title: Validation and evaluation of VapA-specific IgG and IgG subclass enzyme-linked immunosorbent assays (ELISAs) to identify foals with Rhodococcus equi pneumonia.
abstract: BACKGROUND
  Rhodococcus equi (Rhodococcus hoagii/Prescottella equi) is a common cause of foal pneumonia, but its diagnosis remains a challenge for equine veterinarians. While the VapA-specific (virulence-associated protein A) immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) has low sensitivity and specificity for detecting pneumonic foals, little is known about VapA-specific IgG subclasses.
  
  
  OBJECTIVE
  To evaluate the performance of VapA-specific ELISA for IgG and its subclasses IgGa, IgGb and IgG(T) in the early diagnosis of pneumonia caused by R. equi.
  
  
  METHODS
  Assay validation followed by assessment of diagnostic performance using archived samples from animals of known status.
  
  
  METHODS
  Serum samples from exposed (n = 125) and nonexposed adult horses (n = 10) and from experimentally challenged and naturally infected foals were used for ELISA validation. Post mortem and tissue culture records of the last 24 years from the Institute for Experimental Pathology at the University of Iceland in Keldur, Iceland laboratory were evaluated to confirm the absence of R. equi cases in Iceland. The diagnostic performance of VapA-specific IgG and its subclasses was evaluated using banked serum samples from pneumonic (n = 21) and healthy foals (n = 80). To evaluate each IgG assay, a cut-off value was selected based on receiver operating characteristic curve analysis and used to calculate sensitivity and specificity. The intra- and interassay coefficients of variation were calculated for each ELISA.
  
  
  RESULTS
  Using sera from Iceland, where R. equi infection has not been reported, the VapA-specific IgG ELISA differentiated exposed from nonexposed horses. When used to identify infected foals, VapA-specific IgG, IgGa and IgGb had no diagnostic value. In contrast, IgG(T) had high sensitivity and specificity.
  
  
  CONCLUSIONS
  Horses from Iceland are not exposed to VapA(+) R. equi and can serve as negative controls. VapA-specific IgG subclasses, with the exception of IgG(T), are poor predictors of disease. Further investigation on the use of IgG(T) as a diagnostic tool in field conditions is needed.
authors.affiliation: Maxwell H. Gluck Equine Research Center, Lexington, Kentucky, USA.
authors.forename: M G
authors.initials: MG
authors.lastname: Sanz
authors.affiliation: Maxwell H. Gluck Equine Research Center, Lexington, Kentucky, USA.
authors.forename: A F
authors.initials: AF
authors.lastname: Oliveira
authors.affiliation: Department of Veterinary Science, Veterinary Diagnostic Laboratory, Lexington, Kentucky, USA.
authors.forename: A
authors.initials: A
authors.lastname: Loynachan
authors.affiliation: Maxwell H. Gluck Equine Research Center, Lexington, Kentucky, USA.
authors.forename: A
authors.initials: A
authors.lastname: Page
authors.affiliation: Institute for Experimental Pathology, University of Iceland, Reykjavík, Iceland.
authors.forename: V
authors.initials: V
authors.lastname: Svansson
authors.affiliation: Department of Large Animal Medicine, University of Georgia, Athens, USA.
authors.forename: S
authors.initials: S
authors.lastname: Giguère
authors.affiliation: Maxwell H. Gluck Equine Research Center, Lexington, Kentucky, USA.
authors.forename: D W
authors.initials: DW
authors.lastname: Horohov
mesh_terms: D000193:Actinomycetales Infections / Q000097:blood / Q000276:immunology / Q000517:prevention & control / Q000662:veterinary; D000818:Animals; D000831:Animals, Newborn; D000918:Antibody Specificity; D001426:Bacterial Proteins / Q000276:immunology*; D001428:Bacterial Vaccines / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary*; D006734:Horse Diseases / Q000097:blood / Q000175:diagnosis / Q000453:epidemiology / Q000382:microbiology*; D006736:Horses; D007055:Iceland / Q000453:epidemiology; D007074:Immunoglobulin G / Q000097:blood / Q000145:classification*; D018410:Pneumonia, Bacterial / Q000175:diagnosis / Q000453:epidemiology / Q000382:microbiology / Q000662:veterinary*; D016929:Rhodococcus equi / Q000378:metabolism*; D014481:United States / Q000453:epidemiology
chemical_list: D001426:Bacterial Proteins; D001428:Bacterial Vaccines; D007074:Immunoglobulin G; C541879:VapA protein, Rhodococcus equi
keywords: Prescottella equi; Rhodococcus equi; Rhodococcus hoagii; antibody; enzyme-linked immunosorbent assay; foal; immunoglobulin G subclasses