Hemorrhagic fever with renal syndrome (HFRS) is caused by hantaviruses. Data of 34 patients with HFRS hospitalized at Chosun University Hospital, South Korea, between 2010 and 2021 were retrospectively analyzed. Nested reverse transcription polymerase chain reaction (RT-nPCR) targeting the L segment of hantavirus and sequencing were used for diagnosis. Most cases occurred in men and during the months of October through December. Common symptoms were fever, chills, gastrointestinal symptoms, and myalgia. The common laboratory abnormalities were thrombocytopenia, proteinuria, and elevated levels of serum creatinine, aspartate transaminase, alanine transaminase, and lactate dehydrogenase. Approximately 91.2% of patients had the Hantaan virus with a new genotype cluster, whereas 8.8% had the Seoul virus. Seropositivity based on IgM titer >1:32 on admission was noted in 20.6%, and a 4-fold increase in IgG titer of 1:512 was observed in 11.8%. This study demonstrated that RT-nPCR targeting the L segment of hantaviruses is a more reliable diagnostic method compared to serological testing.