Deianiraea, an extracellular bacterium associated with the ciliate Paramecium, suggests an alternative scenario for the evolution of Rickettsiales

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Abstract

Rickettsiales are a lineage of obligate intracellular Alphaproteobacteria, encompassing important human pathogens, manipulators of host reproduction, and mutualists. Here we report the discovery of a novel Rickettsiales bacterium associated with Paramecium, displaying a unique extracellular lifestyle, including the ability to replicate outside host cells. Genomic analyses show that the bacterium possesses a higher capability to synthesise amino acids, compared to all investigated Rickettsiales. Considering these observations, phylogenetic and phylogenomic reconstructions, and re-evaluating the different means of interaction of Rickettsiales bacteria with eukaryotic cells, we propose an alternative scenario for the evolution of intracellularity in Rickettsiales. According to our reconstruction, the Rickettsiales ancestor would have been an extracellular and metabolically versatile bacterium, while obligate intracellularity would have evolved later, in parallel and independently, in different sub-lineages. The proposed new scenario could impact on the open debate on the lifestyle of the last common ancestor of mitochondria within Alphaproteobacteria.

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Data availability

Sequence data have been deposited in GenBank with the accession codes: P. primaurelia CyL4-1 partial 18S-ITS1-5.8S-ITS2-28S—MH185950,P. primaurelia CyL4-1 partial cytochrome oxidase subunit I gene—MH188082, Deianiraea vastatrix CyL4-1 partial 16S rRNA gene—MH197138; Deianiraea vastatrix CyL4-1 genome—CP028925.

References

  1. 1.

    Dumler JS, Walker DH. Rickettsiales. In: Whitman WB, editor. Bergey’s manual of systematics of archaea and bacteria. John Wiley & Sons, Ltd; 2015.

  2. 2.

    Perlman SJ, Hunter MS, Zchori-Fein E. The emerging diversity of Rickettsia. Proc R Soc B. 2006;273:2097–106.

  3. 3.

    Werren JH, Baldo L, Clark ME. Wolbachia: master manipulators of invertebrate biology. Nat Rev Microbiol. 2008;6:741–51.

  4. 4.

    Montagna M, Sassera D, Epis S, Bazzocchi C, Vannini C, Lo N, et al. “Candidatus Midichloriaceae” fam. nov. (Rickettsiales), an ecologically widespread clade of intracellular alphaproteobacteria. Appl Environ Microbiol. 2013;79:3241–8.

  5. 5.

    Castelli M, Sassera D, Petroni G. Biodiversity of “non-model” Rickettsiales and their association with aquatic organisms. In: Thomas S, editor. Rickettsiales—biology, molecular biology, epidemiology, and vaccine development. p. 59–91 (Springer International Publishing, Cham, Switzerland, 2016).

  6. 6.

    Kocan KM, De La Fuente J, Blouin EF, Garcia-Garcia JC. Anaplasma marginale (Rickettsiales: Anaplasmataceae): recent advances in defining host–pathogen adaptations of a tick-borne rickettsia. Parasitology. 2004;129:S285–S300.

  7. 7.

    Weinert LA, Werren JH, Aebi A, Stone GN, Jiggins FM. Evolution and diversity of Rickettsia bacteria. BMC Biol. 2009;7:6.

  8. 8.

    Parola P, Paddock CD, Socolovschi C, Labruna MB, Mediannikov O, Kernif T, et al. Update on tick-borne rickettsioses around the world: a geographic approach. Clin Microbiol Rev. 2013;26:657–702.

  9. 9.

    Taylor MJ, Bandi C, Hoerauf A. Wolbachia endosymbionts of filarial nematodes. Adv Parasitol. 2005;60:245–84.

  10. 10.

    Hosokawa T, Koga R, Kikuchi Y, Meng XY, Fukatsu T. Wolbachia as a bacteriocyte-associated nutritional mutualist. Proc Natl Acad Sci USA. 2010;107:769–74.

  11. 11.

    Andersson SGE, Zomorodipour A, Andersson JO, Sicheritz-Pontén T, Alsmark UCM, Podowski RM, et al. The genome sequence of Rickettsia prowazekii and the origin of mitochondria. Nature. 1998;396:133–43.

