Parasitology Research

, Volume 116, Issue 5, pp 1573–1579 | Cite as

Widespread 5-methylcytosine in the genomes of avian Coccidia and other apicomplexan parasites detected by an ELISA-based method

  • Zhenxing Gong
  • Hao Yin
  • Xueting Ma
  • Baohong Liu
  • Zhenglan Han
  • Lingqiao Gou
  • Jianping Cai
Original Paper

Abstract

To date, little is known about cytosine methylation in the genomic DNA of apicomplexan parasites, although it has been confirmed that this important epigenetic modification exists in many lower eukaryotes, plants, and animals. In the present study, ELISA-based detection demonstrated that low levels of 5-methylcytosine (5-mC) are present in Eimeria spp., Toxoplasma gondii, Cryptosporidium spp., and Neospora caninum. The proportions of 5-mC in genomic DNA were 0.18 ± 0.02% in E tenella sporulated oocysts, 0.19 ± 0.01% in E. tenella second-generation merozoites, 0.22 ± 0.04% in T. gondii tachyzoites, 0.28 ± 0.03% in N. caninum tachyzoites, and 0.06 ± 0.01, 0.11 ± 0.01, and 0.09 ± 0.01% in C. andersoni, C. baileyi, and C. parvum sporulated oocysts, respectively. In addition, we found that the percentages of 5-mC in E. tenella varied considerably at different life stages, with sporozoites having the highest percentage of 5-mC (0.78 ± 0.10%). Similar stage differences in 5-mC were also found in E. maxima, E. necatrix, and E. acervulina, the levels of 5-mC in their sporozoites being 4.3-, 1.8-, 2.5-, and 2.0-fold higher than that of sporulated oocysts, respectively (p < 0.01). Furthermore, a total DNA methyltransferase-like activity was detected in whole cell extracts prepared from E. tenella sporozoites. In conclusion, genomic DNA methylation is present in these apicomplexan parasites and may play a role in the stage conversion of Eimeria.

Keywords

Eimeria spp. Apicomplexan parasites 5-Methylcytosine DNA methyltransferase-like activity Genomic DNA 

