Development of a defined media for culture of Leishmania amastigotes

Abstract

Leishmaniasis is caused by parasites belonging to the genus Leishmania. It is transmitted to different hosts including humans, dogs and rodents through the bite of sand flies. The existing insect and mammalian models to study Leishmania’s biology are restrained by issues of the contaminations, validation and phenotypic variations. Serum-based culturing systems gained momentum to investigate the pathogenicity of the Leishmania by maintaining the parasites over multiple passages, but their use could not be extrapolated to carry the metabolic and proteomic studies in Leishmania due to phenotypic complexities. This study will try to test the feasibility of growing Leishmania amastigotes over multiple passages in the serum-free, simple and defined culture medium at different pH levels. We used the Nayak medium (NM), which was previously developed to support the growth of Leishmania promastigotes, to investigate the growth of L. mexicana amastigotes and promastigotes under different pH and temperature conditions. HOMEM and SDM media were used as control media for promastigotes and amastigotes, respectively. The results showed that NM can support the survival and duplication of the promastigotes until the sixth day after inoculation at pH 7.4 and 27 0C. The growth of promastigotes in HOMEM was 10-fold better than in the NM. Similarly, amastigotes were survived and replicated till day seven of incubation in NM medium with pH 5.5 and 33 0C as an incubation temperature. This study also revealed the capability of NM to transform promastigotes into amastigote-like forms under different pH conditions. The future work should focus on investigating the protein contents of amastigotes and promastigotes in NM media under different temperature and pH conditions.