[Effects of hypoxia and acidification stress on ion regulation and gill structure of juvenile Larimichthys crocea.] / ä½Žæ°§åŠé ¸åŒ–èƒè¿«å¯¹å¤§é»„é±¼å¹¼é±¼ç¦»å­è°ƒèŠ‚ä¸Žé³ƒç»„ç»‡ç»“æž„çš„å½±å“.

To investigate the ion regulation of large yellow croaker (Larimichthys crocea) under hypoxia and acidification stresses, we investigated the effects of hypoxia (dissolved oxygen DO 3.5 mg·L-1, pH 8.1), acidification (DO 7.0 mg·L-1, pH 7.35) and combined stresses of hypoxia and acidification (DO 3.5 mg·L-1, pH 7.35) on gill tissue structure and physiological indices related to ion regulation of juvenile L. croaker. The results showed that, under hypoxia stress, gill Na+/K+-ATPase activity, serum Na+, Ca2+ and Cl- contents of juvenile L. croaker decreased first and then increased. Under acidification stress, gill Ca2+-ATPase activity, serum Na+ and Ca2+ contents of juvenile L. croaker increased first and then decreased. Under the combined stresses of hypoxia and acidification, Na+/K+-ATPase activity and Na+, K+ and Ca2+ contents increased first and then decreased, while Ca2+-ATPase activity and Cl- content decreased first and then increased. The results of gill histology showed that hypoxia and acidification stresses led to the detachment of gill epithelial cells, and the combined stresses of hypoxia and acidification led to proliferation, hypertrophy and swelling of gill epithelial cells. Comprehensive analysis showed that hypoxia and acidification stress affected the activities of major ion regulatory enzymes in juvenile L. croaker and caused different degrees of damage to gill tissue, resulting in imbalanced ion regulation in juvenile L. croaker.

SpSR-B2 functions as a potential pattern recognition receptor involved in antiviral and antibacterial immune responses of mud crab Scylla paramamosain.

Although class B scavenger receptors (SR-Bs) in mammals are multifunctional molecules, the functions of SR-Bs in invertebrates remain largely unknown. In this study, we characterized an SR-B homolog, namely SpSR-B2, from Scylla paramamosain. SpSR-B2 shared high similarity with mammalian SR-Bs, and exhibited specific binding activity to ac-LDL, indicating that it may be a new member of SR-B class in invertebrates. SpSR-B2 was upregulated after challenge with white spot syndrome virus (WSSV) or bacteria. Binding assays showed that SpSR-B2 specifically interacted with WSSV envelope protein VP24. Besides, SpSR-B2 could bind to all tested bacterial cells and agglutinate these bacteria. SpSR-B2 also exhibited a strong binding activity to LPS but weak binding activities to other tested polysaccharides. These findings indicated that SpSR-B2 was a potential recognition molecule for viral protein VP24 and bacterial LPS. Knockdown of SpSR-B2 resulted in dramatically decreased expressions of certain antimicrobial peptides (AMPs), and overexpression of SpSR-B2 led to the increased expression of the AMP of SpALF2, suggesting that SpSR-B2 could regulate the expression of AMPs. Taken together, this study revealed that SpSR-B2 functioned as a potential pattern recognition receptor participating in antiviral and antibacterial immunity, and provided new insights into the immune functions of invertebrate SR-Bs.

Characterization of the complete mitochondrial DNA sequence of the Ariosoma meeki (Anguilliformes, Congridae).

In this study, the complete mitochondrial genome of Ariosoma meeki was sequenced, assembled and annotated. The circular genome is 16,154 bp in length with nucleotide composition is 28.42% A, 26.53% T, 19.65% G, and 25.40% C and contains 13 protein-coding genes (PCGs), 21 transfer RNA genes (tRNAs), 2 ribosomal RNA unit genes and a large non-coding region (putative control region). To further explore the evolution relationship of the Anguilliformes, we constructed the phylogenetic tree and found that the A. meeki had closer relationship with Ariosoma shiroanago. This study provided the valuable evidence on phylogenetic relationship of the A. meeki at the molecular level and essential resource for further study the molecular phylogenetic, biogeography and adaptive evolution of this lineage.

Evolutionarily conserved vasa identifies embryonic and gonadal germ cells in spinyhead croaker Collichthys lucidus.

