Optimization of Aotus derived Plasmodium vivax ex-vivo cultures

Background: Plasmodium vivax has proven difficult to maintain in continuous culture, mainly to special culture conditions and its preference for invading reticulocytes. In so far, different empirical culture media have been tried in P. vivax ex-vivo cultures with relative success, but a definitive P. vivax culture media composition had remained elusive. In order to improve the viability of P. vivaxex-vivo cultures, this study examines the effect of McCoy’s culture media supplemented with Lipid concentrate or GlutaMax™ (McCoy’s modified), alone or in combination over the viability of P. vivax SAL-1 infected Aotus erythrocytes. Materials and Methods: For this purpose, we tested the effect of modified McCoy’s media on ex-vivo cultures of P. vivax SAL-1 Aotus infected red blood cells (iRBCs) obtained from twenty-two (22) spleen intact animals. The iRBCs suspension was further plated and incubated at 37 Celsius in a Gas Chamber under a 5% O2 + 5% CO2 and 90 % N2 gas atmosphere and static conditions. Media was changed at 24 hours and cells were harvested at 48 hours. Thin smears were prepared at at 0, 24 and 48 hours PI and stained with Giemsa for parasitemia determination (%) and differential staging. At the end of the 48 hours of incubation, samples were preserved in Trizol and IFA slides prepared for gene expression studies further analysis. Enhance viability was defined as ex-vivo cultures with parasitemias statistically different (p < 0.05) compared to the referral group (Sera only) by 48 hours of incubation. Results: Data obtained from these experiments indicated that the viability of ex-vivo cultures was enhanced by McCoy’s modified media irrespective of their WBCs filtration status. Moreover, the data suggests that media supplementation with sera plus Lipid concentrate plus GlutaMaxTM was superior to human sera alone or when in combination with either Lipid concentrate or GlutaMaxTM. Conclusions: Supplementation of McCoy’sculture media with Lipid concentrate alone or in combination with GlutaMaxTM was superior to McCoy’sculture supplemented with human sera alone at enhancing the viability of P. vivax Aotus iRBCs ex-vivo cultures.