Evaluation of poliovirus vaccines for pestivirus contamination: non-specific amplification of poliovirus sequences by pan-pestivirus primers.
Two lots of polyvalent live vaccines for human use against poliovirus were tested by reverse transcriptase (RT) and nested PCR for the presence of contaminating pestivirus RNA. By RT-PCR, samples from both lots showed a band of approximately 450 bp instead of 300 bp for the reference pestivirus strain used as positive control. After nested PCR, the template DNA (450 bp product) was not amplified, suggesting non-specificity of the previous amplification. Sequencing analysis confirmed the non-specificity of the 450 bp bands and revealed, respectively, 80 and 77% homology with a region in the VP1 gene of poliovirus type 1 in samples 1 and 2. This suggests that more caution should be taken in interpreting the results obtained by PCR, and that they should be confirmed by nested PCR or sequencing.
Zanotto
Carlo
C
Department of Pharmacology, University of Milan, Via Vanvitelli, 32, 20129, Milan, Italy.
Giangaspero
Massimo
M
Büttner
Mathias
M
Braun
Angelika
A
Morghen
Carlo De Giuli
Cde G
Elli
Veronica
V
Panuccio
Alfonso
A
Radaelli
Antonia
A
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D004340:Drug Contamination; D004341:Drug Evaluation; D000882:Haplorhini; D006801:Humans; D008969:Molecular Sequence Data; D010576:Pestivirus / Q000235:genetics / Q000302:isolation & purification*; D023321:Poliovirus Vaccines; D016133:Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid
D017931:DNA Primers; D023321:Poliovirus Vaccines