A polydnavirus, Cotesia plutella bracovirus (CpBV), encodes host translation inhibitory factors (HTIFs). Two HTIFs have been identified and named as CpBV15α and CpBV15β. HTIFs specifically inhibit host gene expression at post transcriptional level. This suggests that HTIFs can discriminate host mRNAs for their inhibition. To test this hypothesis, we chose a storage protein 1 (SP1) for a target and CpBV15β for non-target genes. Both target and non-target genes were constructed in a cloning vector by cloning their 5'UTR+ORF and in vitro transcribed. The capped partial mRNAs were in vitro translated using rabbit reticulocyte lysate in the presence or absence of HTIFs. Translation of SP1 mRNA was significantly inhibited by the HTIF, but those of CpBV15α and CpBV15β were not. We prepared chimeric mRNAs by exchanging 5'UTR and ORF of both target and non-target genes. Inhibitory activity of HTIFs was dependent on the 5'UTR structure. Target 5'UTR had more complicated secondary structure than non-target 5'UTR in terms of free energy required for unwinding. To identify the target molecule of HTIF to discriminate 5'UTR, an immunoprecipitation using HTIF antiserum was conducted. HTIF was co-precipitated with eIF4A that would be required for unwinding the secondary structure of mRNA. These results suggest that HTIF of CpBV can sequester host eIF4A, any mRNAs possessing complicated secondary structures in their 5'UTRs would be difficult to recruit initiation complex for efficient translation.