Candidatus Mesenet longicola”: Novel Endosymbionts of Brontispa longissima that Induce Cytoplasmic Incompatibility

Abstract

Intracellular bacteria that are mainly transmitted maternally affect their arthropod hosts’ biology in various ways. One such effect is known as cytoplasmic incompatibility (CI), and three bacterial species are known to induce CI: Wolbachia, Cardinium hertigii, and a recently found alphaproteobacterial symbiont. To clarify the taxonomic status and provide the foundation for future studies to reveal CI mechanisms and other phenotypes, we investigated genetic and morphological properties of the third CI inducer that we have previously reported inducing CI in the coconut beetle Brontispa longissima. The draft genome of the bacteria was obtained from the oocytes of two isofemale lines of B. longissima infected with the bacteria: one from Japan (GL2) and the other from Vietnam (L5). Genome features of the symbionts (sGL2 and sL5) were highly similar, showing 1.3 Mb in size, 32.1% GC content, and 99.83% average nucleotide sequence. A phylogenetic study based on 43 universal and single-copy phylogenetic marker genes indicates that they formed a distinct clade in the family Anaplasmataceae. 16S rRNA gene sequences indicate that they are different from the closest known relatives, at least at the genus level. Therefore, we propose a new genus and species, “Candidatus Mesenet longicola”, for the symbionts of B. longissima. Morphological analyses showed that Ca. M. longicola is an intracellular bacterium that is ellipsoidal to rod-shaped and 0.94 ± 0.26 μm (mean ± SD) in length, and accumulated in the anterior part of the oocyte. Candidates for the Ca. M. longicola genes responsible for CI induction are also described.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Data Availability

The draft genomes of Ca. M. longicola have been deposited in the DNA Data Bank of Japan (DDBJ)/European Molecular Biology Laboratory (EMBL)/GenBank database with accession number DRR226614-DRR226615.

References

  1. 1.

    Laven H (1967) Eradication of Culex pipiens fatigans through cytoplasmic incompatibility. Nature 216:383–384

    CAS  Article  Google Scholar 

  2. 2.

    Boller EF, Russ K, Vallo V, Bush GL (1976) Incompatible races of European cherry fruit fly, Rhagoletis cerasi (Diptera: Tephritidae), their origin and potential use in biological control. Ent Exp Appl 20:237–247

    Article  Google Scholar 

  3. 3.

    Yen JH, Barr RA (1971) New hypothesis of the cause of cytoplasmic incompatibility in Culex pipiens L. Nature 232:657–658

    CAS  Article  Google Scholar 

  4. 4.

    Yen JH, Barr RA (1973) The etiological agent of cytoplasmic incompatibility in Culex pipiens. J Invertebr Pathol 22:242–250

    CAS  Article  Google Scholar 

  5. 5.

    Hunter MS, Perlman SJ, Kelly SE (2003) A bacterial symbiont in the Bacteroidetes induces cytoplasmic incompatibility in the parasitoid wasp Encarsia pergandiella. Proc R Soc Lond B 270:2185–2190

    Article  Google Scholar 

  6. 6.

    Takano SI, Tuda M, Takasu K, Furuya N, Imamura Y, Kim S, Tashiro K, Iiyama K, Tavares M, Amaral AC (2017) Unique clade of alphaproteobacterial endosymbionts induces complete cytoplasmic incompatibility in the coconut beetle. Proc Natl Acad Sci U S A 114:6110–6115

    CAS  Article  Google Scholar 

  7. 7.

    Beckmann JF, Ronau JA, Hochstrasser M (2017) A Wolbachia deubiquitylating enzyme induces cytoplasmic incompatibility. Nat Microbiol 2:17007. https://doi.org/10.1038/nmicrobiol.2017.7

    Article  PubMed  PubMed Central  Google Scholar 

  8. 8.

