title: Development of a recombinant protein-based ELISA for diagnosis of larval cyathostomin infection.
abstract: Cyathostomins are ubiquitous nematodes of horses. Once ingested, they can spend a substantial time as encysted larvae in the intestinal wall. The larvae can comprise up to 90% of the total burden, with up to several million worms reported in individuals. These stages can emerge in large numbers to cause life-threatening colitis. Direct methods for detection of encysted larval burdens in live horses do not exist. Previously, two antigen complexes were identified as promising markers for infection. A component of these, cyathostomin gut associated larval antigen-1 (Cy-GALA-1), was identified following immunoscreening of a complementary DNA library. Serum immunoglobulin G(T) (IgG(T)) responses to Cy-GALA-1 were shown to inform on larval infection. Sequence analysis of polymerase chain reaction products amplified from individual worms indicated that Cy-GALA-1 was derived from Cyathostomum pateratum. As cyathostomin infections always comprise multiple species, a diagnostic test must account for this. Here, segments of the Cy-gala gene were isolated from four common species, Cyathostomum catinatum, Cylicocyclus ashworthi, Cylicostephanus goldi and Cylicostephanus longibursatus, and the associated proteins expressed in recombinant form. The specificity and immunogenicity of each protein was confirmed. Each protein was assessed by enzyme linked immuno sorbent assay (ELISA) for its ability for informing on the presence of encysted larval infection and the level of burden.
authors.affiliation: Moredun Research Institute,Pentlands Science Park,Edinburgh EH26 0PZ,Scotland,UK.
authors.forename: Mairi C
authors.initials: MC
authors.lastname: Mitchell
authors.affiliation: Moredun Research Institute,Pentlands Science Park,Edinburgh EH26 0PZ,Scotland,UK.
authors.forename: Thomas
authors.initials: T
authors.lastname: Tzelos
authors.affiliation: Royal Dick School of Veterinary Studies,University of Edinburgh,Edinburgh EH25 9RG,Scotland,UK.
authors.forename: Ian
authors.initials: I
authors.lastname: Handel
authors.affiliation: Moredun Research Institute,Pentlands Science Park,Edinburgh EH26 0PZ,Scotland,UK.
authors.forename: Hamish E G
authors.initials: HE
authors.lastname: McWilliam
authors.affiliation: Department of Infection Biology,Institute of Infection and Global Health,University of Liverpool,Liverpool L69 7ZJ,England,UK.
authors.forename: Jane E
authors.initials: JE
authors.lastname: Hodgkinson
authors.affiliation: Moredun Research Institute,Pentlands Science Park,Edinburgh EH26 0PZ,Scotland,UK.
authors.forename: Alasdair J
authors.initials: AJ
authors.lastname: Nisbet
authors.affiliation: I.I. Schmalhausen Institute of Zoology NAS of Ukraine,vul B Khmelnytskogo 15,Kyiv 01030,Ukraine.
authors.forename: Vitaliy O
authors.initials: VO
authors.lastname: Kharchenko
authors.affiliation: Moredun Research Institute,Pentlands Science Park,Edinburgh EH26 0PZ,Scotland,UK.
authors.forename: Stewart T G
authors.initials: ST
authors.lastname: Burgess
authors.affiliation: Moredun Research Institute,Pentlands Science Park,Edinburgh EH26 0PZ,Scotland,UK.
authors.forename: Jacqueline B
authors.initials: JB
authors.lastname: Matthews
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000918:Antibody Specificity; D000947:Antigens, Helminth / Q000235:genetics / Q000276:immunology*; D018076:DNA, Complementary / Q000235:genetics; D018509:DNA, Helminth / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D015801:Helminth Proteins / Q000235:genetics / Q000276:immunology*; D006734:Horse Diseases / Q000175:diagnosis* / Q000469:parasitology; D006736:Horses; D007814:Larva; D011994:Recombinant Proteins; D016415:Sequence Alignment; D013319:Strongyle Infections, Equine / Q000175:diagnosis* / Q000469:parasitology; D013320:Strongyloidea / Q000276:immunology* / Q000302:isolation & purification
chemical_list: D000947:Antigens, Helminth; D018076:DNA, Complementary; D018509:DNA, Helminth; D015801:Helminth Proteins; D011994:Recombinant Proteins
keywords: ELISA; cyathostomin; diagnosis; encysted larvae; nematode