Sensitive Detection of Plasmodium vivax Using a High-Throughput, Colourimetric Loop Mediated Isothermal Amplification (HtLAMP) Platform

A Potential Novel Tool for Malaria Elimination

S Britton, Q Cheng, Matthew Grigg, CB Poole, Cielo Pasay, Timothy William, K Fornace, Nicholas Anstey, Colin Sutherland, Chris Drakeley, James McCarthy

Research output: Contribution to journalArticleResearchpeer-review

10 Downloads (Pure)

Abstract

Introduction: Plasmodium vivax malaria has a wide geographic distribution and poses challenges to malaria elimination that are likely to be greater than those of P. falciparum. Diagnostic tools for P. vivax infection in non-reference laboratory settings are limited to microscopy and rapid diagnostic tests but these are unreliable at low parasitemia. The development and validation of a high-throughput and sensitive assay for P. vivax is a priority.

Methods: A high-throughput LAMP assay targeting a P. vivax mitochondrial gene and deploying colorimetric detection in a 96-well plate format was developed and evaluated in the laboratory. Diagnostic accuracy was compared against microscopy, antigen detection tests and PCR and validated in samples from malaria patients and community controls in a district hospital setting in Sabah, Malaysia.

Results: The high throughput LAMP-P. vivax assay (HtLAMP-Pv) performed with an estimated limit of detection of 1.4 parasites/ μL. Assay primers demonstrated cross-reactivity with P. knowlesi but not with other Plasmodium spp. Field testing of HtLAMP-Pv was conducted using 149 samples from symptomatic malaria patients (64 P. vivax, 17 P. falciparum, 56 P. knowlesi, 7 P. malariae, 1 mixed P. knowlesi/P. vivax, with 4 excluded). When compared against multiplex PCR, HtLAMP-Pv demonstrated a sensitivity for P. vivax of 95% (95% CI 87–99%); 61/64), and specificity of 100% (95% CI 86–100%); 25/25) when P. knowlesi samples were excluded. HtLAMP-Pv testing of 112 samples from asymptomatic community controls, 7 of which had submicroscopic P. vivax infections by PCR, showed a sensitivity of 71% (95% CI 29–96%; 5/7) and specificity of 93% (95% CI87-97%; 98/105).

Conclusion: This novel HtLAMP-P. vivax assay has the potential to be a useful field applicable molecular diagnostic test for P. vivax infection in elimination settings.

Original languageEnglish
Pages (from-to)1-16
Number of pages16
JournalPLoS Neglected Tropical Diseases
Volume10
Issue number2
DOIs
Publication statusPublished - 2016

Fingerprint

Plasmodium vivax
Malaria
Malaysia
Routine Diagnostic Tests
Microscopy
Vivax Malaria
Polymerase Chain Reaction
Mitochondrial Genes
Plasmodium
Parasitemia
Molecular Pathology
Multiplex Polymerase Chain Reaction
Limit of Detection
Parasites
Antigens
Infection

