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Synchronous development of Eimeria tenella in chicken caeca and utility of laser microdissection for purification of single stage schizont RNA

Published online by Cambridge University Press:  20 August 2012

M. MATSUBAYASHI
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
T. HATTA
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
T. MIYOSHI
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
M. A. ALIM
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
K. YAMAJI
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
K. SHIMURA
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
T. ISOBE
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan
N. TSUJI*
Affiliation:
National Institute of Animal Health, National Agricultural and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan Department of Global Agricultural Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan
*
*Corresponding author: Laboratory of Parasitic Diseases, National Institute of Animal Health, National Agricultural and Food Research Organization, 3-1-5, Kannondai, Tsukuba, Ibaraki 305-0856, Japan. Tel: +81 29 838 7749. Fax: +81 29 838 7749. E-mail: tsujin@affrc.go.jp.

Summary

Eimeria tenella is recognized worldwide as a significant pathogen in the poultry industry. However, a lack of methods for isolating developing schizonts has hindered the use of transcriptome analyses to discover novel and developmentally regulated genes. In the present study, we characterized the long-term successive development of E. tenella in infected chicken caeca and assessed the utility of laser microdissection (LMD) for the isolation of schizont RNA. Developmental stages, including those of the first, second, and third-generation schizonts and gametocytes, were synchronous. Using LMD, only the mature second-generation schizonts were successfully excised from the lamina propria, and non-degraded RNA was purified from the schizonts. E. tenella-specific genes were amplified by reverse transcription polymerase chain reaction (RT-PCR). These results augment our understanding of the E. tenella life cycle, and reveal LMD as a potentially useful tool for gene expression analyses of the intracellular stages of E. tenella.

