Comparison of loop-mediated isothermal amplification (LAMP) and PCR for the diagnosis of infection with Trypanosoma brucei ssp. in equids in The Gambia

Gummery, L., Jallow, S., Raftery, A. G. , Bennet, E. , Rodgers, J. and Sutton, D. G.M. (2020) Comparison of loop-mediated isothermal amplification (LAMP) and PCR for the diagnosis of infection with Trypanosoma brucei ssp. in equids in The Gambia. PLoS ONE, 15(8), e0237187. (doi: 10.1371/journal.pone.0237187) (PMID:32833981) (PMCID:PMC7444819)

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Abstract

Introduction: Infection of equids with Trypanosoma brucei (T. brucei) ssp. is of socioeconomic importance across sub-Saharan Africa as the disease often progresses to cause fatal meningoencephalitis. Loop-mediated isothermal amplification (LAMP) has been developed as a cost-effective molecular diagnostic test and is potentially applicable for use in field-based laboratories. Part I: Threshold levels for T. brucei ssp. detection by LAMP were determined using whole equine blood specimens spiked with known concentrations of parasites. Results were compared to OIE antemortem gold standard of T. brucei-PCR (TBR-PCR). Results I: Threshold for detection of T. brucei ssp. on extracted DNA from whole blood was 1 parasite/ml blood for LAMP and TBR-PCR, and there was excellent agreement (14/15) between tests at high (1 x 103/ml) concentrations of parasites. Detection threshold was 100 parasites/ml using LAMP on whole blood (LWB). Threshold for LWB improved to 10 parasites/ml with detergent included. Performance was excellent for LAMP at high (1 x 103/ml) concentrations of parasites (15/15, 100%) but was variable at lower concentrations. Agreement between tests was weak to moderate, with the highest for TBR-PCR and LAMP on DNA extracted from whole blood (Cohen’s kappa 0.95, 95% CI 0.64–1.00). Part II: A prospective cross-sectional study of working equids meeting clinical criteria for trypanosomiasis was undertaken in The Gambia. LAMP was evaluated against subsequent TBR-PCR. Results II: Whole blood samples from 321 equids in The Gambia were processed under field conditions. There was weak agreement between LWB and TBR-PCR (Cohen’s kappa 0.34, 95% CI 0.19–0.49) but excellent agreement when testing CSF (100% agreement on 6 samples). Conclusions: Findings support that LAMP is comparable to PCR when used on CSF samples in the field, an important tool for clinical decision making. Results suggest repeatability is low in animals with low parasitaemia. Negative samples should be interpreted in the context of clinical presentation.

Item Type:Articles
Additional Information:Funding: This work was supported by The Royal Society of Tropical Medicine and Hygiene (Awarded to LG; Grant no. GR000850; https://rstmh.org/), The Donkey Sanctuary (grant awarded to DGMS; https://www.thedonkeysanctuary.org.uk/); and The University of Glasgow Small Grants Fund (DGMS).
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Bennet, Dr Euan and Gummery, Lauren and Raftery, Alexandra and Sutton, Professor David and Rodgers, Dr Jean
Creator Roles:
Gummery, L.Conceptualization, Formal analysis, Funding acquisition, Investigation, Methodology, Visualization, Writing – original draft, Writing – review and editing
Raftery, A. G.Methodology, Resources, Writing – review and editing
Bennet, E.Formal analysis
Rodgers, J.Methodology, Supervision, Writing – review and editing
Sutton, D. G.M.Conceptualization, Funding acquisition, Investigation, Methodology, Supervision, Writing – review and editing
Authors: Gummery, L., Jallow, S., Raftery, A. G., Bennet, E., Rodgers, J., and Sutton, D. G.M.
College/School:College of Medical Veterinary and Life Sciences > Institute of Biodiversity Animal Health and Comparative Medicine
College of Medical Veterinary and Life Sciences > School of Biodiversity, One Health & Veterinary Medicine
Journal Name:PLoS ONE
Publisher:Public Library of Science
ISSN:1932-6203
ISSN (Online):1932-6203
Copyright Holders:Copyright © 2020 Gummery et al.
First Published:First published in PLoS ONE 15(8):e0237187
Publisher Policy:Reproduced under a Creative Commons license
Data DOI:10.5525/gla.researchdata.976

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