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

Abstract

Genome sequence analysis of Middle East respiratory syndrome coronavirus (MERS-CoV) variants from patient specimens has revealed the evolutionary dynamics and mechanisms of pathogenesis of the virus. However, most studies have analyzed the consensus sequences of MERS-CoVs, precluding an investigation of intrapatient heterogeneity. Here, we analyzed non–consensus sequences to characterize intrapatient heterogeneity in cases associated with the 2015 outbreak of MERS in South Korea. Deep-sequencing analysis of MERS-CoV genomes performed on specimens from eight patients revealed significant intrapatient variation; therefore, sequence heterogeneity was further analyzed using targeted deep sequencing. A total of 35 specimens from 24 patients (including a super-spreader) were sequenced to detect and analyze variants displaying intrapatient heterogeneity. Based on the analysis of non–consensus sequences, we demonstrated the intrapatient heterogeneity of MERS-CoVs, with the highest level in the super-spreader specimen. The heterogeneity could be transmitted in a close association with variation in the consensus sequences, suggesting the occurrence of multiple MERS-CoV infections. Analysis of intrapatient heterogeneity revealed a relationship between D510G and I529T mutations in the receptor-binding domain (RBD) of the viral spike glycoprotein. These two mutations have been reported to reduce the affinity of the RBD for human CD26. Notably, although the frequency of both D510G and I529T varied greatly among specimens, the combined frequency of the single mutants was consistently high (87.7% ± 1.9% on average). Concurrently, the frequency of occurrence of the wild type at the two positions was only 6.5% ± 1.7% on average, supporting the hypothesis that selection pressure exerted by the host immune response played a critical role in shaping genetic variants and their interaction in human MERS-CoVs during the outbreak.

Sequential Emergence and Wide Spread of Neutralization Escape Middle East Respiratory Syndrome Coronavirus Mutants, South Korea, 2015

“…Moreover, the second mutation (D510G) was first detected in P014. The respiratory sample collected from P014 on June 1, 2015, included mixed spike sequences (wild-type, I529T, or D510G) and a rare double mutation (D510G-I529T) (11). Even though most of the patients infected by P014 carried MERS-CoV with the I529T mutation, the second mutant D510G was transiently or consistently observed in some of the tertiary and quaternary cases (P050, P066, P080, P122, P155, and P168) (11).…”

“…The respiratory sample collected from P014 on June 1, 2015, included mixed spike sequences (wild-type, I529T, or D510G) and a rare double mutation (D510G-I529T) (11). Even though most of the patients infected by P014 carried MERS-CoV with the I529T mutation, the second mutant D510G was transiently or consistently observed in some of the tertiary and quaternary cases (P050, P066, P080, P122, P155, and P168) (11). These results indicate that P014 initially harbored mixed wild-type and I529T mutant viruses and generated the D510G mutation, but the I529T mutant virus dominated the exposure period.…”

“…We collected genetic information on spike genes of MERS-CoV analyzed in the patients' specimens from the National Center for Biotechnology Information (http://www.ncbi. nlm.nih.gov/) and a previous analysis by Park et al (11). As of January 31, 2018, a total of 75 spike gene sequences from 48 patients in South Korea were available for analysis (Appendix Tables 1, 2, https://wwwnc.cdc.gov/EID/ article/25/6/18-1722-App1.pdf).…”

Temporal profiles of viral load in posterior oropharyngeal saliva samples and serum antibody responses during infection by SARS-CoV-2: an observational cohort study

“…However, single nucleotide polymorphisms were shown to emerge during hospitalisation for MERS-CoV infection, using a targeted sequencing approach. 27 Further studies in more patients with samples obtained at longer intervals could be more informative.…”

Prolonged Shedding of Human Coronavirus in Hematopoietic Cell Transplant Recipients: Risk Factors and Viral Genome Evolution

“…Genetic variability of HCoV OC43 at the community level and intrahost heterogeneity of MERS coronavirus have been reported [22][23][24][25]. Such variability might have important implications in viral disease pathogenesis, and the study of viral genome evolution within a host can provide vital information important in developing and assessing antiviral agents [26].…”