Analysis of intrapatient heterogeneity uncovers the microevolution of Middle East respiratory syndrome coronavirus

Park, Donghyun; Huh, Hee Jae; Kim, Yeon Jeong; Son, Dae-Soon; Jeon, Hyo-Jeong; Im, Eu-Hyun; Kim, Jong-Won; Lee, Nam Yong; Kang, Eun-Suk; Kang, Cheol In; Chung, Doo Ryeon; Ahn, Jin-Hyun; Peck, Kyong Ran; Choi, Sun Shim; Kim, Yae-Jean; Ki, Chang-Seok; Park, Woong-Yang

Specimens from eight patients who had been identified as MERS-CoV-positive were sequenced. The samples were found positive by both upstream-of-the-envelope gene (upE) and open reading frame 1a (orf1a) real-time polymerase chain reaction (PCR) assays, with cycle threshold (Ct) values averaging 17.3 (15.4-22.6) and 18.3 (15.4-23 .0), respectively (Supplemental Table S1 ). For each patient, a MERS-CoV consensus sequence that indicated the major nucleotide at each genomic position was generated. The genome sequences of the eight isolates differed from each other at only seven positions ( Table 2) . The nucleotide substitutions were all nonsynonymous and occurred in the orf1ab (n = 3) and S (n = 4) genes ( Table 2 ). The sequences of the eight isolates had high nucleotide identities (ranging from 99.96% to 100%, with 99.98% to 100% sequence identities in ORF 1a and 1b [orf1ab] and 99.85% to 100% identities in the spike glycoprotein gene) with recently published sequences of MERS-CoV isolates from the outbreak in South Korea Lu et al. 2015; Seong et al. 2016) . The E, M, and N genes were 100% identical to the previously described MERS-CoV isolates from South Korea. The sequence from Patient 14 in this study differed from the sequence from a previous study at two positions (Seong et al. 2016) .