Background To date it is unclear whether SARS-CoV-2 is present in spent dialysate from peritoneal dialysis (PD) patients with COVID-19. Our aim was to assess the presence or absence of SARS-CoV-2 in spent dialysate from chronic PD patients with confirmed diagnosis of COVID-19. Methods Spent PD dialysate samples from COVID-19 positive PD patients were collected between March and August 2020. The multiplexed real-time reverse transcriptase-polymerase chain reaction assay contained primer/probe sets specific to different SARS-CoV-2 genomic regions and to bacteriophage MS2 as internal process control for nucleic acid extraction. Demographic and clinical data were obtained from patients' electronic health records. Results A total of 26 spent PD dialysate samples were collected from 11 patients from 10 dialysis centers. Spent PD dialysate samples were collected on average 25±13 days (median 20, range 10 to 45) after onset of symptoms. The temporal distance of PD effluent collection relative to the closest positive nasal swab RT PCR was 15±11 days (median 14; range 1 to 41). All 26 PD effluent samples tested negative at three SARS-CoV-2 genomic regions. Conclusions Our findings indicate the absence of SARS-CoV-2 in spent PD dialysate collected 10 days or later after the onset of COVID-19 symptoms. We cannot rule out presence of SARS-CoV-2 in spent PD dialysate in the early stage of COVID-19.