  12. 12.

    Fitzpatrick DA, Creevey CJ, McInerney JO. Genome phylogenies indicate a meaningful α-proteobacterial phylogeny and support a grouping of the mitochondria with the Rickettsiales. Mol Biol Evol. 2006;23:74–85.

  13. 13.

    Wang Z, Wu M. An integrated phylogenomic approach toward pinpointing the origin of mitochondria. Sci Rep. 2015;5:7949.

  14. 14.

    Esser C, Ahmadinejad N, Wiegand C, Rotte C, Sebastiani F, Gelius-Dietrich G, et al. A genome phylogeny for mitochondria among α-proteobacteria and a predominantly eubacterial ancestry of yeast nuclear genes. Mol Biol Evol. 2004;21:1643–60.

  15. 15.

    Martijn J, Vosseberg J, Guy L, Offre P, Ettema TJG. Deep mitochondrial origin outside the sampled alphaproteobacteria. Nature. 2018;557:101–5.

  16. 16.

    Epis S, Sassera D, Beninati T, Lo N, Beati L, Piesman J, et al. Midichloria mitochondrii is widespread in hard ticks (Ixodidae) and resides in the mitochondria of phylogenetically diverse species. Parasitology. 2008;135:485–94.

  17. 17.

    Driscoll T, Gillespie JJ, Nordberg EK, Azad AF, Sobral BW. Bacterial DNA sifted from the Trichoplax adhaerens (Animalia: Placozoa) genome project reveals a putative rickettsial endosymbiont. Genome Biol Evol. 2013;5:621–45.

  18. 18.

    Dantas-Torres F, Chomel BB, Otranto D. Ticks and tick-borne diseases: a One Health perspective. Trends Parasitol. 2012;28:437–46.

  19. 19.

    Schrallhammer M, Ferrantini F, Vannini C, Galati S, Schweikert M, Görtz HD, et al. “Candidatus Megaira polyxenophila” gen. nov., sp. nov.: considerations on evolutionary history, host range and shift of early divergent rickettsiae. PLoS ONE. 2013;8:e72581.

  20. 20.

    Senra MVX, Dias RJP, Castelli M, Silva-Neto ID, Verni F, Soares CAG, et al. A house for two-double bacterial infection in Euplotes woodruffi Sq1 (Ciliophora, Euplotia) sampled in southeastern Brazil. Microb Ecol. 2016;71:505–17.

  21. 21.

    Matsuura Y, Kikuchi Y, Meng XY, Koga R, Fukatsu T. Novel clade of alphaproteobacterial endosymbionts associated with stinkbugs and other arthropods. Appl Environ Microbiol. 2012;78:4149–56.

  22. 22.

    Braig HR, Guzman H, Tesh RB, O’Neill SL. Replacement of the natural Wolbachia symbiont of Drosophila simulans with a mosquito counterpart. Nature. 1994;367:453–5.

  23. 23.

    Caspi-Fluger A, Inbar M, Mozes-Daube N, Katzir N, Portnoy V, Belausov E, et al. Horizontal transmission of the insect symbiont Rickettsia is plant-mediated. Proc R Soc B. 2012;279:1791–6.

  24. 24.

    Schulz F, Martijn J, Wascher F, Lagkouvardos I, Kostanjšek R, Ettema TJG, et al. A Rickettsiales symbiont of amoebae with ancient features. Environ Microbiol. 2016;18:2326–42.

  25. 25.

    Casiraghi M, Anderson TJ, Bandi C, Bazzocchi C, Genchi C. A phylogenetic analysis of filarial nematodes: comparison with the phylogeny of Wolbachia endosymbionts. Parasitology. 2001;122:93–103.

  26. 26.

    Walker DH, Ismail N. Emerging and re-emerging rickettsioses: endothelial cell infection and early disease events. Nat Rev Microbiol. 2008;6:375–86.

  27. 27.

    Rikihisa Y. Anaplasma phagocytophilum and Ehrlichia chaffeensis: subversive manipulators of host cells. Nat Rev Microbiol. 2010;8:328–39.

  28. 28.