    LePage DP, Metcalf JA, Bordenstein SR, On J, Perlmutter JI, Shropshire JD, Layton EM, Funkhouser-Jones LJ, Beckmann JF, Bordenstein SR (2017) Prophage WO genes recapitulate and enhance Wolbachia-induced cytoplasmic incompatibility. Nature 543:243–247

    CAS  Article  Google Scholar 

  9. 9.

    Chen H, Ronau JA, Beckmann JF, Hochstrasser M (2019) A Wolbachia nuclease and its binding partner provide a distinct mechanism for cytoplasmic incompatibility. Proc Natl Acad Sci U S A 116:22314–22321

    CAS  Article  Google Scholar 

  10. 10.

    Shropshire JD, On J, Layton EM, Zhou H, Bordenstein SR (2018) One prophage WO gene rescues cytoplasmic incompatibility in Drosophila melanogaster. Proc Natl Acad Sci U S A 115:4987–4991

    CAS  Article  Google Scholar 

  11. 11.

    Min KT, Benzer S (1997) Wolbachia, normally a symbiont of Drosophila, can be virulent, causing degeneration and early death. Proc Natl Acad Sci U S A 94:10792–10796

    CAS  Article  Google Scholar 

  12. 12.

    McMeniman CJ, Lane RV, Cass BN, Fong AWC, Sidhu M, Wang YF, O’Neill SL (2009) Stable introduction of a life-shortening Wolbachia infection into the mosquito Aedes aegypti. Science 323:141–144

    CAS  Article  Google Scholar 

  13. 13.

    Hoffmann AA, Ross PA, Rašić G (2015) Wolbachia strains for disease control: ecological and evolutionary considerations. Evol Appl 8:751–768

    Article  Google Scholar 

  14. 14.

    Beckmann JF, Bonneau M, Chen H, Hochstrasser M, Poinsot D, Merçot H, Weill M, Sicard M, Charlat S (2019) The toxin–antidote model of cytoplasmic incompatibility: genetics and evolutionary implications. Trends Genet 35:175–185

    CAS  Article  Google Scholar 

  15. 15.

    Takano S, Mochizuki A, Konishi K, Takasu K, Alouw JC, Pandin DS, Nakamura S (2011) Two cryptic species in Brontispa longissima (Coleoptera: Chrysomelidae): evidence from mitochondrial DNA analysis and crosses between the two nominal species. Ann Entomol Soc Am 104:121–131

    Article  Google Scholar 

  16. 16.

    Kang DD, Li F, Kirton E, Thomas A, Egan R, An H, Wang Z (2019) MetaBAT 2: an adaptive binning algorithm for robust and efficient genome reconstruction from metagenome assemblies. PeerJ 7:e7359. https://doi.org/10.7717/peerj.7359

    Article  PubMed  PubMed Central  Google Scholar 

  17. 17.

    Parks DH, Imelfort M, Skennerton CT, Hugenholtz P, Tyson GW (2015) Checkm: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res 25:1043–1055

    CAS  Article  Google Scholar 

  18. 18.

    Tanizawa Y, Fujisawa T, Kaminuma E, Nakamura Y, Arita M (2016) DFAST and DAGA: Web-based integrated genome annotation tools and resources. Biosci Microbiota Food Health 35:173–184

    CAS  Article  Google Scholar 

  19. 19.

    Ohtsubo Y, Ikeda-Ohtsubo W, Nagata Y, Tsuda M (2008) GenomeMatcher: a graphical user interface for DNA sequence comparison. BMC Bioinf 9:376. https://doi.org/10.1186/1471-2105-9-376

    CAS  Article  Google Scholar 

  20. 20.

    Kozlov A, Darriba D, Flouri T, Morel B, Stamatakis A (2019) RAxML-NG: a fast, scalable, and user-friendly tool for maximum likelihood phylogenetic inference. Bioinformatics 35:4453–4455

    CAS  Article  Google Scholar 

  21. 21.

