Many animals are dependent on microbial partners that provide essential nutrients lacking from their diet. Ticks, whose diet consists exclusively on vertebrate blood, rely on maternally inherited bacterial symbionts to supply B vitamins. While previously studied tick species consistently harbor a single lineage of those nutritional symbionts, we evidence here that the invasive tick Hyalomma marginatum harbors a unique dual-partner nutritional system between an ancestral symbiont, Francisella, and a more recently acquired symbiont, Midichloria. Using metagenomics, we show that Francisella exhibits extensive genome erosion that endangers the nutritional symbiotic interactions. Its genome includes folate and riboflavin biosynthesis pathways but deprived functional biotin biosynthesis on account of massive pseudogenization. Co-symbiosis compensates this deficiency since the Midichloria genome encompasses an intact biotin operon, which was primarily acquired via lateral gene transfer from unrelated intracellular bacteria commonly infecting arthropods. Thus, in H. marginatum, a mosaic of co-evolved symbionts incorporating gene combinations of distant phylogenetic origins emerged to prevent the collapse of an ancestral nutritional symbiosis. Such dual endosymbiosis was never reported in other blood feeders but was recently documented in agricultural pests feeding on plant sap, suggesting that it may be a key mechanism for advanced adaptation of arthropods to specialized diets.

Center of Rickettsiosis and Arthropod Borne Diseases Logroño Spain

Centre of Research in Ecology and Evolution of Diseases Montpellier France Montpellier France

Department of Biology and Biotechnology L Spallanzani University of Pavia Pavia Italy

Department of Biomedical and Clinical Sciences L Sacco and Pediatric Clinical Research Center University of Milan Milan Italy

Department of Chemical Biological Pharmaceutical and Environmental Sciences University of Messina Messina Italy

Department of Veterinary Medicine University of Milan Lodi Italy

Faculty of Science University of South Bohemia České Budějovice Czech Republic

Institute of Infection Veterinary and Ecological Sciences University of Liverpool Liverpool United Kingdom

Koret School of Veterinary Medicine The Robert H Smith Faculty of Agriculture Food and Environment The Hebrew University of Jerusalem Rehovot Israel

MIVEGEC Montpellier France

Zobrazit více v PubMed

Akhter S, Aziz RK, Edwards RA. PhiSpy: a novel algorithm for finding prophages in bacterial genomes that combines similarity- and composition-based strategies. Nucleic Acids Research. 2012;40:1–13. doi: 10.1093/nar/gks406. PubMed DOI PMC

Alonge M, Soyk S, Ramakrishnan S, Wang X, Goodwin S, Sedlazeck FJ, Lippman ZB, Schatz MC. RaGOO: fast and accurate reference-guided scaffolding of draft genomes. Genome Biology. 2019;20:1–17. doi: 10.1186/s13059-019-1829-6. PubMed DOI PMC

Andrews S. FastQC: A Quality Control Tool for High Throughput Sequence Data. ScienceOpen, Inc; 2010.

Azagi T, Klement E, Perlman G, Lustig Y, Mumcuoglu KY, Apanaskevich DA, Gottlieb Y. Francisella-Like Endosymbionts and Rickettsia Species in Local and Imported Hyalomma Ticks. Applied and Environmental Microbiology. 2017;83:1–14. doi: 10.1128/AEM.01302-17. PubMed DOI PMC

Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin AV, Vyahhi N, Tesler G, Alekseyev MA, Pevzner PA. SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. Journal of Computational Biology. 2012;19:455–477. doi: 10.1089/cmb.2012.0021. PubMed DOI PMC

Bates D, Mächler M, Bolker BM, Walker SC. Fitting Linear Mixed-Effects Models Using lme4. Journal of Statistical Software. 2015;67:i01. doi: 10.18637/jss.v067.i01. DOI

Ben-Yosef M, Rot A, Mahagna M, Kapri E, Behar A, Gottlieb Y. Coxiella-Like Endosymbiont of Rhipicephalus sanguineus Is Required for Physiological Processes During Ontogeny. Frontiers in Microbiology. 2020;11:1–16. doi: 10.3389/fmicb.2020.00493. PubMed DOI PMC

Bennett GM, Moran NA. Heritable symbiosis: The advantages and perils of an evolutionary rabbit hole. PNAS. 2015;112:10169–10176. doi: 10.1073/pnas.1421388112. PubMed DOI PMC

