Abstract
Porcine reproductive and respiratory syndrome virus (PRRSV) and African swine fever virus (ASFV) cause serious economic losses to the swine industry worldwide. Both viruses show a tropism for macrophages, based on the use of specific entry mediators (e.g., Siglec-1 and CD163). Identifying additional mediators of viral entry is essential for advancing antiviral and vaccine development. In this context, monoclonal antibodies (mAbs) are valuable tools. This study employed a library of 166 mAbs targeting porcine alveolar macrophages (PAMs) to identify candidates capable of blocking early infection stages, including viral binding, internalization, and fusion. Immunofluorescence analysis revealed 74 mAbs with cytoplasmic staining and 70 mAbs with membrane staining. Fifteen reacted with blood monocytes as determined by flow cytometry. mAb blocking assays were performed at 4 °C and 37 °C to analyze the ability of mAbs to block PRRSV and/or ASFV infections in PAMs. The mAb 28C10 significantly blocked PRRSV (96% at 4 °C and 80% at 37 °C) and ASFV (64% at 4 °C and 81% at 37 °C) infections. The mAb 28G10B6 significantly blocked PRRSV (86% at 4 °C and 74% at 37 °C) and partially blocked ASFV (35% at 4 °C and 64% at 37 °C) infections. mAb 26B8F5-I only partially blocked PRRSV infection (65% at 4 °C and 46% at 37 °C). Western blotting and mass spectrometry identified the corresponding proteins as Siglec-1 (28C10; 250 kDa), MYH9 (28G10B6; 260 kDa), and ANXA1 (26B8F5-I; 37 kDa). Our findings are indicative that Siglec-1, MYH9, and ANXA1 play a role in PRRSV/ASFV entry into macrophages.