In this study, a 2198 bp full-length cDNA of spinyhead croaker Collichthys lucidus vasa gene encoding 616 amino-acid residues was obtained. Multiple alignment revealed that C. lucidus vasa has eight conserved characteristic motifs of the DEAD box protein family and has the highest identity to large yellow croaker Larimichthys croceas. Reverse-transcription (RT)-PCR and Western blot analyses indicated that the vasa messenger (m)RNA and Vasa protein are specifically expressed in the gonads in both sexes. In situ hybridisation (ISH) demonstrated that vasa RNA is exclusively detected in the germ cells in C. lucidus gonads and its temporospatial expression reveals a dynamic pattern during oogenesis. Surprisingly, C. lucidus vasa 3'UTR can direct stable and specific GFP expression in the primordial germ cells (PGC) of medaka Oryzias latipes embryos. Taken together, these results suggest that because C. lucidus vasa expression delineates critical stages of oogenesis, it may be a useful molecular marker for the identification of gonadal germ cells, facilitating the isolation and utilization of germ cells in future study.

Complete mitochondrial genome of the mudskipper Boleophthalmus pectinirostris (Perciformes, Gobiidae): repetitive sequences in the control region.

The mudskipper, Boleophthalmus pectinirostris (Perciformes, Gobiidae), is an amphibious gobioid fish. In this paper, the complete mitochondrial genome of B. pectinirostris was firstly determined. The mitogenome (17,111 bp) comprises 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and 1 putative control region. 130-bp tandem repeat was identified in the control region, which was almost identical among the 10 individuals examined, and three different frequencies of the repeat unit (five, six or seven) were found among these individuals.

[Multiple nuclear pseudogenes of mitochondrial cytochrome oxidase I gene in Scylla paramamosain (Decapoda : Portunidae)].

Nuclear mitochondrial pseudogenes (Numts), widely existing in nuclear genomes of many organisms, are those nuclear DNA sequences which have high similarity with mtDNAs. In this study, we identified 14 different multiple nuclear pseudogenes of mitochondrial cytochrome oxidase I gene from an individual Scylla paramamosain (Decapoda: Portunidae). Of these sequences, ninety-six variable sites were detected, accounting for 15.7%. Nucleotide diversity (Pi) index and Haplo-type diversity (Hd) index were 0.05682 and 0.8800, respectively. Among the 14 pseudogenes, the copies of sequence one were the most, occupying 30.8% of total copies, and sequence two run the second one (occupying 19.2%). These sequences were divided into two groups compared with their homologues. In Group 1, there were no insertion or deletion sites, while 8 deletion and 5 insertion sites were detected in Group 2, resulting in frame-shift mutations. It was proved that these two groups of pseudogenes were evolved from at least two independent mtDNA integrations into S. paramamosain nuclear genomes through Chi-square and homogeneity tests.

[Separation and characterization of mitochondrial COI pseudogenes in Scylla paramamosain].

Mitochondrial DNA marker has been applied in genetic structure and phylogenetic analysis, but the differences between nuclear mitochondrial pseudogenes (Numts) and mitochondrial DNA complicate phylogenetic analysis. Using the universal and special primers, 34 pseudogene sequences (Numts) and 5 real mtDNA COI sequences were amplified respectively from 34 individuals of Scylla paramamosain from the coast of southeast China. Twenty-nine haplotypes were identified and two groups (Group I and Group II) were determined according to the homology of the 34 pseudogene sequences. Two insertions and an 8-bp deletion were found in some sequences of Group I, but no indel (insertion and deletion) was found in Group II. Compared to mtDNA COI sequences, the two groups had their conservative sites respectively. At least two independent nuclear transfer events are required to explain the codon position biases revealed through the pairwise comparisons of the 34 Numts, one for Group I and one for Group II. The results showed the necessity of great care in interpreting PCR-generated mtDNA sequences using universal primers when used in population or evolutionary studies in genus Scylla.

Molecular cloning and characterization of Stevia rebaudiana UDP-glucosyltransferase.

We report here the cloning and characterization of a UDP-glucose flavonoid glucosyltransferase (srUFGT) in Stevia rebaudiana. The isolated cDNA was 1419 bp in length encoding 473 deduced amino acids with a predicted molecular mass of 53.2 kDa. The products of in vitro translation from an expression vector had anthocyanidins and steviol glucosyltransferase activity. Comparison of the activity of the recombinant UDP-glucosyltransferase toward a range of acceptor substrates suggests that it may participate in the synthesis of steviol glycosides. The results support the hypothesis that the flavonoid glucosyltransferases, which have a broad substrate specificity, may be not only involved in flavonoid glucosylation but also play a role in producing the water-soluble steviol-glycosides in S. rebaudiana.