Advertisement

Current Microbiology

, Volume 62, Issue 5, pp 1510–1520 | Cite as

Diversity of Endosymbionts in the Potato Psyllid, Bactericera cockerelli (Hemiptera: Triozidae), Vector of Zebra Chip Disease of Potato

  • Punya Nachappa
  • Julien Levy
  • Elizabeth Pierson
  • Cecilia TamborindeguyEmail author
Article

Abstract

Zebra chip disease is an emerging, serious disease of solanaceous crops and the causal agent is a bacterium “Candidatus Liberibacter solanacearum” (CLs), also known as “Candidatus Liberibacter psyllaurous”, which is transmitted by the potato psyllid, Bactericera cockerelli (Šulc). We performed bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP) of the 16S rDNA genes to determine the bacterial microbiota in adult insects from CLs-uninfected and CLs-infected strains of B. cockerelli and potato leaf samples. We obtained sequences from five bacterial species among the two psyllid strains, including “Candidatus Carsonella ruddii”, Wolbachia, CLs, and two transient bacteria, Acinetobacter and Methylibium. We did not detect any common bacteria between psyllids and potato leaf samples using pyrosequencing. We performed PCR analysis using species-specific 16S rDNA primers to confirm pyrosequencing results in individual psyllids including eggs, early-instars, late-instars, and adults of both sexes from both CLs-uninfected and CLs-infected psyllid strains. The primary endosymbiont, “Candidatus Carsonella ruddii” and Wolbachia were detected in all life-stages and sexes of both strains using PCR analyses. The percentage of CLs-infected individuals increased from early-instar (0%), late-instar (40%) until adulthood (60%) in the CLs-infected strain. We believe that CLs levels in early-instars are probably too low to be detected by standard PCR. Using PCR analyses, we confirmed the presence of Acinetobacter in CLs-uninfected and CLs-infected adults (75 and 25%, respectively) but not Methylibium. Further, we detected Acinetobacter in potato leaves using PCR indicating that the psyllids may have acquired this bacterium via feeding on the host plant.

Keywords

Acinetobacter Polymerase Chain Reaction Analysis Multiplex Polymerase Chain Reaction Potato Psyllid Asian Citrus Psyllid 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

We like to thank Elena Lyuksyutova for helping with PCR analyses and Dr. Anne M. Estes for helpful comments. This work was supported by Texas Department of Agriculture.