Binetruy F, Buysse M, Lejarre Q, Barosi R, Villa M, Rahola N, Paupy C, Ayala D, Duron O. Microbial community structure reveals instability of nutritional symbiosis during the evolutionary radiation of Amblyomma ticks. Molecular Ecology. 2020;29:1016–1029. doi: 10.1111/mec.15373. PubMed DOI

Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014;30:2114–2120. doi: 10.1093/bioinformatics/btu170. PubMed DOI PMC

Bonnet SI, Pollet T. Update on the intricate tango between tick microbiomes and tick-borne pathogens. Parasite Immunology. 2021;43:1–12. doi: 10.1111/pim.12813. PubMed DOI

Buysse M, Plantard O, McCoy KD, Duron O, Menard C. Tissue localization of Coxiella-like endosymbionts in three European tick species through fluorescence in situ hybridization. Ticks and Tick-Borne Diseases. 2019;10:798–804. doi: 10.1016/j.ttbdis.2019.03.014. PubMed DOI

Buysse M, Duron O. Evidence that microbes identified as tick-borne pathogens are nutritional endosymbionts. Cell. 2021;184:2259–2260. doi: 10.1016/j.cell.2021.03.053. PubMed DOI

Buysse M, Nardi T, Floriano MF. Hmar-2021. a459aeeGitHub. 2021 https://github.com/mariebuysse/Hmar-2021

Campbell MA, Van Leuven JT, Meister RC, Carey KM, Simon C, McCutcheon JP. Genome expansion via lineage splitting and genome reduction in the cicada endosymbiont Hodgkinia. PNAS. 2015;112:10192–10199. doi: 10.1073/pnas.1421386112. PubMed DOI PMC

Capella-Gutiérrez S, Silla-Martínez JM, Gabaldón T. trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics. 2009;25:1972–1973. doi: 10.1093/bioinformatics/btp348. PubMed DOI PMC

Castresana J. Selection of conserved blocks from multiple alignments for their use in phylogenetic analysis. Molecular Biology and Evolution. 2000;17:540–552. doi: 10.1093/oxfordjournals.molbev.a026334. PubMed DOI

Chen H, Boutros PC. VennDiagram: a package for the generation of highly-customizable Venn and Euler diagrams in R. BMC Bioinformatics. 2011;12:35. doi: 10.1186/1471-2105-12-35. PubMed DOI PMC

Darling ACE, Mau B, Blattner FR, Perna NT. Mauve: multiple alignment of conserved genomic sequence with rearrangements. Genome Research. 2004;14:1394–1403. doi: 10.1101/gr.2289704. PubMed DOI PMC

Darriba D, Posada D, Kozlov AM, Stamatakis A, Morel B, Flouri T. ModelTest-NG: A New and Scalable Tool for the Selection of DNA and Protein Evolutionary Models. Molecular Biology and Evolution. 2020;37:291–294. doi: 10.1093/molbev/msz189. PubMed DOI PMC

Di Lecce I, Bazzocchi C, Cecere JG, Epis S, Sassera D, Villani BM, Bazzi G, Negri A, Saino N, Spina F, Bandi C, Rubolini D. Patterns of Midichloria infection in avian-borne African ticks and their trans-Saharan migratory hosts. Parasites & Vectors. 2018;11:106. doi: 10.1186/s13071-018-2669-z. PubMed DOI PMC

Driscoll TP, Verhoeve VI, Brockway C, Shrewsberry DL, Plumer M, Sevdalis SE, Beckmann JF, Krueger LM, Macaluso KR, Azad AF, Gillespie JJ. Evolution of Wolbachia mutualism and reproductive parasitism: insight from two novel strains that co-infect cat fleas. PeerJ. 2020;8:1–39. doi: 10.7717/peerj.10646. PubMed DOI PMC

Duron O, Bouchon D, Boutin S, Bellamy L, Zhou L, Engelstädter J, Hurst GD. The diversity of reproductive parasites among arthropods: Wolbachia do not walk alone. BMC Biology. 2008;6:1–12. doi: 10.1186/1741-7007-6-27. PubMed DOI PMC

Duron O, Binetruy F, Noël V, Cremaschi J, McCoy KD, Arnathau C, Plantard O, Goolsby J, Pérez de León AA, Heylen DJA, Van Oosten AR, Gottlieb Y, Baneth G, Guglielmone AA, Estrada-Peña A, Opara MN, Zenner L, Vavre F, Chevillon C. Evolutionary changes in symbiont community structure in ticks. Molecular Ecology. 2017;26:2905–2921. doi: 10.1111/mec.14094. PubMed DOI

