Cross-protection induced by cross-reactive antigen against Fasciola gigantica and Trichinella spiralis infections

Trichinella spiralis/Fasciola gigantica cross-reactive fraction was purified from T. spiralis larval extract by affinity column chromatography in which CNBr-Sepharose 4B was coupled with F. gigantica antibodies. The fraction consisted of six polypeptides of 191KDa, 178KDa, 149KDa, 106KDa. 101 KDa and 32KDa as revealed by SDS-PAGE. Analysis of the free amino acids of the fraction revealed 17 amino acids with high proportions of tyrosine and glutamic. Immunization of rabbits subcutaneously with the cross-reactive fraction in Freund's adjuvant, followed by challenge with F. gigantica metacercariae resulted in reduction in worm burdens reached to 47.8%. While, immunization of rats with the same fraction in Freund's adjuvant, followed by infection with T. spiralis larvae resulted in reduction in worm count reached to 74%. IgG antibody response in rabbits increased due to immunization to reach its maximum value at the time of infection and then decreased gradually up to the end of the experiments; but remained higher than the level in non vaccinated control animals. In rat sera, IgG level increased due to vaccination but the level recorded its optimum value one week post infection and then decreased. Thus, the cross-reactive antigen proved cross-protection with the protection inducing capability against both diseases

Trichinella spiralis: affinity purified antigen based diagnosis and immunoprophylaxis

The current research introduces a trial to develop vaccine candidate against trichenosis. A method of affinity chromatography was adopted to purify a Trichinella spiralis larval extract. The isolated fraction resolved into six bands of 148 KDa, 133 KDa, 118.5 KDa, 101 KDa 98.5 KDa and 79.5 KDa as observed by SDS-PAGE. The diagnostic value of this fraction was checked against antibodies regularly collected from rats experimentally infected with trichinosis compared with that of crude larval extract by ELISA. The crude extract detected the antibodies as early as one week post infection and the maximum level was recorded four weeks post infection. The advantage of the isolated fraction over the crude extract in trichinosis diagnosis was clearly observed at high serum dilution reached to 1:4000. The protective value of the isolated fraction was also investigated. Rats immunized subcutaneously with affinity purified larval extract with Freund's adjuvant showed reduction in worm burden reached to 82%. IgG antibody response in immunized rats was higher than that of control infected animals as measured by ELISA. This response might be partially responsible for the observed protection

Identification and diagnostec evaluation of cross-reactive antigen between hydatid cyst fluid of echinococcusgranulosus and trichinella spiralis larval extract

A cross reactive fraction was isolated from hydatid cyst fluid antigen of E. granulosus using CNBr Sepharose 4B affinity chromatography in which anti-T. spiralis antibodies were coupled with the column. Biochemical characterization of the isolated fraction included the use of SDS-PAGE, isoelectric focusing and amino acid analysis. The fraction showed 5 polypeptides of 165, 95.5, 63.5, 30.6 and 24 KDa. The isoelectric points of these polypeptides were 7.8, 7.2, 6.7, 6.2 and 5.7. The fraction exhibited 17 amino acids and was rich in tyrosine [20.81] and glutamic [15.28] micro gram/100 mg. The fraction proved higher potency in the diagnosis of experimental trichinellosis in rats than echinococcosis in dogs by ELISA. All experimentally infected animals reacted positively, recording 100% diagnostic sensitivity. Collectively, the present study proved that the hydatid cyst fluid cross-reactive fraction could be used in the diagnosis of trichinellosis at different intervals of infection and as early as 1 week post infection

Partial purification and characterization of ascaridia galli diagnostic warm antigen

Partial purification of Ascaridia galli whole worm extract was conducted by Cyanogen bromide Sepharose 4B immunoaffinity column chromatography. The resulted fraction was characterized by sodium dodecyle sulphate polyacrylamide gel electrophoresis [SDS-PAGE] and isoelectric focusing. The fraction was found to be consisted of six bands of 207 KDa, 157 KDa, 110 KDa, 103 KDa, 76 KDa and 41 KDa. This profile was compared with that of whole worm and excretory-secretory antigens. Both antigens were resolved into multiple bands in both high and low molecular weight ranges. The isoelectric focusing of the fraction displayed 8 bands of isoelectric points 7.5, 7.0, 6.8, 6.5, 6.2, 5.8. 5.3 and 4.6. The potency of this fraction in the diagnosis of natural ascaridiosis in chickens was assessed by ELISA compared with that of whole worm and ES antigens. The affinity purified fraction showed higher potentials in the diagnosis of A. galli infection in chickens than whole worm antigen at any sera dilution and than ES antigen at high sera dilutions. While ES antigen of the worms revealed higher diagnostic capabilities than whole worm extract. The current research recommends utilization of the affinity isolated fraction in the diagnosis of natural ascaridiosis in chickens