    Ludwig W, Strunk O, Westram R, Richter L, Meier H, Yadhukumar BA, Lai T, Steppi S, Jobb G, Förster W, Brettske I, Gerber S, Ginhart AW, Gross O, Grumann S, Hermann S, Jost R, König A, Liss T, Lüßmann R, May M, Nonhoff B, Reichel B, Strehlow R, Stamatakis A, Stuckmann N, Vilbig A, Lenke M, Ludwig T, Bode A, Schleifer KH (2004) ARB: a software environment for sequence data. Nucleic Acids Res 32:1363–1371

    CAS  Article  Google Scholar 

  22. 22.

    Cole JR, Wang Q, Fish JA, Chai B, McGarrell DM, Sun Y, Brown CT, Porras-Alfaro A, Kuske CR, Tiedje JM (2014) Ribosomal Database Project: data and tools for high throughput rRNA analysis. Nucleic Acids Res 42(Database issue):D633–D642

    CAS  Article  Google Scholar 

  23. 23.

    Sanguin H, Herrera A, Oger-Desfeux C, Dechesne A, Simonet P, Navarro E, Vogel TM, Moënne-Locoz Y, Nesme X, Grundmann GL (2006) Development and validation of a prototype 16S rRNA-based taxonomic microarray for Alphaproteobacteria. Environ Microbiol 8:289–307

    CAS  Article  Google Scholar 

  24. 24.

    Klasson L, Walker T, Sebaihia M, Sanders MJ, Quail MA, Lord A, Sanders S, Earl J, O’Neill SL, Thomson N, Sinkins SP, Parkhill J (2008) Genome evolution of Wolbachia strain wPip from the Culex pipiens group. Mol Biol Evol 25:1877–1887

    CAS  Article  Google Scholar 

  25. 25.

    Wu M, Sun LV, Vamathevan J, Riegler M, Deboy R, Brownlie JC, McGraw EA, Martin W, Esser C, Ahmadinejad N, Wiegand C, Madupu R, Beanan MJ, Brinkac LM, Daugherty SC, Scott Durkin A, Kolonay JF, Nelson WC, Mohamoud Y, Lee P, Berry K, Young MB, Utterback T, Weidman J, Nierman WC, Paulsen IT, Nelson KE, Tettelin H, O’Neill SL, Eisen JA (2004) Phylogenomics of the reproductive parasite Wolbachia pipientis wMel: a streamlined genome overrun by mobile genetic elements. PLoS Biol 2:327–341

    CAS  Article  Google Scholar 

  26. 26.

    Brown AMV, Wasala SK, Howe DK, Peetz AB, Zasada IA, Denver DR (2016) Genomic evidence for plant-parasitic nematodes as the earliest Wolbachia hosts. Sci Rep 6:34955. https://doi.org/10.1038/srep34955

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  27. 27.

    diCenzo GC, Finan TM (2017) The divided bacterial genome: structure, function, and evolution. Microbiol Mol Biol Rev 81:e00019–e00017. https://doi.org/10.1128/27.MMBR.00019-17

    Article  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Andersson SGE, Zomorodipour A, Andersson JO, Sicheritz-Pontén T, Alsmark UCM, Podowski RM, Näslund AK, Eriksson AS, Winkler HH, Kurland CG (1998) The genome sequence of Rickettsia prowazekii and the origin of mitochondria. Nature 396:133–140

    CAS  Article  Google Scholar 

  29. 29.

    Shigenobu S, Watanabe H, Hattori M, Sakaki Y, Ishikawa H (2000) Genome sequence of the endocellular bacterial symbiont of aphids Buchnera sp. APS Nat 407:81–86

    CAS  Google Scholar 

  30. 30.

    Nakabachi A, Yamashita A, Toh H, Ishikawa H, Dunbar HE, Moran NA, Hattori M (2006) The 160-kilobase genome of the bacterial endosymbiont Carsonella. Science 314:267

    CAS  Article  Google Scholar 

  31. 31.

    Sun L, Foster J, Tzertzinis G, Ono M, Bandi C, Slatko B, O’Neill S (2001) Determination of Wolbachia genome size by pulsed-field gel electrophoresis. J Bacteriol 183:2219–2225

    CAS  Article  Google Scholar 

  32. 32.