    Rasgon JL, Gamston CE, Ren X. Survival of Wolbachia pipientis in cell-free medium. Appl Environ Microbiol. 2006;72:6934–7.

  29. 29.

    Philip RN, Casper EA. Serotypes of spotted fever group rickettsiae isolated from Dermacentor andersoni (Stiles) ticks in western Montana. Am J Trop Med Hyg. 1981;30:230–8.

  30. 30.

    Labruna MB, Whitworth T, Horta MC, Bouyer DH, McBride JW, Pinter A, et al. Rickettsia species infecting Amblyomma cooperi ticks from an area in the state of São Paulo, Brazil, where Brazilian spotted fever is endemic. J Clin Microbiol. 2004;42:90–98.

  31. 31.

    Sunyakumthorn P, Bourchookarn A, Pornwiroon W, David C, Barker SA, Macaluso KR. Characterization and growth of polymorphic Rickettsia felis in a tick cell line. Appl Environ Microbiol. 2008;74:3151–8.

  32. 32.

    Muñoz-Gómez S, Hess S, Burger G, Lang BF, Susko E, Slamovits CH, et al. An updated phylogeny of the Alphaproteobacteria reveals that the parasitic Rickettsiales and Holosporales have independent origins. eLife 2019;8:e42535.

  33. 33.

    Georgiades K, Madoui MA, Le P, Robert C, Raoult D. Phylogenomic analysis of Odyssella thessalonicensis fortifies the common origin of Rickettsiales, Pelagibacter ubique and Reclimonas americana mitochondrion. PLoS ONE. 2011;5:e24857.

  34. 34.

    Ferla MP, Thrash JC, Giovannoni SJ, Patrick WM. New rRNA gene-based phylogenies of the Alphaproteobacteria provide perspective on major groups, mitochondrial ancestry and phylogenetic instability. PLoS ONE. 2013;8:e83383.

  35. 35.

    Hess S, Suthaus A, Melkonian M. “Candidatus Finniella” (Rickettsiales, Alphaproteobacteria), novel endosymbionts of viridiraptorid amoeboflagellates (Cercozoa, Rhizaria). Appl Environ Microbiol. 2016;82:659–70.

  36. 36.

    Parks DH, Chuvochina M, Waite DW, Rinke C, Skarshewski A, Chaumell PA, et al. A standardized bacterial taxonomy based on genome phylogeny substantially revises the tree of life. Nat Biotechnol. 2018;36:996–1004.

  37. 37.

    Szokoli F, Castelli M, Sabaneyeva E, Schrallhammer M, Krenek S, Doak TG, et al. Disentangling the taxonomy of Rickettsiales and description of two novel symbionts (“Candidatus Bealeia paramacronuclearis” and “Candidatus Fokinia cryptica”) sharing the cytoplasm of the ciliate protist Paramecium biaurelia. Appl Environ Microbiol. 2016;82:7236–47.

  38. 38.

    Floriano AM, Castelli M, Krenek S, Berendonk TU, Bazzocchi C, Petroni G, et al. The genome sequence of “Candidatus Fokinia solitaria”: insights on reductive evolution in Rickettsiales. Genome Biol Evol. 2018;10:1120–6.

  39. 39.

    Darby AC, Cho NH, Fuxelius HH, Westberg J, Andersson SG. Intracellular pathogens go extreme: genome evolution in the Rickettsiales. Trends Genet. 2007;23:511–20.

  40. 40.

    Sassera D, Lo N, Epis S, D’Auria G, Montagna M, Comandatore F, et al. Phylogenomic evidence for the presence of a flagellum and cbb3 oxidase in the free-living mitochondrial ancestor. Mol Biol Evol. 2011;28:3285–96.

  41. 41.

    Szokoli F, Sabaneyeva E, Castelli M, Krenek S, Schrallhammer M, Soares CAG, et al. “Candidatus Fokinia solitaria”, a novel “stand-alone” symbiotic lineage of Midichloriaceae (Rickettsiales). PLOS ONE. 2016;11:e0145743.

  42. 42.

    Lanzoni O, Fokin SI, Lebedeva N, Migunova A, Petroni G, Potekhin A. Rare freshwater ciliate Paramecium chlorelligerum Kahl, 1935 and its macronuclear symbiotic bacterium “Candidatus Holospora parva”. PLOS ONE. 2016;11:e0167928.

