Assessing anti-malarial drug effects ex vivo using the haemozoin detection assay
Autor: | Martin P. Grobusch, José Francisco Fernandes, Thomas Hänscheid, Maria Rebelo, Carolina Tempera |
---|---|
Přispěvatelé: | Repositório da Universidade de Lisboa, Amsterdam institute for Infection and Immunity, Amsterdam Public Health, Infectious diseases |
Jazyk: | angličtina |
Rok vydání: | 2015 |
Předmět: |
Hemeproteins
030231 tropical medicine Resistance Plasmodium falciparum Drug resistance Pharmacology Flow cytometry Haemozoin 03 medical and health sciences chemistry.chemical_compound Antimalarials Inhibitory Concentration 50 0302 clinical medicine field trial Chloroquine parasitic diseases medicine Humans Artemisinin Malaria Falciparum 0303 health sciences Blood Cells medicine.diagnostic_test biology 030306 microbiology Methodology Anti-malarial sensitivity testing biology.organism_classification Flow Cytometry Artemisinins 3. Good health Malaria Infectious Diseases Parasitology chemistry Artesunate Ex vivo medicine.drug |
Zdroj: | Malaria Journal Malaria journal, 14(1). BioMed Central |
ISSN: | 1475-2875 |
Popis: | © 2015 Rebelo et al.; licensee BioMed Central. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Background: In vitro sensitivity assays are crucial to detect and monitor drug resistance. Plasmodium falciparum has developed resistance to almost all anti-malarial drugs. Although different in vitro drug assays are available, some of their inherent characteristics limit their application, especially in the field. A recently developed approach based on the flow cytometric detection of haemozoin (Hz) allowed reagent-free monitoring of parasite maturation and detection of drug effects in culture-adapted parasites. In this study, the set-up, performance and usefulness of this novel assay were investigated under field conditions in Gabon. Methods: An existing flow cytometer (Cyflow Blue) was modified on site to detect light depolarization caused by Hz. Blood from malaria patients was incubated for 72 hrs with increasing concentrations of chloroquine, artesunate and artemisinin. The percentage of depolarizing red blood cells (RBC) was used as maturation indicator and measured at 24, 48 and 72 hrs of incubation to determine parasite growth and drug effects. Results: The flow cytometer was easily adapted on site to detect light depolarization caused by Hz. Analysis of ex vivo cultures of parasites, obtained from blood samples of malaria patients, showed four different growth profiles. In 39/46 samples, 50% inhibitory concentrations (IC50) were successfully determined. IC50 values for chloroquine were higher than 200 nM in 70% of the samples, indicating the presence of chloroquine-resistant parasites. For artesunate and artemisinin, IC50 values ranged from 0.9 to 60 nM and from 2.2 nM to 124 nM, respectively, indicating fully sensitive parasites. Conclusion: Flow cytometric detection of Hz allowed the detection of drug effects in blood samples from malaria patients, without using additional reagents or complex protocols. Adjustment of the initial parasitaemia was not required, which greatly simplifies the protocol, although it may lead to different IC50 values. Further investigation of set-up conditions of the Hz assay, as well as future studies in various settings should be performed to further determine the usefulness of this assay as a tool for rapid resistance testing in malaria-endemic countries. This work was supported by the Luso-American Foundation (FLAD-LACR grant: B-A.V-109-09/07). MR acknowledges Fundação para a Ciência e a Tecnologia for doctoral grant (SFRH/BD/84530/2012) and Fundação Calouste Gulbenkian for the Award CAML/Gulbenkian for Travel ACGT fellowship. |
Databáze: | OpenAIRE |
Externí odkaz: |