References

  1. Blake DP, Smith AL, Shirley MW (2003) Amplified fragment length polymorphism analyses of Eimeria spp.: an improved process for genetic studies of recombinant parasites. Parasitol Res 90:473–475CrossRefPubMedGoogle Scholar
  2. Bollati V, Schwartz J, Wright R, Litonjua A, Tarantini L, Suh H, Sparrow D, Vokonas P, Baccarelli A (2009) Decline in genomic DNA methylation through aging in a cohort of elderly subjects. Mech Ageing Dev 130:234–239CrossRefPubMedGoogle Scholar
  3. Braun R, Shirley MW (1995) Eimeria species and strains of chicken. In: Eckert J, Braun R, Shirley MW, Coudert P (eds) Guidelines on techniques in coccidiosis research. European Commission, Luxemburg, pp 146–147Google Scholar
  4. Capuano F, Mulleder M, Kok R, Blom HJ, Ralser M (2014) Cytosine DNA methylation is found in Drosophila melanogaster but absent in Saccharomyces cerevisiae, Schizosaccharomyces pombe, and other yeast species. Anal Chem 86:3697–3702CrossRefPubMedPubMedCentralGoogle Scholar
  5. Choi SW, Keyes MK, Horrocks P (2006) LC/ESI-MS demonstrates the absence of 5-methyl-2′-deoxycytosine in Plasmodium falciparum genomic DNA. Mol Biochem Parasitol 150:350–352CrossRefPubMedGoogle Scholar
  6. Feng S, Cokus SJ, Zhang X, Chen PY, Bostick M, Goll MG, Hetzel J, Jain J, Strauss SH, Halpern ME, Ukomadu C, Sadler KC, Pradhan S, Pellegrini M, Jacobsen SE (2010) Conservation and divergence of methylation patterning in plants and animals. Proc Natl Acad Sci U S A 107:8689–8694CrossRefPubMedPubMedCentralGoogle Scholar
  7. Field LM, Lyko F, Mandrioli M, Prantera G (2004) DNA methylation in insects. Insect Mol Biol 13:109–115CrossRefPubMedGoogle Scholar
  8. Fisher O, Siman-Tov R, Ankri S (2004) Characterization of cytosine methylated regions and 5-cytosine DNA methyltransferase (Ehmeth) in the protozoan parasite Entamoeba histolytica. Nucleic Acids Res 32:287–297CrossRefPubMedPubMedCentralGoogle Scholar
  9. Fisher O, Siman-Tov R, Ankri S (2006) Pleiotropic phenotype in Entamoeba histolytica overexpressing DNA methyltransferase (Ehmeth). Mol Biochem Parasitol 147:48–54CrossRefPubMedGoogle Scholar
  10. Gao F, Liu X, Wu XP, Wang XL, Gong D, Lu H, Xia Y, Song Y, Wang J, Du J, Liu S, Han X, Tang Y, Yang H, Jin Q, Zhang X, Liu M (2012) Differential DNA methylation in discrete developmental stages of the parasitic nematode Trichinella spiralis. Genome Biol 13:R100–R113CrossRefPubMedPubMedCentralGoogle Scholar
  11. Geyer KK, Chalmers IW, Mackintosh N, Hirst JE, Geoghegan R, Badets M, Brophy PM, Brehm K, Hoffmann KF (2013) Cytosine methylation is a conserved epigenetic feature found throughout the phylum Platyhelminthes. BMC Genomics 14:462–475CrossRefPubMedPubMedCentralGoogle Scholar
  12. Geyer KK, Rodriguez Lopez CM, Chalmers IW, Munshi SE, Truscott M, Heald J, Wilkinson MJ, Hoffmann KF (2011) Cytosine methylation regulates oviposition in the pathogenic blood fluke Schistosoma mansoni. Nat Commun 2:424–434CrossRefPubMedPubMedCentralGoogle Scholar
  13. Gissot M, Choi SW, Thompson RF, Greally JM, Kim K (2008) Toxoplasma gondii and Cryptosporidium parvum lack detectable DNA cytosine methylation. Eukaryot Cell 7:537–540CrossRefPubMedPubMedCentralGoogle Scholar
  14. Gissot M, Kelly KA, Ajioka JW, Greally JM, Kim K (2007) Epigenomic modifications predict active promoters and gene structure in Toxoplasma gondii. PLoS Pathog 3:e77. doi:10.1371/journal.ppat.0030077 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Glastad KM, Hunt BG, Yi SV, Goodisman MA (2011) DNA methylation in insects: on the brink of the epigenomic era. Insect Mol Biol 20:553–565CrossRefPubMedGoogle Scholar
  16. Goll MG, Bestor TH (2005) Eukaryotic cytosine methyltransferases. Annu Rev Biochem 74:481–514CrossRefPubMedGoogle Scholar
  17. Gowher H, Ehrlich KC, Jeltsch A (2001) DNA from Aspergillus flavus contains 5-methylcytosine. FEMS Microbiol Lett 205:151–155CrossRefPubMedGoogle Scholar
  18. Hahn MA, Pfeifer GP (2010) Methods for genome-wide analysis of DNA methylation in intestinal tumors. Mutat Res 693:77–83CrossRefPubMedGoogle Scholar
  19. Hakimi MA, Deitsch KW (2007) Epigenetics in Apicomplexa: control of gene expression during cell cycle progression, differentiation and antigenic variation. Curr Opin Microbiol 10:357–362CrossRefPubMedGoogle Scholar
  20. Hu CW, Chen JL, Hsu YW, Yen CC, Chao MR (2015) Trace analysis of methylated and hydroxymethylated cytosines in DNA by isotope-dilution LC-MS/MS: first evidence of DNA methylation in Caenorhabditis elegans. Biochem J 465:39–47CrossRefPubMedGoogle Scholar
  21. Jaenisch R, Bird A (2003) Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nat Genet 33(Suppl):245–254CrossRefPubMedGoogle Scholar
  22. Katrib M, Ikin RJ, Brossier F, Robinson M, Slapetova I, Sharman PA, Walker RA, Belli SI, Tomley FM, Smith NC (2012) Stage-specific expression of protease genes in the apicomplexan parasite, Eimeria tenella. BMC Genomics 13:685–703CrossRefPubMedPubMedCentralGoogle Scholar
  23. Lee TF, Zhai J, Meyers BC (2010) Conservation and divergence in eukaryotic DNA methylation. Proc Natl Acad Sci U S A 107:9027–9028CrossRefPubMedPubMedCentralGoogle Scholar
