Applied Entomology and Zoology

, Volume 47, Issue 1, pp 1–8 | Cite as

Gut symbiotic bacteria in the cabbage bugs Eurydema rugosa and Eurydema dominulus (Heteroptera: Pentatomidae)

  • Yoshitomo Kikuchi
  • Takahiro Hosokawa
  • Naruo Nikoh
  • Takema Fukatsu
Original Research Paper


The cabbage bugs Eurydema rugosa Motschulsky and Eurydema dominulus (Scopoli) (Heteroptera: Pentatomidae: Strachiini) possess a number of crypts in a posterior region of the midgut, which are filled with bacterial symbiont cells. Here we characterized the gut symbionts of Eurydema stinkbugs using molecular phylogenetic and histological techniques. Specific gammaproteobacteria were consistently identified from the posterior midgut of E. rugosa representing nine populations and E. dominulus representing six populations, respectively. The bacterial 16S rRNA gene sequences were identical within the species but slightly different (98.2% sequence identity) between the species. Molecular phylogenetic analysis revealed that the Eurydema symbionts formed a well-defined monophyletic group in the Gammaproteobacteria. The symbionts were phylogenetically distinct from the gut symbionts of the stinkbug families Acanthosomatidae, Plataspidae, Parastrachiidae, Scutelleridae, and other pentatomid species, suggesting multiple evolutionary origins of the gut symbiotic bacteria among diverse stinkbugs. In situ hybridization confirmed that the symbiont is located in the cavity of the midgut crypts. Aposymbiotic insects of E. rugosa, which were produced by egg surface sterilization, were viable but suffered retarded growth, reduced body weight, and abnormal body color, suggesting the biological importance of the symbiont for the host.


Eurydema rugosa Eurydema dominulus Symbiotic bacteria Midgut crypts Gammaproteobacteria 