Cite this

Britton, S ; Cheng, Q ; Grigg, Matthew ; Poole, CB ; Pasay, Cielo ; William, Timothy ; Fornace, K ; Anstey, Nicholas ; Sutherland, Colin ; Drakeley, Chris ; McCarthy, James. / Sensitive Detection of Plasmodium vivax Using a High-Throughput, Colourimetric Loop Mediated Isothermal Amplification (HtLAMP) Platform : A Potential Novel Tool for Malaria Elimination. In: PLoS Neglected Tropical Diseases. 2016 ; Vol. 10, No. 2. pp. 1-16.
@article{a0dbf4f377d345468b4fc179eaa1be93,
title = "Sensitive Detection of Plasmodium vivax Using a High-Throughput, Colourimetric Loop Mediated Isothermal Amplification (HtLAMP) Platform: A Potential Novel Tool for Malaria Elimination",
abstract = "Introduction: Plasmodium vivax malaria has a wide geographic distribution and poses challenges to malaria elimination that are likely to be greater than those of P. falciparum. Diagnostic tools for P. vivax infection in non-reference laboratory settings are limited to microscopy and rapid diagnostic tests but these are unreliable at low parasitemia. The development and validation of a high-throughput and sensitive assay for P. vivax is a priority. Methods: A high-throughput LAMP assay targeting a P. vivax mitochondrial gene and deploying colorimetric detection in a 96-well plate format was developed and evaluated in the laboratory. Diagnostic accuracy was compared against microscopy, antigen detection tests and PCR and validated in samples from malaria patients and community controls in a district hospital setting in Sabah, Malaysia. Results: The high throughput LAMP-P. vivax assay (HtLAMP-Pv) performed with an estimated limit of detection of 1.4 parasites/ μL. Assay primers demonstrated cross-reactivity with P. knowlesi but not with other Plasmodium spp. Field testing of HtLAMP-Pv was conducted using 149 samples from symptomatic malaria patients (64 P. vivax, 17 P. falciparum, 56 P. knowlesi, 7 P. malariae, 1 mixed P. knowlesi/P. vivax, with 4 excluded). When compared against multiplex PCR, HtLAMP-Pv demonstrated a sensitivity for P. vivax of 95{\%} (95{\%} CI 87–99{\%}); 61/64), and specificity of 100{\%} (95{\%} CI 86–100{\%}); 25/25) when P. knowlesi samples were excluded. HtLAMP-Pv testing of 112 samples from asymptomatic community controls, 7 of which had submicroscopic P. vivax infections by PCR, showed a sensitivity of 71{\%} (95{\%} CI 29–96{\%}; 5/7) and specificity of 93{\%} (95{\%} CI87-97{\%}; 98/105). Conclusion: This novel HtLAMP-P. vivax assay has the potential to be a useful field applicable molecular diagnostic test for P. vivax infection in elimination settings.",
author = "S Britton and Q Cheng and Matthew Grigg and CB Poole and Cielo Pasay and Timothy William and K Fornace and Nicholas Anstey and Colin Sutherland and Chris Drakeley and James McCarthy",
note = "Australian National Health and Medical Research Council (Program grant #1037304, Project Grant #1045156",
year = "2016",
doi = "10.1371/journal.pntd.0004443",
language = "English",
volume = "10",
pages = "1--16",
journal = "PLoS Neglected Tropical Diseases",
issn = "1935-2727",
publisher = "Public Library of Science (PLoS)",
number = "2",

}

Sensitive Detection of Plasmodium vivax Using a High-Throughput, Colourimetric Loop Mediated Isothermal Amplification (HtLAMP) Platform : A Potential Novel Tool for Malaria Elimination. / Britton, S; Cheng, Q; Grigg, Matthew; Poole, CB; Pasay, Cielo; William, Timothy; Fornace, K; Anstey, Nicholas; Sutherland, Colin; Drakeley, Chris; McCarthy, James.

In: PLoS Neglected Tropical Diseases, Vol. 10, No. 2, 2016, p. 1-16.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Sensitive Detection of Plasmodium vivax Using a High-Throughput, Colourimetric Loop Mediated Isothermal Amplification (HtLAMP) Platform

T2 - A Potential Novel Tool for Malaria Elimination

AU - Britton, S

AU - Cheng, Q

AU - Grigg, Matthew

AU - Poole, CB

AU - Pasay, Cielo

AU - William, Timothy

AU - Fornace, K

AU - Anstey, Nicholas

AU - Sutherland, Colin

AU - Drakeley, Chris

AU - McCarthy, James

N1 - Australian National Health and Medical Research Council (Program grant #1037304, Project Grant #1045156