  43. 43.

    Boscaro V, Petroni G, Ristori A, Verni F, Vannini C. “Candidatus Defluviella procrastinata” and “Candidatus Cyrtobacter zanobii”, two novel ciliate endosymbionts belonging to the “Midichloria clade”. Microb Ecol. 2013;65:302–10.

  44. 44.

    Cole JR, Wang Q, Fish JA, Chai B, McGarrell DM, Sun Y, et al. Ribosomal Database Project: data and tools for high throughput rRNA analysis. Nucleic Acids Res. 2014;42:D633–D642.

  45. 45.

    Quast C, Pruesse E, Yilmaz P, Gerken J, Schweer T, Yarza P, et al. The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucl Acids Res. 2013;41:D590–D596.

  46. 46.

    Westram R, Bader K, Prüsse E, Kumar Y, Meier H, Glöckner FO, et al. ARB: a software environment for sequence data. In de Bruijn FJ, editor. Handbook of molecular microbial ecology I: metagenomics and complementary approaches. p. 399–406 (John Wiley & Sons, Hoboken, New Jersey, 2011).

  47. 47.

    Sabaneyeva E, Castelli M, Szokoli F, Benken K, Lebedeva N, Salvetti A, et al. Host and symbiont intraspecific variability: The case of Paramecium calkinsi and “Candidatus Trichorickettsia mobilis”. Eur J Protistol. 2018;62:79–94.

  48. 48.

    Wright ES, Yilmaz LS, Noguera DR. DECIPHER, a search-based approach to chimera identification for 16S rRNA sequences. Appl Environ Microbiol. 2012;78:717–25.

  49. 49.

    Lagkouvardos I. IMNGS: a comprehensive open resource of processed 16S rRNA microbial profiles for ecology and diversity studies. Sci Rep. 2016;6:33721.

  50. 50.

    Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010;26:2460–1.

  51. 51.

    Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, et al. SPAdes: A new genome assembly algorithm and its applications to single-cell sequencing. J Comp Biol. 2012;19:455–77.

  52. 52.

    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. Front Genet. 2013;4:237.

  53. 53.

    Seemann T. Prokka: rapid prokaryotic genome annotation. Bioinformatics. 2014;30:2068–9.

  54. 54.

    Emanuelsson O, Brunak S, von Heijne G, Nielsen H. Locating proteins in the cell using TargetP, SignalP and related tools. Nat Protoc. 2007;2:953–71.

  55. 55.

    Krogh A, Larsson B, von Heijne G, Sonnhammer EL. Predicting transmembrane protein topology with a hidden markov model: application to complete genomes. J Mol Biol. 2001;305:567–80.

  56. 56.

    Siguier P, Perochon J, Lestrade L, Mahillon J, Chandler M. ISfinder: the reference centre for bacterial insertion sequences. Nucleic Acids Res. 2006;34:D32–6.

  57. 57.

    Zhou Y, Liang Y, Lynch KH, Dennis JJ, Wishart DS. PHAST: a fast phage search tool. Nucleic Acids Res. 2011;39:W347–52.

  58. 58.

    Nakamura T, Yamada KD, Tomii K, Katoh K. Parallelization of MAFFT for large-scale multiple sequence alignments. Bioinformatics. 2018;34:2490–2.

  59. 59.

    Darriba D, Taboada GL, Doallo R, Posada D. ProtTest 3: fast selection of best-fit models of protein evolution. Bioinformatics. 2011;27:1164–5.

  60. 60.

    Stamatakis A. Using RAxML to infer phylogenies. Curr Protoc Bioinform. 2015;51:6.14.1–6.14.14.

  61. 61.

    Ronquist F, Teslenko M, Van Der Mark P, Ayres DL, Darling A, Höhna S, et al. Mrbayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol. 2012;61:539–42.

  62. 62.

    Karp PD, Latendresse M, Paley SM, Krummenacker M, Ong QD, Billington R, et al. Pathway tools version 19.0: integrated software for pathway/genome informatics and systems biology. Brief Bioinform. 2016;17:877–90.

  63. 63.

