Skip to main content
SpringerLink
Log in

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

  • Original Paper
  • Published:
Parasitology Research Aims and scope Submit manuscript

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.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  • 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 90:473–475

    Article  CAS  PubMed  Google Scholar 

  • 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 130:234–239

    Article  CAS  PubMed  Google Scholar 

  • 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 146–147

    Google Scholar 

  • 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 86:3697–3702

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 150:350–352

    Article  CAS  PubMed  Google Scholar 

  • 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 107:8689–8694

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Field LM, Lyko F, Mandrioli M, Prantera G (2004) DNA methylation in insects. Insect Mol Biol 13:109–115

    Article  CAS  PubMed  Google Scholar 

  • 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 32:287–297

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Fisher O, Siman-Tov R, Ankri S (2006) Pleiotropic phenotype in Entamoeba histolytica overexpressing DNA methyltransferase (Ehmeth). Mol Biochem Parasitol 147:48–54

    Article  CAS  PubMed  Google Scholar 

  • 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–R113

    Article  PubMed  PubMed Central  Google Scholar 

  • 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 14:462–475

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 2:424–434

    Article  PubMed  PubMed Central  Google Scholar 

  • Gissot M, Choi SW, Thompson RF, Greally JM, Kim K (2008) Toxoplasma gondii and Cryptosporidium parvum lack detectable DNA cytosine methylation. Eukaryot Cell 7:537–540

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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

    Article  PubMed  PubMed Central  Google Scholar 

  • Glastad KM, Hunt BG, Yi SV, Goodisman MA (2011) DNA methylation in insects: on the brink of the epigenomic era. Insect Mol Biol 20:553–565

    Article  CAS  PubMed  Google Scholar 

  • Goll MG, Bestor TH (2005) Eukaryotic cytosine methyltransferases. Annu Rev Biochem 74:481–514

    Article  CAS  PubMed  Google Scholar 

  • Gowher H, Ehrlich KC, Jeltsch A (2001) DNA from Aspergillus flavus contains 5-methylcytosine. FEMS Microbiol Lett 205:151–155

    Article  CAS  PubMed  Google Scholar 

  • Hahn MA, Pfeifer GP (2010) Methods for genome-wide analysis of DNA methylation in intestinal tumors. Mutat Res 693:77–83

    Article  CAS  PubMed  Google Scholar 

  • Hakimi MA, Deitsch KW (2007) Epigenetics in Apicomplexa: control of gene expression during cell cycle progression, differentiation and antigenic variation. Curr Opin Microbiol 10:357–362

    Article  CAS  PubMed  Google Scholar 

  • 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 465:39–47

    Article  CAS  PubMed  Google Scholar 

  • Jaenisch R, Bird A (2003) Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nat Genet 33(Suppl):245–254

    Article  CAS  PubMed  Google Scholar 

  • 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 13:685–703

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lee TF, Zhai J, Meyers BC (2010) Conservation and divergence in eukaryotic DNA methylation. Proc Natl Acad Sci U S A 107:9027–9028

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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):2012–2016

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 15:47–56

    Article  PubMed  PubMed Central  Google Scholar 

  • 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

    Article  PubMed  PubMed Central  Google Scholar 

  • 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 155:81–93

    Article  CAS  PubMed  Google Scholar 

  • Pegoraro M, Bafna A, Davies NJ, Shuker DM, Tauber E (2016) DNA methylation changes induced by long and short photoperiods in Nasonia. Genome Res 26:203–210

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 14:696–706

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 110:8627–8631

    Article  PubMed  PubMed Central  Google Scholar 

  • 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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 1–24

    Google Scholar 

  • 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 14:6711–6719

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Suzuki MM, Bird A (2008) DNA methylation landscapes: provocative insights from epigenomics. Nat Rev Genet 9:465–476

    Article  CAS  PubMed  Google Scholar 

  • 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 24:821–830

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 84:7249–7255

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tomley F (1997) Techniques for isolation and characterization of apical organelles from Eimeria tenella sporozoites. Methods 13:171–176

    Article  CAS  PubMed  Google Scholar 

  • 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 78:82–86

    Article  CAS  PubMed  Google Scholar 

  • 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 13:174–176

    Article  PubMed  PubMed Central  Google Scholar 

  • Zhao X, Duszynski DW, Loker ES (2001) A simple method of DNA extraction for Eimeria species. J Microbiol Methods 44:131–137

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This research was supported by the National Natural Science Foundation of China (NSFC Grant No. 31272554) and the Agricultural Science and Technology Innovation Program (CAAS-ASTIP-2014-LVRI-09) to JPC. Authors are also indebted to Prof. Xingquan Zhu, Longxian Zhang, and Qun Liu for kindly providing the T. gondii, Cryptosporidium spp., and N. caninum genomic DNA.

Author information

Authors and Affiliations

  1. State Key Laboratory of Veterinary Etiological Biology; Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Science, Lanzhou, Gansu Province, 730046, People’s Republic of China

    Zhenxing Gong, Hao Yin, Xueting Ma, Baohong Liu, Zhenglan Han, Lingqiao Gou & Jianping Cai

  2. Jiangsu Co-Innovation Center for Prevention and Control of Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu Province, 225009, People’s Republic of China

    Zhenxing Gong, Hao Yin, Xueting Ma, Baohong Liu, Zhenglan Han, Lingqiao Gou & Jianping Cai

Authors
  1. Zhenxing Gong
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hao Yin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xueting Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Baohong Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Zhenglan Han
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Lingqiao Gou
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jianping Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

ZXG, HY, and XTM performed the experiments. BHL and ZLH analyzed the data. LQG contributed the reagents/materials. ZXG and HY wrote the paper. JPC designed the experiments and revised the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Jianping Cai.

Ethics declarations

Consent for publication

Not applicable.

Availability of data and materials

The dataset supporting the conclusions of this article is included within the article.

Conflict of interest

The authors declare that they have no competing interests.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Gong, Z., Yin, H., Ma, X. et al. Widespread 5-methylcytosine in the genomes of avian Coccidia and other apicomplexan parasites detected by an ELISA-based method. Parasitol Res 116, 1573–1579 (2017). https://doi.org/10.1007/s00436-017-5434-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00436-017-5434-x

Keywords

Search

Navigation