CONTINUOUS CULTURES OF PLASMODIUM FALCIPARUM ESTABLISHED IN TANZANIA FROM PATIENTS WITH ACUTE MALARIA In vitro Malaria cultures

Main Article Content

Florence Urio
Matilda Mkombachepa
Gration Rwegasira
Twilumba Makene
Billy Ngasala
Teddy Mselle
Julie Makani
Lucio Luzzatto

Keywords

Plasmodium falciparum, in vitro cultures, Albumax II, Human sera

Abstract

Background: Malaria morbidity and mortality, almost entirely from Plasmodium falciparum, are still rampant in Africa: therefore, it is important to study the biology of the parasite and the parasite-host cell interactions. In vitro cultivation of Plasmodium falciparum is most useful for this purpose, as well as for investigating drug resistance and possible new therapies. Here we report that the Trager & Jensen continuous culture of P. falciparum can be established in a laboratory in Tanzania with minimal facilities and with modest expenditure.


Methodology: This was an in-vitro set up of continuous culture of Plasmodium falciparum study, carried out in 2016-2020 at Muhimbili university of health and allied sciences, Dar-es salaam. Parasite samples were obtained from patients with acute malaria, frozen parasites, and live cultures. Data was collected and analyzed using GraphPad Prism version 8.


Results: We have successfully achieved exponential growth of existing strains that are used worldwide, as well as of parasites in clinical samples from patients with acute malaria. In the aim to optimize growth we have compared human serum and bovine serum albumin as components of the culture media. Additionally, culture synchronization has been achieved using sorbitol.


Conclusion: This experimental system is now available to our institution and to researchers aiming at investigating drug sensitivity and mechanisms of protection against Plasmodium falciparum that accrue from various genes expressed in red cells.


 

Downloads

Download data is not yet available.