    Kanehisa M, Sato Y, Kawashima M, Furumichi M, Tanabe M. KEGG as a reference resource for gene and protein annotation. Nucleic Acids Res. 2016;44:D457–62.

  64. 64.

    Galperin MY, Makarova KS, Wolf YI, Koonin EV. Expanded microbial genome coverage and improved protein family annotation in the COG database. Nucleic Acids Res. 2015;43:D261–9.

  65. 65.

    Modeo L, Fokin SI, Boscaro V, Andreoli I, Ferrantini F, Rosati G, et al. Morphology, ultrastructure, and molecular phylogeny of the ciliate Sonderia vorax with insights into the systematics of order Plagiopylida. BMC Microbiol. 2013;13:40.

  66. 66.

    Nitla V, Serra V, Fokin SI, Modeo L, Verni F, Sandeep BV, et al. Critical revision of the family Plagiopylidae (Ciliophora: Plagiopylea), including the description of two novel species, Plagiopyla ramani and Plagiopyla narasimhamurtii, and redescription of Plagiopyla nasuta Stein, 1860 from India. Zool J Linn Soc. 2018. https://doi.org/10.1093/zoolinnean/zly041

  67. 67.

    Davidson SK, Powell R, James S. A global survey of the bacteria within earthworm nephridia. Mol Phylogenet Evol. 2013;67:188–200.

  68. 68.

    Fraune S, Bosch TCG. Long-term maintenance of species-specific bacterial microbiota in the basal metazoan Hydra. Proc Natl Acad Sci USA. 2007;104:13146–51.

  69. 69.

    Fraune S, Augustin R, Anton-Erxleben F, Wittlieb J, Gelhaus C, Klimovich VB, et al. In an early branching metazoan, bacterial colonization of the embryo is controlled by maternal antimicrobial peptides. Proc Natl Acad Sci USA. 2010;107:18067–72.

  70. 70.

    Bauermeister J, Ramette A, Dattagupta S. Repeatedly evolved host-specific ectosymbioses between sulfur-oxidizing bacteria and amphipods living in a cave ecosystem. PloS ONE. 2012;7:e50254.

  71. 71.

    Dishaw LJ, Flores-Torres J, Lax S, Gemayel K, Leigh B, Melillo D, et al. The gut of geographically disparate Ciona intestinalis harbors a core microbiota. PloS ONE. 2014;9:e93386.

  72. 72.

    Kong HH, Oh J, Deming C, Conlan S, Grice EA, Beatson MA, et al. Temporal shifts in the skin microbiome associated with disease flares and treatment in children with atopic dermatitis. Genome Res. 2012;22:850–9.

  73. 73.

    Shinkai T, Enishi O, Mitsumori M, Higuchi K, Kobayashi Y, Takenaka A, et al. Mitigation of methane production from cattle by feeding cashew nut shell liquid. J Dairy Sci. 2012;95:5308–16.

  74. 74.

    Wang Z, Wu M. Complete genome sequence of the endosymbiont of Acanthamoeba strain UWC8, an amoeba endosymbiont belonging to the “Candidatus Midichloriaceae” family in Rickettsiales. Genome Announc. 2014;2:e00791–14.

  75. 75.

    Martijn J, Schulz F, Zaremba-Niedzwiedzka K, Viklund J, Stepanauskas R, Andersson SG, et al. Single-cell genomics of a rare environmental alphaproteobacterium provides unique insights into Rickettsiaceae evolution. ISME J. 2015;9:2373–85.

  76. 76.

    Kühn J, Briegel A, Mörschel E, Kahnt J, Leser K, Wick S, et al. Bactofilins, a ubiquitous class of cytoskeletal proteins mediating polar localization of a cell wall synthase in Caulobacter crescentus. EMBO J. 2010;29:327–39.

  77. 77.

    Dunning Hotopp JC, Lin M, Madupu R, Crabtree J, Angiuoli SV, Eisen J, et al. Comparative genomics of emerging human ehrlichiosis agents. PLoS Genet. 2006;2:e21.

  78. 78.

    Gillespie JJ, Kaur SJ, Rahman MS, Rennoll-Bankert K, Sears KT, Beier-Sexton M, et al. Secretome of obligate intracellular Rickettsia. FEMS Microbiol Rev. 2015;39:47–80.

