The Efficacy of Photochemical Treatment with Amotosalen and UVA (INTERCEPT) for Inactivation of T. cruzi In Buffy Coat Platelet Components.

Abstract Introduction.Transfusion-associated Chagas’ disease has been transmitted by labile blood components in endemic countries. In addition, at least 6 cases have occurred in the USA and Canada, due to platelets. In many countries, no test to prevent transfusion-transmission of T. cruzi is in routine use. This study evaluated the efficacy of photochemical treatment (PCT) to inactivate T. cruzi in contaminated platelet components. Methods. Paired platelet components were prepared as follows. 3 pooled ABO matched buffy-coat platelet components (BC-PLT) were combined, and then re-divided into 3 BC-PLT units of approximately 300 mL. Each BC-PLT unit was inoculated with a high dose (5x103/mL) of viable T. cruzi metacyclic trypomastigotes obtained from mice infected with either the G, Tulahuen (T) or Y strains. Five pools of 3 BC-PLT units each were assayed for infectivity before and after PCT with the INTERCEPT system (150 μM amotosalen + 3 J/cm2). Infectivity was determined with 3 different methods by assaying 10 mL samples for parasite viability as follows: 1) in vitro culture to detect viable epimastigotes; 2) 3H-Thymidine incorporation in culture; and 3) in vivo inoculation into Interferon-γ Receptor (IFN-γR) deficient mice, highly susceptible to T. cruzi infection. For all BC-PLT units, 10 mL aliquots of each platelet unit were taken before inoculation as negative controls (NC), after parasite inoculation before PCT as positive controls (PC), and after PCT. In vitro cultures were seeded, observed weekly and parasitemia calculated at 30 days post-inoculation(d.p.i) as parasites/mL (p/mL) by limiting dilution analysis. At 30 d.p.i, cultures were pulsed with 3H-Thymidine for 16 hours, cultures were harvested and counts per minute (c.p.m.) measured. In the in vivo study, mice were inoculated intraperitoneally with BC-PLT (10 mL concentrated to 1 mL) and parasitemia was monitored at 7, 14, 21 and 28 d.p.i. Survival of mice was followed up for more than 2 months post inoculation. Results. After 30 days follow up, no viable parasites were detected in PCT BC-PLT by any of the 3 viability assays, and with any of the 3 T. cruzi strains tested. Only the untreated PC samples showed the presence of the replicative form of the parasite by in vitro cultures derived from the 3 different T. cruzi strains. PC from BC-PLT contaminated with the different strains contained 7812, 550, and 100 parasites/mL for the Y, T and G strains, respectively, by in vitro cultures. PC, but not PCT BC-PLT samples, showed significant incorporation of 3H-Thymidine. In addition, in vivo studies showed that only mice inoculated with PC died as a result of the infection. Moreover, 100% of mice died at 14, 28 and 38 days after inoculation with PC containing the Y, T and G strains, respectively. All mice inoculated with NC and PCT BC-PLT aliquots were alive after more than 2 months post-exposure. Conclusions.The pathogen reduction system using Amotosalen HCl and UVA demonstrated robust efficacy for inactivation of high doses of 3 different strains of T. cruzi . This PCT system offers the potential to make the platelet supply safer in countries with T. cruzi endemicity and in countries with growing immigrant blood donor populations from Latin America and other regions in which T. cruzi is endemic.

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