plateau.