  79. 79.

    Gillespie JJ, Phan IQ, Driscoll TP, Guillotte ML, Lehman SS, Rennoll-Bankert KE, et al. The Rickettsia type IV secretion system: Unrealized complexity mired by gene family expansion. Path Dis. 2016;74:ftw058.

  80. 80.

    Gillespie JJ, Ammerman NC, Dreher-Lesnick SM, Rahman MS, Worley MJ, Setubal JC, et al. An anomalous type IV secretion system in Rickettsia is evolutionarily conserved. PLoS ONE. 2009;4:e4833.

  81. 81.

    Korotkov KV, Sandkvist M, Hol WGJ. The type II secretion system: biogenesis, molecular architecture and mechanism. Nat Rev Microbiol. 2012;10:336–51.

  82. 82.

    Guérin J, Bigot S, Schneider R, Buchanan SK, Jacob-Dubuisson F. Efficiency and simplicity in the secretion of large proteins for bacteria–host and bacteria–bacteria interactions. Front Cell Infect Microbiol. 2017;7:148.

  83. 83.

    Manson McGuire A, Cochrane K, Griggs AD, Haas BJ, Abeel T, Zeng Q, et al. Evolution of invasion in a diverse set of Fusobacterium species. Mbio. 2014;5:e01864–14.

  84. 84.

    Soo RM, Woodcroft BJ, Parks DH, Tyson GW, Hugenholtz P. Back from the dead; the curious tale of the predatory cyanobacterium Vampirovibrio chlorellavorus. PeerJ. 2015;3:e968.

  85. 85.

    Mattick JS. Type IV pili and twitching motility. Annu Rev Microbiol. 2002;56:289–314.

  86. 86.

    Ge Y, Rikihisa Y. Subversion of host cell signaling by Orientia tsutsugamushi. Microbes Infect. 2011;13:638–48.

  87. 87.

    Cardwell MM, Martinez JJ. The Sca2 autotransporter protein from Rickettsia conorii is sufficient to mediate adherence to and invasion of cultured mammalian cells. Infect Immun. 2009;77:5272–80.

  88. 88.

    Gouin E, Egile C, Dehoux P, Villiers V, Adams J, Gertler F, et al. The RickA protein of Rickettsia conorii activates the Arp2/3 complex. Nature. 2004;427:457–61.

  89. 89.

    Mohan Kumar D, Yamaguchi M, Miura K, Lin M, Los M, Coy JF, et al. Ehrlichia chaffeensis uses its surface protein EtpE to bind GPI-anchored protein DNase X and trigger entry into mammalian cells. PloS Pathog. 2013;9:e1003666.

  90. 90.

    Renesto P, Dehoux P, Gouin E, Touqui L, Cossart P, et al. Identification and characterization of a phospholipase D‐superfamily gene in rickettsiae. J Infect Dis. 2003;188:1276–83.

  91. 91.

    Kahlon A, Ojogun N, Ragland SA, Seidman D, Troese MJ, Ottens AK, et al. Anaplasma phagocytophilum Asp14 is an invasin that interacts with mammalian host cells via its C terminus to facilitate infection. Infect Immun. 2013;81:65–79.

  92. 92.

    Seidman D, Ojogun N, Walker NJ, Mastronunzio J, Kahlon A, Hebert KS, et al. Anaplasma phagocytophilum surface protein AipA mediates invasion of mammalian host cells. Cell Microbiol. 2014;16:1133–45.

  93. 93.

    Vannini C, Ferrantini F, Schleifer KH, Ludwig W, Verni F, Petroni G. “Candidatus Anadelfobacter veles” and “Candidatus Cyrtobacter comes,” two new Rickettsiales species hosted by the protist ciliate Euplotes harpa (Ciliophora, Spirotrichea). Appl Environ Microbiol. 2010;76:4047–54.

  94. 94.

    Schmitz-Esser S, Linka N, Collingro A, Beier CL, Neuhaus HE, Wagner M, et al. ATP/ADP translocases: a common feature of obligate intracellular amoebal symbionts related to Chlamydiae and Rickettsiae. J Bacteriol. 2004;186:683–91.