    Fenollar F, La Scola B, Inokuma H, Dumler JS, Taylor MJ, Raoult D (2003) Culture and phenotypic characterization of a Wolbachia pipientis isolate. J Clin Microbiol 41:5434–5441

    CAS  Article  Google Scholar 

  33. 33.

    Moran NA (1996) Accelerated evolution and Muller’s ratchet in endosymbiotic bacteria. Proc Natl Acad Sci U S A 93:2873–2878

    CAS  Article  Google Scholar 

  34. 34.

    Singhal K, Mohanty S (2019) Genome organisation and comparative genomics of four novel Wolbachia genome assemblies from Indian Drosophila host. Funct Integr Genom 19:617–632

    CAS  Article  Google Scholar 

  35. 35.

    Siozios S, Pilgrim J, Darby AC, Baylis M, Hurst GDD (2019) The draft genome of strain cCpun from biting midges confirms insect Cardinium are not a monophyletic group and reveals a novel gene family expansion in a symbiont. PeerJ 7:e6448. https://doi.org/10.7717/peerj.6448

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Yarza P, Yilmaz P, Pruesse E, Glockner FO, Ludwig W, Schleifer KH, Whitman WB, Euzeby J, Amann R, Rossello-Mora R (2014) Uniting the classification of cultured and uncultured bacteria and archaea using 16S rRNA gene sequences. Nat Rev Microbiol 12:635–645

    CAS  Article  Google Scholar 

  37. 37.

    Li K, Stanojević M, Stamenković G, Ilić B, Paunović M, Lu M, Pešić B, Maslovara IĐ, Siljic M, Cirkovic V, Zhang Y (2019) Insight into diversity of bacteria belonging to the order Rickettsiales in 9 arthropods species collected in Serbia. Sci Rep 9:18680. https://doi.org/10.1038/s41598-019-55077-y

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Murray RGE, Schleifer KH (1994) A proposal for recording the properties of putative taxa of procaryotes. Int J Syst Bacteriol 44:174–176

    CAS  Article  Google Scholar 

  39. 39.

    Murray RGE, Stackebrandt E (1995) Implementation of the provisional status Candidatus for incompletely described prokaryotes. Int J Syst Bacteriol 45:186–187

    CAS  Article  Google Scholar 

  40. 40.

    Veneti Z, Clark ME, Karr TL, Savakis C, Bourtzis K (2004) Heads or tails: host-parasite interactions in the Drosophila-Wolbachia system. Appl Environ Microbiol 70:5366–5372

    CAS  Article  Google Scholar 

  41. 41.

    Feree PM, Frydman HM, Li JM, Cao J, Wieschaus E, Sullivan W (2005) Wolbachia utilizes host microtubules and Dynein for anterior localization in the Drosophila oocyte. PLoS Pathog 1:e14. https://doi.org/10.1371/journal.ppat.0010014

    CAS  Article  Google Scholar 

  42. 42.

    Shi P, He Z, Li S, An X, Lv N, Ghanim M, Cuthbertson AGS, Ren SX, Qiu BL (2016) Wolbachia has two different localization patterns in whitefly Bemisia tabaci AsiaII7 species. PLoS ONE 11:e0162558. https://doi.org/10.1371/journal.42.pone.0162558

    Article  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Guo Y, Hoffmann AA, Xu XQ, Mo PW, Huang HJ, Gong JT, Ju JF, Hong XY (2018) Vertical transmission of Wolbachia is associated with host vitellogenin in Laodelphax striatellus. Front Microbiol 9:2016. https://doi.org/10.3389/fmicb.2018.02016

    Article  PubMed  PubMed Central  Google Scholar 

  44. 44.

    Hildebrandt PK, Conroy JD, McKee AE, Nyindo MBA, Huxsoll DL (1973) Ultrastructure of Ehrlichia canis. Infect Immun 7:265–271

    CAS  Article  Google Scholar 

  45. 45.