Duron O, Morel O, Noël V, Buysse M, Binetruy F, Lancelot R, Loire E, Ménard C, Bouchez O, Vavre F, Vial L. Tick-Bacteria Mutualism Depends on B Vitamin Synthesis Pathways. Current Biology. 2018;28:1896–1902. doi: 10.1016/j.cub.2018.04.038. PubMed DOI

Duron O, Gottlieb Y. Convergence of Nutritional Symbioses in Obligate Blood Feeders. Trends in Parasitology. 2020;36:816–825. doi: 10.1016/j.pt.2020.07.007. PubMed DOI

Edgar RC. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research. 2004;32:1792–1797. doi: 10.1093/nar/gkh340. PubMed DOI PMC

Emms DM, Kelly S. OrthoFinder: phylogenetic orthology inference for comparative genomics. Genome Biology. 2019;20:1–14. doi: 10.1186/s13059-019-1832-y. PubMed DOI PMC

Galperin MY, Makarova KS, Wolf YI, Koonin EV. Expanded microbial genome coverage and improved protein family annotation in the COG database. Nucleic Acids Research. 2015;43:D261–D269. doi: 10.1093/nar/gku1223. PubMed DOI PMC

Gerhart JG, Moses AS, Raghavan R. A Francisella-like endosymbiont in the Gulf Coast tick evolved from a mammalian pathogen. Scientific Reports. 2016;6:1–6. doi: 10.1038/srep33670. PubMed DOI PMC

Gerhart JG, Auguste Dutcher H, Brenner AE, Moses AS, Grubhoffer L, Raghavan R. Multiple Acquisitions of Pathogen-Derived Francisella Endosymbionts in Soft Ticks. Genome Biology and Evolution. 2018;10:607–615. doi: 10.1093/gbe/evy021. PubMed DOI PMC

Gerth M, Bleidorn C. Comparative genomics provides a timeframe for Wolbachia evolution and exposes a recent biotin synthesis operon transfer. Nature Microbiology. 2016;2:16241. doi: 10.1038/nmicrobiol.2016.241. PubMed DOI

Giladi M, Altman-Price N, Levin I, Levy L, Mevarech M. FolM, a new chromosomally encoded dihydrofolate reductase in Escherichia coli. Journal of Bacteriology. 2003;185:7015–7018. doi: 10.1128/JB.185.23.7015-7018.2003. PubMed DOI PMC

Gillespie JJ, Joardar V, Williams KP, Driscoll T, Hostetler JB, Nordberg E, Shukla M, Walenz B, Hill CA, Nene VM, Azad AF, Sobral BW, Caler E. A Rickettsia genome overrun by mobile genetic elements provides insight into the acquisition of genes characteristic of an obligate intracellular lifestyle. Journal of Bacteriology. 2012;194:376–394. doi: 10.1128/JB.06244-11. PubMed DOI PMC

Gottlieb Y, Lalzar I, Klasson L. Distinctive Genome Reduction Rates Revealed by Genomic Analyses of Two Coxiella-Like Endosymbionts in Ticks. Genome Biology and Evolution. 2015;7:1779–1796. doi: 10.1093/gbe/evv108. PubMed DOI PMC

Greay TL, Gofton AW, Paparini A, Ryan UM, Oskam CL, Irwin PJ. Recent insights into the tick microbiome gained through next-generation sequencing. Parasites & Vectors. 2018;11:12. doi: 10.1186/s13071-017-2550-5. PubMed DOI PMC

Guizzo MG, Parizi LF, Nunes RD, Schama R, Albano RM, Tirloni L, Oldiges DP, Vieira RP, Oliveira WHC, Leite MDS, Gonzales SA, Farber M, Martins O, Vaz IDS, Oliveira PL. A Coxiella mutualist symbiont is essential to the development of Rhipicephalus microplus. Scientific Reports. 2017;7:1–10. doi: 10.1038/s41598-017-17309-x. PubMed DOI PMC

Gulia-Nuss M. Genomic insights into the Ixodes scapularis tick vector of Lyme disease. Nature Communications. 2016;7:10507. doi: 10.1038/ncomms10507. PubMed DOI PMC

Gurevich A, Saveliev V, Vyahhi N, Tesler G. QUAST: Quality assessment tool for genome assemblies. Bioinformatics. 2013;29:1072–1075. doi: 10.1093/bioinformatics/btt086. PubMed DOI PMC

