Emergence of SARS-CoV-2 with Dual-Drug Resistant Mutations During a Long-Term Infection in a Kidney Transplant Recipient

Yoko Tanino,1,2,* Keisuke Nishioka,1,* Chie Yamamoto,2,* Yohei Watanabe,1,3 Tomo Daidoji,1,4 Masataka Kawamoto,5 Sayaka Uda,6 Shoko Kirito,1 Yuta Nakagawa,2 Yu Kasamatsu,2 Yoshiyuki Kawahara,7 Yuri Sakai,7 Shuji Nobori,8 Tohru Inaba,2 Bon Ota,9 Naohisa Fujita,7 Atsushi Hoshino,10 Yoko Nukui,2 Takaaki Nakaya1 1Department of Infectious Diseases, Kyoto Prefectural University of Medicine, Kyoto, Japan; 2Department of Infection Control and Laboratory Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan; 3JST, MIRAI, Tokyo, Japan; 4School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan; 5Department of Forensics Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan; 6Department of Pulmonary Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan; 7Kyoto Prefectural Institute of Public Health and Environment, Kyoto, Japan; 8Department of Organ Transplantation and General Surgery, Kyoto Prefectural University of Medicine, Kyoto, Japan; 9Department of Emergency Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan; 10Department of Cardiovascular Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan*These authors contributed equally to this workCorrespondence: Takaaki Nakaya; Yohei Watanabe, Department of Infectious Diseases, Kyoto Prefectural University of Medicine, 465 Kajii-cho, Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto, 602-8566, Japan, Tel +81-75-251-5325, Fax +81-75-251-5328, Email tnakaya@koto.kpu-m.ac.jp; nabe@koto.kpu-m.ac.jpIntroduction: Various therapeutic agents are being developed for the treatment of coronavirus disease 2019 (COVID-19). Therefore, it is crucial to accumulate information regarding the features of drug-resistant viruses to these antiviral drugs.Methods: We investigated the emergence of dual-drug resistance in a kidney transplant recipient who received sotrovimab (from day 0) and remdesivir (RDV) (from day 8 to day 17). We sequenced the whole viral genomes from nasopharyngeal swabs taken on day 0 and seven points after starting treatment (on days 12, 19, 23, 37, 43, 48, and 58). The genetic traits of the wild-type (day 0) and descendant viruses (after day 12) were determined by comparing the genomes with those of a Wuhan strain and the day 0 wild-type strain, respectively. Three viral isolates (from samples collected on days 0, 23, and 37) were investigated for their escape ability and growth kinetics in vitro.Results: The sotrovimab resistant mutation (S:E340K) and the RDV resistant mutation RdRp:V792I (nt: G15814A) emerged within 12 days (day 12) and 11 days (day 19) after the treatment, respectively. The day 23 isolate harboring S:E340K/RdRp:V791I was resistant to both sotrovimab and RDV, showing 364- and 2.73-fold higher resistance respectively, compared with the wild-type. Moreover, compared with the day 23 isolate, the day 37 isolate accumulated multiple additional mutations and had a higher level of resistance to both drugs.Conclusion: Drug-resistant variants with double mutations (S:E340K/RdRp:V791I) became dominant within 23 days after starting treatment, suggesting that even a combination therapy involving sotrovimab and RDV, dual-drug resistant viruses may emerge rapidly in immunocompromised patients. The dual-resistant variants had lower virus yields than those of the wild-type virus in vitro, suggesting that they paid a fitness cost.Keywords: SARS-CoV-2, sotrovimab, remdesivir, drug resistance, immunosuppression therapy