Rescue of a bovine respiratory syncytial virus genomic RNA analog by bovine, human and ovine respiratory syncytial viruses confirms the "functional integrity" and "cross-recognition'' of BRSV cis-acting elements by HRSV and ORSV

The nucleotide sequences of the 3' leader and 5' trailer regions were deter mined for genomic RNA of bovine respiratory syncytial virus (BRSV) strain A -51908. The leader and trailer sequences are '45' and '161' nucleotides in length, respectively The functionality of BRSV leader and trailer sequences and their recognition by HRSV and ovine respiratory syncytial virus (ORSV) proteins were examined with a in vitro transcribed BRSV genomic RNA analog carrying the bacterial chloramphenicol acetyl transferase (CAT) gene under the control of BRSV transcription signals. Upon transfection into BRSV, HR SV or ORSV infected cells, the BRSV minireplicons were 'rescued' such that the reporter gene was expressed, the minigenome was replicated and packaged into micrococcal nuclease resistant-infectious minireplicons. The passage of infectious minireplicons could be blocked by a polyclonal BRSV neutraliz ing antiserum. Bovine parainfluenza virus-3, a heterologous paramyxovirus w as inactive in rescuing BRSV genomic RNA analog. Mutational substitution of the G residue at position 4 of leader sequence in the BRSV genomic RNA ana log, with an A or U residue inhibited its transcription and replication, wh ile replacement with a C residue had no significant effect on rescue. These results show that the cis-acting elements of BRSV are functional and are a lso recognized by the proteins of HRSV and ORSV. The helper virus complemen ted rescue system developed here will be useful for characterizing the cis- acting elements of BRSV.