In vitro Sensitivity Test of Metronidazole by Using Clinical Isolates of E. histolytica and E. bangladeshi

Abstract

Infectious diseases remain the major cause of morbidity and mortality worldwide including Bangladesh. Entamoeba histolytica is the etiological agent of amoebic dysentery and amoebic liver abscess (ALA). Worldwide, 40–50 million symptomatic cases of amoebiasis occur annually and 70,000 to 100,000 deaths are due to this infection. In amoebiasis Metronidazole, a 5-nitroimidazole drug is the drug of choice for intestinal disease caused by Entamoeba histolytica. In 2010-2011 a new species of Entamoeba named E. bangladeshi has been identified, which was positive for Entamoeba organism but negative for E. histolytica, E. dispar and E. moshkovskii by PCR. It was found in the feces of children living in Mirpur, Dhaka, Bangladesh. Till now no studies have been carried out to determine the sensitivity of Metronidazole against this new species. Therefore, the objective of the study is to determine the sensitivity of metronidazole against clinical isolate of E. bangladeshi. The clinical isolates of E. histolytica and E. bangladeshi were treated with metronidazole at different concentrations (1.28, 0.64, 0.32, 0.16, 0.08, and 0.04 µg/ml). Drug sensitivity assay of the samples was carried out by using microtiter plates containing 100 µg/ml of parasite suspension (1×106 parasites/ml). Plates were incubated at 370C. After 4 hours incubation the viable parasites were counted by haemocytometer under the microscope. Viable counts of the E. histolytica and E. bangladeshi in each concentration of drugs were compared to the control. Result showed that viable cell count is significantly reduced in the treatment groups when compared with the control (p=<0.05). However, the viable cell count of E. bangladeshi is similar when it was compared with cell count of E. histolytica and no significant difference has been observed between these two groups. We have also found that inhibition of parasite is occurred in a dose dependent manner. Cell inhibition is maximum at the highest concentration of Metrondazole. The cell inhibition is 32% and 18% when Metronidazole treated against E. histolytica and E. bangladeshi respectively. We can conclude that treatment with Metronidazole drug may be a useful treatment to inhibit the growth of E. bangladeshi. Further study is needed to explore different drugs at different concentrations and time intervals to determine the in vitro sensitivity against E. bangladeshi isolates.