  1. Abe Y, Mishiro K, Takanashi M (1995) Symbiont of brown-winged green bug, Plautia stali Scott. Jpn J Appl Entomol Zool 39:109–115CrossRefGoogle Scholar
  2. Bellostas N, Sørensen AD, Sørensen JC, Sørensen H (2007) Genetic variation and metabolism of glucosinolates. Adv Bot Res 45:369–415CrossRefGoogle Scholar
  3. Buchner P (1965) Endosymbiosis of animals with plant microorganisms. Interscience, New YorkGoogle Scholar
  4. Douglas AE (1998) Nutritional interactions in insect-microbial symbioses: aphids and their symbiotic bacteria Buchnera. Annu Rev Entomol 43:17–37PubMedCrossRefGoogle Scholar
  5. Folmer O, Black M, Hoeh W, Lutz R, Vrijenhoek R (1994) DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol Mar Biol Biotechnol 3:294–299PubMedGoogle Scholar
  6. Fukatsu T, Hosokawa T (2002) Capsule-transmitted gut symbiotic bacterium of the Japanese common plataspid stinkbug, Megacopta punctatissima. Appl Environ Microbiol 68:389–396PubMedCrossRefGoogle Scholar
  7. Fukatsu T, Nikoh N (1998) Two intracellular symbiotic bacteria from the mulberry psyllid Anomoneura mori (Insecta, Homoptera). Appl Environ Microbiol 64:3599–3606PubMedGoogle Scholar
  8. Galtier N, Gouy M, Gautier C (1996) SEAVIEW and PHYLO_WIN: two graphic tools for sequence alignment and molecular phylogeny. Bioinformatics 12:543–548Google Scholar
  9. Glasgow H (1914) The gastric caeca and the caecal bacteria of the Heteroptera. Biol Bull 3:101–171CrossRefGoogle Scholar
  10. Guindon S, Dufayard JF, Lefort V, Anisimova M, Hordijk W, Gascuel O (2010) New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst Biol 59:307–321PubMedCrossRefGoogle Scholar
  11. Hosokawa T, Kikuchi Y, Meng XY, Fukatsu T (2005) The making of symbiont capsule in the plataspid stinkbug Megacopta punctatissima. FEMS Microbiol Ecol 54:471–477PubMedCrossRefGoogle Scholar
  12. Hosokawa T, Kikuchi Y, Nikoh N, Shimada M, Fukatsu T (2006) Strict host–symbiont cospeciation and reductive genome evolution in insect gut bacteria. PLoS Biol 4:e337PubMedCrossRefGoogle Scholar
  13. Kaiwa N, Hosokawa T, Kikuchi Y, Nikoh N, Meng XY, Kimura N, Ito M, Fukatsu T (2010) Primary gut symbiont and secondary Sodalis-allied symbiont in the scutellerid stinkbug Cantao ocellatus. Appl Environ Microbiol 76:3486–3494PubMedCrossRefGoogle Scholar
  14. Kaiwa N, Hosokawa T, Kikuchi Y, Nikoh N, Meng XY, Kimura N, Ito M, Fukatsu T (2011) Bacterial symbionts of the giant jewel stinkbug Eucorysses grandis (Hemiptera: Scutelleridae). Zool Sci 28:169–174PubMedCrossRefGoogle Scholar
  15. Kaltenpoth M, Winter SA, Kleinhammer A (2009) Localization and transmission route of Coriobacterium glomerans, the endosymbiont of pyrrhocorid bugs. FEMS Microbiol Ecol 69:373–383PubMedCrossRefGoogle Scholar
  16. Kikuchi Y (2009) Endosymbiotic bacteria in insects: their diversity and culturability. Microbes Environ 24:195–204PubMedCrossRefGoogle Scholar
  17. Kikuchi Y, Hosokawa T, Fukatsu T (2007) Insect–microbe mutualism without vertical transmission: a stinkbug acquires beneficial gut symbiont from environment every generation. Appl Environ Microbiol 73:4308–4316PubMedCrossRefGoogle Scholar
  18. Kikuchi Y, Hosokawa T, Fukatsu T (2008) Diversity of bacterial symbiosis in stinkbugs. In: Dijk TV (ed) Microbial ecology research trends. Nova Science Publishers, Inc, New York, pp 39–63Google Scholar
  19. Kikuchi Y, Hosokawa T, Nikoh N, Meng XY, Kamagata Y, Fukatsu T (2009) Host–symbiont co-speciation and reductive genome evolution in gut symbiotic bacteria of acanthosomatid stinkbugs. BMC Biol 7:2PubMedCrossRefGoogle Scholar
  20. Kimura M (1980) A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120PubMedCrossRefGoogle Scholar
  21. Nikoh N, Hosokawa T, Oshima K, Hattori M, Fukatsu T (2011) Reductive evolution of bacterial genome in insect gut environment. Genome Biol Evol 3:702–714Google Scholar
  22. Prado SS, Almeida RPP (2009a) Phylogenetic placement of pentatomid stink bug gut symbionts. Curr Microbiol 58:64–69PubMedCrossRefGoogle Scholar
  23. Prado SS, Almeida RPP (2009b) Role of symbiotic gut bacteria in the development of Acrosternum hilare and Murgantia histrionica. Entomol Exp Appl 132:21–29CrossRefGoogle Scholar
  24. Prado SS, Rubinoff D, Almeida RPP (2006) Vertical transmission of a pentatomid caeca-associated symbiont. Ann Entomol Soc Am 99:577–585CrossRefGoogle Scholar
  25. Prado SS, Hung KY, Daugherty MP, Almeida RPP (2010) Indirect effects of temperature on stink bug fitness, via maintenance of gut-associated symbionts. Appl Environ Microbiol 76:1261–1266PubMedCrossRefGoogle Scholar
  26. R Development Core Team (2010) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna. ISBN 3-900051-07-0.
  27. Robinson-Rechavi M, Huchon D (2000) RRTree: relative-rate tests between groups of sequences on a phylogenetic tree. Bioinformatics 16:296–297PubMedCrossRefGoogle Scholar
  28. Rosenkranz W (1939) Die Symbiose der Pentatomiden. Z Morph Ökol Tiere 36:279–309CrossRefGoogle Scholar
  29. Schaefer CW, Panizzi AR (2000) Heteroptera of economic importance. CRC Press, FloridaCrossRefGoogle Scholar
  30. Shigenobu S, Watanabe H, Hattori M, Sakaki Y, Ishikawa H (2000) Genome sequence of the endocellular bacterial symbiont of aphids Buchnera sp. APS. Nature 407:81–86PubMedCrossRefGoogle Scholar
  31. Swain T (1977) Secondary compounds as protective agents. Annu Rev Plant Physiol 28:479–501CrossRefGoogle Scholar
  32. Swofford DL (2001) PAUP* Version 4.0b10 (computer program). Sinauer, SunderlandGoogle Scholar
  33. Tada A, Kikuchi Y, Hosokawa T, Musolin DL, Fujisaki K, Fukatsu T (2011) Obligate association with gut bacterial symbiont in Japanese populations of the southern green stinkbug Nezara viridula (Heteroptera: Pentatomidae). Appl Entomol Zool. doi:10.1007/s13355-011-0066-6
  34. Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680PubMedCrossRefGoogle Scholar
  35. Tomokuni M, Yasunaga T, Takai M, Yamashita I, Kawamura M, Kawasawa T (1993) A field guide to Japanese bugs. Zenkoku Noson Kyoiku Kyoukai, TokyoGoogle Scholar
  36. Weirauch C, Schuh RT (2011) Systematics and evolution of Heteroptera: 25 years of progress. Annu Rev Entomol 56:487–510PubMedCrossRefGoogle Scholar

Copyright information

© The Japanese Society of Applied Entomology and Zoology 2011

Authors and Affiliations

  • Yoshitomo Kikuchi
    • 1
    • 2
  • Takahiro Hosokawa
    • 3
  • Naruo Nikoh
    • 4
  • Takema Fukatsu
    • 3
  1. 1.National Institute of Advanced Industrial Science and Technology (AIST), Hokkaido CenterSapporoJapan
  2. 2.Graduate School of AgricultureHokkaido UniversitySapporoJapan
  3. 3.National Institute of Advanced Industrial Science and Technology (AIST)TsukubaJapan
  4. 4.Department of Liberal ArtsThe Open University of JapanChibaJapan

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