Highly sensitive quantitative real-time PCR for the detection of Plasmodium liver-stage parasite burden following low-dose sporozoite challenge

Schussek, Sophie, Groves, Penny L., Apte, Simon H., and Doolan, Denise L. (2013) Highly sensitive quantitative real-time PCR for the detection of Plasmodium liver-stage parasite burden following low-dose sporozoite challenge. PLoS ONE, 8 (10). e77811. pp. 1-10.

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The pre-erythrocytic stages of Plasmodium spp. are increasingly recognised as ideal targets for prophylactic vaccines and drug treatments. Intense research efforts in the last decade have been focused on in vitro culture and in vivo detection and quantification of liver stage parasites to assess the effects of candidate vaccines or drugs. Typically, the onset of blood stage parasitaemia is used as a surrogate endpoint to estimate the efficacy of vaccines and drugs targeting pre-erythrocytic parasite stages in animal models. However, this provides no information on the parasite burden in the liver after vaccination or treatment and therefore does not detect partial efficacy of any vaccine or drug candidates. Herein, we describe a quantitative RT-PCR method adapted to detect and quantitate Plasmodium yoelii liver stages in mice with increased sensitivity even after challenge with as few as 50 cryopreserved sporozoites (corresponding to approximately 5-10 freshly isolated sporozoites). We have validated our quantitative RT-PCR assay according to the MIQE (Minimum Information for Publication of Quantitative Real-Time PCR Experiments) guidelines and established high reproducibility and accuracy. Our assay provides a rapid and reproducible assessment of liver stage parasite burden in rodent malaria models, thereby facilitating the evaluation of the efficacy of anti-malarial drugs or prophylactic vaccines with high precision and efficacy.

Item ID: 41430
Item Type: Article (Research - C1)
ISSN: 1932-6203
Additional Information:

© 2013 Schussek et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Funders: National Health and Medical Research Council (NHMRC) Australia
Projects and Grants: NHMRC Program Grant 496600
Date Deposited: 24 Mar 2016 05:01
FoR Codes: 11 MEDICAL AND HEALTH SCIENCES > 1107 Immunology > 110799 Immunology not elsewhere classified @ 100%
SEO Codes: 92 HEALTH > 9201 Clinical Health (Organs, Diseases and Abnormal Conditions) > 920109 Infectious Diseases @ 100%
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