Abstract Nephrolithiasis is a prevalent urological disease with complicated pathogenesis. Accumulating evidence suggests microbes may participate in stone formation. JC polyomavirus (JCPyV) commonly infects humans and persists latently in renal tubular epithelial cells with intermittent urinary shedding. However, the association between JCPyV viruria and nephrolithiasis remains unclear. We enrolled 188 nephrolithiasis patients and 194 stone-free controls. Urinary JCPyV DNA was detected by qPCR.