Efforts were begun to adapt MAPREC as a potentially useful regulatory method for control of consistency of production of live viral vaccines other than OPV. High-passage samples of yellow fever virus vaccine (YFV) and of a cloned Jeryl-Lynn strain of mumps virus were obtained and subjected to sequence heterogeneity assay. Two nucleotide differences were identified in serially passaged YFV, and amounts of those mutations in various passage levels, and in commercial batches of YFV vaccine are now being quantified by MAPREC. A nucleotide sequence difference between the original strain of YFV and the passaged virus stocks was detected. Different methods for quantificaiton of this mutation were evaluated. They include hybridization with oligonucleotide microchips and single-base extension reaction. Quantitative aspects of these tests are now being compared. A new mumps candidate vaccine strain, derived from the licensed Jeryl-Lynn vaccine strain, was completely sequenced, and that sequence was compared to another mumps strain, the nucleotide sequence of which has been published. A high passage of the new Jeryl-Lynn strain was also sequenced. Serially passaged preparations of mumps vaccine viruses are now being compared to unpassaged vaccine virus preparations to identify unstable genomic sites suitable for monitoring the consistency of mumps vaccine production. A MAPREC test for quantification of sequence heterogeneities in Mumps vaccine batches was developed. In addition, other quantitaitve techniques including MALDI-TOF mass-spectrometry and hybridization with oligonucleotide microchips were also used. Another part of this project deals with determination of ratio of two genetic sub-components of Jeryl Lynn vaccine. The strain contains two distinct strains, relative contents of which changes depending on growth condition. In order to ensure consistency of production, a method for quantificaiton of the sub-strains is highly desirable. The complete nucleotide sequences of both sub-strains were determined, and methods for their quantificaiton developed. The results of the studies with mumps vaccine strains were presented at the ASV Meeting.
