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WWARN’s In Vitro partnership with the Institut Pasteur in Cambodia

5 March 2014

The Institut Pasteur recently released two laboratory and field-based tests that correlate with the slow parasite clearance that has defined artemisinin-resistance. These assays were a key to the identification of a genetic marker of artemisinin resistance in collaboration with colleagues at the Institut Pasteur in Paris, CNRS in Toulouse and the US National Institute of Allergy & Infectious Diseases (NIAD).

We are delighted to announce a new partnership with the Institut Pasteur in Phnom Penh, Cambodia to lead WWARN’s In Vitro Module. Didier Menard will direct WWARN’s in vitro activities together with the support of Benoit Witkowski.

Dr Menard confirms “The Institut Pasteur Cambodia is pleased to be part of a new partnership with the WWARN network. We look forward to strengthening our experience and developing new opportunities to work on the implementation of in vitro phenotypic tools to guide the use of artemisinin derivatives and non-artemisinin components in ACTs, the developments of collaborative studies through multi-disciplinary approaches, and the opportunity to share expertise in order to provide evidence on the alternative choices of treatment if there is suspected drug resistance.”

The first initiative will start this month with a training programme in partnership with the Kenya Medical Research Initiative (KEMRI) and colleagues from the Centers for Disease Control & Prevention (CDC) in Kisumu, Kenya from 1st- 11th March. This training will strengthen the skills of researchers to use a new procedure which describes two assays to evaluate the susceptibility of Plasmodium falciparum malaria parasites to artemisinin drug treatment.

The first in vitro Ring‐stage Survival Assay (RSA0‐3h) test is performed on 0‐3 hour post‐invasion rings obtained from culture‐adapted parasites. The second ex vivo Ring‐stage Survival Assay (ex vivo RSA) approach is performed on parasite isolates that are freshly collected from patients diagnosed with malaria. In both assays, parasites are exposed to a drug dose of 700 nM dihydroartemisinin (DHA) for 6 hours; their survival rate is then assessed 72 hours later.

Over a number of days, the KEMRI team will receive in-depth guidance and support from Benoit Witkowski to define the microscopic readout of RSA and perform both in vitro and ex vivo RSA analysis, respectively, on 6 adapted strains and 6-10 fresh isolates from KEMRI’s Kisumu research site.

It is expected that the group will then be able to detect the presence of artemisinin resistant parasites if they appear in their site, which should in future reinforce staff research capacity to carry out malaria surveillance trials. It is hoped it will also encourage further opportunities to work together with KEMRI, and other research institutes, to monitor the eventual emergence or spread of malaria resistance worldwide.

Download and share the Standard Operating Procedure (SOP) entitled: Ring-stage Survival Assays (RSA) to evaluate the in-vitro and ex-vivo susceptibility of Plasmodium falciparum to artemisinins.

You can also access the SOP PCR and sequencing for genotyping  of candidate Plasmodium falciparum artemisinin resistance SNPs in the Kelch 13 gene.

Artemisinin resistant in vitro culture adapted parasites from Didier Menard at the Institut Pasteur Cambodia have been deposited in MR4 for investigators to use as controls for the ring-stage assay and other applications.  Six culture isolates, starting with MRA-1236 Plasmodium falciparum IPC3445 Pailin Cambodia 2010, will be available to order in the BEI Resources online catalogue beginning in April, 2014.  To request these strains, investigators need to register with BEI Resources for access to the online catalog at Level 2. Information on registration for online access to all MR4 and BEI Resources cultures and reagents, available for free to investigators world-wide through NIAID, can be found here.

Visit the Institut Pasteur in Cambodia and KEMRI to find out more about their work. If you’re interested in joining an RSA training session please email bwitkowski(at)pasteur-kh.org with your details.