Yago Pico de Coaña | PubMed Health

PubMed2003

A highly infective Plasmodium vivax strain adapted to Aotus monkeys: quantitative haematological and molecular determinations useful for P. vivax malaria vaccine development.

The New World primate Aotus nancymaae is susceptible to infection by the human malaria parasite Plasmodium vivax and has therefore been recommended by the World Health Organization as a model for malaria vaccine candidate evaluation. We report the isolation, adaptation, titration and genetic characterization of a P. vivax wild strain in splenectomized A. nancymaae monkeys. Parasitemia remained high after 22 passages, reaching 7.88% by Giemsa and Acridine Orange staining and Real-Time PCR determination, making this P. vivax strain a highly infective and reliable asset to be used in P. vivax biological studies and vaccine development.

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Quantifying Aotus monkey cytokines by real-time quantitative RT-PCR.

Aotus spp. monkeys are considered the ideal model for studying the progress of malarial infection and the immune response it elicits. We describe the use of a recently developed technique, real-time quantitative RT-PCR, to quantify several Aotus monkey cytokine mRNAs involved in Th1/Th2 responses (IL-4, IL-10, TNF-beta and IFN-gamma). Specific primers were designed for each cytokine and standard curves were constructed using serial dilutions of pDNA containing each target sequence. Results were normalized to GAPDH housekeeping gene expression levels. Standard curves showed high correlation coefficients and were linear over a wide range of copy numbers. Quantification of Aotus samples showed little intra- and inter-experiment variation, thus, the technique has proven to be highly reproducible and sensitive allowing us to detect as little as 25 copies/microl of target DNA. This technique will allow studying Th1 and Th2 cytokine patterns elicited in response to infection for prospectively evaluating the efficacy of malarial vaccines.

Animals↗