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Pine wilt disease, which is caused by the pine wood nematode (PWN), Bursaphelenchus xylophilus , has caused huge damage to pine forests around the world. In this study, we analysed the PWN transcriptome to investigate the expression of genes related to the associated bacterial species Pseudomonas fluorescens and found that the gene adh- 1 encoding alcohol dehydrogenase (ADH) was upregulated. The open reading frame of adh -1, which encoded a protein of 352 amino acid residues, was cloned from B. xylophilus. Recombinant ADH with a relative molecular weight of 39 kDa, was present mainly in inclusion bodies and was overexpressed in Escherichia coli BL21 (DE3) and purified after refolding. The biochemical assay revealed that recombinant ADH could catalyse the dehydrogen reaction of eight tested alcohols including ethanol in the presence of NAD+. Quantitative real-time RT-PCR analysis indicated that ethanol upregulated adh -1 expression in PWN. Results of RNA interference and inhibition of ADH treatment indicated that downregulating expression of adh -1 or inhibition of ADH could reduce ethanol tolerance and the vitality and reproduction ability of B. xylophilus , suggesting that adh -1 is involved in pathogenicity of PWN. [ABSTRACT FROM AUTHOR]

Lignin is the main component of stone cells, and stone cell content is one of the crucial factors for fruit quality in chinese white pear (Pyrus x bretschneideri). The lignin biosynthesis pathway is complex and involves many enzymatic reactions. Cinnamate-4-hydroxylase [C4H (EC.1.14.13.11)] is an essential enzyme in lignin metabolism. This study was conducted to investigate the effect of bagging on lignin metabolism during fruit development in chinese white pear. The study showed that bagging had little effect on stone cell content, lignin content, C4H activity, and C4H gene expression and that there was a positive correlation between C4H gene expression and lignin content as well as stone cell content. Moreover, a full-length complementary DNA (cDNA) encoding C4H (PbrC4H, GenBank accession number KJ577541.1) was isolated from chinese white pear fruit. The cDNA is 1515 bp long and encodes a protein of 504 amino acids. Sequence alignment suggested that the deduced protein belongs to the P450 gene family and that C4H might be located subcellularly in the cell membrane. The results indicate that bagging cannot change the lignin and stone cell content significantly and that C4H catalyzes a step in lignin biosynthesis. These findings provide certain theoretical references and practical criteria for improving the quality of chinese white pear. [ABSTRACT FROM AUTHOR]