Fasciola gigantica: Immunization of rabbits with proteins isolated from copropantigen

Two fractions were isolated from coproantigen by ion-exchange chromatography, in which DEAE cellulose was utilized. Both fractions and crude antigen were characterized by SDS-polyacrylamide gel electrophoresis, which revealed 13 bands of molecular weight ranged from 205-31 in crude coproantigen. While, fraction I was resolved into 6 bands of molecular weights of 198, 178, 148, 111, 101 and 45. Fraction II showed seven bands of 191 kDa, 178 kDa, 166 kDa, 118 kDa, 98.5 kDa, 72 kDa and 32 kDa. Fraction II showed higher immunoreactivity than fraction I by ELISA. Three immunoreactive bands of 191 kDa, 118 kDa and 98.5 kDa were identified in fraction II using immunoblot assay. Five bands of 178 kDa, 148 kDa, 111 kDa, 101 kDa and 45 kDa were detected in fraction I. Immunization of rabbits twice with fraction II in Freund's adjuvant with two-week interval, followed by challenge with F. gigantica metacercariae resulted in 66.6% protection from infection. The protection was assessed by the detection of hepatic damage, worm recoveries and antibody response. A high level of IgG response in vaccinated rabbits than control infected ones occurred and was found to be responsible for the recorded protection

Cross - reaction: a common trait among helminthes

The cross-reaction between three important zoonotic helminths, Fasciola gigantica, Trichinella spiralis and Echinococcus granulosus was proved by ELISA. Cross-binding activities in the prepared antisera were strongly directed towards protoscolices and hydatid fluid antigens of E. Granulosus rather than to F. gigantica and T. spiralis antigens. Two sets of polypeptides were identified in each antigen by immunoblot [species-specific and cross-reactive]. The cross-reactive components in F. gigantica antigen were 205 KD, 178 KD, 166 KD, 106 KD, 100 KD, 65 KD, 45 KD and 32 KD. While, cross-reactive molecules in T. spiralis antigen were 205 KD, 191 KD, 166 KD, 148 KD, 132 KD and 32 KD. In protoscolices antigen, six cross-reactive components were identified [205 KD, 191 KD, 149 KD, 106 KD, 45 KD and 32 KD]. Moreover, 205 KD, 190 KD, 177 KD, 149 KD, 103 KD and 33 KD were detected in hydatid fluid antigen by heterologous antisera. Three polypeptides of 205 KD, 149 KD and 32 KD showed broad immunogenicity with the developed antisera raising the prospect of being putative common immunoprophylactic

Comparative immunodiagnostic approach of toxocariasis in buffalo calves [Jorunal Article]

Five Toxocara vitulorum antigens were utilized to diagnose natural toxocariasis in buffalo calves by ELISA. Adult antigen was proved to be the most potent one. The second potent antigen was egg antigen, followed by excretory-secretory antigen of male worms, then female worms. The coproantigen was the least potent one. The electrophoretic make-up of the antigens, examined by SDS-PAGE, revealed different patterns of separation. Common as well as specific component[s] to each antigen were identified. Employing naturally infected buffalo calf sera in immunoblot assay, five immunogenic components were detected in the adult antigen of molecular weight 191 KD, 166 KD, 102 KD, 65 KD and 54 KD. The reactive polypeptides in egg antigen were 191 KD, 105 KD, 99 KD and 79 KD. In coproantigen, six bands were identified. These components were of a molecular weight 191 KD, 178 KD, 166 KD, 124 KD, 96 KD and 65 KD. Five components of molecular weight 191 KD, 166 KD, 102 KD, 96 KD and 65 KD were immunogenic in excretory/secretory antigen of male worms. Only four polypeptides of 191 KD, 166 KD, 102 KD and 66 KD were identified in excretory/secretory female antigen

