The study was conducted in accordance with the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) statement.20. To achieve a comprehensive review of the published literature on microbiome diversity by NGS in field-collected ticks, a literature search was conducted on Chinese and English databases using a set of terms and Boolean operators, primarily via PubMed, Web of Science (WOS), China National Knowledge Infrastructure (CNKI) and WanFang databases until April 1, 2022, with no language or publication type restrictions. In the first step, general search terms were applied, including: “tick”, “Amblyomma”, “Archaeocroton”, “Bothriocroton”, “Dermacentor”, “Haemaphysalis”, “Hyalomme”, “Ixodes”, “Nosomma”, “Rhipicephalus”, “Rhipicenter”, “Robertsique”, “Anticola”, “Argas”, “carios”, “Nothoaspis”, “Ornithodoros“, “next generation sequencing”, “high throughput sequencing”, “deep sequencing”, “Roche 454”, “Illumina”, “Ion Torrent”, “SOLiD” in the search of English literature databases, and the keywords (“tick”, “virome”, “microbiome”, “metagenome”, “high throughput sequencing”, “deep sequencing”, “next generation sequencing”) were used in the database search of Chinese literature. Data on all types of microorganisms including viruses, bacteria and eukaryotes were included. Emerging pathogens were defined as those that were first isolated or discovered after 1980. Ticks can feed on a wide range of vertebrates. Therefore, to highlight the presence of tick-specific pathogens, we chose to include data collected in the field. live ticks, although they do not include data from detached ticks, as the latter could represent a complex microbiome derived from both the tick and the host animal. We excluded the following studies: (i) data obtained from experimentally fed ticks or detached ticks taken from animals; (ii) studies on the evaluation of methods for the isolation and propagation of laboratory strains; (iii) a review article and (iv) studies that only tested the specific microorganism in ticks (Fig. 1a).
A total of 2797 studies were retrieved for screening, including 2070 from the English database and 727 from the Chinese database. The title and abstract of retrieved studies were independently reviewed by three reviewers (MC L, JT Z and YZ) to identify studies potentially eligible for inclusion, which was narrowed down to 362 studies. For the third step, the full texts of the remaining studies were retrieved and independently assessed for eligibility by two reviewers (ZY H and BK F). Finally, a total of 7 Chinese studies and 69 English studies were eligible for data extraction (Fig. 1a). The first was published in 2011, and the number of publications has increased over the years, with a remarkable increase from the year 2017 (Fig. 1b). Of all the selected studies, 69 (90.8%) used the Illumina sequencing platform and 5.3% used the Ion Torrent sequencing platform (Fig. 1c). Data came from 46 tick species of 7 genera collected from 24 countries on 6 continents, and the geographical distribution of tick genera was shown in Fig. 2a. Viral metagenomic profiling, eukaryotic and bacterial microbiome profiling corresponding to various tick genera were displayed in all countries (Fig. 2b,c).
The full text of all selected articles was reviewed and data extracted into a standardized dataset in Microsoft Excel 2019 which primarily includes: (i) identification of tested ticks at family, genus and species, (ii) methods for tick species identification, (iii) life cycle stages of ticks tested, (iv) geographic location of ticks at country and province level, (v ) taxonomic annotations of microorganisms at family, genus, species level, (vi) platforms used for NGS. A double check by two people (MC L and JT Z) was carried out to correct errors and remove duplicates. All conflicts of opinion and uncertainties were discussed and resolved by consensus with a third reviewer (JJ C). The primary variable of interest was the viral/bacterial/eukaryotic component of the microbiome, determined for specific tick species at a specific site over time. All data was entered into the resultant by trained co-authors.
Information on the location of the tick collection site was extracted at the province level from the selected literatures. If no longitude or latitude data was reported, or location information was provided only at a large scale, such as scenic area, mountainous region, ArcGIS 10.7 software was used to extract the geographical coordinates of the central points of the corresponding administrative areas from the digital map, which was obtained from GADM (World Administrative Areas Database) and standard map service system. If the collection site could not be determined by any of these means, the authors were contacted for further information. We used R Studio Version 4.1.2 software and ArcGIS 10.7 software to statistically analyze and visualize the geographical data obtained.