PCR detection of S. calchasi DNA in brains of pigeons with specific nested primer pairs targeting the ITS1 region. All brains of pigeons of group A (no. 2–9) and pigeons of group B (no. 10–12, 15, L60 and L74) except for no. 14 were positive. Five reference animals (r1-r5) used for RT-qPCR as well as negative controls and two processing controls were negative.