Low-cost liquid medium for in vitro cultivation of Leishmania parasites in low-income countries

Endalamaw Gadisa,Sven Britton,Hannah Akuffo,Amha Kebede,Liv Eidsmo,Dawit Wolday,Geremew Tasew

doi:10.3402/gha.v2i0.2046

Abstract

Background: Prompt laboratory diagnosis and initiation of treatment are effective components of leishmaniasis control. Detection of Leishmania parasites by ex-vivo culture of lesion scrapings is considered a definitive diagnostic method preceding initiation of treatment.Objective: A pilot study to find alternative medium that could reduce the cost of culturing from patient lesions for diagnosing leishmaniasis.Method: GALF-1 medium was formulated in our lab from locally available inexpensive solutions and powders in the presence of urine from healthy individuals. Amastigote to promastigote transformation, recovery of parasites after cryopreservation, cost and mass cultivation was compared using the following media: GALF-1, RPMI 1640, and conventional Locke's semi-solid medium (LSSM), a modifications of Novy–MacNeal–Nicolle culture media, which uses Locke's solution as an overlayResults: GALF-1 preparation was cheap and the components available in low-income countries such as Ethiopia. Preparation was simple, not requiring autoclaving and extra distilled water. GALF-1 was able to transform amastigotes from Ethiopian patients’ samples and could be used to cultivate promastigotes in large quantities. GALF-1 decreased Leishmania culture costs by ∼80–95% compared to LSSM and RPMI 1640, respectively. Promastigotes cultured with GALF-1 could be cryopreserved in liquid nitrogen with comparable re-culture potential.Conclusion: Affordability of diagnostic assays is a key issue for endemic resource-poor countries and the possibility to cut the cost of the efficient culture method for diagnosis through the use of inexpensive, locally formulated reagents could improve the diagnosis of leishmaniasis in Ethiopia and in other low-income countries.

Highlights

Prompt laboratory diagnosis and initiation of treatment are effective components of leishmaniasis control
We describe a pilot study using a new low-cost liquid medium for Leishmania culture designated GALF-1
Parasites inoculated into RPMI reached stationary phase (Â7.1 )106/ml) within an average of 4Á5 days followed by a declining phase (Â5.6 )105/ ml) between the sixth and ninth day

Summary

Introduction

Prompt laboratory diagnosis and initiation of treatment are effective components of leishmaniasis control. Method: GALF-1 medium was formulated in our lab from locally available inexpensive solutions and powders in the presence of urine from healthy individuals. Amastigote to promastigote transformation, recovery of parasites after cryopreservation, cost and mass cultivation was compared using the following media: GALF-1, RPMI 1640, and conventional Locke’s semi-solid medium (LSSM), a modifications of NovyÁMacNealÁNicolle culture media, which uses Locke’s solution as an overlay. Results: GALF-1 preparation was cheap and the components available in low-income countries such as Ethiopia. Conclusion: Affordability of diagnostic assays is a key issue for endemic resource-poor countries and the possibility to cut the cost of the efficient culture method for diagnosis through the use of inexpensive, locally formulated reagents could improve the diagnosis of leishmaniasis in Ethiopia and in other low-income countries

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Results

Conclusion