  95. 95.

    Vannini C, Petroni G, Verni, Rosati G. A bacterium belonging to the Rickettsiaceae family inhabits the cytoplasm of the marine ciliate Diophrys appendiculata (Ciliophora, Hypotrichia). Microb Ecol. 2004;49:434–42.

  96. 96.

    Ogata H, La Scola B, Audic S, Renesto P, Blanc G, Robert C, et al. Genome sequence of Rickettsia bellii illuminates the role of amoebae in gene exchanges between intracellular pathogens. PloS Genet. 2006;2:e76.

  97. 97.

    Kang YJ, Diao XN, Zhao GY, Chen MH, Xiong Y, Shi M, et al. Extensive diversity of Rickettsiales bacteria in two species of ticks from China and the evolution of the Rickettsiales. BMC Evol Biol. 2014;14:167.

  98. 98.

    Driscoll TP, Verhoeve VI, Guillotte ML, Lehman SS, Rennoll SA, Beier-Sexton M, et al. Wholly Rickettsia! Reconstructed metabolic profile of the quintessential bacterial parasite of eukaryotic cells. Mbio. 2017;8:e00859–17.

  99. 99.

    Vannini C, Boscaro V, Ferrantini F, Benken KA, Mironov TI, Schweikert M, et al. Flagellar movement in two bacteria of the family Rickettsiaceae: a re-evaluation of motility in an evolutionary perspective. PLoS ONE. 2014;9:e8771.

  100. 100.

    Kwan JC, Schmidt EW. Bacterial endosymbiosis in a chordate host: long-term co-evolution and conservation of secondary metabolism. PLoS ONE. 2013;8:e80822.

  101. 101.

    Castelli M, Serra V, Senra MVX, Basuri CK, Soares CAG, Fokin SI, et al. The hidden world of Rickettsiales symbionts: “Candidatus Spectririckettsia obscura,” a novel bacterium found in Brazilian and Indian Paramecium caudatum. Microb Ecol 2018. https://doi.org/10.1007/s00248-018-1243-8

  102. 102.

    Yurchenko T, Ševčíková T, Přibyl P, El Karkouri K, Klimeš V, Amaral R, et al. A gene transfer event suggests a long-term partnership between eustigmatophyte algae and a novel lineage of endosymbiotic bacteria. ISME J. 2018;12:2163–75.

  103. 103.

    Amann RI, Binder BJ, Olson RJ, Chisholm SW, Devereux R, Stahl DA. Combination of 16S ribosomal-RNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations. Appl Environ Microbiol. 1990;56:1919–25.

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Acknowledgements

This work was funded by the European Commission FP7-PEOPLE-2009-IRSES grant 247658 (project CINAR-PATHOBACTER) to GP, the Human Frontier Science Programme Grant RGY0075/2017 to DS, the Italian Ministry of Education, University and Research (MIUR): Dipartimenti di Eccellenza Programme (2018–2022)—Department of Biology and Biotechnology “L. Spallanzani”, University of Pavia to DS, and, for the work of the group from St. Petersburg, by the RSF grant 16-14-10157 to AP. We would like to thank A. Oren for advice in bacterial nomenclature and species description. S. Gabrielli and S. Lometto are acknowledged for assistance in graphical artwork and in phylogenomic analyses, respectively. Cultures were maintained in Culture Collection of Microorganisms (RC CCM), Saint Petersburg State University.

Author information

NL isolated the host cells and performed live experiments. ES performed TEM and fluorescence microscopy. KB performed AFM. OL, AP and GP performed molecular experiments and probe design. MC, OL and GP performed phylogeny. MC and OL performed IMNGS analyses. MC, AMF and DS performed genome assembly and analyses. MC, AMF, SG and DS performed metabolic and evolutionary analyses. General interpretation of the results and manuscript writing were performed by MC, DS and GP, and contributed by CB, LM, ES and AP. All the authors contributed to the interpretation and writing of the specific results, and to the discussions. GP planned and coordinated the whole project, with DS planning and coordinating the genomic analyses, and ES coordinating the microscopy.

Correspondence to Davide Sassera or Giulio Petroni.

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