Assessment of an automated capillary system for Plasmodium vivax microsatellite genotyping

Malaria Journal

Research

Margaret N. Kosek¹  Pablo Yori¹  Robert H. Gilman¹  Martiza Calderón²  Manuel Fasabi³  Oscar Nolasco⁴  Mari Hoshi⁴  Paulo Manrique⁴  Dionicia Gamboa⁵  Joseph M. Vinetz⁶ 

[1] Department of International Health, Johns Hopkins School of Public Health, Baltimore, MD, USA;Division of Infectious Diseases, Department of Medicine, University of California San Diego, La Jolla, CA, USA;Instituto Nacional de Salud, Lima, Peru;Malaria Laboratory, Institute of Tropical Medicine Alexander von Humboldt, Universidad Peruana Cayetano Heredia, Lima, Peru;Malaria Laboratory, Institute of Tropical Medicine Alexander von Humboldt, Universidad Peruana Cayetano Heredia, Lima, Peru;Departamento de Ciencias Celulares y Moleculares, Universidad Peruana Cayetano Heredia, Lima, Peru;Malaria Laboratory, Institute of Tropical Medicine Alexander von Humboldt, Universidad Peruana Cayetano Heredia, Lima, Peru;Division of Infectious Diseases, Department of Medicine, University of California San Diego, La Jolla, CA, USA;

关键词: Malaria;    Molecular biology;    Genotyping;    Microsatellites;    Molecular epidemiology;    Technology;   

DOI  :  10.1186/s12936-015-0842-9

 received in 2015-03-24, accepted in 2015-08-08,  发布年份 2015

来源: Springer

【 摘 要 】

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.

【 授权许可】

CC BY   
© Manrique et al. 2015