Guy L. GenoPlotR: comparative gene and genome visualization in R. Bioinformatics. 2010;27:2334–2335. doi: 10.1093/bioinformatics/btq413. PubMed DOI PMC

Hugoson E, Lam WT, Guy L. MiComplete: Weighted quality evaluation of assembled microbial genomes. Bioinformatics. 2020;36:936–937. doi: 10.1093/bioinformatics/btz664. PubMed DOI PMC

Husnik F, McCutcheon JP. Repeated replacement of an intrabacterial symbiont in the tripartite nested mealybug symbiosis. PNAS. 2016;113:E5416–E5424. doi: 10.1073/pnas.1603910113. PubMed DOI PMC

Jia N, Wang J, Shi W, Du L, Sun Y, Zhan W, Jiang J-F, Wang Q, Zhang B, Ji P, Bell-Sakyi L, Cui X-M, Yuan T-T, Jiang B-G, Yang W-F, Lam TT-Y, Chang Q-C, Ding S-J, Wang X-J, Zhu J-G, Ruan X-D, Zhao L, Wei J-T, Ye R-Z, Que TC, Du C-H, Zhou Y-H, Cheng JX, Dai P-F, Guo W-B, Han X-H, Huang E-J, Li L-F, Wei W, Gao Y-C, Liu J-Z, Shao H-Z, Wang X, Wang C-C, Yang T-C, Huo Q-B, Li W, Chen H-Y, Chen S-E, Zhou L-G, Ni X-B, Tian J-H, Sheng Y, Liu T, Pan Y-S, Xia L-Y, Li J, Tick Genome and Microbiome Consortium (TIGMIC) Zhao F, Cao W-C. Large-Scale Comparative Analyses of Tick Genomes Elucidate Their Genetic Diversity and Vector Capacities. Cell. 2020;182:1328–1340. doi: 10.1016/j.cell.2020.07.023. PubMed DOI

Ju JF, Bing XL, Zhao DS, Guo Y, Xi Z, Hoffmann AA, Zhang KJ, Huang HJ, Gong JT, Zhang X, Hong XY. Wolbachia supplement biotin and riboflavin to enhance reproduction in planthoppers. The ISME Journal. 2020;14:676–687. doi: 10.1038/s41396-019-0559-9. PubMed DOI PMC

Kanehisa M, Sato Y, Morishima K. BlastKOALA and GhostKOALA: KEGG Tools for Functional Characterization of Genome and Metagenome Sequences. Journal of Molecular Biology. 2016;428:726–731. doi: 10.1016/j.jmb.2015.11.006. PubMed DOI

Katoh K, Misawa K, Kuma K, Miyata T. MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform. Nucleic Acids Research. 2002;30:3059–3066. doi: 10.1093/nar/gkf436. PubMed DOI PMC

Kent BN, Bordenstein SR. Phage WO of Wolbachia: lambda of the endosymbiont world. Trends in Microbiology. 2010;18:173–181. doi: 10.1016/j.tim.2009.12.011. PubMed DOI PMC

Kumar S, Jones M, Koutsovoulos G, Clarke M, Blaxter M. Blobology: exploring raw genome data for contaminants, symbionts and parasites using taxon-annotated GC-coverage plots. Frontiers in Genetics. 2013;4:1–12. doi: 10.3389/fgene.2013.00237. PubMed DOI PMC

Li H, Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics. 2009;25:1754–1760. doi: 10.1093/bioinformatics/btp324. PubMed DOI PMC

Lukasik P, Nazario K, Van Leuven JT, Campbell MA, Meyer M, Michalik A, Pessacq P, Simon C, Veloso C, McCutcheon JP. Multiple origins of interdependent endosymbiotic complexes in a genus of cicadas. PNAS. 2018;115:E226–E235. doi: 10.1073/pnas.1712321115. PubMed DOI PMC

Machado-Ferreira E, Vizzoni VF, Balsemão-Pires E, Moerbeck L, Gazeta GS, Piesman J, Voloch CM, Soares CAG. Coxiella symbionts are widespread into hard ticks. Parasitology Research. 2016;115:4691–4699. doi: 10.1007/s00436-016-5230-z. PubMed DOI