Potency of three Haemonchus contortus antigens in the diagnosis of ovine haemonchosis

One hundred faecal specimens and corresponding blood samples were evaluated for Haemonchus contortus infection in sheep by ELISA utilizing somatic, circulating and copro-antigens. Results proved that coproantigen was more potent than the others in diagnosis of sheep haemonchosis. SDS-poly-acrylamide gel electrophoresis demonstrated that the three antigens are structurally related. Immunoblot analysis revealed that somatic antigen contained 6 polypeptides of molecular weights 234, 205, 178, 83, 48 and 14 kDa, while, coproantigen showed 4 immunogenic polypeptides of 234, 205, 85 and 45 kDa. In circulating antigen only 2 polypeptides of molecular weights 165 and 99 kDa were immunoreactive. The potency of coproantigen may be attributed partially to one polypeptide of molecular weight 45 kDa. This low molecular weight polypeptide was only expressed in coproantigen as an immunogen, probably increases the immunogenicity, and may have potential diagnostic and protective values

Isolation and structural characterization of toxocara vitulorum specific antigen and its potency in diagnosis of toxocariasis among buffalo calves

A method of affinity chromatography purification of species-specific antigen from Toxocara vitulorum adult worm was described. The purification process resulted in a fraction with 9315 fold increased in specific activity compared to crude extract. Structural characterization of the isolated fraction by SDS polyacrylamide gel electrophoresis, isoelectric focusing and amino acid analysis showed that the fraction consists of only two polypeptides of 92 kD and 87 kD with isoeletric points of 8.8 and 8.9. Moreover, 17 amino acids were identified in the fraction with high proportions of only three of them [tyrosine, glycine and glutamic]. The isolated antigen proved potency in the diagnosis of toxocariasis among buffalo claves using ELISA which recorded 100% sensitivity

Molecular identity of major cross-reactive adult antigens in Fasciola gigantica, Toxocara vitulorum and Monieza expansa

Cross reactivity between Fasciola gigantica, Toxocara vitulorum and Moniezia expansa, whole worm extracts was proved by ELISA. Intense cross-reaction was observed between F. gigantica and M. expansa rather than between each of them and T. vitulorum. As judged by immunoblot, the cross-reactive antigens in F. gigantica which recognized by T. vitulorum antisera was 109 kD, while this component in addition to another one of 52 kD were detected by M. Expansa sera in the same extract. Furthermore, T. vitulorum antigen which cross-reacted with F. gigantica, was 133 kD and with M. expansa was 143 kD. Antigens responsible for cross-reactivity in M. expansa were 130 kD and 210 kD to T. vitulorum and F. Gigantica, respectively

Structural characterization and immunolocalization of egg antigens cross react with Toxicara vitulorum, Fasciola gigantica and moniezia expansa mature flukes

A structural homology between eggs of Toxocara vitulorum, Fasciola gigantica and Moniezia expansa was proved by the use of SDS-PAGE. In immunoblot, nine, eleven and seven polypeptides were recognized in F. gigantica, M. expansa and T. vitulorum eggs, respectively, by their respective rabbit anti-adult antisera. Moreover, components of 240 KD and 206 KD were recognized in the three eggs by different anti-adult antisera. The anatomic localization of the cross-reactive epitopes in eggs was determined by indirect immunofluorescence microscopy. The cross- reactive epitopes were mainly associated with embryonated cells of F. gigantica, egg shell, larvae and vitelline membranes of T. vitulorum and egg shell and granular layer of M. expansa

Field application of bacillus thuringiensis var kurstaki [dipel-2x] against soft and hard ticks

Field application of Bacillus thuringiensis [B.t.] var. kurstaki [Dipel-2X] against hard tick, Boophilus annulatus and soft tick, Argas [percicargas] persicus was probed. Hard tick died on infected calves after 6 days post-spraying with Dipel-2X, while soft tick succumbed in its cracks after 6 weeks post treatment. The indirect effect of Dipel-2X on Argas [p.] persicus was also investigated. Egg laying of tick fed on hens injected with 20 and 40 mg Dipel-2X was clearly affected. Molecular structures of hemolymphs collected from ticks got Dipel-2X infected blood meals did not change as demonstrated by SDS-PAGE