Transmission electron microscope study of the ultrastructural changes induced in the tegument and gut of Fasciola hepatica following in vivo drug treatment with clorsulon

Abstract

Using transmission electron microscopy (TEM), both the tegument and gut of Fasciola hepatica were examined in an effort to identify and characterise the ultrastructural changes induced following treatment with the flukicidal drug clorsulon. Male Sprague-Dawley rats infected with F. hepatica were dosed orally at 8-8.5 weeks post-infection with clorsulon at a concentration of 12.5 mg/kg body weight. After 24, 48 and 72 h, rats were sacrificed by cervical dislocation and mature flukes recovered from the bile ducts. After 24 h treatment in vivo, disruption of the tegumental syncytium was concentrated at the apex of the syncytium where a dark band consisting of numerous secretory bodies was present. Some blebbing of the apex had also occurred, "open" bodies were present in this region and the mitochondria were slightly swollen. In the cell bodies, swelling of the mitochondria and their cristae had also occurred and the Golgi complexes appeared to be smaller than normal. The disruption seen after 48 h treatment in vivo was similar but more severe: the frequency of blebbing had increased, as had the number of "open" bodies and the swelling of the mitochondria. Vacuoles had begun to appear in the syncytium-both autophagic and electron-lucent-and swelling of the mucopolysaccharide masses around the basal infolds had occurred. Lipid droplets were observed occasionally. In the cell bodies, autophagic vacuoles had begun to appear and swelling of the mitochondria had increased in severity. After 72 h treatment in vivo, more severe disruption was seen in the tegumental syncytium in which widespread swelling and blebbing of the apex was apparent. The basal infolds had become very badly swollen in a number of specimens and damage to the spines was evident. The mitochondria remained swollen, as did the mucopolysaccharide masses around the basal infolds. Lipid droplets were more frequently observed in the syncytium. In the tegumental cells, swelling of the mitochondria was greater and an increase in the number of autophagic vacuoles was apparent. The gut showed signs of disruption after 24 h treatment in vivo, in that the surface lamellae were disrupted and a build-up of autophagic vacuoles at the apex of the cells had taken place. Swelling of the mitochondria and the cisternae of granular endoplasmic reticulum (gER) was evident. There was a decrease in the number of secretory bodies. After 48 h treatment in vivo, the number of autophagic vacuoles in the gastrodermal cells had increased, the mitochondria and gER remained swollen and the disruption seen to the lamellae was still evident. In the 72 h-treated specimens, the disruption seen in the gastrodermal cells had increased significantly, with severe vacuolation of the apical cytoplasm. An increase in the number of autophagic vacuoles was evident, the mitochondria and the gER remained swollen and lipid droplets were present in the cells.

Electron microscopical study to assess thein vitroeffects of the synthetic trioxolane OZ78 against the liver fluke,Fasciola hepatica

“…This may be due to inhibition of the energy-dependent ion pumps on the tegumental membranes and is evidenced by the swollen mitochondria and cisternae of ger and the presence of lipid droplets, together with the swelling of the basal infolds. Similar changes have been seen following treatment with fasciolicides that are believed to target energy metabolism, including clorsulon, closantel and nitroxynil (Skuce and Fairweather, 1990 ;Meaney et al 2004Meaney et al , 2005Meaney et al , 2007McKinstry et al 2007McKinstry et al , 2009. Alternatively, the flooding may stem from damage to the surface membrane as a result of impairment of production of secretory bodies in the tegumental cells.…”

“…Greater swelling of the basal infolds would lead to the sloughing of the tegument, as has been observed following treatment with other fasciolicides (e.g. Fairweather et al 1986 ;Skuce and Fairweather, 1990 ;Stitt and Fairweather, 1993 ;Anderson and Fairweather, 1995 ;Meaney et al 2004). Clearly, there is disruption of the tegumental osmoregulatory system, which spreads internally to affect the subtegumental tissues.…”

“…There was little secretory activity and the ger was swollen and vesiculated. Vesiculation of the ger has been linked to a state of starvation or stress in the fluke (Robinson and Threadgold, 1975 ;Skuce and Fairweather, 1990 ;Meaney et al 2004Meaney et al , 2005McKinstry et al 2007). Moreover, there were signs that the cells were beginning to break down, as evidenced by the presence of autophagic vacuoles and degenerating mitochondria.…”

Biochemical Characterization and Differential Expression of a 16.5-Kilodalton Tegument-Associated Antigen from the Liver Fluke Fasciola hepatica

“…The tegument carries out several functions, including osmoregulation, secretion, uptake of nutrients, and evasion from host immune responses (22,23). Tegumental proteins of trematodes have been shown to be targets of several anthelmintic drugs (51,52) and vaccine candidates (46,57,65). The importance of the tegumental surface for parasite functions is clear, and due to their accessibility in worms, tegumental proteins are potential targets not only for vaccines but also for new immunodiagnostic tools.…”

Metabolic enzymes of helminth parasites: potential as drug targets

“…It also has weak activity against carbonic anhydrase (EC 2.4.1.1) in the host and, most likely, the parasite [52,53]. The drug causes widespread damage to the ultrastructure of F. hepatica [54][55][56][57].…”