Type
Research Article
Copyright
Copyright © Cambridge University Press 2012

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References

REFERENCES

Amiruddin, N., Lee, X. W., Blake, D. P., Suzuki, Y., Tay, Y. L., Lim, L. S., Tomley, F. M., Watanabe, J., Sugimoto, C. and Wan, K. L. (2012). Characterisation of full-length cDNA sequences provides insights into the Eimeria tenella transcriptome. BMC Genomics 13, 21.CrossRefGoogle ScholarPubMed
Ball, S. J., Daszak, P., Pittilo, R. M. and Norton, C. C. (1995). Ultrastructural observations on the third-generation merozoites of Eimeria tenella in chicks. Acta Veterinaria Hungarica 43, 139144.Google ScholarPubMed
Bonner, R. F., Emmert-Buck, M., Cole, K., Pohida, T., Chuaqui, R., Goldstein, S. and Liotta, L. A. (1997). Laser capture microdissection: molecular analysis of tissue. Science 278, 14811483.CrossRefGoogle ScholarPubMed
Chapman, H. D. and Shirley, M. W. (2003). The Houghton strain of Eimeria tenella: a review of the type strain selected for genome sequencing. Avian Pathology 32, 115127.CrossRefGoogle ScholarPubMed
Daszak, P., Ball, S. J., Pittilo, R. M. and Norton, C. C. (1993). Ultrastructural observations on caecal epithelial cells invaded by first-generation merozoites of Eimeria tenella in vivo. Annals of Tropical Medicine and Parasitology 87, 359364.CrossRefGoogle ScholarPubMed
Ding, X., Lillehoj, H. S., Dalloul, R. A., Min, W., Sato, T., Yasuda, A. and Lillehoj, E. P. (2005) In ovo vaccination with the Eimeria tenella EtMIC2 gene induces protective immunity against coccidiosis. Vaccine 23, 37333740.CrossRefGoogle ScholarPubMed
Emmert-Buck, M. R., Bonner, R. F., Smith, P. D., Chuaqui, R. F., Zhuang, Z., Goldstein, S. R., Weiss, R. A. and Liotta, L. A. (1996). Laser capture microdissection. Science 274, 9981001.CrossRefGoogle ScholarPubMed
Fernando, M. A. (1982). Pathology and pathogenicity. In The Biology of the Coccidia (ed. Long, P. L.), pp. 287327. University Park Press, London, UK.Google Scholar
Geysen, J., Ausma, J. and Vanden Bossche, H. (1991). Simultaneous purification of merozoites and schizonts of Eimeria tenella (Apicomplexa) by Percoll flotation and assessment of cell viability with a double fluorescent dye assay. Journal of Parasitology 77, 989993.CrossRefGoogle ScholarPubMed
Jones, M. K., Higgins, T., Stenzel, D. J. and Gobert, G. N. (2007). Towards tissue specific transcriptomics and expression pattern analysis in schistosomes using laser microdissection microscopy. Experimental Parasitology 117, 259266.CrossRefGoogle ScholarPubMed
Kheysin, Y. M. (1972). Species properties of coccidian of domestic animals. In Life Cycles of Coccidia of Domestic Animals (ed. Todd, K. S. Jr.), pp. 218228. University Park Press, Baltimore, MD, USA.Google Scholar
Levine, N. D. (1973). The Apicomplexa and the coccidian proper. In Protozoan Parasites of Domestic Animals and of Man, 2nd Edn, pp. 197200. Burgess Publishing Company, Minneapolis, MN, USA.Google Scholar
Li, L., Brunk, B. P., Kissinger, J. C., Pape, D., Tang, K., Cole, R. H., Martin, J., Wylie, T., Dante, M., Fogarty, S. J., Howe, D. K., Liberator, P., Diaz, C., Anderson, J., White, M., Jerome, M. E.Johnson, E. A., Radke, J. A., Stoeckert, C. J. Jr., Waterston, R. H., Clifton, S. W., Roos, D. S. and Sibley, L. D. (2003). Gene discovery in the apicomplexa as revealed by EST sequencing and assembly of a comparative gene database. Genome Research 13, 443454.CrossRefGoogle ScholarPubMed
McDonald, V. and Shirley, M. W. (1987). The endogenous development of virulent strains and attenuated precocious lines of Eimeria tenella and E. necatrix. Journal of Parasitology 73, 993997.CrossRefGoogle ScholarPubMed
McDonald, V. and Rose, M. E. (1987). Eimeria tenella and E. necatrix: a third generation of schizogony is an obligatory part of the developmental cycle. Journal of Parasitology 73, 617622.CrossRefGoogle Scholar
Miska, K. B., Fetterer, R. H. and Rosenberg, G. H. (2008). Analysis of transcripts from intracellular stages of Eimeria acervulina using expressed sequence tags. Journal of Parasitology 94, 462466.CrossRefGoogle ScholarPubMed
Ng, S. T., Sanusi Jangi, M., Shirley, M. W., Tomley, F. M. and Wan, K. L. (2002). Comparative EST analyses provide insights into gene expression in two asexual developmental stages of Eimeria tenella. Experimental Parasitology 101, 168173.CrossRefGoogle ScholarPubMed
Novaes, J., Rangel, L. T., Ferro, M., Abe, R. Y., Manha, A. P., de Mello, J. C., Varuzza, L., Durham, A. M., Madeira, A. M. and Gruber, A. (2012). A comparative transcriptome analysis reveals expression profiles conserved across three Eimeria spp. of domestic fowl and associated with multiple developmental stages. International Journal for Parasitology 42, 3948.CrossRefGoogle ScholarPubMed
Ortega-Mora, L. M., Troncoso, J. M., Rojo-Vázquez, F. A. and Gómez-Bautista, M. (1992). Cross-reactivity of polyclonal serum antibodies generated against Cryptosporidium parvum oocysts. Infection and Immunity 60, 34423445.CrossRefGoogle ScholarPubMed
Otto, T. D., Wilinski, D., Assefa, S., Keane, T. M., Sarry, L. R., Böhme, U., Lemieux, J., Barrell, B., Pain, A., Berriman, M., Newbold, C. and Llinás, M. (2010). New insights into the blood-stage transcriptome of Plasmodium falciparum using RNA-Seq. Molecular Microbiology 76, 1224.CrossRefGoogle ScholarPubMed
Ouarzane, M., Labbé, M. and Péry, P. (1998). Purification of first-generation Eimeria tenella schizonts. Journal of Parasitology 84, 10271031.CrossRefGoogle ScholarPubMed
Radke, J. R., Behnke, M. S., Mackey, A. J., Radke, J. B., Roos, D. S. and White, M. W. (2005). The transcriptome of Toxoplasma gondii. BMC Biology 3, 26.CrossRefGoogle ScholarPubMed
Roberts, L., Bowers, J., Sensinger, K., Lisowski, A., Getts, R. and Anderson, M. G. (2009). Identification of methods for use of formalin-fixed, paraffin-embedded tissue samples in RNA expression profiling. Genomics 94, 341348.CrossRefGoogle ScholarPubMed
Ryan, R., Shirley, M. and Tomley, F. (2000). Mapping and expression of microneme genes in Eimeria tenella. International Journal for Parasitology 30, 14931499.CrossRefGoogle ScholarPubMed
Sacci, J. B. Jr., Ribeiro, J. M., Huang, F., Alam, U., Russell, J. A., Blair, P. L., Witney, A., Carucci, D. J., Azad, A. F. and Aguiar, J. C. (2005). Transcriptional analysis of in vivo Plasmodium yoelii liver stage gene expression. Molecular and Biochemical Parasitology 142, 177183.CrossRefGoogle ScholarPubMed
Schaap, D., Arts, G., van Poppel, N. F. and Vermeulen, A. N. (2005). De novo ribosome biosynthesis is transcriptionally regulated in Eimeria tenella, dependent on its life cycle stage. Molecular and Biochemical Parasitology 139, 239248.CrossRefGoogle ScholarPubMed
Semblat, J. P., Silvie, O., Franetich, J. F., Hannoun, L., Eling, W. and Mazier, D. (2002). Laser capture microdissection of Plasmodium falciparum liver stages for mRNA analysis. Molecular and Biochemical Parasitology 121, 179183.CrossRefGoogle ScholarPubMed
Vaitukaitis, J. L. (1981). Production of antisera with small doses of immunogen: multiple intradermal injections. Methods in Enzymology 73, 4652.CrossRefGoogle ScholarPubMed
Wan, K. L., Chong, S. P., Ng, S. T., Shirley, M. W., Tomley, F. M. and Jangi, M. S. (1999). A survey of genes in Eimeria tenella merozoites by EST sequencing. International Journal for Parasitology 29, 18851892.CrossRefGoogle ScholarPubMed