Antigenic profile, isolation and characterization of whole body extract of Paramphistomum gracile

Abstract

An antigenic component of adult Paramphistomum gracile was characterized by means of indirect enzyme-linked immunosorbent assay (indirect ELISA), sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting using sera from cattle naturally infected with P. gracile, Eurytrema pancreaticum, Fasciola gigantica, Moniezia benedeni, strongylids, Trichuris sp. and Strongyloides sp. The whole body (WB) extracts of P. gracile were fractionated by gel filtration chromatography in a Sephadex G-200 column. It was found that the WB extract fractions, F1-F3 were highly antigenic, F5 was moderately antigenic and F4 was poorly antigenic. For SDS-PAGE and immunoblotting, the antigenic molecules of WB extract and all five fractions were mostly at molecular weights (MW) ranging from 12 to 150 kDa. One antigenic protein of 16 kDa detected in WB extract and F1-F3 was found to give a consistent reaction with sera from infected cattle. The antigenicity of the purified 16 kDa protein was confirmed by immunoblotting and indirect ELISA using a pool of sera and individual serum samples from infected cattle (at 1 : 78 125 dilution) and hyperimmunized rabbit (at 1 : 390 625 dilution). This finding suggests that the 16 kDa protein may be a potential antigen for the immunodiagnosis of cattle paramphistomosis caused by P. gracile.

The delicate balance between Entamoeba histolytica, mucus and microbiota

“…Further investigation will be needed to better understand the connection between commensal microbiota and the immune response to this parasite. Intestinal bacteria can affect the outcome of parasitic helminths infections (reviewed in 42 ) as well as those associated with some parasitic protozoa (reviewed in 43 ). Studies done in germ free mice have showed that microbiota is important for protozoa to mount a characteristic immune response with Giardia duodenalis, 44 Leishmania amazonensis 45 and Schistosoma mansoni infection.…”

Paramphistomosis of Ruminants: An Emerging Parasitic Disease in Europe

“…Conducting drug efficacy tests in vivo against immature paramphistomes is challenging, owing to the current lack of a diagnostic test for pre-patent infections or a suitable animal model for experimental infection. Some efforts towards diagnostic development have been made recently: Anuracpreeda et al developed a diagnostic sandwich ELISA, based on detection of a 16 kDa protein from a Paramphistomum gracile whole worm extract[37,63,64]. This offers hope for the development of antibody-based diagnostic tools (e.g.…”

Immunodiagnosis of paramphistomosis using monoclonal antibody-based sandwich ELISA for detection ofParamphistomum gracilecirculating 16 kDa antigen

“…The 16 kDa antigen of P. gracile (16 kDaAgPg) was obtained as per the method described earlier by Anuracpreeda et al . (2016 a , 2017). Protein concentrations of antigen preparations were determined by Lowry's method (Lowry et al 1951).…”

“…Hybridoma clone (1D10) producing a high antibody titre against 16 kDaAgPg was selected and the specific MoAb isotypes were determined by indirect ELISA. In the cross-reactivity study, proteins (10 µ g) in WB from P. gracile as well as WB from other trematode, cestode and nematode parasites were separated using 12·5% SDS–PAGE, and transferred onto nitrocellulose (NC) membranes for immunoblotting by the selected MoAb 1D10 according to the method of Anuracpreeda et al (2016 a ). Polyclonal anti-16 kDaAgPg for the detection of antigen captured by the immobilized MoAb was produced by immunizing New Zealand White rabbits with 16 kDa antigen of P. gracile as per the method of Anuracpreeda et al (2016 c ).…”