After collecting calcareous corpuscles from plerocercoid of Spirometra mansoni (sparganum), we evaluated the antigenic values of calcareous corpuscles binding proteins obtained from the cyst fluid of Taenia solium metacestodes. Immunoblot analysis revealed that cysticercosis patient sera strongly recognized 10 and 95 kDa calcareous corpuscles binding proteins. This result demonstrated that calcareous corpuscles are bound with major secretory antigenic proteins, which is possibly involved in the secretory pathways of the 10 and 95 kDa proteins presenting in the cyst fluid of T. solium metacestodes.
Animals ; Antigens, Helminth/*analysis ; Carrier Proteins/*immunology ; Cysticercosis/diagnosis/immunology ; Helminth Proteins/*immunology ; Humans ; Immunoblotting/methods ; Molecular Weight ; Serologic Tests ; Sparganum ; Taenia solium/*chemistry/immunology

We purified specific 31/36 kDa antigenic molecules from sparganum in different intermediate hosts (snakes and mice) and analyzed their monosaccharide compositions. Compositional analysis showed that glucose and mannose concentrations were 2-3 fold higher in the 31/36 kDa molecule purified from snakes than those from mice. This result implies that antigenic glycoproteins of sparganum from snakes might be modified in mammalian sparganosis with respect to their carbohydrate composition.
Animals ; Antigens, Helminth/*chemistry ; Human ; Mice ; Monosaccharides/*analysis ; Snakes/parasitology ; Sparganum/immunology ; Spirometra/*immunology ; Support, Non-U.S. Gov't

Three dogs were experimentally infected with Dirofilaria immitis. All dogs were euthanised at 30, 36 and 37 weeks after inoculation of D. immitis for the recovery of adult worms. Three cases accounted to 42.91 % recovery of inoculated worms. Serum samples from dogs experimentally inoculated with D. immitis were analyzed by ELISA and immunoblotting methods. Antibody titers of dogs detected by ELISA peaked between 7 and 14 weeks then decreased between weeks 15 to 24 followed by another increase during weeks 25 to 30 and persisted throughout the remainde of the experiment period. Analysis of adult D. immitis protein stained with Coomassie brilliant blue R-250 indicated separately more than 10 bands, and the major bands were 22, 40, 46, 56, 70, 72 and 89 kDa. Antigenic identification of extracts antigens of adults D. immitis by immunoblotting analysis revealed several bands from pooled sera of patent infection (30 weeks after inoculation). The detected bands were 24, 70, 80 and 110 kDa, 22, 72 and 84 kDa, and 58 and 72 kDa in dogs 1, 2 and 3, respectively. Results of antibody titers reached high levels on the 4th molting stage after inoculation of infective larva (L3), and reinforced previous findings that high molecular weight regions are detected in young animals.
Animals ; Antibodies, Helminth/analysis ; Antigens, Helminth/analysis/chemistry ; Autopsy ; Dirofilaria immitis/chemistry/*immunology ; Dirofilariasis/*immunology/parasitology ; Dog Diseases/*immunology/*parasitology ; Dogs/*immunology/*parasitology ; Electrophoresis, Polyacrylamide Gel ; Enzyme-Linked Immunosorbent Assay ; Female ; Immunoblotting ; Male ; Time Factors

The antigenic characterizations and serological reactions of human liver flukes, Clonorchis sinensis and Opisthorchis viverrini, were analyzed by immunoblot. The antigenic profiles of the crude extract of Clonorchis contained major proteins of 8, 26-28, 34-37, 43, and 70 kDa, and those of Opisthorchis 34-37, 43, 70, and 100 kDa. Of these, the 8, 26-28 and 34-37 kDa bands of Clonorchis and the 100 kDa of Opisthorchis were major components of each excretory-secretory antigen. The 8 and 26-28 kDa bands were specific to Clonorchis but the 100 kDa of Opisthorchis cross-reacted with the sera of clonorchiasis, and the 34-37, 70 and 100 kDa bands cross-reacted with sera of other helminthiases. The frequency and intensity of the immunoblot reactions were positively correlated with the intensity of the liver fluke infection.
Animals ; Antigens, Helminth/analysis/*chemistry/immunology ; Clonorchiasis/*diagnosis ; Clonorchis sinensis/*immunology ; Cross Reactions ; Electrophoresis, Polyacrylamide Gel/veterinary ; Humans ; Immunoblotting/veterinary ; Molecular Weight ; Opisthorchiasis/*diagnosis ; Opisthorchis/*immunology

