Background: We aimed to assess the in vitro effects of the green synthesized silver nanoparticles (Ag NPs) via Thymus vulgaris (thyme) against Leishmania major infection.
Methods: We have prepared T. vulgalis silver nanoparticles (TSNPs) by adding thyme extract to the silver nitrate aqueous solution (0.2 mM), and evaluated their antileishmanial activity. The viability of L. major promastigotes was assessed in the presence of various concentrations of TSNPs by direct counting after 24 h. The MTT assay was used to identify the viability of promastigotes. The same procedures were assessed in uninfected macrophage cells. The apoptotic effects of nanoparticles on L. major promastigotes were determined by flow cytometry assay using annexin staining. To evaluate anti-amastigotes activity of TSNPs, light microscopic observation was used to determine the number of parasites within the macrophages in each well.
Results: The effect of TSNPs on promastigotes and amastigotes of L. major was effective and had a reverse relationship with its concentration. TSNPs, inhibited the growth rate of L. major amastigotes and, the IC50 value of these nanoparticles was estimated 3.02 μg/mL (28 μM) after 72h. The results of flow cytometry showed that the toxic effects of TSNPs on promastigotes after 24 hours were statistically significant (P<0.05) and showed 69.51% of apoptosis.
Conclusion: TSNPs had an inhibitor effect on promastigote and amastigote forms of L. major in vitro. It might be considered as a candidate for the treatment of this infection.
Background: Resistance to artemisinin has threatened major achievements in malaria control, more investigations is needed about resistant strains and related genes. We aimed to induce resistance to artesunate in the Plasmodium falciparum 3D7 strain using intermittent exposure method and comparing P.fk13 gene sequence between susceptible and resistance strains.
Methods: P. falciparum 3D7 strain was cultured according to Trager & Jensen method with some modifications. Serial concentrations between 10-2 mol/l, to 10-7mol/l were prepared, then P. falciparum 3D7 was exposed to each of the dilution to determine IC50 and lethal dose. Sensitivity reduction process was started from the concentration of 10-7mol/l and ended at 10-2mol/l. Exposed parasites were collected after at least 27 days after cultivation in each drug concentration. DNA extraction, PCR and sequencing process were performed to investigate any possible mutations in the P.fk13 gene sequence.
Results: Effectiveness of 10-2mol/l concentration of artemisinin was found as a lethal dose. IC50 value was equal to 5×10-4 mol/l. The resistant strain was provided in the lab, sequenced and registered in the gene bank as P.f Art -2, (accession number MH796123. 1). Alignment of this registered sample showed no mutation in P.f kelch13 gene in comparison with standard strain submitted in the GenBank.
Conclusion: Resistance to artesunate in malaria parasite may occur but with no mutation in the P.f kelch13 gene. Therefore, whole genome sequencing should be applied to determine mutations in resistant strains.
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