Evaluation of diagnostic assays for the serological detection of Actinobacillus pleuropneumoniae on samples of known or unknown exposure.

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 enzyme-linked 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.

Efficient surveillance of pig populations using oral fluids.

Currently virus surveillance in swine herds is constrained by the cost-effectiveness and efficiency of sampling methods. The objective of this study was to assess the value of using oral fluids collected by barn personnel as a method of surveillance based on PCR testing. Approximately 12,150 pigs in 10 wean-to-finish barns on 10 farms were monitored for the presence of porcine circovirus type 2 (PCV2), porcine reproductive and respiratory syndrome virus (PRRSV), influenza A virus (IAV), and Torque teno virus genogroups 1 (TTV1) and 2 (TTV2) by sampling oral fluid specimens. Oral fluid samples were collected from 6 pens at each site starting at the time of pig placement (∼3 weeks of age) and continuing thereafter at 2-week intervals for a period of 18 weeks. Data were analyzed both on a pen basis and barn basis. Overall, 508 (85%) samples were positive for PCV2, 73 (12%) for PRRSV, 46 (8%) for IAV, 483 (81%) for TTV2, and 155 (26%) for TTV1 during the study period. The estimated arithmetic means of the quantitative PCR-positive oral fluids for PCV2, PRRSV, and IAV were 1×10(4.62), 1×10(4.97), and 1×10(5.49)per ml. With a single exception, all barns were positive for PCV2 and TTV2 at every sampling point in the study. Virus detection varied among barns, particularly for IAV and PRRSV. The pen level, cumulative distribution of agent combinations between all 10 barns were statistically different. The most commonly observed patterns were PCV2+TTV2 (239 pen samples, 40%), PCV2+TTV1+TTV2 (88 pen samples, 15%), and PCV2 alone (66 pen samples, 11%). This "proof-of-concept" project showed that a variety of viruses could be detected either intermittently or continuously in pig populations and demonstrated that barn herd virus status is highly variable, even among barns in the same production system. Oral fluid sampling is a promising approach for increasing the efficiency and cost effectiveness of virus surveillance in swine herds.

Interlaboratory comparison of Porcine circovirus-2 indirect immunofluorescent antibody test and enzyme-linked immunosorbent assay results on experimentally infected pigs.

A blinded interlaboratory assessment of the diagnostic agreement and accuracy of serologic tests for routine detection of antibodies against Porcine circovirus-2 (PCV-2), including indirect fluorescent antibody tests (IFATs) and enzyme-linked immunosorbent assays (ELISAs) was conducted in 7 North American laboratories. Serum samples were collected weekly, on trial days 0, 7, 14, 21, 28, 35, 42, and 49, from the following groups of animals: 1) negative controls (n  =  7), 2) PCV-2a (n  =  8), 3) PCV-2b (n  =  8), 4) PCV-1 (n  =  8), 5) PCV-2 vaccine A (n  =  8; Ingelvac® CircoFLEX™), 6) PCV-2 vaccine B (n  =  8; Circumvent® PCV2), and 7) PCV-2 vaccine C (n  =  8; Suvaxyn® PCV2 One Dose). Results from each laboratory were analyzed by kappa and receiver operating characteristic (ROC) analysis. Kappa analysis indicated that, by trial day 49, IFATs had almost perfect agreement, in-house ELISAs had fair to almost perfect agreement, and commercially available anti-PCV-2 immunoglobulin G ELISAs (I or S) had moderate to substantial agreement. From trial days 14-49, the area under the ROC curve for the 2 laboratories that offered IFATs, the 4 laboratories that offered in-house ELISAs, and the 3 laboratories that used commercially available ELISAs ranged from 0.94 to 1.00, 0.72 to 1.00, and 0.95 to 1.00, respectively. However, test sensitivities varied based on laboratory-specific cutoffs that were used to dichotomize test results.