Fasciola gigantica: the in vitro effects of artesunate as compared to triclabendazole on the 3-weeks-old juvenile.

The in vitro effect of artesunate (ATS) on the 3-week-old juveniles of Fasciola gigantica was compared with triclabendazole (TCZ) by incubating the parasites in M-199 medium containing the drugs at concentrations of 20, 40, and 80 µg/ml for 1, 3, 6, 12, and 24h. The anthelmintic activities of these drugs were evaluated based on the relative motility value (RM) and the alterations of the tegument as observed by scanning (SEM) and transmission (TEM) electron microscopy. The RM values of TCZ-treated flukes decreased significantly from 6 to 24h for all dosages. For ATS-treated flukes, RM value decreased markedly from 12 to 24h, but the rates of decline were less than TCZ at the same doses. When observed by SEM, the tegument showed similar sequence of morphological changes after treatments with both drugs, comprising of swelling of tegumental ridges, followed by blebbing and later rupturing of the blebs, leading to erosion and lesion, and disruption of the tegument. When examined by TEM, ultrastructural changes in the tegument and associated structures after treatments with TCZ and ATS were similar which comprised of swelling, blebbing of the tegument, dilation of basal infoldings, and depolymerization of the microtrabecular network. After a longer incubation time, the tegument was completely sloughed off and the tegument cell bodies became necrotic. Additionally, in ATS-treated flukes, mitochondria showed severe swelling, rupturing of outer membrane, and their interior filled with flocculent materials.

Fasciola gigantica and Schistosoma mansoni: vaccine potential of recombinant glutathione S-transferase (rFgGST26) against infections in mice.

Recombinant Fasciola gigantica glutathione S-transferase (rFgGST26) was expressed in Escherichia coli. This protein had 86% and 56% sequence identity with 26 kDa GST from Fasciola hepatica and Schistosoma mansoni, respectively. Polyclonal antibody raised in ICR mice against rFgGST26 recognized immunoblotted 26 kDa native GSTs from F. gigantica and S. mansoni. rFgGST26 was used as a vaccine in combination with Freund's adjuvant to evaluate the induction of immune responses and protection against F. gigantica and S. mansoni infection in mice. Mice were immunized via subcutaneous (s.c.), intramuscular (i.m.) or intradermal (i.d.) routes. Strong protection (77-84%) against F. gigantica was observed in all routes. Immunization via s.c. route induced immune response with IgG1 isotype predominating, while i.m. and i.d. routes resulted in mixed IgG1/IgG2a immune responses. Passive intraperitoneal transfer of IgG1 predominating antisera from s.c. rFgGST26-immunized donors to naive recipient mice resulted in 47% protection against F. gigantica infection. This suggests that the mechanism of resistance depends on the presence of specific antibody against rFgGST26. Immunization with rFgGST26 via i.m. and i.d. routes resulted in significant cross protection (55%) against S. mansoni infection in the i.d. route with mixed IgG1/IgG2a response with IgG1 isotype predominating. This indicated that rFgGST26 is a good vaccine candidate against F. gigantica in mice and could also provide cross protection against S. mansoni.

Toxicity of copper in butterfish (Poronotus triacanthus): tissues accumulation and ultrastructural changes.

The present study investigated the accumulation of copper in various tissues and ultrastructural alterations in butterfish, Poronotus triacanthus. In acute toxicity test, butterfish were exposed to 250 microg/L copper for 24 h. In subacute toxicity test, fish were exposed to 25 microg/L copper for 7 days and then returned to normal water for 48 h. The levels of copper accumulation in the tissues were determined by using an atomic absorption spectrometer. After the 7 day exposure, the highest level of copper was found in the liver, followed by kidney, gills, and muscle tissues (3.64, 0.62, 0.59, and 0.34 microg/L, respectively). The recovery group has shown some reduction in copper level in these tissues when compared with those of the 7 day exposure group (3.09, 0.34, 0.30, and 0.27 microg/L, respectively). In gills, the major changes such as filament cell proliferation, increase in intercellular spaces, epithelial lifting, and thickening of the filament and lamellar epithelium were observed. In liver, the major changes such as swollen mitochondria, fragmented in rough endoplasmic reticulum, increases in number and size of lysosomes and lipid droplets. Infiltration of leukocytes, increasing hepatocyte size with pyknotic nuclei, and presence of vacuoles were also observed. In kidney, the changes included alterations of the first proximal tubules, as well as vacuolization of the cytoplasm, proliferation of lysosomes and mitochondria, dilation of endoplasmic reticulum, and finally, cell necrosis. The transmission electron microscopic analysis revealed that the 7 day exposure group had more severe effect in tissue alterations than the 24 h exposure group. Tissue regeneration was also observed in the 48 h recovery group.

