To construct a rhMPVΔM2-2 virus that can replicate efficiently in Vero cells without trypsin supplementation, we engineered the M2-2 deletion in a different subtype A hMPV strain. This recombinant strain is derived from hMPV/NL/1/00, and contains F 2 /F 1 cleavage-enhancing mutations in the F gene, a property that could facilitate the testing and manufacture of potential live hMPV vaccine candidates [16, 17] . The impact of the physical deletion of M2-2 on hMPV replication, and genetic stability in tissue culture were evaluated. rhMPV/ΔM2-2 exhibited somewhat restricted replication in Vero cells, but was significantly attenuated in hamsters. Hamsters immunized with rhMPV/ΔM2-2 were protected from experimental challenge with wthMPV/NL/1/00. The deletion of M2-2 resulted in higher levels of viral mRNA transcripts in tissue culture, giving rise to aberrant ratios of genomic RNA to viral transcripts. In addition, previously unreported genetic instability was observed, resulting in a higher frequency of point mutations and random insertions of U nucleotides in poly-U tracts of the rhMPV/ΔM2-2 genomic RNA.