  24. Liu SY, Lin JQ, Wu HL, Wang CC, Huang SJ, Luo YF, Sun JH, Zhou JX, Yan SJ, He JG, Wang J, He ZM (2012) Bisulfite sequencing reveals that Aspergillus flavus holds a hollow in DNA methylation. PLoS One 7:e30349. doi:10.1371/journal.pone.0030349 CrossRefPubMedPubMedCentralGoogle Scholar
  25. Lou J, Wang Y, Yao C, Jin L, Wang X, Xiao Y, Wu N, Song P, Song Y, Tan Y, Gao M, Liu K, Zhang X (2013) Role of DNA methylation in cell cycle arrest induced by Cr (VI) in two cell lines. PLoS One 8:e71031. doi:10.1371/journal.pone.0071031 CrossRefPubMedPubMedCentralGoogle Scholar
  26. Militello KT, Wang P, Jayakar SK, Pietrasik RL, Dupont CD, Dodd K, King AM, Valenti PR (2008) African trypanosomes contain 5-methylcytosine in nuclear DNA. Eukaryot Cell 7(11):2012–2016CrossRefPubMedPubMedCentralGoogle Scholar
  27. Motohashi K (2015) A simple and efficient seamless DNA cloning method using SLiCE from Escherichia coli laboratory strains and its application to SLiP site-directed mutagenesis. BMC Biotechnol 15:47–56CrossRefPubMedPubMedCentralGoogle Scholar
  28. Osabe K, Clement JD, Bedon F, Pettolino FA, Ziolkowski L, Llewellyn DJ, Finnegan EJ, Wilson IW (2014) Genetic and DNA methylation changes in cotton (Gossypium) genotypes and tissues. PLoS One 9:e86049. doi:10.1371/journal.pone.0086049 CrossRefPubMedPubMedCentralGoogle Scholar
  29. Padmanabhan N, Jia D, Geary-Joo C, Wu X, Ferguson-Smith AC, Fung E, Bieda MC, Snyder FF, Gravel RA, Cross JC, Watson ED (2013) Mutation in folate metabolism causes epigenetic instability and transgenerational effects on development. Cell 155:81–93CrossRefPubMedGoogle Scholar
  30. Pegoraro M, Bafna A, Davies NJ, Shuker DM, Tauber E (2016) DNA methylation changes induced by long and short photoperiods in Nasonia. Genome Res 26:203–210CrossRefPubMedPubMedCentralGoogle Scholar
  31. Ponts N, Fu L, Harris EY, Zhang J, Chung DW, Cervantes MC, Prudhomme J, Atanasova-Penichon V, Zehraoui E, Bunnik EM, Rodrigues EM, Lonardi S, Hicks GR, Wang Y, Le Roch KG (2013) Genome-wide mapping of DNA methylation in the human malaria parasite Plasmodium falciparum. Cell Host Microbe 14:696–706CrossRefPubMedPubMedCentralGoogle Scholar
  32. Raddatz G, Guzzardo PM, Olova N, Fantappie MR, Rampp M, Schaefer M, Reik W, Hannon GJ, Lyko F (2013) Dnmt2-dependent methylomes lack defined DNA methylation patterns. Proc Natl Acad Sci U S A 110:8627–8631CrossRefPubMedPubMedCentralGoogle Scholar
  33. Schulz EC, Roth HM, Ankri S, Ficner R (2012) Structure analysis of Entamoeba histolytica DNMT2 (EhMeth). PLoS One 7:e38728. doi:10.1371/journal.pone.0038728 CrossRefPubMedPubMedCentralGoogle Scholar
  34. Shirley MW (1995) Eimeria species and strains of chicken. In: Eckert J, Braun R, Shirley MW, Coudert P (eds) Guidelines on techniques in coccidiosis research. European Commission, Luxemburg, pp 1–24Google Scholar
  35. Simpson VJ, Johnson TE, Hammen RF (1986) Caenorhabditis elegans DNA does not contain 5-methylcytosine at any time during development or aging. Nucleic Acids Res 14:6711–6719CrossRefPubMedPubMedCentralGoogle Scholar
  36. Suzuki MM, Bird A (2008) DNA methylation landscapes: provocative insights from epigenomics. Nat Rev Genet 9:465–476CrossRefPubMedGoogle Scholar
  37. Takayama S, Dhahbi J, Roberts A, Mao G, Heo SJ, Pachter L, Martin DI, Boffelli D (2014) Genome methylation in D. melanogaster is found at specific short motifs and is independent of DNMT2 activity. Genome Res 24:821–830CrossRefPubMedPubMedCentralGoogle Scholar
  38. Tang Y, Gao XD, Wang Y, Yuan BF, Feng YQ (2012) Widespread existence of cytosine methylation in yeast DNA measured by gas chromatography/mass spectrometry. Anal Chem 84:7249–7255CrossRefPubMedPubMedCentralGoogle Scholar
  39. Tomley F (1997) Techniques for isolation and characterization of apical organelles from Eimeria tenella sporozoites. Methods 13:171–176CrossRefPubMedGoogle Scholar
  40. Xie M, Gilbert JM, McDougald LR (1992) Electrophoretic and immunologic characterization of proteins of merozoites of Eimeria acervulina, E. maxima, E. necatrix, and E. tenella. J Parasitol 78:82–86CrossRefPubMedGoogle Scholar
  41. Yi S (2012) Birds do it, bees do it, worms and ciliates do it too: DNA methylation from unexpected corners of the tree of life. Genome Biol 13:174–176CrossRefPubMedPubMedCentralGoogle Scholar
  42. Zhao X, Duszynski DW, Loker ES (2001) A simple method of DNA extraction for Eimeria species. J Microbiol Methods 44:131–137CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2017

Authors and Affiliations

  • Zhenxing Gong
    • 1
    • 2
  • Hao Yin
    • 1
    • 2
  • Xueting Ma
    • 1
    • 2
  • Baohong Liu
    • 1
    • 2
  • Zhenglan Han
    • 1
    • 2
  • Lingqiao Gou
    • 1
    • 2
  • Jianping Cai
    • 1
    • 2
  1. 1.State Key Laboratory of Veterinary Etiological Biology; Key Laboratory of Veterinary Parasitology of Gansu ProvinceLanzhou Veterinary Research Institute, Chinese Academy of Agricultural ScienceLanzhouPeople’s Republic of China
  2. 2.Jiangsu Co-Innovation Center for Prevention and Control of Animal Infectious Diseases and ZoonosesJiangsu ProvincePeople’s Republic of China

Personalised recommendations