Abstract
BackgroundSeveral platforms have been used to generate the primary data for microsatellite analysis of malaria parasite genotypes. Each has relative advantages but share a limitation of being time- and cost-intensive. A commercially available automated capillary gel cartridge system was assessed in the microsatellite analysis of Plasmodium vivax diversity in the Peruvian Amazon.MethodsThe reproducibility and accuracy of a commercially-available automated capillary system, QIAxcel, was assessed using a sequenced PCR product of 227 base pairs. This product was measured 42 times, then 27 P. vivax samples from Peruvian Amazon subjects were analyzed with this instrument using five informative microsatellites. Results from the QIAxcel system were compared with a Sanger-type sequencing machine, the ABI PRISM® 3100 Genetic Analyzer.ResultsSignificant differences were seen between the sequenced amplicons and the results from the QIAxcel instrument. Different runs, plates and cartridges yielded significantly different results. Additionally, allele size decreased with each run by 0.045, or 1 bp, every three plates. QIAxcel and ABI PRISM systems differed in giving different values than those obtained by ABI PRISM, and too many (i.e. inaccurate) alleles per locus were also seen with the automated instrument.ConclusionsWhile P. vivax diversity could generally be estimated using an automated capillary gel cartridge system, the data demonstrate that this system is not sufficiently precise for reliably identifying parasite strains via microsatellite analysis. This conclusion reached after systematic analysis was due both to inadequate precision and poor reproducibility in measuring PCR product size.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-015-0842-9) contains supplementary material, which is available to authorized users.
Highlights
Several platforms have been used to generate the primary data for microsatellite analysis of malaria parasite genotypes
Analysis of Plasmodium vivax samples from the Peruvian Amazon A total of 271 P. vivax single infection samples were analyzed using samples obtained from three rural communities around Iquitos (Padrecocha, San José de Lupuna and Santo Tomás) from the years 2005 and 2008
This study aimed to explore the utility of a non-fluorophore-dependent, automated microsatellite analysis system that purportedly had two major advantages—higher throughput and lower cost—to obtain raw microsatellite marker data
Summary
Several platforms have been used to generate the primary data for microsatellite analysis of malaria parasite genotypes. Each has relative advantages but share a limitation of being time- and cost-intensive. A commercially available automated capillary gel cartridge system was assessed in the microsatellite analysis of Plasmodium vivax diversity in the Peruvian Amazon. Molecular markers are widely used in the study of malaria and other infectious diseases [1]. Such markers allow for the differentiation of new and recurrent infections, tracing spread of clones of a pathogenic microorganism and discovering new introductions of infection
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