Validating the use of Loop-mediated isothermal amplification (LAMP) to detect Histoplasma capsulatum in equine clinical samples stored on FTA cards

GRANT PROPOSAL

CONTENTS

TITLE Page Number
INTRODUCTION 3
LITERATURE REVIEW 4-5
HYPOTHESIS & OBJECTIVES 6
METHODOLOGY 7-9
REQUIREMENTS 10
PROJECT IMPACT 11
REFERENCES 12-13

INTRODUCTION
In a broad range of point-of-care applications, that impacts the health of both animals and humans, there is a need for rapid detection techniques, to detect and report pathogenic infections. The purpose of this project is to investigate the possibility of adapting the LAMP method in detection of Histoplasmosis directly from the samples collected from equines and stored on Whatmann FTA cards.
Histoplasma capsulatum, a dimorphic fungus causes Histoplasmosis in both humans and animals. As very few rapid assays are present for this disease, timely detection is hence problematic in resource-challenged countries (1). The available identification test kits from Gen-Probe can rapidly identify H. capsulatumn cultures but are very expensive and not easily accessible in developing countries. Several other detection techniques for Histoplasmosishave been established (2, 3, 4), but failed in subjecting to large-scale inter-laboratory evaluation. This leads to the development of an alternative detection technique for histoplasmosis, which is an affordable molecular identiﬁcation tool, a loop-mediated isothermal ampliﬁcation (LAMP) assay, which do not require expensive types of equipment and so is easy to perform in developing or underdeveloped countries.
Epizootic lymphangitis (EZL) caused by H. capsulatum is traditionally a disease of equids, specifically horses, in the endemic parts of Africa. Due to lack of recourses and infrastructure, blood samples and pus samples of infected animals have collected and preserved on FTA cards and transferred to the University of Liverpool (5). These samples can be used to confirm the presence of disease by different amplification techniques. This project will provide further insight into the applicability of LAMP technology to samples stored on FTA cards, and take steps to examine its feasibility in a low resource setting.

LITERATURE REVIEW
Notomi et al in 2000 first stated novel technique termed loop-mediated isothermal ampliﬁcation (LAMP) (6). It is a single step nucleic acid amplification reaction, in which amplification of target nucleic acid sequence occurs with high sensitivity and specificity under isothermal condition thereby reducing risk of contamination; therefore, it is proved to be faster and convenient

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