Development Of A Rapid Test For Toxocariasis Using Recombinant Toxocara Cati Tes-120 Protein

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dc.contributor.authorTan Farrizam, Siti Naqiuyah
dc.date.accessioned2022-11-15T03:16:57Z
dc.date.available2022-11-15T03:16:57Z
dc.date.issued2020-01
dc.description.abstractToxocariasis is a neglected zoonotic disease with worldwide distribution and has been shown to be especially prevalent among children from socio-economically disadvantaged populations. The causative agents are Toxocara canis and Toxocara cati in which the definitive hosts are dogs and cats, respectively. Although it is acknowledged that human toxocariasis is caused by both T. canis and T. cati, however, the importance of T. cati as a zoonotic agent of the disease has been under-recognized since T. cati TES is rarely used for Toxocara serodiagnosis and T. canis TES antigens are generally thought to be able to detect infections by both species. Thus, it is highly likely that some cases of T. cati infection may be missed by tests using only T. canis antigen. Therefore, it is an urgent need for the development of diagnostic tools that use both T. cati and T. canis recombinant antigens to enable detection of infections caused by both species. Previously, a T. cati recombinant protein TES-120 was successfully produced and cloned into a pGEX-4T-1 expression vector which showed good diagnostic potential in western blot. However, the yield was low and unsuitable for rapid test development. In this study, the T. cati DNA insert was cloned into a pET32 expression vector and two short peptides tagged variants [five residues each of aspartic acid (D) and lysine(K)] were designed to increase the chances of obtaining a good yield of the soluble recombinant protein. All three variants namely TES-120 cati/pET32a, TES-120 5D cati/pET32a, and TES-120 5K cati/pET32a were expressed and purified. After the protein identities were confirmed by anti-histidine western blot xxiii and MALDI TOF/TOF, the immunoreactivities of all three variants against serum samples were tested by western blot and the best variant was found to be TES- 120 cati/pET32a. Lateral flow dipstick rapid test was then developed and tested against a panel of serum samples, with anti-human IgG4 gold conjugate as the detector. Diagnostic seroreactivity of the newly developed recombinant TES-120 cati lateral flow dipstick was then compared to previously developed prototype rapid test which comprised three dipsticks separately dotted with T. canis recombinant antigens i.e. TES-26, TES-30, and TES-120. In this study, the diagnostic sensitivity of rTES-120 cati, rTES-26, rTES-30 and rTES-120 was found to be 64.0% (14/22), 82.0% (18/22), 77.0% (17/22) and 50.0% (11/22), respectively. When the results of all four dipsticks were combined, the rapid test successfully detected 95.0% (21/22) of the positive serum samples. In comparison, a sensitivity of 86% (19/22) was obtained when results of the T. cati dipstick was excluded. All four recombinant dipsticks produced 100% specificity when tested against healthy and other helminthic infection serum samples. Thus, as a conclusion, the inclusion of T. cati TES-120 in the lateral flow rapid test for toxocariasis increased its diagnostic sensitivity and enabling the detection of Toxocara infection from both species T. cati and T. canis.en_US
dc.identifier.urihttp://hdl.handle.net/123456789/16648
dc.language.isoenen_US
dc.publisherUniversiti Sains Malaysiaen_US
dc.subjectToxocariasisen_US
dc.subjectProteinen_US
dc.titleDevelopment Of A Rapid Test For Toxocariasis Using Recombinant Toxocara Cati Tes-120 Proteinen_US
dc.typeThesisen_US
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