Dirofilaria immitis (heartworm) infections affect domestic dogs, cats, and various wild mammals with increasing incidence in temperate and tropical areas. More sensitive antibody detection methodologies are required to diagnose asymptomatic dirofilariasis with low worm burdens. Applying current transcriptomic technologies would be useful to discover potential diagnostic markers for D. immitis infection. A filarial homologue of the mammalian translationally controlled tumor protein (TCTP) was initially identified by screening the assembled transcriptome of D. immitis (DiTCTP). A BLAST analysis suggested that the DiTCTP gene shared the highest similarity with TCTP from Loa loa at protein level (97%). A histidine-tagged recombinant DiTCTP protein (rDiTCTP) of 40 kDa expressed in Escherichia coli BL21 (DE3) showed immunoreactivity with serum from a dog experimentally infected with heartworms. Localization studies illustrated the ubiquitous presence of rDiTCTP protein in the lateral hypodermal chords, dorsal hypodermal chord, muscle, intestine, and uterus in female adult worms. Further studies on D. immitis-derived TCTP are warranted to assess whether this filarial protein could be used for a diagnostic purpose.
Animal Structures/chemistry ; Animals ; Antibodies, Helminth/blood ; Antigens, Helminth/chemistry/*genetics/immunology/*isolation & purification ; Cloning, Molecular ; Dirofilaria immitis/chemistry/*genetics/immunology ; Disease Models, Animal ; Dogs ; Escherichia coli/genetics ; Gene Expression ; Molecular Sequence Data ; Molecular Weight ; Recombinant Fusion Proteins/chemistry/genetics/immunology/isolation & purification ; Sequence Analysis, DNA ; Tumor Markers, Biological/chemistry/*genetics/immunology/*isolation & purification

A highly specific antigenic protein of 31 kDa from plerocercoid of Spirometra mansoni (sparganum) was obtained by gelatin affinity and Mono Q anion-exchange column chromatography. The purified 31 kDa protein was subjected to N-glycan enzymatic digestion for structural analysis. The relative electrophoretic mobility was analyzed by SDS-PAGE, before and after digestion. On SDS-PAGE after enzymatic digestion, the 31 kDa protein showed a molecular shift of approximately 2 kDa, which indicated the possession of complex N-linked oligosaccharides (N-glycosidase F sensitive) but not of high-mannose oligosaccharides (endo-beta-N-acetylglucosaminidase H, non-sensitive). Chemically periodated 31 kDa protein showed statistically non-significant changes with human sparganosis sera by enzyme linked immunosorbent assay (ELISA). Therefore, the dominant epitopes of the 31 kDa molecule in human sparganosis were found to be mainly polypeptide, while N-glycans of the antigenic molecule in sparganum was minimal in anti-carbohydrate antibody production.
Animals ; Antigens, Helminth/*analysis/chemistry/immunology/metabolism ; Carbohydrates/analysis/immunology ; Chromatography, Affinity ; Chromatography, Ion Exchange ; Electrophoresis, Polyacrylamide Gel ; Enzyme-Linked Immunosorbent Assay ; Epitopes/analysis/immunology ; Glucosaminidase/metabolism ; Human ; Peptide-N4- (N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism ; Periodic Acid/chemistry ; Sparganosis/*parasitology ; Sparganum/immunology/metabolism ; Spirometra/immunology/*metabolism ; Support, Non-U.S. Gov't

Many immune down-regulatory molecules have been isolated from parasites, including cystatin (cystain protease inhibitor). In a previous study, we isolated and characterized Type I cystatin (CsStefin-1) of the liver fluke, Clonorchis sinensis. To investigate whether the CsStefin-1 might be a new host immune modulator, we induced intestinal inflammation in mice by dextran sodium sulfate (DSS) and treated them with recombinant CsStefin-1 (rCsStefin-1). The disease activity index (DAI) increased in DSS only-treated mice. In contrast, the DAI value was significantly reduced in rCsStefin-1-treated mice than DSS only-treated mice. In addition, the colon length of DSS only-treated mice was shorter than that of rCsStefin-1 treated mice. The secretion levels of IFN-gamma and TNF-alpha in the spleen and mesenteric lymph nodes (MLNs) were significantly increased by DSS treatment, but the level of TNF-alpha in MLNs was significantly decreased by rCsStefin-1 treatment. IL-10 production in both spleen and MLNs was significantly increased, and IL-10+F4/80+ macrophage cells were significantly increased in the spleen and MLNs of rCsStefin-1 treated mice after DSS treatment. In conclusion, rCsStefin-1 could reduce the intestinal inflammation occurring after DSS treatment, these effects might be related with recruitment of IL-10 secreting macrophages.
Animals ; Antigens, Differentiation/analysis ; Clonorchis sinensis/*enzymology ; Colon/pathology ; Cystatins/*metabolism ; Cytokines/secretion ; Dextran Sulfate/toxicity ; Female ; Helminth Proteins/*metabolism ; Immunologic Factors/*metabolism ; Inflammation/chemically induced/*pathology ; Interleukin-10/analysis ; Intestines/*drug effects/pathology ; Lymph Nodes/immunology ; Macrophages/chemistry/*immunology ; Mice ; Mice, Inbred C57BL ; Severity of Illness Index ; Spleen/immunology