A novel enolase from Taenia solium metacestodes and its evaluation as an immunodiagnostic antigen for porcine cysticercosis

for the Cysticercosis Working Group in Peru

Research output: Contribution to journalArticle

Abstract

Cysticercosis is a worldwide parasitic disease of humans and pigs principally caused by infection with the larvae of the pork tapeworm Taenia solium. Through the use of the recently-made-available T. solium genome, we identified a gene within a novel 1448 bp ORF that theoretically encodes for a 433 amino acid-long protein and predicted to be an α-enolase closely related to enolases of other flatworms. Additional bioinformatic analyses revealed a putative plasminogen-binding region on this protein, suggesting a potential role for this protein in pathogenesis. On this basis, we isolated the mRNA encoding for this presumptive enolase from T. solium metacestodes and reverse-transcribed it into cDNA before subsequently cloning and expressing it in both E. coli (rEnoTs) and insect cells (rEnoTsBac), in a 6xHis tagged manner. The molecular weights of these two recombinant proteins were ∼48 and ∼50 kDa, respectively, with the differences likely attributable to differential glycosylation. We used spectrophotometric assays to confirm the enolase nature of rEnoTs as well as to measure its enzymatic activity. The resulting estimates of specific activity (60.000 U/mg) and Km (0.091 mM) are quite similar to the catalytic characteristics of enolases of other flatworms. rEnoTs also exhibited high immunogenicity, eliciting a strong polyclonal antibody response in immunized rabbits. We subsequently employed rEnoTsBac for use in an ELISA aimed at discriminating between healthy pigs and those infected with T. solium. This diagnostic assay exhibited a sensitivity of 88.4% (95% CI, 74.92%–96.11%) and a specificity of 83.7% (95% CI: 69.29%–93.19%). In conclusión, this study reports on and enzymatically characterizes a novel enolase from T. solium metacestode, and shows a potential use as an immunodiagnostic for porcine cysticercosis.

Original languageEnglish (US)
Pages (from-to)44-54
Number of pages11
JournalExperimental Parasitology
Volume191
DOIs
StatePublished - Aug 1 2018

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Taenia solium
Cysticercosis
Phosphopyruvate Hydratase
Swine
Antigens
Platyhelminths
Parasitic Diseases
Proteins
Plasminogen
Computational Biology
Glycosylation
Recombinant Proteins
Open Reading Frames
Antibody Formation
Larva
Insects
Organism Cloning
Complementary DNA
Molecular Weight
Enzyme-Linked Immunosorbent Assay

Keywords

  • Baculovirus
  • Cysticercosis
  • E. coli
  • Enolase
  • Immunodiagnosis
  • Peru
  • Taenia solium

ASJC Scopus subject areas

  • Parasitology
  • Immunology

Cite this

A novel enolase from Taenia solium metacestodes and its evaluation as an immunodiagnostic antigen for porcine cysticercosis. / for the Cysticercosis Working Group in Peru.

In: Experimental Parasitology, Vol. 191, 01.08.2018, p. 44-54.

Research output: Contribution to journalArticle

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title = "A novel enolase from Taenia solium metacestodes and its evaluation as an immunodiagnostic antigen for porcine cysticercosis",
abstract = "Cysticercosis is a worldwide parasitic disease of humans and pigs principally caused by infection with the larvae of the pork tapeworm Taenia solium. Through the use of the recently-made-available T. solium genome, we identified a gene within a novel 1448 bp ORF that theoretically encodes for a 433 amino acid-long protein and predicted to be an α-enolase closely related to enolases of other flatworms. Additional bioinformatic analyses revealed a putative plasminogen-binding region on this protein, suggesting a potential role for this protein in pathogenesis. On this basis, we isolated the mRNA encoding for this presumptive enolase from T. solium metacestodes and reverse-transcribed it into cDNA before subsequently cloning and expressing it in both E. coli (rEnoTs) and insect cells (rEnoTsBac), in a 6xHis tagged manner. The molecular weights of these two recombinant proteins were ∼48 and ∼50 kDa, respectively, with the differences likely attributable to differential glycosylation. We used spectrophotometric assays to confirm the enolase nature of rEnoTs as well as to measure its enzymatic activity. The resulting estimates of specific activity (60.000 U/mg) and Km (0.091 mM) are quite similar to the catalytic characteristics of enolases of other flatworms. rEnoTs also exhibited high immunogenicity, eliciting a strong polyclonal antibody response in immunized rabbits. We subsequently employed rEnoTsBac for use in an ELISA aimed at discriminating between healthy pigs and those infected with T. solium. This diagnostic assay exhibited a sensitivity of 88.4{\%} (95{\%} CI, 74.92{\%}–96.11{\%}) and a specificity of 83.7{\%} (95{\%} CI: 69.29{\%}–93.19{\%}). In conclusi{\'o}n, this study reports on and enzymatically characterizes a novel enolase from T. solium metacestode, and shows a potential use as an immunodiagnostic for porcine cysticercosis.",
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AU - Gilman, Robert H

AU - Liendo, Ruddy

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AU - Luis, Sueline

AU - Quiñones-Garcia, Stefany

AU - Silverstein, Zach

AU - García, Hector H.

AU - Gonzalez, Armandoe

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