References

  1. 1.
    Abad JA, Bandla M, French-Monar RD, Liefting LW, Clover GRG (2009) First report of the detection of Candidatus Liberibacter species in zebra chip disease infected potato plants in the United States. Plant Dis 93:108CrossRefGoogle Scholar
  2. 2.
    Baumann P (2005) Biology bacteriocyte-associated endosymbionts of plant sap-sucking insects. Annu Rev Entomol 59:155–189Google Scholar
  3. 3.
    Baumann P, Baumann L, Lai Y, Rouhbakhsh D, Moran NA, Clark MA (1995) Genetics, physiology, and evolutionary relationships of the genus Buchnera: intracellular symbionts of aphids. Annu Rev Microbiol 49:55–94PubMedCrossRefGoogle Scholar
  4. 4.
    Baumann P, Moran NA, Baumann L (2000) Bacteriocyte-associated endosymbionts of insects. In: Dworkin M (ed) The prokaryotes. Springer, New York, NY, pp 1–55. http://link.springer.de/link/service/books/10125
  5. 5.
    Brownlie JC, Cass BN, Riegler M, Witsenburg JJ, Iturbe-Ormaetxe I, McGraw EA, O’Neill SL (2009) Evidence for metabolic provisioning by a common invertebrate endosymbiont, Wolbachia pipientis, during periods of nutritional stress. PLoS Pathog 5:e1000368PubMedCrossRefGoogle Scholar
  6. 6.
    Buchner P (1965) Endosymbiosis of animals with plant microorganisms. Wiley, New York, NYGoogle Scholar
  7. 7.
    Chang KP, Musgrave AJ (1969) Histochemistry and ultrastructure of the mycetome and its ‘symbiotes’ in the pear psylla, Psylla pyricola Foerster (Homoptera). Tissue Cell 1:597–606PubMedCrossRefGoogle Scholar
  8. 8.
    Cowan D, Meyer Q, Stafford W, Muyanga S, Cameron R, Wittwer P (2005) Metagenomic gene discovery: past, present and future. Trends Biotechnol 23:321–329PubMedCrossRefGoogle Scholar
  9. 9.
    Cranshaw WS (2001) Diseases caused by insect toxin: psyllid yellows. In: Stevenson WR, Loria R, Franc GD, Weingartner DP (eds) Compendium of potato diseases, 2nd edn. APS, St. Paul, MN, pp 73–74Google Scholar
  10. 10.
    Crosslin JM, Bester G (2009) First report of ‘Candidatus Liberibacter psyllaurous’ in zebra chip symptomatic potatoes from California. Plant Dis 93:551CrossRefGoogle Scholar
  11. 11.
    Crosslin JM, Munyaneza JE (2009) Evidence that the zebra chip disease and the putative causal agent can be maintained in potatoes by grafting and in vitro. Am J Potato Res 86:183–187CrossRefGoogle Scholar
  12. 12.
    Dale C, Moran NA (2006) Molecular interactions between bacterial symbionts and their hosts. Cell 126:453–465PubMedCrossRefGoogle Scholar
  13. 13.
    Douglas AE (1998) Nutritional interactions in insect–microbial symbioses: aphids and their symbiotic bacteria Buchnera. Annu Rev Entomol 43:17–37PubMedCrossRefGoogle Scholar
  14. 14.
    Douglas AE (2003) The nutritional physiology of aphids. Adv Insect Physiol 31:73–140CrossRefGoogle Scholar
  15. 15.
    Dowd SE, Callaway TR, Wolcott RD, McKeehan T, Hagevoort RG, Edrington TS (2008) Evaluation of the bacterial diversity in the feces of cattle using 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP). BMC Microbiol 8:125PubMedCrossRefGoogle Scholar
  16. 16.
    Engelbrektson A, Kunin V, Wrighton KC, Zvenigorodsky N, Chen F, Ochman H, Hugenholtz P (2010) Experimental factors affecting PCR-based estimates of microbial species richness and evenness. ISME J 4:642–647PubMedCrossRefGoogle Scholar
  17. 17.
    Fukatsu T, Nikoh N (1998) Two intracellular symbiotic bacteria from the mulberry psyllid Anomoneura mori (Insecta, Homoptera). Appl Environ Microbiol 64:3599–3606PubMedGoogle Scholar
  18. 18.
    Fukatsu T, Nikoh N, Kawai R, Koga R (2000) The secondary endosymbiotic bacterium of the pea aphid Acyrthosiphon pisum (Insecta, Homoptera). Appl Environ Microbiol 66(7):2748–2758PubMedCrossRefGoogle Scholar
  19. 19.
    Fukatsu T, Watanabe K, Sekiguchi Y (1998) Specific detection of intracellular symbiotic bacteria of aphids by oligonucleotide-probed in situ hybridization. Appl Entomol Zool 33:461–472Google Scholar
  20. 20.
    Gontcharova V, Youn E, Sun Y, Wolcott RD, Dowd SE (2010) A comparison of bacterial composition in diabetic ulcers and contralateral intact skin. Open Microbiol J 4:8–19PubMedCrossRefGoogle Scholar
