BackgroundDespite its economic importance, we have a limited understanding of the molecular mechanisms underlying shell formation in pearl oysters, wherein the calcium carbonate crystals, nacre and prism, are formed in a highly controlled manner. We constructed comprehensive expressed gene profiles in the shell-forming tissues of the pearl oyster Pinctada fucata and identified novel shell formation-related genes candidates.Principal FindingsWe employed the GS FLX 454 system and constructed transcriptome data sets from pallial mantle and pearl sac, which form the nacreous layer, and from the mantle edge, which forms the prismatic layer in P. fucata. We sequenced 260477 reads and obtained 29682 unique sequences. We also screened novel nacreous and prismatic gene candidates by a combined analysis of sequence and expression data sets, and identified various genes encoding lectin, protease, protease inhibitors, lysine-rich matrix protein, and secreting calcium-binding proteins. We also examined the expression of known nacreous and prismatic genes in our EST library and identified novel isoforms with tissue-specific expressions.ConclusionsWe constructed EST data sets from the nacre- and prism-producing tissues in P. fucata and found 29682 unique sequences containing novel gene candidates for nacreous and prismatic layer formation. This is the first report of deep sequencing of ESTs in the shell-forming tissues of P. fucata and our data provide a powerful tool for a comprehensive understanding of the molecular mechanisms of molluscan biomineralization.
A strain of Vibrio (KC13.17.5) causing acute hepatopancreatic necrosis disease (AHPND) in shrimp in northern Vietnam was isolated. Normally, AHPND is caused by Vibrio parahaemolyticus, but the genomic sequence of the strain indicated that it belonged to Vibrio harveyi. The sequence data included plasmid-like sequences and putative virulence genes.
Cultured shrimp have a high economic value. Shrimp production had been steadily increasing until 2011, when the outbreak of a new disease, called early mortality syndrome (EMS) or acute hepatopancreatic necrosis disease (AHPND), caused severe damage to the industry (1, 2). AHPND causes mortality of as high as 100%, and the causative agent of AHPND is a strain of Vibrio parahaemolyticus. The sequenced genomes of strains from Thailand and Mexico have a plasmid that might contain putative virulence genes (3, 4). Virulent strains can be identified by PCR using primers based on the virulence genes (5).Because the virulence genes were on a plasmid, virulence might be transferred not only among V. parahaemolyticus strains but also to different bacterial species. In Vietnam, an AHPND strain, KC13.17.5, was isolated and identified to be Vibrio sp., but the 16S rRNA gene sequence showed that it was not V. parahaemolyticus. To further characterize this strain, its genome was sequenced.Bacterial DNA was prepared according to Sambrook and Russell (6). A mate-pair library was generated from DNA using an Illumina Nextera XT DNA sample preparation kit. The library was sequenced using Illumina MiSeq and MiSeq reagent kit version 2 (300 cycles). Sequence data were assembled with CLC Genomics Workbench version 6.5.1, and then the scaffolds were analyzed on the RAST server (7). BLASTn searches of the genome were used to find sequences homologous to known plasmid-like and putative virulence genes (8).The genome of the strain was assembled into 42 scaffolds. Annotation by the RAST server identified 5,288 genes. By further characterization, the closest neighbor of this strain was predicted to be Vibrio harveyi ATCC BAA-1116. A homology search using the predicted genes also showed the higher similarity to the genes of V. harveyi and Vibrio campbellii than those of V. parahaemolyticus.We previously found that contig 4 of TUMSAT_D06_S3 (63 kbp) was highly conserved among 3 AHPND strains but was not conserved in 3 non-AHPND strains (3). This sequence is likely a plasmid. In the present study, some of the scaffolds, such as scaffold 21 (29 kbp) and scaffold 27 (16 kbp), were almost identical to contig 4 of TUMSAT_D06_S3. The putative virulence genes were identified and used to diagnose AHPND by PCR (5). Because scaffold 25 (3.8 kbp) was identical to the virulence genes, we speculate that this strain was made virulent by acquiring the plasmid.Here, we sequenced an AHPND-causing species of Vibrio that was not V. parahaemolyticus. This strain also possesses the putative toxin genes and related plasmid-like sequences. The strain was closest to V. harveyi, meaning that the toxin genes could be transm...
Some strains of Vibrio parahaemolyticus cause acute hepatopancreatic necrosis disease (AHPND) in shrimp. We sequenced 3 AHPND and 3 non-AHPND strains and found that all of them lacked the pathogenicity island relevant to human infection. A unique sequence encoding a type IV pilus/type IV secretion system was found in 3 AHPND strains.
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