PY - 2016

Y1 - 2016

N2 - Introduction: Plasmodium vivax malaria has a wide geographic distribution and poses challenges to malaria elimination that are likely to be greater than those of P. falciparum. Diagnostic tools for P. vivax infection in non-reference laboratory settings are limited to microscopy and rapid diagnostic tests but these are unreliable at low parasitemia. The development and validation of a high-throughput and sensitive assay for P. vivax is a priority. Methods: A high-throughput LAMP assay targeting a P. vivax mitochondrial gene and deploying colorimetric detection in a 96-well plate format was developed and evaluated in the laboratory. Diagnostic accuracy was compared against microscopy, antigen detection tests and PCR and validated in samples from malaria patients and community controls in a district hospital setting in Sabah, Malaysia. Results: The high throughput LAMP-P. vivax assay (HtLAMP-Pv) performed with an estimated limit of detection of 1.4 parasites/ μL. Assay primers demonstrated cross-reactivity with P. knowlesi but not with other Plasmodium spp. Field testing of HtLAMP-Pv was conducted using 149 samples from symptomatic malaria patients (64 P. vivax, 17 P. falciparum, 56 P. knowlesi, 7 P. malariae, 1 mixed P. knowlesi/P. vivax, with 4 excluded). When compared against multiplex PCR, HtLAMP-Pv demonstrated a sensitivity for P. vivax of 95% (95% CI 87–99%); 61/64), and specificity of 100% (95% CI 86–100%); 25/25) when P. knowlesi samples were excluded. HtLAMP-Pv testing of 112 samples from asymptomatic community controls, 7 of which had submicroscopic P. vivax infections by PCR, showed a sensitivity of 71% (95% CI 29–96%; 5/7) and specificity of 93% (95% CI87-97%; 98/105). Conclusion: This novel HtLAMP-P. vivax assay has the potential to be a useful field applicable molecular diagnostic test for P. vivax infection in elimination settings.

AB - Introduction: Plasmodium vivax malaria has a wide geographic distribution and poses challenges to malaria elimination that are likely to be greater than those of P. falciparum. Diagnostic tools for P. vivax infection in non-reference laboratory settings are limited to microscopy and rapid diagnostic tests but these are unreliable at low parasitemia. The development and validation of a high-throughput and sensitive assay for P. vivax is a priority. Methods: A high-throughput LAMP assay targeting a P. vivax mitochondrial gene and deploying colorimetric detection in a 96-well plate format was developed and evaluated in the laboratory. Diagnostic accuracy was compared against microscopy, antigen detection tests and PCR and validated in samples from malaria patients and community controls in a district hospital setting in Sabah, Malaysia. Results: The high throughput LAMP-P. vivax assay (HtLAMP-Pv) performed with an estimated limit of detection of 1.4 parasites/ μL. Assay primers demonstrated cross-reactivity with P. knowlesi but not with other Plasmodium spp. Field testing of HtLAMP-Pv was conducted using 149 samples from symptomatic malaria patients (64 P. vivax, 17 P. falciparum, 56 P. knowlesi, 7 P. malariae, 1 mixed P. knowlesi/P. vivax, with 4 excluded). When compared against multiplex PCR, HtLAMP-Pv demonstrated a sensitivity for P. vivax of 95% (95% CI 87–99%); 61/64), and specificity of 100% (95% CI 86–100%); 25/25) when P. knowlesi samples were excluded. HtLAMP-Pv testing of 112 samples from asymptomatic community controls, 7 of which had submicroscopic P. vivax infections by PCR, showed a sensitivity of 71% (95% CI 29–96%; 5/7) and specificity of 93% (95% CI87-97%; 98/105). Conclusion: This novel HtLAMP-P. vivax assay has the potential to be a useful field applicable molecular diagnostic test for P. vivax infection in elimination settings.

UR - http://www.scopus.com/inward/record.url?scp=84959290432&partnerID=8YFLogxK

U2 - 10.1371/journal.pntd.0004443

DO - 10.1371/journal.pntd.0004443

M3 - Article

VL - 10

SP - 1

EP - 16

JO - PLoS Neglected Tropical Diseases

JF - PLoS Neglected Tropical Diseases

SN - 1935-2727

IS - 2

ER -