Serological Profiling of PRRS and PCV2

The presence of PRRSv-specific antibodies (Abs) was assessed by means of two commercially available ELISA kits. The IDEXX PRRS X3 Ab test (ELISA 1) (IDEXX Laboratories, Westbrook, ME, USA) was used to analyze the presence of PRRSv-specific Abs directed against recombinant PRRSv-1 and PRRSv-2 ORF7 antigens and is considered the gold standard for PRRSv Ab testing [4 ]. Additionally, Abs directed against a specific PRRSv-1 antigen-gylcoprotein-rich extract were analyzed using the CIVTEST SUIS PRRS E/S test (ELISA 2) (HIPRA, Amer, Spain) [18 (link)]. The presence of porcine circovirus type 2 (PCV2)-specific Abs was assessed using the Biochek PCV2 Antibody Test (Biochek, Reeuwijk, The Netherlands). All ELISA tests were performed according to the manufacturers’ guidelines, without modifications. All washing steps were performed using the WellWash Versa Microplate Washer (Thermo Fisher Scientific, Waltham, MA, USA). Absorbance at 650 nm (ELISA 1), 450 nm (ELISA 2) and 405 nm (PCV2 ELISA) were measured using the MultiSkan FC MicroPlate Photometer (Thermo Fisher Scientific, Waltham, MA, USA). Samples with a sample-to-positive (S/p) ratio ≥ 0.4 or ≥ 0.5 were considered to be seropositive in ELISA 1 or the PCV2 ELISA, respectively. Samples with a relative index percent (IRPC value) > 20 were considered to be seropositive in ELISA 2.