Manzano-Marın A, Coeur d’acier A, Clamens AL, Orvain C, Cruaud C, Barbe V, Jousselin E. Serial horizontal transfer of vitamin-biosynthetic genes enables the establishment of new nutritional symbionts in aphids’ di-symbiotic systems. The ISME Journal. 2020;14:259–273. doi: 10.1038/s41396-019-0533-6. PubMed DOI PMC

Martin M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet.Journal. 2011;17:10. doi: 10.14806/ej.17.1.200. DOI

Matsuura Y, Moriyama M, Łukasik P, Vanderpool D, Tanahashi M, Meng XY, McCutcheon JP, Fukatsu T. Recurrent symbiont recruitment from fungal parasites in cicadas. PNAS. 2018;115:E5970–E5979. doi: 10.1073/pnas.1803245115. PubMed DOI PMC

McCutcheon JP, Boyd BM, Dale C. The Life of an Insect Endosymbiont from the Cradle to the Grave. Current Biology. 2019;29:R485–R495. doi: 10.1016/j.cub.2019.03.032. PubMed DOI

Narasimhan S, Fikrig E. Tick microbiome: the force within. Trends in Parasitology. 2015;31:315–323. doi: 10.1016/j.pt.2015.03.010. PubMed DOI PMC

Narasimhan S, Swei A, Abouneameh S, Pal U, Pedra JHF, Fikrig E. Grappling with the tick microbiome. Trends in Parasitology. 2021;37:722–733. doi: 10.1016/j.pt.2021.04.004. PubMed DOI PMC

Nardi T, Olivieri E, Kariuki E, Sassera D, Castelli M. Sequence of a Coxiella Endosymbiont of the Tick Amblyomma nuttalli Suggests a Pattern of Convergent Genome Reduction in the Coxiella Genus. Genome Biology and Evolution. 2021;13:1–7. doi: 10.1093/gbe/evaa253. PubMed DOI PMC

Nikoh N, Hosokawa T, Moriyama M, Oshima K, Hattori M, Fukatsu T. Evolutionary origin of insect-Wolbachia nutritional mutualism. PNAS. 2014;111:10257–10262. doi: 10.1073/pnas.1409284111. PubMed DOI PMC

Penz T, Schmitz-Esser S, Kelly SE, Cass BN, Müller A, Woyke T, Malfatti SA, Hunter MS, Horn M. Comparative genomics suggests an independent origin of cytoplasmic incompatibility in Cardinium hertigii. PLOS Genetics. 2012;8:e1003012. doi: 10.1371/journal.pgen.1003012. PubMed DOI PMC

Perner J, Sobotka R, Sima R, Konvickova J, Sojka D, Oliveira P de, Hajdusek O, Kopacek P. Acquisition of exogenous haem is essential for tick reproduction. eLife. 2016;5:e12318. doi: 10.7554/eLife.12318. PubMed DOI PMC

Ríhová J, Nováková E, Husník F, Hypša V. Legionella Becoming a Mutualist: Adaptive Processes Shaping the Genome of Symbiont in the Louse Polyplax serrata. Genome Biology and Evolution. 2017;9:2946–2957. doi: 10.1093/gbe/evx217. PubMed DOI PMC

Santos-Garcia D, Rollat-Farnier PA, Beitia F, Zchori-Fein E, Vavre F, Mouton L, Moya A, Latorre A, Silva FJ. The genome of Cardinium cBtQ1 provides insights into genome reduction, symbiont motility, and its settlement in Bemisia tabaci. Genome Biology and Evolution. 2014;6:1013–1030. doi: 10.1093/gbe/evu077. PubMed DOI PMC

Santos-Garcia D, Juravel K, Freilich S, Zchori-Fein E, Latorre A, Moya A, Morin S, Silva FJ. To B or Not to B: Comparative Genomics Suggests Arsenophonus as a Source of B Vitamins in Whiteflies. Frontiers in Microbiology. 2018;9:1–16. doi: 10.3389/fmicb.2018.02254. PubMed DOI PMC

Sassera D, Beninati T, Bandi C, Bouman EAP, Sacchi L, Fabbi M, Lo N. “Candidatus Midichloria mitochondrii”, an endosymbiont of the tick Ixodes ricinus with a unique intramitochondrial lifestyle. International Journal of Systematic and Evolutionary Microbiology. 2006;56:2535–2540. doi: 10.1099/ijs.0.64386-0. PubMed DOI

