Malaria pp 385-400 | Cite as

Quantification of Sporozoite Invasion, Migration, and Development by Microscopy and Flow Cytometry

  • Photini Sinnis
  • Patricia De La Vega
  • Alida Coppi
  • Urszula Krzych
  • Maria M. Mota
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 923)

Abstract

There is an important role for in vitro assays to better understand the initial steps of malaria infection. In this section, we describe both microscopy-based and flow cytometry-based sporozoite invasion, migration and development assays with the rodent malaria parasites, Plasmodium berghei and Plasmodium yoelii, and the human malaria parasite, Plasmodium falciparum.

Key words

Invasion Attachment Cell traversal Exoerythrocytic stages Sporozoites Malaria Plasmodium Flow cytometry 

References

  1. 1.
    Hollingdale MR et al (1983) Entry of Plasmodium berghei sporozoites into cultured cells and their transformation into trophozoites. Am J Trop Med Hyg 32:685–690PubMedGoogle Scholar
  2. 2.
    Hollingdale MR et al (1983) In vitro cultivation of the exoerythrocytic stage of Plasmodium berghei in a hepatoma cell line. Am J Trop Med Hyg 32:682–684PubMedGoogle Scholar
  3. 3.
    Mazier D et al (1985) Complete development of hepatic stages of Plasmodium falciparum in vitro. Science 227:440–442PubMedCrossRefGoogle Scholar
  4. 4.
    Renia L et al (1988) Malaria sporozoite penetration: a new approach by double staining. J Immunol Meth 112:201–205CrossRefGoogle Scholar
  5. 5.
    Mota M et al (2001) Migration of Plasmodium sporozoites through cells before infection. Science 291:141–144PubMedCrossRefGoogle Scholar
  6. 6.
    VanBuskirk KM et al (2009) Preerythrocytic, live-attenuated Plasmodium falciparum vaccine candidates by design. Proc Natl Acad Sci USA 106:13004–13009PubMedCrossRefGoogle Scholar
  7. 7.
    Prudencio M et al (2008) Dissecting in vitro host cell infection by Plasmodium sporozoites using flow cytometry. Cell Microbiol 10:218–224PubMedGoogle Scholar
  8. 8.
    Coppi A et al (2011) The malaria circumsporozoite protein has two functional domains, each with distinct roles as sporozoites journey from mosquito to mammalian host. J Exp Med 208:341–356PubMedCrossRefGoogle Scholar
  9. 9.
    Yu M et al (2008) The fatty acid biosynthesis enzyme FabI plays a key role in the development of liver stage malarial parasites. Cell Host Microbe 4:567–578PubMedCrossRefGoogle Scholar
  10. 10.
    Haussig JM et al (2011) Inactivation of a Plasmodium apicoplast protein attenuates formation of liver merozoites. Mol Microbiol 81:1511–1525PubMedCrossRefGoogle Scholar
  11. 11.
    Mueller AK et al (2005) Plasmodium liver stage developmental arrest by depletion of a protein at the parasite-host interface. Proc Natl Acad Sci USA 102:3022–3027PubMedCrossRefGoogle Scholar
  12. 12.
    Hobbs CV et al (2009) HIV protease inhibitors inhibit the development of preerythrocytic-stage Plasmodium parasites. J Infect Dis 199:134–141PubMedCrossRefGoogle Scholar
  13. 13.
    Parvanova I et al (2009) A small molecule inhibitor of signal peptide peptidase inhibits Plasmodium development in the liver and decreases malaria severity. PLoS One 4:e5078PubMedCrossRefGoogle Scholar
  14. 14.
    Albuquerque SS et al (2009) Host cell transcriptional profiling during malaria liver stage infection reveals a coordinated and sequential set of biological events. BMC Genomics 10:270PubMedCrossRefGoogle Scholar
  15. 15.
    Rodrigues CD et al (2008) Host scavenger receptor SR-B1 plays a dual role in the establishment of malaria parasite liver infection. Cell Host Microbe 4:271–282PubMedCrossRefGoogle Scholar
  16. 16.
