Alirahmi Heshmatollah, Farahnak Ali, Golmohamadi Taghi, Esharghian Mohammad Reza
Department of Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
Acta Med Iran. 2010 Nov-Dec;48(6):367-70.
Fascioliasis is a worldwide parasitic disease in human and domestic animals. The causative agents of fascioliasis are Fasciola hepatica and Fasciola gigantica. In the recent years, fasciola resistance to drugs has been reported in the many of publications. Fasciola spp has detoxification system including GST enzyme which may be responsible for its resistance. Therefore , the aim of the study was to assay of GST enzyme activity in fasciola parasites. Fasciola gigantica and Fasciola hepatica helminths were collected from abattoir as a live and cultured in buffer media for 4 h at 37 °C. Excretory-Secretory products were collected and stored in -80◦C. F. gigantica and Fasciola hepatica were homogenized with homogenizing buffer in a glass homogenizer to prepare of somatic extract. Suspension was then centrifuged and supernatant was stored at -80°C. In order to assay the enzyme activity, excretory-secretory and somatic extracts in the form of cocktails (potassium phosphate buffer, reduced glutathione and 1-chloro-2,4-dinitrobenzene substrates) were prepared and their absorbance recorded for 5 minutes at 340 nm. The total and specific GST activity of F. gigantica somatic and ES products were obtained as 2916.00, 272.01 micromole/minute and 1.33, 1.70 micromole/minute/mg protein, respectively. Fasciola hepatica also showed 2705.00, 276.86 micromole/minute and 1.33, 1.52 micromole/minute/mg protein, respectively. These results are important for analysis of parasite survival / resistance to drugs which use for treatment of fascioliasis.
肝片吸虫病是一种在人类和家畜中广泛传播的寄生虫病。肝片吸虫病的病原体是肝片吸虫和巨片吸虫。近年来,许多出版物都报道了片吸虫对药物产生耐药性。片吸虫属具有包括谷胱甘肽S-转移酶(GST)在内的解毒系统,这可能是其产生耐药性的原因。因此,本研究的目的是测定片吸虫寄生虫中的GST酶活性。从屠宰场收集活的巨片吸虫和肝片吸虫,并在缓冲培养基中于37℃培养4小时。收集排泄-分泌产物并储存在-80℃。将巨片吸虫和肝片吸虫在玻璃匀浆器中用匀浆缓冲液匀浆以制备体细胞提取物。然后将悬浮液离心,上清液储存在-80℃。为了测定酶活性,制备了呈混合液形式(磷酸钾缓冲液、还原型谷胱甘肽和1-氯-2,4-二硝基苯底物)的排泄-分泌和体细胞提取物,并在340nm处记录其吸光度5分钟。巨片吸虫体细胞和排泄-分泌产物的总GST活性和比GST活性分别为2916.00、272.01微摩尔/分钟和1.33、1.70微摩尔/分钟/毫克蛋白质。肝片吸虫也分别显示为2705.00、276.86微摩尔/分钟和1.33、1.52微摩尔/分钟/毫克蛋白质。这些结果对于分析用于治疗肝片吸虫病的寄生虫存活/耐药性具有重要意义。