Inactivation of Plasmodium falciparum in whole blood using the amustaline and glutathione pathogen reduction technology

Abstract

BACKGROUND Risk of transfusion‐transmitted (TT) malaria is mainly associated with whole blood (WB) or red blood cell (RBC) transfusion. Risk mitigation relies mostly on donor deferral while a limited number of countries perform blood testing, both negatively impacting blood availability. This study investigated the efficacy of the pathogen reduction system using amustaline and glutathione (GSH) to inactivate Plasmodium falciparum in WB. STUDY DESIGN AND METHODS WB units were spiked with ring stage P. falciparum infected RBCs. Parasite loads were measured in samples at time of infection, after 24 hours at room temperature (RT), and after a 24‐hour incubation at RT post‐treatment with 0.2 mM amustaline and 2 mM GSH. Serial 10‐fold dilutions of the samples were inoculated to RBC cultures and maintained up to 4 weeks. Parasitemia was quantified by cytometry. RESULTS The P. falciparum viability assay has a limit of detection of a single live parasite per sample. Input parasite titer was >5.7 log10 TCID50 per mL. A 24‐hour incubation at RT paused parasite development in controls, but they retained viability and infectivity when tested in culture. In contrast, no infectious parasites were detected in the amustaline/GSH‐treated sample after 4 weeks of culture. CONCLUSION A robust level of P. falciparum inactivation was achieved in WB using amustaline/GSH treatment. Parasite log reduction was >5.7 log10 TCID50 per mL. Development of such a pathogen reduction system may provide an opportunity to reduce the risk of TT malaria and improve blood availability.

Robust inactivation of Plasmodium falciparum in red blood cell concentrates using amustaline and glutathione pathogen reduction

“…An alternative nucleic acid targeted PRT system for RBC based on a chemical treatment using amustaline and glutathione (GSH) has been developed to overcome the problem of high UV light absorption by RBC when attempting to inactivate pathogens in WB and RBC concentrates and is under clinical evaluation. Amustaline–GSH treatment has been shown to efficiently inactivate various pathogens, including viruses, parasites, and bacteria in RBC, 21,23 and P. falciparum in WB units 24 . In the present study, we extended our previous investigations 24,25 to the inactivation of P. falciparum in RBC in additive solutions AS‐1 (Adsol) or AS‐5 (Optisol) and examined the treatment efficiency according to parasite load, stage, and strains from different origins.…”

“…P. falciparum strains of different origins and susceptibilities (sensitive (s) or resistant (r)) to antimalarial drugs chloroquine (CQ), pyrimethanime (PYR), and mefloquine (MF) were provided from the Unicellular Eukaryote Collection (MNHN, France): FCB1/Colombia (CQr, PYRr, MFs), 3D7 (CQs, MFs, PYRs, derived from the NF54 strain, a Schipol Airport case, The Netherlands), W2/Indochina (CQr, PYRr, MFs), and its DD2 derived clone (CQr, PYRr, MFr). Strains were maintained in culture on human RBC (A/O groups) provided by the Etablissement Français du Sang (EFS) in RPMI 1640 medium supplemented with 0.5% Albumax‐II, at 37°C, under 5% CO 2 as described 24 . Parasites were synchronized by sorbitol treatment 26 and ~20% ring cultures were obtained as described 27 .…”

“…Parasite development was routinely followed on Diff‐Quik‐stained smears (Medion Diagnostics). Parasitemia was determined by cytometry using the fluorescent DNA probeYOYO‐1 as described 24,28 …”

“…A culture was considered positive when parasitemia reached 1%. Parasite loads were calculated as described 29 and were expressed as the log 10 of the median Tissue Culture Infectious Dose (log 10 TCID 50 ) 24 using the formula:…”

Malaria

“…In einer randomisierten doppelt-blinden klinischen Studie (NCT02118428) konnte unter endemischen Bedingungen der Nutzen der Pathogeninaktivierung (Mirasol) gezeigt werden: Bei Transfusion von parasitämischen Vollblutkonserven konnte mit Hilfe der Pathogeninaktivierung eine deutliche Reduktion der transfusionsbedingten Malaria bei den Empfängerinnen und Empfängern erreicht werden (1/28 versus 8/37 Patienten) [114]. Sow et al untersuchten die Wirksamkeit des Amustalin/Glutathion-Systems (Cerus) zur Inaktivierung von P. falciparum in gespikten Proben und fanden eine Reduzierung der TCID50 um mindestens 5,7 log-Stufen [123]. In einer weiteren Studie wurde errechnet, dass die Kombination von Malariaschnelltest und Pathogeninaktivierung mit der Riboflavin/ UVB-Methode (Mirasol) das Risiko, eine malariainfizierte, infektiöse Blutkonserve zu transfundieren, gegenüber dem jetzigen Standard um 83 % senken könnte [124].…”