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Background  Confirmation of artemisinin-delayed parasite clearance in  Plasmodium falciparum  along the Thai-Myanmar border has inspired a global response to contain and monitor drug resistance to avert the disastrous consequences of a potential spread to Africa. However, resistance data from Myanmar are sparse, particularly from high-risk areas where limited health services and decades of displacement create conditions for resistance to spread. Subclinical infections may represent an important reservoir for resistance genes that confer a fitness disadvantage relative to wild-type alleles. This study estimates the prevalence of resistance genotypes in three previously unstudied remote populations in Myanmar and tests the  a priori  hypothesis that resistance gene prevalence would be higher among isolates collected from subclinical infections than isolates collected from febrile clinical patients. A systematic review of resistance studies is provided for context.    Methods  Community health workers in Karen and Kachin States and an area spanning the Indo-Myanmar border collected dried blood spots from 988 febrile clinical patients and 4,591 villagers with subclinical infection participating in routine prevalence surveys. Samples positive for  P. falciparum  18 s ribosomal RNA by real-time PCR were genotyped for  P. falciparum  multidrug resistance protein ( pfmdr1)  copy number and the  pfcrt  K76T polymorphism using multiplex real-time PCR.    Results   Pfmdr1  copy number increase and the  pfcrt  K76 polymorphism were determined for 173 and 269 isolates, respectively. Mean  pfmdr1  copy number was 1.2 (range: 0.7 to 3.7).  Pfmdr1  copy number increase was present in 17.5%, 9.6% and 11.1% of isolates from Karen and Kachin States and the Indo-Myanmar border, respectively.  Pfmdr1  amplification was more prevalent in subclinical isolates (20.3%) than clinical isolates (6.4%, odds ratio 3.7, 95% confidence interval 1.1 - 12.5). P fcrt  K76T prevalence ranged from 90-100%.    Conclusions  Community health workers can contribute to molecular surveillance of drug resistance in remote areas of Myanmar. Marginal and displaced populations under-represented among previous resistance investigations can and should be included in resistance surveillance efforts, particularly once genetic markers of artemisinin-delayed parasite clearance are identified. Subclinical infections may contribute to the epidemiology of drug resistance, but determination of gene amplification from desiccated filter samples requires further validation when DNA concentration is low.

malaria journal

Molecular surveillance for drug-resistant Plasmodium falciparum in clinical and subclinical populations from three border regions of Burma/Myanmar: cross-sectional data and a systematic review of resistance studies