title: Loop mediated isothermal amplification (LAMP) accurately detects malaria DNA from filter paper blood samples of low density parasitaemias.
abstract: BACKGROUND
  Loop mediated isothermal amplification (LAMP) provides an opportunity for improved, field-friendly detection of malaria infections in endemic areas. However data on the diagnostic accuracy of LAMP for active case detection, particularly low-density parasitaemias, are lacking. We therefore evaluated the performance of a new LAMP kit compared with PCR using DNA from filter paper blood spots.
  
  
  RESULTS
  Samples from 865 fever patients and 465 asymptomatic individuals collected in Zanzibar were analysed for Pan (all species) and Pf (P. falciparum) DNA with the Loopamp MALARIA Pan/Pf kit. Samples were amplified at 65°C for 40 minutes in a real-time turbidimeter and results were compared with nested PCR. Samples with discordant results between LAMP and nested PCR were analysed with real-time PCR. The real-time PCR corrected nested PCR result was defined as gold standard. Among the 117 (13.5%) PCR detected P. falciparum infections from fever patients (mean parasite density 7491/µL, range 6-782,400) 115, 115 and 111 were positive by Pan-LAMP, Pf-LAMP and nested PCR, respectively. The sensitivities were 98.3% (95%CI 94-99.8) for both Pan and Pf-LAMP. Among the 54 (11.6%) PCR positive samples from asymptomatic individuals (mean parasite density 10/µL, range 0-4972) Pf-LAMP had a sensitivity of 92.7% (95%CI 80.1-98.5) for detection of the 41 P. falciparum infections. Pan-LAMP had sensitivities of 97% (95%CI 84.2-99.9) and 76.9% (95%CI 46.2-95) for detection of P. falciparum and P. malariae, respectively. The specificities for both Pan and Pf-LAMP were 100% (95%CI 99.1-100) in both study groups.
  
  
  CONCLUSIONS
  Both components of the Loopamp MALARIA Pan/Pf detection kit revealed high diagnostic accuracy for parasite detection among fever patients and importantly also among asymptomatic individuals of low parasite densities from minute blood volumes preserved on filter paper. These data support LAMPs potential role for improved detection of low-density malaria infections in pre-elimination settings.
authors.affiliation: Malaria Research, Department of Microbiology, Tumor and Cellbiology, Karolinska Institutet, Stockholm, Sweden; Unit of infectious diseases, Karolinska University Hospital, Stockholm, Sweden.
authors.forename: Berit
authors.initials: B
authors.lastname: Aydin-Schmidt
authors.affiliation: Malaria Research, Department of Microbiology, Tumor and Cellbiology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Weiping
authors.initials: W
authors.lastname: Xu
authors.affiliation: Department of Clinical Parasitology, Foundation for Innovative New Diagnostics (FIND), Geneva, Switzerland.
authors.forename: Iveth J
authors.initials: IJ
authors.lastname: González
authors.affiliation: Hospital for Tropical Diseases, University College London Hospitals, London, United Kingdom; NIHR Biomedical Research Centre at University College London Hospitals NHS Foundation Trust and University College, London, United Kingdom.
authors.forename: Spencer D
authors.initials: SD
authors.lastname: Polley
authors.affiliation: Global Good Fund, Bellevue, Washington, United States of America.
authors.forename: David
authors.initials: D
authors.lastname: Bell
authors.affiliation: Malaria Research, Department of Microbiology, Tumor and Cellbiology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Delér
authors.initials: D
authors.lastname: Shakely
authors.affiliation: Zanzibar Malaria Elimination Program, Ministry of Health, Zanzibar, Tanzania.
authors.forename: Mwinyi I
authors.initials: MI
authors.lastname: Msellem
authors.affiliation: Malaria Research, Department of Microbiology, Tumor and Cellbiology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Anders
authors.initials: A
authors.lastname: Björkman
authors.affiliation: Malaria Research, Department of Microbiology, Tumor and Cellbiology, Karolinska Institutet, Stockholm, Sweden; Global Health, Department of Public Health Sciences, Karolinska Institutet, Stockholm, Sweden; Center for Clinical Research, Sörmland County Council, Sörmland, Sweden.
authors.forename: Andreas
authors.initials: A
authors.lastname: Mårtensson
mesh_terms: D017931:DNA Primers / Q000235:genetics; D016054:DNA, Protozoan / Q000097:blood / Q000302:isolation & purification*; D006801:Humans; D057230:Limit of Detection; D008288:Malaria / Q000097:blood* / Q000175:diagnosis* / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D013636:Tanzania
chemical_list: D017931:DNA Primers; D016054:DNA, Protozoan
references.citation: J Clin Microbiol. 2010 Aug;48(8):2866-71
references.pmid: 20554824
references.citation: Infect Genet Evol. 2008 May;8(3):340-5
references.pmid: 18359278
references.citation: Clin Chem Lab Med. 2003 Jan;41(1):68-73
references.pmid: 12636052
references.citation: PLoS One. 2013 Aug 29;8(8):e71539
references.pmid: 24009663
references.citation: Trends Parasitol. 2002 Sep;18(9):395-8
references.pmid: 12377256
references.citation: J Infect Dis. 2013 Aug 15;208(4):637-44
references.pmid: 23633403
references.citation: PLoS One. 2010 Oct 29;5(10):e13733
references.pmid: 21060829
references.citation: Malar J. 2009 Apr 29;8:86
references.pmid: 19402894
references.citation: PLoS Med. 2011 Jan 25;8(1):e1000396
references.pmid: 21311583
references.citation: Parasitol Today. 1993 Aug;9(8):303-5
references.pmid: 15463789
references.citation: J Clin Microbiol. 2007 Aug;45(8):2521-8
references.pmid: 17567794
references.citation: PLoS One. 2013 Sep 04;8(9):e72912
references.pmid: 24023791
references.citation: J Infect Dis. 2014 Oct 15;210(8):1180-7
references.pmid: 24795480
references.citation: J Clin Microbiol. 2009 Apr;47(4):975-80
references.pmid: 19244467
references.citation: Lancet. 2010 Nov 6;376(9752):1592-603
references.pmid: 21035841
references.citation: J Infect Dis. 2013 Aug 15;208(4):645-52
references.pmid: 23633405
references.citation: Acta Trop. 2012 Jun;122(3):233-40
references.pmid: 22366670
references.citation: Clin Microbiol Infect. 2011 Nov;17(11):1624-31
references.pmid: 21910780
references.citation: Bull World Health Organ. 1999;77(7):553-9
references.pmid: 10444878
references.citation: Clin Chem. 2006 Feb;52(2):303-6
references.pmid: 16339303
references.citation: Nat Commun. 2012;3:1237
references.pmid: 23212366
references.citation: PLoS Negl Trop Dis. 2012;6(6):e1572
references.pmid: 22745836
references.citation: J Clin Microbiol. 2010 Oct;48(10):3539-43
references.pmid: 20686079
references.citation: Malar J. 2011 Dec 16;10:370
references.pmid: 22177014