  21. 21.
    Hansen AK, Trumble JT, Stouthamer R, Paine TD (2008) A New Huanglongbing (HLB) species, “Candidatus Liberibacter solanacearum”, found to infect tomato and potato, is vectored by the psyllid Bactericera cockerelli (Sulc). Appl Environ Microbiol 74:5862–5865PubMedCrossRefGoogle Scholar
  22. 22.
    Hilgenboecker K, Hammerstein P, Schlattmann P, Telschow A, Werren JH (2008) How many species are infected with Wolbachia?—A statistical analysis of current data. FEMS Microbiol Lett 281:215–220PubMedCrossRefGoogle Scholar
  23. 23.
    Hoffmann AA, Turelli M (1997) Cytoplasmic incompatibility in insects. In: O’Neill SL, Hoffmann AA, Werren JH (eds) Influential passengers: inherited microorganisms and arthropod reproduction. Oxford University Press, Oxford, UK, pp 42–80Google Scholar
  24. 24.
    Huigens ME, Hohmann C, Luck RF, Gort G, Stouthamer R (2004) Reduced competitive ability due to Wolbachia infection in the parasitoid wasp Trichogramma kaykai. Entomol Exp Appl 110:115–123CrossRefGoogle Scholar
  25. 25.
    Jeyaprakash A, Hoy MA (2000) Long PCR improves Wolbachia DNA amplification: wsp sequences found in 76% of sixty-three arthropod species. Insect Mol Biol 9:393–405PubMedCrossRefGoogle Scholar
  26. 26.
    Kimura M (1980) A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120PubMedCrossRefGoogle Scholar
  27. 27.
    Kremer N, Charif D, Henri H, Bataille M, Prevost G, Kraaijeveld K, Vavre F (2009) A new case of Wolbachia dependence in the genus Asobara: evidence for parthenogenesis induction in Asobara japonica. Heredity 103:248–256PubMedCrossRefGoogle Scholar
  28. 28.
    Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, Thompson JD, Gibson TJ, Higgins DG (2007) Clustal W and Clustal X version 2.0. Bioinformatics 23:2947–2948PubMedCrossRefGoogle Scholar
  29. 29.
    Li W, Abad JA, French-Monar RD, Rascoe J, Wen A, Gudmestad NC, Secor GA, Lee I-M, Duan Y, Levy L (2009) Multiplex real-time PCR for detection, identification and quantification of ‘Candidatus Liberibacter solanacearum’ in potato plants with zebra chip. J Microbiol Methods 78:59–65PubMedCrossRefGoogle Scholar
  30. 30.
    Liefting LW, Perez-Egusquiza ZC, Clover GRG, Anderson JAD (2008) A new ‘Candidatus Liberibacter’ species in Solanum tuberosum in New Zealand. Plant Dis 92:1474CrossRefGoogle Scholar
  31. 31.
    Lin H, Doddapaneni H, Munyaneza JE, Civerolo EL, Sengoda VG, Buchman JL, Stenger DC (2009) Molecular characterization and phylogenetic analysis of 16S rDNA from a new “Candidatus Liberibacter” strain associated with zebra chip disease of potato (Solanum tuberosum L.) and the potato psyllid (Bactericera cockerelli Sulc). J Plant Pathol 91:215–219Google Scholar
  32. 32.
    Liu D, Trumble JT (2004) Tomato psyllid behavioral responses to tomato plant lines and interactions of plant lines with insecticides. J Econ Entomol 97:1078–1085PubMedCrossRefGoogle Scholar
  33. 33.
    Meyer J, Hoy MA (2008) Molecular survey of endosymbionts in florida populations Diaphorina citri (Hemiptera: Psyllidae) and its parasitoids Tamarixia radiata (Hymenoptera: Eulophidae) and Diaphorencyrtus aligarhensis (Hymenoptera: Encyrtidae). Florida Entomologist 91(2):294–304CrossRefGoogle Scholar
  34. 34.
    Möller EM, Bahnweg G, Sandermann H, Geiger HH (1992) A simple and efficient protocol for isolation of high molecular weight DNA from filamentous fungi, fruit bodies, and infected plant tissues. Nucleic Acids Res 20:6115–6116PubMedCrossRefGoogle Scholar
  35. 35.
    Moran N, Baumann P (1994) Phylogenetics of cytoplasmically inherited microorganisms of arthropods. Trends Ecol Evol 9:15–20PubMedCrossRefGoogle Scholar
  36. 36.
    Moreira LA, Iturbe-Ormaetxe I, Jeffery JA, Lu G, Pyke AT, Hedges LM, Rocha BC, Hall-Mendelin S, Day A, Riegler M, Hugo LE, Johnson KN, Kay BH, McGraw EA, van den Hurk AF, Ryan PA, O’Neill SL (2009) A Wolbachia symbiont in Aedes aegypti limits infection with dengue, Chikungunya, and Plasmodium. Cell 139(7):1268–1278PubMedCrossRefGoogle Scholar
  37. 37.
