Leader switching occurs during the rescue of defective RNAs by heterologous strains of the coronavirus infectious bronchitis virus

A defective RNA (D-RNA), CD-61, derived from the Beaudette strain of the avian coronavirus infectious bronchitis virus (IBV), was rescued (replicated and packaged) using four heterologous strains of IBV as helper virus. Sequence analysis of the genomic RNA from the four heterologous IBV strains (M41, H120, HV10 and D207) identified nucleotide differences of up to 17% within the leader sequence and up to 4.3% within the whole of the adjacent 5' untranslated region (UTR). Analysis of the 5' ends of the rescued D-RNAs showed that the Beaudette leader sequence, present on the initial CD-61, had been replaced with the corresponding leader sequence from the helper IBV strain but the adjacent 5' UTR sequence of the rescued D-RNAs corresponded to the original CD-61 Beaudette sequence. These results demonstrated that the phenomenon of leader switching previously identified for the coronaviruses murine hepatitis virus and bovine coronavirus (BCoV) also occurred during the replication of IBV D-RNAs. Three predicted stem-loop structures were identified within the 5' UTR of IBV. Stem-loop I showed a high degree of covariance amongst the IBV strains providing phylogenetic evidence that this structure exists and is potentially involved in replication, supporting previous observations that a BCoV stem-loop homologue was essential for replication of BCoV defective interfering RNAs.