Human and camel cystic echinococcosis - a polyclonal antibody-based sandwich ELISA for its serodiagnosis with molecular identification.

Cystic echinococcosis (CE) is an emergent neglected disease affecting human and animals in Egypt with a wide distribution and incidence. This study aimed to evaluate the use of a polyclonal antibody-based sandwich ELISA in the detection of Echinococcus granulosus antigen in human and camel sera. Hydatid cyst protoscoleces antigen (PsAg) was isolated from hydatid cysts collected from naturally infected camel livers and lungs. PsAg was used for immunization of rabbits to raise IgG polyclonal antibodies (IgG PsAb). IgG PsAb were then precipitated, purified using Protein-A Sepharose gel and labeled with horseradish peroxidase enzyme. We assayed the purity of the IgG PsAb, and the two prepared E. granulosus antigens CPsAg from camel cysts and HPsAg from human cysts by Sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The resulted protein bands of the prepared CPsAg appeared at different molecular weights: 180, 90, 68, 54, 42 and 22 kDa while, HPsAg shared with it in 4 common bands at 68, 54, 42, and 22 kDa. The purified IgG PsAb had been resolved at two bands at 52 kDa and at 32 kDa. Sandwich ELISA were performed for the detection of circulating E. granulosus antigens in sera of human (n = 183) and camels (n = 190). The purified IgG PsAb showed strong reactivity against E. granulosus infected human and camel samples and no cross reactivity neither with free-healthy negative sera nor with others parasitic diseases (Schistosomiasis, Fascioliasis, Toxoplasmosis, Ancylostomiasis for human samples and Fascioliasis, ticks' infestation, Eimeriosis, Cryptosporidiosis, Nasal myiasis, Toxoplasmosis for camel samples). The sensitivity of the assay was 98.25% (56/57) and 96.9% (31/32) against human and camel samples, respectively. Specificity was 100% in both human and camel samples. Sandwich ELISA detected CE in 33.3% (24/72) and 55.6% (50/90) random human and camel samples, respectively. Indirect ELISA, using CPsAg, was used for detection of antibodies in positive human and camels' sera and detected 96.5% (55/57) and 93.8% (30/32) of human and camel samples, respectively. In our study, Genomic DNA was extracted from protoscoleces fluid of human liver hydatid cysts to identify the Echinococcus sp. isolate based on NADH dehydrogenase subunit 1 (NAD1) gene by Polymerase Chain Reaction (PCR) and the isolate (GenBank: OP785689.1) were identified as E. granulosus sensu lato genotype. In conclusion, Sandwich ELISA technique was found to be a potent and sensitive assay for detection of hydatid antigen in both human and camel samples.

Therapeutic effect of Moringa oleifera and Thymus vulgaris oils against hepatic coccidiosis in experimentally infected rabbits.

The present study was conducted to detect the therapeutic effect of Moringa oleifera and Thymus vulgaris oils on hepatic coccidiosis in experimentally infected rabbits. Also, immunomodulatory effect of the two oils was detected. Twenty-four Newzealand rabbits were used in this study and divided into 4 groups; healthy rabbits, experimentally infected rabbits with Eimeria stiedae oocysts, and two infected treated groups (one with moringa (200 mg/kg) and the other with thyme (500 mg/kg) oils). The results showed highly significant reduction in oocysts shedding (P<0.001 and P<0.05) in the two infected and treated rabbits than the infected non-treated rabbits in almost all days post infection (PI). Thyme oil was more potent and stopped oocysts shedding earlier at the day 34 PI compared to moringa oil at the day 41 PI. Microscopically, there was a damage in the oocysts shed by treated rabbits. Macroscopically, the livers of thyme oil treated rabbits showed more enhancement with protection percentage 75% than those treated with moringa oil in which protection percentage was 55%. The highest titer of antibodies was detected in moringa oil treated rabbits. It was concluded that both moringa and thyme oils had an anti-coccidial effect with thyme oil superiority. So, thyme oil could be useful as an alternative product for the control of rabbit coccidiosis.

Anticryptosporidial effect of pomegranate peels water extract in experimentally infected mice with special reference to some biochemical parameters and antioxidant activity.

Cryptosporidiosis is a zoonotic disease caused by a well-known parasitic protozoan called Cryptosporidium. Infection in livestock causes important economic losses among farm animals and its control has a global concern. In this study, internal white and external red layers were separated from pomegranate peels (Punica granatum) then; they were grinded to reach Nano form. Anticryptosporidial effect of their water extracts was investigated in experimentally infected mice. Also, their antioxidant activity, biochemical and histopathological changes were studied. Briefly, hot aqueous extracts of pomegranate peels were prepared regarding its good sensory attributes at concentration of 10% W/V. Analysis of total phenolics, individual phenolics by HPLC-DAD and antioxidant activities have been done. Forty-five mice were divided into five groups each one containing nine mice. The first group was healthy mice and the 2nd one was infected orally with 104 Cryptosporidium parvum (C. parvum) oocysts/mice and not treated. The other 3 groups were infected and orally treated with Nitazoxanide (NTZ) for the 3rd group, pomegranate red peel extract for the 4th group and pomegranate white peel extract for the 5th group. Blood samples were collected after 1 and 2 weeks post treatment for protein profile, liver enzymes and antioxidant activity evaluation. After 3 weeks, all animals were sacrificed and ileal tissues were embedded in paraffin for histopathological examination. The results showed that pomegranate peel extracts were rich in phenolic compounds, had high antioxidant activity and therapeutic effect on C. parvum in experimentally infected mice. Red peel extract diminished C. parvum oocysts count significantly in experimentally infected mice than white peel and NTZ treatments. Also, the histopathological examination revealed that red peel treated mice ileal sections showed a great enhancement in the shape and structure of villi towards normal structure than other treated groups. Most of the measured biochemical parameters after 2 weeks' treatment with red pomegranate peel and NTZ were enhanced in their concentrations towards the healthy normal status. In conclusion, this study showed the effectiveness of Nano-form of pomegranate white and red peel extracts against C. parvum oocysts. Pomegranate red peel extract was found to have antioxidant activity that could significantly enhance the serum biochemical parameters and oxidative stress towards the healthy normal status. Furthermore, it is suggested that pomegranate peel should be separated and used in the daily animal diet or as a functional beavarage for human as accepted from the panelists to give protective effects against this parasite as well as to improve health benefits.