The family Polyomaviridae is characterized by small non-enveloped icosahedral virions that encapsidate a circular double-stranded DNA genome encoding five to eight proteins. Presently, 14 human polyomaviruses have been identified. Of these, at least six may lead to disease in immunocompromised individuals: BK Polyomavirus (BKPyV) causes nephropathy, hemorrhagic cystitis, and urothelial carcinoma, while JC Polyomavirus (JCPyV) is the etiological agent behind progressive multifocal leukoencephalopathy and JCPyV-nephropathy. Merkel cell Polyomavirus (MCPyV) and Trichodysplasia spinulosa Polyomavirus (TSPyV) can cause Merkel cell carcinoma and trichodysplasia spinulosa, respectively. Human polyomavirus 6 and 7 infection (HPyV6/HPyV7) can manifest as hyperproliferative keratinopathy. In adults, the seroprevalence ranges between 30% and 90%. Transmission occurs from an early age, possibly via the skin or the oropharyngeal, respiratory, or gastrointestinal tract. After a primary infection without specific symptoms, a latent or persistent infection is established in the reno-urinary tract, the skin, and other sites. In healthy individuals, BKPyV and JCPyV are intermittently reactivated in the reno-urinary tract and shed in urine without causing any symptoms. However, in kidney transplant patients, uncontrolled high-level virus replication may cause BKPyV- or JCPyV-nephropathy. Polyomavirus pathogenesis is driven by viral cytopathic effects, inflammation, or a combination of these, or the triggering of a high proliferation rate combined with limited cytopathic effects. The diseases are diagnosed by the detection of polyomavirus DNA and/or proteins in combination with cytopathic effects or hyperproliferation. Magnetic resonance imaging is used to detect demyelination and organ damage due to PML. Regrettably, no efficient anti-viral therapies are available, and treatment is based on reducing immunosuppressive treatment and strengthening the polyomavirus-specific immune response.