Evaluation of a Western Blot Method for the Detection of Yersinia Antibodies: Evidence of Serological Cross-Reactivity between Yersinia Outer Membrane Proteins and Borrelia burgdorferi

Abstract

Yersinia enterocolitica and Yersinia pseudotuberculosis have been identified as causative organisms of reactive arthritis in humans. We evaluated a Western blot assay which uses Yersinia outer membrane proteins as antigens for the detection of Yersinia antibodies as a replacement for the complement fixation (CF) assay. Clinical agreement, sensitivity, and specificity were determined by testing 19 positive and 21 negative serum samples by the CF assay, Western blot assay, and enzyme-linked immunosorbent assay (ELISA). The CF assay and ELISA were compared to the Western blot assay, which was the reference method used in this study. Sera with antibodies that could potentially cross-react with Yersinia were also tested by the Western blot assay. The agreement, sensitivity, and specificity of the CF method were 61%, 26%, and 95%, respectively; and those for the ELISA were 89%, 95%, and 82%, respectively. The prevalences of Yersinia antibodies in 50 healthy donors were 6% for immunoglobulin G (IgG), 2% for IgA, and 2% for IgM. Sera positive for Bartonella henselae, Brucella, Chlamydia pneumoniae, and Rickettsia rickettsii antibodies showed cross-reactivity by the Western blot assay. The highest cross-reactivity was observed with Borrelia burgdorferi; 5 of 11 (45%) specimens were cross-reactive by the IgM-specific assay. Overall, the Western blot assay performs acceptably and is more sensitive than the CF assay, warranting replacement of the CF assay in the laboratory. Due to the evidence of cross-reactivity, particularly with B. burgdorferi, which can cause an oligoarthritis similar to reactive arthritis, the diagnosis of reactive arthritis should be based on clinical findings and complete serologic analysis of the potential causative infectious pathogens.

Lyme Borreliosis - the Utility of Improved Real-Time PCR Assay in Detection of Borrelia burgdorferi Infections

“…The diagnostic problem of ELISA tests is also the large cross-reactivity to antigens commonly observed in other bacteria, viruses and mammalian tissues such as p41, p58-60, p66, p68, p71, p73, flagellin etc. [26,27]. The line blot tests confirmed that more than 8% of the positive results of the ELISA in class IgG and [30,[32][33][34][35][36].…”

Too early to dismiss Yersinia enterocolitica infection in the aetiology of Graves’ disease: evidence from a twin case–control study

“…Although immunoblotting is a very sensitive and specific method for detecting antibodies against YOPs, cross‐reactivity to other bacterial species has been reported 36 . To minimize this source of bias we have used molecular‐defined single YOPs as antigens for the Western blotting.…”

“…35 Although immunoblotting is a very sensitive and specific method for detecting antibodies against YOPs, cross-reactivity to other bacterial species has been reported. 36 To minimize this source of bias we have used molecular-defined single YOPs as antigens for the Western blotting. It is important to realize that outer membrane proteins of all YE serotypes and other plasmid-containing enterobacteria are identical, although the plasmids vary in their DNA sequence homologies.…”

“…Specific IgA and IgG antibodies towards YOPs were demonstrated by immunoblotting with a quantitative YOP-antibody assay (AID, Strassberg, Germany). In brief, YOP antigens (25,34,36,37,39,40,46 and 48 kDa) are blotted onto nitrocellulose. Next, sera are diluted 1 : 51 in phosphatebuffered saline (PBS)-Tween and incubated with antigen-coated nitrocellulose strips overnight at 22 ° C. The IgA and IgG antibodyantigen complexes formed are quantified after immunostaining with the AID Scan System.…”

Cross-reaction between Yersinia outer membrane proteins and anti-Borreli antibodies in sera of patients with Lyme disease

“…[12] and Rawlins et al. [13]. The most highly cross‐reactive proteins of Borrelia were p66 (a common bacterial antigen), p41 (flagelin), p32 (OspA) and a protein of c. 20 kDa.…”