Immunodiagnosis of fascioliasis using sandwich enzyme-linked immunosorbent assay for detection of Fasciola gigantica paramyosin antigen

Abou-Elhakam, Hany Mohamed Adel; Bauomy, Ibraheem Rabia; Deeb, Somaya O. El; Amir, Azza M. El

doi:10.4103/2229-5070.113907

Cited by 5 publications

(3 citation statements)

References 27 publications

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“…Our investigations using synthetic peptides or peptide cocktails from Ov58GPCR-ECD as well as the recombinant ECD led to results that match these main requirements. Synthetic peptides have been used in the sero-diagnosis of many infectious disease with good performance [ 32 – 34 ] with their main advantage being the potential for reduced cross-reactivity often observed with recombinant and native antigens [ 93 ]. The Ov58GPCR peptides and peptide cocktails in this preliminary study were all able to discriminate between actively-infected onchocerciasis subjects and treated/non-infected individuals although to different extents.…”

Section: Discussionmentioning

confidence: 99%

Prediction and validation of the structural features of Ov58GPCR, an immunogenic determinant of Onchocerca volvulus

et al. 2018

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Onchocerciasis is a severely debilitating yet neglected tropical disease (NTD) that creates social stigma, generates and perpetuates poverty, and leads ultimately in some cases to irreversible unilateral or bilateral blindness if untreated. Consequently, the disease is a major impediment to socioeconomic development. Many control programs have been launched for the disease with moderate successes achieved. This mitigated hit is partially due to the lingering need for reliable, non-invasive and easily applicable tools for mapping endemic regions and post-elimination surveillance. In this work, bioinformatics analyses combined with immunological assays were applied in a bid to develop potential tools for diagnosis and assessing the success of drug treatment programs. We report that (i) the O. volvulus antigen, Ov58GPCR is a G-protein coupled receptor (GPCR) conserved in related nematodes, (ii) synthetic peptides predicted to be in the extracellular domain (ECD) of Ov58GPCR are indeed immunogenic epitopes in actively-infected individuals, (iii) synthetic peptide cocktails discriminate between actively-infected individuals, treated individuals and healthy African controls, (iv) polyclonal antibodies against one of the peptides or against the bacterially-expressed ECD reacted specifically with the native antigen of O. volvulus total and surface extracts, (v) Ov58GPCR is transcribed in both larvae and adult parasite stages, (vi) IgG and IgE responses to the recombinant ECD decline with ivermectin treatment. All these findings suggest that the extracellular domain and synthetic peptides of Ov58GPCR, as well as the specific immune response generated could be harnessed in the context of disease diagnosis and surveillance.

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Section: Discussionmentioning

confidence: 99%

Prediction and validation of the structural features of Ov58GPCR, an immunogenic determinant of Onchocerca volvulus

et al. 2018

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“…However, the immunological potential of linear antigenic epitopes has been demonstrated by microarray screening of the entire proteome of different pathogens [61][62][63], as it is the case of O. volvulus for which three peptide motifs with an IgG1, IgG3, IgE and IgM response profile were identified [64]. Furthermore, though Paramyosin is a widely conserved protein across species with well-known antigenic potential [65,66], the immunological properties of Ol-PARA [25] have been herein investigated by the identification of the three linear epitopes (LEEARRRLE, SRLQSEVEVLIVDL, MQVDEEHKMF), which showed a high intensity of reactivity when tested against serum sample of O. lupi infected dog.…”

Section: Plos Neglected Tropical Diseasesmentioning

confidence: 99%

Major antigen and paramyosin proteins as candidate biomarkers for serodiagnosis of canine infection by zoonotic Onchocerca lupi

et al. 2021

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Onchocerca lupi (Spirurida: Onchocercidae) is a filarial worm parasitizing domestic carnivores and humans. Adult nematodes usually localize beneath in the sclera or in the ocular retrobulbar of infected animals, whilst microfilariae are found in the skin. Therefore, diagnosis of O. lupi is achieved by microscopic and/or molecular detection of microfilariae from skin biopsy and/or surgical removal of adults from ocular tissues of infected hosts. An urgent non-invasive diagnostic tool for the diagnosis of O. lupi in dog is mandatory. In this study, an immunoproteomic analyses was performed using a combination of immunoblotting and mass spectrometry techniques. Onchocerca lupi major antigen (Ol-MJA) and paramyosin (Ol-PARA) proteins were identified as potential biomarkers for serodiagnosis. Linear epitopes were herein scanned for both proteins using high-density peptide microarray. Sera collected from dog infected with O. lupi and healthy animal controls led to the identification of 11 immunodominant antigenic peptides (n = 7 for Ol-MJA; n = 4 for Ol-PARA). These peptides were validated using sera of dogs uniquely infected with the most important filarioids infesting dogs either zoonotic (Dirofilaria repens, Dirofilaria immitis) or not (Acanthocheilonema reconditum and Cercopithifilaria bainae). Overall, six antigenic peptides, three for Ol-MJA and for Ol-PARA, respectively, were selected as potential antigens for the serological detection of canine O. lupi infection. The molecular and proteomic dataset herein reported should provide a useful resource for studies on O. lupi toward supporting the development of new interventions (drugs, vaccines and diagnostics) against canine onchocercosis.

