Evaluation of diagnostic assays for the serological detection ofActinobacillus pleuropneumoniaeon samples of known or unknown exposure

Abstract

Abstract. Accurate diagnosis of exposure to Actinobacillus pleuropneumoniae is important for maintaining negative farms. In the present study, the ability of a dual-plate complement fixation (CF) assay and 3 commercially available enzymelinked immunosorbent assays (ELISAs; quad-plate ELISA-1, single-plate ELISA-2, and single-plate ELISA-3) in detecting serological evidence of A. pleuropneumoniae exposure was compared using serum samples of experimentally infected or vaccinated pigs, or field samples from the United States. Forty-two pigs were divided into groups of 2 pigs and were inoculated with 1 of 15 A. pleuropneumoniae strains representing all known serovars of A. pleuropneumoniae, or with Actinobacillus suis, or were vaccinated with a bacterin containing A. pleuropneumoniae serovar 1, 3, 5, or 7. Serum samples collected at the day of inoculation or vaccination and 7, 14, 21, and 28 days later were used to compare the assays. On samples from experimentally infected pigs, the dual-plate CF assay, quad-plate ELISA-1, single-plate ELISA-2, and single-plate ELISA-3 had sensitivities of 0.46, 0.74, 0.13, and 0.13 and specificities of 0.90, 1.0, 1.0, and 1.0, respectively. Vaccinated pigs were identified only by the dual-plate CF assay and the quad-plate ELISA-1. In addition, 90 serum samples with unknown A. pleuropneumoniae exposure collected under field conditions were tested with all assays. The agreement of the 4 assays on field samples was slight to fair. While several assays are available for demonstration of A. pleuropneumoniae exposure, differences in assay targets complicate test choices. Decisions on which assay or combination of assays to use depend on the specific reasons for running the assays.

Simultaneous Detection of Antibodies against Apx Toxins ApxI, ApxII, ApxIII, and ApxIV in Pigs with Known and Unknown Actinobacillus pleuropneumoniae Exposure Using a Multiplexing Liquid Array Platform

“…(i) Experimental samples of known A. pleuropneumoniae exposure. Experimental blood serum samples were used from a previous challenge study, the details of which have been described (3). All the original samples, including those that were excluded in the previous study due to the lack of any detectable antibodies, were tested by the novel 4-plex FMIA.…”

“…In addition, eight pigs were vaccinated with in-houseproduced bacterins based on serovar 1, 3, 5, or 7. Blood serum samples were collected at weekly intervals from the time of inoculation or vaccination until day 28 (3). Pigs infected with serovar 13 never seroconverted and were excluded from the study.…”

“…Specifically, a dual-plate CFT for serovars 1, 3, 5, and 7 (29,30), a quadplate indirect ELISA (ELISA-1; Swinecheck mix-APP 1-2-9-11, 3-6-8-15, and 4-5-7, and Swinecheck mix-APP 10-12 antibody test kits; Biovet, Inc., Quebec, Canada) composed of two kits with plates coated with lipopolysaccharide of certain serovars capable of detecting serovars 1 to 12, a single-plate indirect ELISA (ELISA-2; Civtest Suis APP; Laboratories Hipra S.A., Girona, Spain) based on ApxI and surface transferring-binding protein 2 (Tbp2), and a single-plate indirect ELISA (ELISA-3; IDEXX APP-ApxIV antibody [Ab] test; IDEXX Laboratories, Inc., Westbrook, ME) based on a recombinant ApxIV antigen were used. The FMIA and ELISA-3 were directly compared, as both specifically targeted the detection of anti-ApxIV IgG antibodies (3).…”

“…Among the different ELISAs, those targeting ApxIV toxin to quantify circulating levels of the antibody to A. pleuropneumoniae (20), those targeting ApxI, ApxII, or ApxIII for the detection of specific antibodies to Apx exotoxins (21), those targeting capsular polysaccharides of A. pleuropneumoniae to identify antibodies against groups of its serovars (22), and serovar-specific ELISAs based on long-chain lipopolysaccharides (23) have been widely used and offer better sensitivities and specificities than the CFT. It has also been demonstrated that ELISAs utilizing different antigens or that are performed under different assay conditions may have conflicting results and cross-reactions between A. pleuropneumoniae serovars and other bacterial species, and this may be problematic (2,3). The test specificity greatly varies depending on the serologic assay performed.…”

“…A. pleuropneumoniae can infect pigs of all ages, but clinical disease is mainly seen in growing pigs Ͼ12 weeks of age (2,3). The disease is transmitted by the aerosol route or by direct contact and is often highly contagious.…”