Schistosoma mekongi: the in vitro effect of praziquantel and artesunate on the adult fluke.

The efficacy and tolerance of 80 microg/ml praziquantel (PZQ) and 40 microg/ml artesunate (ATS) against adult stage Schistosoma mekongi in vitro were investigated after 3, 6, 12, and 24h incubation by monitoring worm motility and compared tegumental changes using scanning electron microscopy (SEM). Thirty mice were infected with S. mekongi cercaria for 49 days. Adult worms were collected by perfusion method and prepared for in vitro study. Contraction and decreased motor activity were observed after as little as 3h incubation with PZQ and ATS. Some of the worms were immobile 12h after exposure, and died within 24h. The tegument of S. mekongi showed severe swelling, vacuolization and disruption, fusion of the tegumental ridges, collapse and peeling. After 12-24h incubation, PZQ induced similar but they less severe, tegumental changes to those observed after exposure to ATS. The direct observation of the fluke motility and SEM study suggest that ATS is more effective than PZQ in causing tegumental damage in adult S. mekongi, and provides a basis for subsequent clinical trials.

Eurytrema pancreaticum: the in vitro effect of praziquantel and triclabendazole on the adult fluke.

The efficacy and tolerance of the 80 microg/ml praziquantel (PZQ) and 40 microg/ml triclabendazole (TCZ) against adult stage Eurytrema pancreaticum in vitro were investigated at 3, 12, and 15 h incubation. Motility of the flukes and histopathological changes were studied. Sudden paralysis and death were observed after exposed to PZQ as early as 3h incubation. In contrast, the TCZ treated flukes showed active mobility at all intervals. By light microscopic examination, severe damages in various organs such as tegument, muscle, and testes were observed early at 12h incubation of these drugs. PZQ caused more severe damage to flukes than TCZ. There were vigorous contraction of musculature, progressive shrinkage of circular and longitudinal muscles, vacuolization and disintegration of the tegument disrupting the worms' outer surface including detachment of spines in the PZQ treatment. The cells in testes were slightly increased in size and followed by degeneration leaving several hollow spaces. The uterus and vitelline glands remained unaffected. The direct observation of the fluke motility and light microscopic study highly suggested that PZQ was more effective than TCZ treatment for the eurytremiasis infection.

Effects of praziquantel and artesunate on the tegument of adult Schistosoma mekongi harboured in mice.

The effects of praziquantel and artesunate on the tegument of adult Schistosoma mekongi harboured in mice were compared using scanning electron microscopy (SEM). Forty-two mice infected with S. mekongi for 49 days were treated intragastrically with either 300 mg/kg praziquantel or 300 mg/kg artesunate. Mice were sacrificed 1 or 3 days post-treatment. Worms were collected by perfusion and examined by SEM. One to 3 days after administration of artesunate, the tegument of S. mekongi showed severe swelling, vacuolization, fusion of the tegumental ridges and loss or shortening of the spines on the trabeculae, collapse and peeling. Praziquantel induced similar tegumental alterations as those observed after administration of artesunate, but they were less severe. Three days post-treatment, there was evidence of recovery only in the case of praziquantel. The results of our study suggest that artesunate is more effective than praziquantel in causing tegumental damage in adult S. mekongi, and provides a basis for subsequent clinical trials.

Ultrastructural alterations in the liver and kidney of white sea bass, Lates calcarifer, in acute and subchronic cadmium exposure.

Ultrastructural alterations in the liver and kidney of 3-month-old white sea bass, Latescalcarifer, after cadmium exposure were studied by transmission electron microscopy (TEM). One group of fish was exposed to a cadmium concentration of 10 mg/L (acute) for 96 h in a static system, and another group was exposed to cadmium concentrations of 0.8 and 3 mg/L cadmium (subchronic) for 3 months in a recirculation closed system. Ultrastructural alterations observed in the hepatocytes included mitochondrial condensation, swelling, and lysis. The rough endoplasmic reticulum (RER) showed dilation, fragmentation, and vesiculation. After subchronic exposure there were numerous large lipid droplets and abundant stored glycogen. Ultrastructural alterations observed in the proximal tubules of the kidney included nuclear degeneration, condensation, and massive swelling of the mitochondria; RER fragmentation and vesiculation. Disorganized brush borders and increased numbers of large hydropic vacuoles and lysosomes were also observed.