We synthesized the cDNAs of an insect picornavirus, infectious flacherie virus of silkworm (IFV), genomic RNA and inserted it into a bacterial plasmid (pUC119). The 9,650 nucleotides (nts) sequence except for the poly(A) tail was obtained from the cloned cDNAs, and the sequence integrity was confirmed by primer extension and direct RNA sequencing. The sequence has a large open reading frame (ORF) of 9,255 nts (3,085 codons) flanked by the short 5' non-coding region (156 nts) and by the rather long 3' non-coding (239 nts). The structural proteins VP3, 4, 1 and 2 were located at the N-terminus of the polyprotein in this order and were preceded by a tentative small peptide. Computer analysis identified the sequences similar to the consensus sequences of 2C (helicase?), 3C (protease), and 3D (RNA polymerase) conserved among mammalian and plant picorna(-like) viruses. In addition, the predicted genome organization of IFV was quite similar to those of picornaviruses. Further analyses of the characteristics of the genome structure and a tentative phylogenetic tree constructed on the basis of the amino acid sequence similarity emphasized the evolutionary relationships among the insect and plant viruses.

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We isolated and characterized a novel positive-sense, single-stranded RNA virus from Aedes larvae collected on Okushiri Island, Hokkaido, Japan. This virus, designated Okushiri virus (OKV), replicated in the Aedes albopictus cell line C6/36 with severe cytopathic effects and produced a large number of spherical viral particles that were 50-70 nm in diameter and released into the cell culture medium. The OKV genome consisted of 9,704 nucleotides, excluding the poly(A) tail at the 3'-terminus, and contained three major open reading frames (ORF1, ORF2, and ORF3). ORF1 encoded a putative protein of approximately 268 kDa that included a methyltransferase domain, FtsJ-like methyltransferase domain, helicase domain, and RNA-dependent RNA polymerase domain. The genome organization and results of a phylogenetic analysis based on the amino acid sequence predicted from the nucleotide sequence indicated that OKV is a member of a new insect virus group of negeviruses with a possible evolutionary relationship to some plant viruses. ORF2 and ORF3 were suggested to encode hypothetical membrane-associated proteins of approximately 45 kDa and 22 kDa, respectively. This is the first study on a novel negevirus isolated from mosquito larvae in Japan.

Between June 1987 and December 1990, an iliac crest prosthesis made of bioactive apatite- and wollastonite-containing glass ceramic (A-W.GC) was used in 60 patients for the reconstruction of the iliac crest defect after harvesting autogenous tricortical iliac bone graft. The clinical results of this prosthesis were satisfactory. No patients felt spontaneous pain in the reconstructed area, and 93% of the patients had no tenderness there. In the radiological evaluation at the final follow-up, no apparent "radiolucent clear zone" was detected at the prosthesis-iliac bone junction in 98% of the patients. Excellent new bone formation between the prosthesis and the iliac crest was also noticed in 96% of the patients. The A-W.GC iliac crest prosthesis was beneficial for reconstruction of the iliac crest defect.

In order to improve a virus-independent gene expression system in insect cells, the effect of the repetition of the promoter sequence of a baculovirus immediate early gene, BmNPV ie1, on the gene expression activity was examined. Tandem repetition of ie1 promoter resulted in about a 4-fold increase of the reporter gene expression activity. Primer extension analysis revealed that the repetition increased not only the amount of transcripts (4 to 5-times) but also the number of sites of transcription initiation (at least 3 extra initiation sites). A mutational experiment demonstrated the involvement of independent cis-acting elements in these two different promoter-enhancing effects.