    Prudencio M et al (2008) Kinome-wide RNAi screen implicates at least 5 host hepatocyte kinases in Plasmodium sporozoite infection. PLoS Pathog 4:e1000201PubMedCrossRefGoogle Scholar
  17. 17.
    Pinzon-Ortiz C et al (2001) The binding of the circumsporozoite protein to cell surface heparan sulfate proteoglycans is required for Plasmodium sporozoite attachment to cells. J Biol Chem 276:26784–26791PubMedCrossRefGoogle Scholar
  18. 18.
    Gego A et al (2006) New approach for high-throughput screening of drug activity on Plasmodium liver stages. Antimicrob Agents Chemother 50:1586–1589PubMedCrossRefGoogle Scholar
  19. 19.
    Mueller AK et al (2005) Genetically modified Plasmodium parasites as a protective experimental malaria vaccine. Nature 433:164–167PubMedCrossRefGoogle Scholar
  20. 20.
    van Dijk MR et al (2005) Genetically attenuated, P36p-deficient malarial sporozoites induce protective immunity and apoptosis of infected liver cells. Proc Natl Acad Sci USA 102:12194–12199PubMedCrossRefGoogle Scholar
  21. 21.
    Franke-Fayard B et al (2004) A Plasmodium berghei reference line that constitutively expresses GFP at a high level throughout the complete life cycle. Mol Biochem Parasitol 137:23–33PubMedCrossRefGoogle Scholar
  22. 22.
    Tarun AS et al (2006) Quantitative isolation and in vivo imaging of malaria parasite liver stages. Int J Parasitol 36:1283–1293PubMedCrossRefGoogle Scholar
  23. 23.
    Talman AM et al (2010) A Plasmodium falciparum strain expressing GFP throughout the parasite’s life-cycle. PLoS One 5:e9156PubMedCrossRefGoogle Scholar
  24. 24.
    Ishino T et al (2004) Cell-passage activity is required for the malarial parasite to cross the liver sinusoidal cell layer. PLoS Biol 2:77–84CrossRefGoogle Scholar
  25. 25.
    Amino R et al (2008) Host cell traversal is important for progression of the malaria parasite through the dermis to the liver. Cell Host Microbe 3:88–96PubMedCrossRefGoogle Scholar
  26. 26.
    Moreira CK et al (2008) The Plasmodium TRAP/MIC2 family member, TRAP-Like Protein (TLP), is involved in tissue traversal by sporozoites. Cell Microbiol 10:1505–1516PubMedCrossRefGoogle Scholar
  27. 27.
    Combe A et al (2009) Clonal conditional mutagenesis in malaria parasites. Cell Host Microbe 5:386–396PubMedCrossRefGoogle Scholar
  28. 28.
    Falae A et al (2010) Role of Plasmodium berghei cGMP-dependent protein kinase in late liver stage development. J Biol Chem 285:3282–3288PubMedCrossRefGoogle Scholar
  29. 29.
    Calvo-Calle JM et al (1994) In vitro development of infectious liver stages of P. yoelii and P. berghei malaria in human cell lines. Exp Parasitol 79:362–373PubMedCrossRefGoogle Scholar
  30. 30.
    Silvie O et al (2003) Hepatocyte CD81 is required for Plasmodium falciparum and Plasmodium yoelii sporozoite infectivity. Nat Med 9:93–96PubMedCrossRefGoogle Scholar
  31. 31.
    Sattabongkot J et al (2006) Establishment of a human hepatocyte line that supports in vitro development of the exo-erythrocytic stages of the malaria parasites Plasmodium falciparum and P. vivax. Am J Trop Med Hyg 74:708–715PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Photini Sinnis
    • 1
  • Patricia De La Vega
    • 2
  • Alida Coppi
    • 3
  • Urszula Krzych
    • 2
  • Maria M. Mota
    • 4
  1. 1.Department of Molecular Microbiology and ImmunologyJohn Hopkins Bloomberg School of Public HealthBaltimoreUSA
  2. 2.Department of Cellular ImmunologyWalter Reed Army Institute of ResearchSilver SpringUSA
  3. 3.Regeneron PharmaceuticalsTarrytownUSA
  4. 4.Malaria UnitInstituto de Medicina MolecularLisboaPortugal

Personalised recommendations