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“…Paramyosin is a large structural protein (98–101 kDa) component of invertebrate muscle and found in endo- and ectoparasites of medical importance, but not in vertebrates [ 31 ]. By virtue of its major antigenic properties, this protein has been used for the development of diagnostic assays for filarioids of medical concern such as Wuchereria bancrofti and Brugia malayi [ 27 , 32 , 33 ]. Many studies of the paramyosin protein, as a potential vaccine candidate, have focussed on platyhelminths [ 34 – 39 ], with fewer studies on nematodes [ 40 , 41 ] and ticks [ 42 ].…”

Section: Introductionmentioning

confidence: 99%

Paramyosin of canine Onchocerca lupi: usefulness for the diagnosis of a neglected zoonotic disease

et al. 2016

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BackgroundOf increasing importance to the medical and veterinary communities is the zoonotic filarioid nematode Onchocerca lupi. Onchocercosis, thus far found in wolves, dogs, cats and humans, is diagnosed via skin snips to detect microfilariae and surgical removal of adults from the eye of the host. These methods are time-consuming, laborious and invasive, highlighting the need for new tools for the diagnosis of O. lupi in susceptible hosts. Symptoms related to the presence of the adults in the eye can range from none apparent to severe, including blindness. No reliable chemotherapeutic protocols are available, as yet, to eliminate the infection. Paramyosin, an invertebrate-specific protein, has been well-studied as an allergen, diagnostic marker and vaccine candidate. The aim of this study, therefore, was to isolate and characterise paramyosin from O. lupi to assess its suitability for the development of a serological diagnostic assay.MethodsThe adult and microfilarial stages of O. lupi were isolated from the eyes and skin of a 3-year-old male dog. Total RNA was extracted and reverse transcribed into single stranded cDNA. Reverse-transcription PCR was used to isolate a full-length paramyosin cDNA from adult worms and to investigate the temporal expression patterns of this gene. All amplicons were sequenced using dideoxy chain termination sequencing. Bioinformatics was used to predict the amino acid sequence of the gene, to compare the DNA and protein sequences with those available in public databases and to investigate the phylogenetic relationship of all molecules. Antibody binding sites were predicted using bioinformatics and mapped along with published antigenic epitopes against the O. lupi paramyosin protein. The native protein, and three smaller recombinantly expressed peptides, were subjected to western blot using serum from dogs both positive and negative for O. lupi.ResultsParamyosin of O. lupi was herein molecularly characterized, encoded by a transcript of 2,643 bp and producing a protein of 881 amino acids (101.24 kDa). The paramyosin transcript was detected, by reverse transcription PCR, in adults and microfilariae, but not in eggs. Phylogenetic analysis indicates that this molecule clusters with paramyosins from other filarioids to the exclusion of those from other taxa. A total of 621 unique antibody binding epitopes were predicted for this protein and another 28 were conserved in other organisms. This information was used to design three peptides, for recombinant expression, to identify the antibody binding epitope(s) and reduce potential cross-reactivity with serum from dogs infected with other filarioid nematodes. Native paramyosin, purified from microfilariae and adults, was detected by antibodies present in serum from dogs with known O. lupi infections.ConclusionsData provided herein may assist in the development of a serological diagnostic test, based on antibodies to O. lupi paramyosin, for the diagnosis of this infection, in order to gain more information on the real distribution of this l...

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Immunodiagnosis of fascioliasis using sandwich enzyme-linked immunosorbent assay for detection of Fasciola gigantica paramyosin antigen

Cited by 5 publications

References 27 publications

Prediction and validation of the structural features of Ov58GPCR, an immunogenic determinant of Onchocerca volvulus

Prediction and validation of the structural features of Ov58GPCR, an immunogenic determinant of Onchocerca volvulus

Major antigen and paramyosin proteins as candidate biomarkers for serodiagnosis of canine infection by zoonotic Onchocerca lupi

Paramyosin of canine Onchocerca lupi: usefulness for the diagnosis of a neglected zoonotic disease

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