Abstract 834
PDF Downloads 252
HTML Downloads 119

References

1. https://www.who.int/news-room/feature-stories/detail/world-malaria-report-2019
2. Trager W, Jensen J. Human malaria parasites in continuous culture. Science (80- ) [Internet]. 1976 Aug 20;193(4254):673–5. Available from: https://www.sciencemag.org/lookup/doi/10.1126/science.781840
3. Bhasin VK, Clayton C, Trager W, Cross GAM. Variations in the organization of repetitive DNA sequences in the genomes of Plasmodium falciparum clones. Mol Biochem Parasitol [Internet]. 1985 May;15(2):149–58. Available from: https://linkinghub.elsevier.com/retrieve/pii/0166685185901161
4. Corcoran LM, Forsyth KP, Bianco AE, Brown G V., Kemp DJ. Chromosome size polymorphisms in plasmodium falciparum can involve deletions and are frequent in natural parasite populations. Cell [Internet]. 1986 Jan;44(1):87–95. Available from: https://linkinghub.elsevier.com/retrieve/pii/0092867486904873
5. Summary of discussions on in vitro cultivation of malaria parasites. Bull World Health Organ. 1977;55(2–3):411–9.
6. Desai SA. Insights gained from P. falciparum cultivation in modified media. Sci World J. 2013;2013.
7. Ringwald P, Meche FS, Bickii J, Basco LK. In vitro culture and drug sensitivity assay of Plasmodium falciparum with nonserum substitute and acute-phase sera. J Clin Microbiol. 1999;37(3):700–5.
8. Asahi H, Kanazawa T. Continuous cultivation of intraerythrocytic Plasmodium falciparum in a serum-free medium with the use of a growth-promoting factor. Parasitology [Internet]. 1994 Nov 6;109(4):397–401. Available from: https://www.cambridge.org/core/product/identifier/S0031182000080641/type/journal_article
9. Divo AA, Jensen JB. Studies on serum requirements for the cultivation of Plasmodium falciparum. I. Animal sera. Bull World Health Organ. 1982;60(4):565–9.
10. Ifediba T, Vanderberg JP. Peptones and Calf Serum as a Replacement for Human Serum in the Cultivation of Plasmodium falciparum. J Parasitol [Internet]. 1980 Apr;66(2):236. Available from: https://www.jstor.org/stable/3280810?origin=crossref
11. Lingnau A, Margos G, Maier WA, Seitz HM. Serum-free cultivation of severalPlasmodium falciparum strains. Parasitol Res [Internet]. 1994;80(1):84–6. Available from: http://link.springer.com/10.1007/BF00932631
12. Desjardins RE, Alexander BM, Weatherly NF, Bowdre JH, Oduola AMJ. Use of Non-Human Plasma for in Vitro Cultivation and Antimalarial Drug Susceptibility Testing of Plasmodium Falciparum. Am J Trop Med Hyg [Internet]. 1985 Mar 1;34(2):209–15. Available from: http://www.ajtmh.org/content/journals/10.4269/ajtmh.1985.34.209
13. Sax LJ, Rieckmann KH. Use of Rabbit Serum in the Cultivation of Plasmodium falciparum. J Parasitol [Internet]. 1980 Aug;66(4):621. Available from: https://www.jstor.org/stable/3280518?origin=crossref
14. Willet GP, Canfield CJ. Plasmodium falciparum: Continuous cultivation of erythrocyte stages in plasma-free culture medium. Exp Parasitol [Internet]. 1984 Feb;57(1):76–80. Available from: https://linkinghub.elsevier.com/retrieve/pii/0014489484900651
15. Binh VQ, Luty AJF, Kremsner PG. Differential Effects of Human Serum and Cells on the Growth of Plasmodium falciparum Adapted to Serum-Free in Vitro Culture Conditions. Am J Trop Med Hyg [Internet]. 1997 Nov 1;57(5):594–600. Available from: http://www.ajtmh.org/content/journals/10.4269/ajtmh.1997.57.594
16. Cranmer SL, Magowan C, Liang J, Coppel RL, Cooke BM. An alternative to serum for cultivation of Plasmodium falciparum in vitro. Trans R Soc Trop Med Hyg [Internet]. 1997 May;91(3):363–5. Available from: https://academic.oup.com/trstmh/article-lookup/doi/10.1016/S0035-9203(97)90110-3
17. Flores MVC, Berger-Eiszele SM, Stewart TS. Long-term cultivation of Plasmodium falciparum in media with commercial non-serum supplements. Parasitol Res [Internet]. 1997 Aug 1;83(7):734–6. Available from: http://link.springer.com/10.1007/s004360050330
18. Wang P, Read M, Sims PFG, Hyde JE. Sulfadoxine resistance in the human malaria parasite Plasmodium falciparum is determined by mutations in dihydropteroate synthetase and an additional factor associated with folate utilization. Mol Microbiol [Internet]. 1997 Mar 31;23(5):979–86. Available from: https://onlinelibrary.wiley.com/doi/abs/10.1046/j.1365-2958.1997.2821646.x
19. http://ki.se/sites/default/files/methods_in_malaria_research
20. Yuan L, Hao M, Wu L, Zhao Z, Rosenthal BM, Li X, et al. Refrigeration provides a simple means to synchronize in vitro cultures of Plasmodium falciparum. Exp Parasitol [Internet]. 2014 May;140:18–23. Available from: https://linkinghub.elsevier.com/retrieve/pii/S0014489414000447
21. TRAGER W. A New Method for Intraerythrocytic Cultivation of Malaria Parasites (Plasmodium coatneyi and P. falciparum). J Protozool. 1971;18(2):239–42.
22. Awandare GA, Nyarko PB, Aniweh Y, Ayivor-Djanie R, Stoute JA. Plasmodium falciparum strains spontaneously switch invasion phenotype in suspension culture. Sci Rep. 2018;8(1):1–10.
23. Amoah LE, Kakaney C, Kwansa-Bentum B, Kusi KA. Activity of Herbal Medicines on Plasmodium falciparum Gametocytes: Implications for Malaria Transmission in Ghana. Lanz-Mendoza H, editor. PLoS One [Internet]. 2015 Nov 12;10(11):e0142587. Available from: https://dx.plos.org/10.1371/journal.pone.0142587
24. Lusakibanza M, Mesia G, Tona G, Karemere S, Lukuka A, Tits M, et al. In vitro and in vivo antimalarial and cytotoxic activity of five plants used in congolese traditional medicine. J Ethnopharmacol [Internet]. 2010 Jun;129(3):398–402. Available from: https://linkinghub.elsevier.com/retrieve/pii/S0378874110002382
25. Engelbrecht D, Coetzer TL. Sunlight inhibits growth and induces markers of programmed cell death in Plasmodium falciparum in vitro. Malar J [Internet]. 2015 Dec 29;14(1):378. Available from: http://www.malariajournal.com/content/14/1/378
26. Khandros E, Huang P, Peslak SA, Sharma M, Abdulmalik O, Giardine BM, et al. Rapid emergence of clonal interference during malaria parasite cultivation. Blood [Internet]. 2011;10(1):271. Available from: http://www.malariajournal.com/content/10/1/271
27. Dohutia C, Mohapatra PK, Bhattacharyya DR, Gogoi K, Bora K, Goswami BK. In vitro adaptability of Plasmodium falciparum to different fresh serum alternatives. J Parasit Dis [Internet]. 2017 Jun 30;41(2):371–4. Available from: http://link.springer.com/10.1007/s12639-016-0808-z
28. Sodeinde O, Williams CK. Continuous in-vitro cultivation of Plasmodium falciparum in Ibadan: solutions to scientific and logistical problems. Afr J Med Med Sci [Internet]. 1990 Jun;19(2):71–6. Available from: http://www.ncbi.nlm.nih.gov/pubmed/2115731
29. Djimde AA, Kirkman L, Kassambara L, Diallo M, Plowe C V, Wellems TE, et al. [In vitro cultivation of fields isolates of Plasmodium falciparum in Mali]. Bull Soc Pathol Exot [Internet]. 2007 Feb;100(1):3–5. Available from: http://www.ncbi.nlm.nih.gov/pubmed/17402683
30. Held J, Zanger P, Issifou S, Kremsner PG, Mordmüller B. In vitro activity of tigecycline in Plasmodium falciparum culture-adapted strains and clinical isolates from Gabon. Int J Antimicrob Agents. 2010;35(6).
31. Deans AM, Nery S, Conway DJ, Kai O, Marsh K, Rowe JA. Invasion pathways and malaria severity in Kenyan Plasmodium falciparum clinical isolates. Infect Immun. 2007;75(6):3014–20.
32. Murray L, Stewart LB, Tarr SJ, Ahouidi AD, Diakite M, Amambua-ngwa A, et al. Multiplication rate variation in the human malaria parasite Plasmodium falciparum. Sci Rep [Internet]. 2017;(July):1–8. Available from: http://dx.doi.org/10.1038/s41598-017-06295-9