Sassera D, Lo N, Epis S, D’Auria G, Montagna M, Comandatore F, Horner D, Peretó J, Luciano AM, Franciosi F, Ferri E, Crotti E, Bazzocchi C, Daffonchio D, Sacchi L, Moya A, Latorre A, Bandi C. Phylogenomic evidence for the presence of a flagellum and cbb(3) oxidase in the free-living mitochondrial ancestor. Molecular Biology and Evolution. 2011;28:3285–3296. doi: 10.1093/molbev/msr159. PubMed DOI

Seemann T. Prokka: rapid prokaryotic genome annotation. Bioinformatics. 2014;30:2068–2069. doi: 10.1093/bioinformatics/btu153. PubMed DOI

Selmi R, Ben Said M, Mamlouk A, Ben Yahia H, Messadi L. Molecular detection and genetic characterization of the potentially pathogenic Coxiella burnetii and the endosymbiotic Candidatus Midichloria mitochondrii in ticks infesting camels (Camelus dromedarius) from Tunisia. Microbial Pathogenesis. 2019;136:103655. doi: 10.1016/j.micpath.2019.103655. PubMed DOI

Smith TA, Driscoll T, Gillespie JJ, Raghavan R. A Coxiella-like endosymbiont is a potential vitamin source for the Lone Star tick. Genome Biology and Evolution. 2015;7:831–838. doi: 10.1093/gbe/evv016. PubMed DOI PMC

Stamatakis A. RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics. 2014;30:1312–1313. doi: 10.1093/bioinformatics/btu033. PubMed DOI PMC

Stothard P, Wishart DS. Circular genome visualization and exploration using CGView. Bioinformatics. 2005;21:537–539. doi: 10.1093/bioinformatics/bti054. PubMed DOI

Sudakaran S, Kost C, Kaltenpoth M. Symbiont Acquisition and Replacement as a Source of Ecological Innovation. Trends in Microbiology. 2017;25:375–390. doi: 10.1016/j.tim.2017.02.014. PubMed DOI

Syberg-Olsen M, Garber A, Keeling P, McCutcheon J, Husnik F. Pseudofinder: Detection of Pseudogenes in Prokaryotic Genomes. bioRxiv. 2020 doi: 10.1101/2021.10.07.463580. PubMed DOI PMC

Takeshita K, Yamada T, Kawahara Y, Narihiro T, Ito M, Kamagata Y, Shinzato N. Tripartite Symbiosis of an Anaerobic Scuticociliate with Two Hydrogenosome-Associated Endosymbionts, a Holospora-Related Alphaproteobacterium and a Methanogenic Archaeon. Applied and Environmental Microbiology. 2019;85:1–14. doi: 10.1128/AEM.00854-19. PubMed DOI PMC

Tamas I, Wernegreen JJ, Nystedt B, Kauppinen SN, Darby AC, Gomez-Valero L, Lundin D, Poole AM, Andersson SGE. Endosymbiont gene functions impaired and rescued by polymerase infidelity at poly(A) tracts. PNAS. 2008;105:14934–14939. doi: 10.1073/pnas.0806554105. PubMed DOI PMC

Vautrin E, Vavre F. Interactions between vertically transmitted symbionts: cooperation or conflict? Trends in Microbiology. 2009;17:95–99. doi: 10.1016/j.tim.2008.12.002. PubMed DOI

Vial L, Stachurski F, Leblond A, Huber K, Vourc’h G, René-Martellet M, Desjardins I, Balança G, Grosbois V, Pradier S, Gély M, Appelgren A, Estrada-Peña A. Strong evidence for the presence of the tick Hyalomma marginatum Koch, 1844 in southern continental France. Ticks and Tick-Borne Diseases. 2016;7:1162–1167. doi: 10.1016/j.ttbdis.2016.08.002. PubMed DOI

Wick RR, Schultz MB, Zobel J, Holt KE. Bandage: interactive visualization of de novo genome assemblies. Bioinformatics. 2015;31:3350–3352. doi: 10.1093/bioinformatics/btv383. PubMed DOI PMC

Xie Z, Tang H. ISEScan: automated identification of insertion sequence elements in prokaryotic genomes. Bioinformatics. 2017;33:3340–3347. doi: 10.1093/bioinformatics/btx433. PubMed DOI

Zhong J, Jasinskas A, Barbour AG. Antibiotic treatment of the tick vector Amblyomma americanum reduced reproductive fitness. PLOS ONE. 2007;2:e00405. doi: 10.1371/journal.pone.0000405. PubMed DOI PMC

Ixodes ricinus ticks have a functional association with Midichloria mitochondrii