    Sells DM, Hildebrandt PK, Lewis GE, Nyindo MBA, Ristic M (1976) Ultrastructural observations on Ehrlichia equi organisms in equine granulocytes. Infect Immun 13:273–280

    CAS  Article  Google Scholar 

  46. 46.

    Caturegli P, Asanovich KM, Walls JJ, Bakken JS, Madigan JE, PoPov VL, Dumler JS (2000) ankA: an Ehrlichia phagocytophila group gene encoding a cytoplasmic protein antigen with ankyrin repeats. Infect Immun 68:5277–5283

    CAS  Article  Google Scholar 

  47. 47.

    Beyer AR, VieBrock L, Rodino KG, Miller DP, Tegels BK, Marconi RT, Carlyon JA (2015) Orientia tsutsugamushi strain Ikeda ankyrin repeat-containing proteins recruit SCF1 ubiquitin ligase machinery via poxvirus-like F-box motifs. J Bacteriol 197:3097–3109

    CAS  Article  Google Scholar 

  48. 48.

    VieBrock L, Evans SM, Beyer AR, Larson CL, Beare PA, Ge H, Singh S, Rodino KG, Heinzen RA, Richards AL, Carlyon JA (2014) Orientia tsutsugamushi ankyrin repeat-containing protein family members are Type 1secretion system substrates that traffic to the host cell endoplasmic reticulum. Front Cell Infect Microbiol 4:186. https://doi.org/10.3389/fcimb.2014.00186

    Article  PubMed  Google Scholar 

  49. 49.

    Bordenstein SR, Bordenstein SR (2016) Eukaryotic association module in phage WO genomes from Wolbachia. Nat Commun 7:13155. https://doi.org/10.1038/ncomms13155

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  50. 50.

    Sasaki T, Ishikawa H (1999) Wolbachia infections and cytoplasmic incompatibility in the almond moth and the Mediterranean flour moth. Zool Sci 16:739–744

    Article  Google Scholar 

Download references

Acknowledgements

We thank Naruto Furuya, Kazuhiko Iiyama, and Chisa Yasunaga-Aoki for their suggestions on the study design; Kazunori Matsuo for the suggestions in the nomenclature; Nguyen Tuan Dat and Tran Van Chien for their help in collecting insects in Vietnam; Katsuya Fukami and the Materials Management Center of Kyushu University for arranging transfer of the materials used in this study; Yataro So, Kenji So, Sun Green Co., Ltd., and Futamigaura Cemetery for providing food plants for insect rearing.

Funding

This study was supported by the Japan Society for the Promotion of Science KAKENHI Grant 18H02207 (ST) and 16H06279 (PAGS).

Author information

Affiliations

Authors

Contributions

S. T, Y. G., and T. H. designed the experiments. S. T. and Y. G. performed the experiments and data analyses. S. T. and Y. G. wrote the original draft of the manuscript. All authors reviewed the manuscript and agreed with the final version.

Corresponding author

Correspondence to Shun-ichiro Takano.

Ethics declarations

Conflict of Interest

The authors declare that they have no conflict of interest.

Ethics Approval

Not applicable.

Consent to Participate

All authors reviewed the manuscript and agreed with the final version.

Consent for Publication

All authors reviewed the manuscript and agreed with the final version.

Code Availability

Not applicable.

Supplementary Information

figure7

High resolution image (PNG 94 kb)

ESM 1

(EPS 1307 kb)

ESM 2

(XLSX 15 kb)

ESM 3

(XLSX 11 kb)

ESM 4

(XLSX 20 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Takano, Si., Gotoh, Y. & Hayashi, T. “Candidatus Mesenet longicola”: Novel Endosymbionts of Brontispa longissima that Induce Cytoplasmic Incompatibility. Microb Ecol (2021). https://doi.org/10.1007/s00248-021-01686-y

Download citation

Keywords

  • Anaplasmataceae
  • Biological control
  • Symbiont
  • Wolbachia