Tropical
Journal of Pharmaceutical Research, June 2011;
10(3):
255-263
doi:
10.4314/tjpr.v10i3.2
Abstract
Purpose:
To develop a highly accurate molecular assay for
evaluating the efficacy of metronidazole and some
antifungal agents with meglumine antimoniate against L.
infantum visceral leishmaniasis in different mouse
tissues.
Methods:
The assay was performed with the Light-Cycler system
using SYBR Green I and primers amplifying ca. 120-bp
fragment from minicircles of the kinetoplast DNA (kDNA).The
mice were divided into two groups. Group I served to
evaluate drug activity and parasite load while Group II
was assigned to identify possible synergistic activity
between meglumine (which is highly effective in the
liver but less effective in the spleen) and drugs with
significant activity against spleen infection.
Results:
The assay was able to detect as little as 100 fragments
of
L. major
DNA per reaction, which is equivalent to 0.1 parasites.
The standard curve designed for quantitation of
parasites showed linearity over at least 6-log
DNA
concentration range, corresponding to 0.1 to 104 parasites
per reaction with a correlation coefficient of 0.979.
Metronidazole, ketoconazole, fluconazole, itraconazole
and terbinafine were less effective than antimonial
agents in reducing hepatic parasite load while
ketoconazole potentiated the effect of meglumine
antimoniate reference therapy through its marked
activity against spleen infection (L. infantum visceral
leishmaniasis).
Conclusion:
The assay technique is accurate, sensitive, and rapid
for the detection of kDNA and would be of great help to
scientists who use animals to monitor the efficacy of
anti-leishmanial drugs or vaccines.
Keywords:
Leishmaniasis, Molecular diagnosis, Kinetoplast DNA,
Real-Time LightCycler(LC)-PCR, Spleen infection,
Antifungal agents.