    Munyaneza JE, Buchman JL, Upton JE, Goolsby JA, Crosslin JM, Bester G, Miles GP, Sengoda G (2008) Impact of different potato psyllid populations on zebra chip disease incidence, severity, and potato yield. Subtrop Plant Sci 60:27–37Google Scholar
  38. 38.
    Munyaneza JE, Crosslin JM, Upton JE (2007) Association of Bactericera cockerelli (Homoptera: Psyllidae) with “Zebra Chip”, a new potato disease in southwestern United States and Mexico. J Econ Entomol 100:656–663PubMedCrossRefGoogle Scholar
  39. 39.
    Munyaneza JE, Goolsby JA, Crosslin JM, Upton JE (2007) Further evidence that zebra chip potato disease in the Lower Rio Grande Valley of Texas is associated with Bactericera cockerelli. Subtrop Plant Sci 59:30–37Google Scholar
  40. 40.
    Nakabachi A, Koshikawa S, Miura T, Miyagishima S (2010) Genome size of Pachypsylla venusta (Hemiptera: Psyllidae) and the ploidy of its bacteriocyte, the symbiotic host cell that harbors intracellular mutualistic bacteria with the smallest cellular genome. Bull Entomol Res 100:27–33PubMedCrossRefGoogle Scholar
  41. 41.
    Reineke A, Karlovsky P, Zebitz CPW (1998) Preparation and purification of DNA from insects for AFLP analysis. Insect Mol Biol 7:95–99PubMedCrossRefGoogle Scholar
  42. 42.
    Saito N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425Google Scholar
  43. 43.
    Secor GA, Rivera VV, Abad JA, Lee I-M, Clover GRG, Liefting LW, Li X, De Boer SH (2009) Association of ‘Candidatus Liberibacter solanacearum’ with zebra chip disease of potato established by graft and psyllid transmission, electron microscopy, and PCR. Plant Dis 93:574–583CrossRefGoogle Scholar
  44. 44.
    Sengoda VG, Munyaneza JE, Crosslin JM, Buchman JL, Pappu HR (2010) Phenotypic and etiological differences between psyllid yellows and zebra chip diseases of potato. Am J Pot Res 87:41–49CrossRefGoogle Scholar
  45. 45.
    Shi W, Syrenne R, Sun J-Z, Yuan JS (2010) Molecular approaches to study the insect gut symbiotic microbiota at the ‘omics’ age. Insect Sci 17:199–219CrossRefGoogle Scholar
  46. 46.
    Shigenobu S, Watanabe H, Hattori M, Sakaki Y, Ishikawa H (2000) Genome sequence of the endocellular bacterial symbiont of aphids Buchnera sp. Nature 407:81–86PubMedCrossRefGoogle Scholar
  47. 47.
    Subandiyah SN, Nikoh N, Tsuyumu S, Somowiyarjo S, Fukatsu T (2000) Complex endosymbiotic microbiota of the citrus psyllid Diaphorina citri (Homoptera: Psylloidea). Zool Sci 17:983–989CrossRefGoogle Scholar
  48. 48.
    Tamames J, Gil R, Latorre A, Peretó J, Silva FJ, Moya A (2007) The frontier between cell and organelle: genome analysis of Candidatus Carsonella ruddii. BMC Evol Biol 7:181–188PubMedCrossRefGoogle Scholar
  49. 49.
    Tamura K, Dudley J, Nei M, Kumar S (2007) MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599PubMedCrossRefGoogle Scholar
  50. 50.
    Thao ML, Clark MA, Baumann L, Brennan EB, Moran NA, Baumann P (2000) Secondary endosymbionts of psyllids have been acquired multiple times. Curr Microbiol 41:300–304PubMedCrossRefGoogle Scholar
  51. 51.
    Thao ML, Moran NA, Abbot P, Brennan EB, Burckhardt DH, Baumann P (2000) Cospeciation of psyllids and their prokaryotic primary endosymbionts. Appl Environ Microbiol 2898–2905Google Scholar
  52. 52.
    Tyler HL, Roesch LFW, Gowda S, Dawson WO, Triplett EW (2009) Confirmation of the sequence of ‘Candidatus Liberibacter asiaticus’ and assessment of microbial diversity in Huanglongbing-infected citrus phloem using a metagenomic approach. Mol Plant Microbe Interact 22(12):1624–1634PubMedCrossRefGoogle Scholar
  53. 53.
    Wolcott RD, Gontcharova V, Sun Y, Zischkau AM, Dowd SE (2009) Bacterial diversity in surgical site infections: not just aerobic cocci any more. J Wound Care 18:317–323PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Punya Nachappa
    • 1
  • Julien Levy
    • 2
  • Elizabeth Pierson
    • 2
  • Cecilia Tamborindeguy
    • 1
    Email author
  1. 1.Department of EntomologyTexas A&M UniversityCollege stationUSA
  2. 2.Department of Horticultural SciencesTexas A&M UniversityCollege stationUSA

Personalised recommendations