Asymmetric Radical Oxyboration of ß-Substituted Styrenes via Late-Stage Stereomutation.

Herein, we report an unprecedented copper-catalyzed highly enantio- and diastereoselective radical oxyboration of ß-substituted styrenes. The lynchpin of success is ascribed to the development of a late-stage stereomutation strategy, which enables enantioenriched cis-isomers among a couple of early-generated diastereomers to be converted into trans-isomer counterparts, thus fulfilling high diastereocontrol; while the degree of enantio-differentiation is determined by the borocupration process of the C=C bond. This reaction provides an efficient protocol to access enantioenriched trans-1,2- dioxygenation products. The value of this method has further been highlighted in a diversity of follow-up stereospecific transformations and further modifying complex molecules.

Copper-Catalyzed Defluorinative Boroarylation of Alkenes with Polyfluoroarenes.

A catalytic defluorinative boroarylation of alkenes with polyfluoroarenes and B2pin2 with the assistance of a PCy3-ligated copper catalyst was developed. Taking advantage of bench-stable alkenes as latent nucleophiles and avoiding traditional reliance on stoichiometric quantities of organometallics, this method showcased good functional group compatibility and proceeded under very mild reaction conditions. A series of valuable boronate-containing polyfluoroarenes including all-carbon quaternary carboncenter-containing triaryl alkylboronates that are otherwise difficult to access were efficiently prepared.

Copper-Catalyzed Defluorinative Hydroarylation of Alkenes with Polyfluoroarenes.

A catalytic defluorinative hydroarylation of alkenes with polyfluoroarenes in the presence of dppbz-ligated Cu catalyst and silanes was developed. This method provides a straightforward and alternative avenue to synthetic important polyfluorinated arenes with readily available and bench-stable alkenes as latent nucleophiles, and therefore avoids conventional reliance on stoichiometric quantities of organometallic reagents. This reaction proceeds under very mild conditions and exhibits good functional group compatibility and high level of regioselectivity. The synthetic potential of this method was further demonstrated by a gram-scale synthesis, and an array of experimental studies were also carried out to elaborate the probable mechanism.

Copper-Catalyzed Asymmetric Reductive Allylation of Ketones with 1,3-Dienes.

A catalytic chemo-, regio-, and enantioselective allylation of ketones with 1,3 dienes in the presence of ( R, R)-Ph-BPE ligated Cu catalyst and hydrosilane is presented. This method provides a straightforward and alternative avenue to synthesize chiral homoallylic tertiary alcohols with 1,3-dienes as the latent allylic nucleophiles and avoids the traditional reliance on stoichiometric quantities of allylmetal reagents. This transformation proceeds under very mild conditions and displays good functional group tolerance and could be performed on a gram-scale.

[Effects of brief exposure to high temperature on Neoseiulus californicus].

To determine the influence of high temperature shock on the survival characteristics and population development of N. californicus, the hatching rate, survival rate and developmental duration were investigated after the eggs, larvae and adults of N. californicus were exposed to high temperatures (35, 38, 42, 45 °C) for 1-8 h. The results showed that with higher temperature and longer time, the survival rate of eggs and larvae would be lower, and their developmental duration declined firstly and then increased. The hatching rate of eggs treated at 42 °C for 8 h was significantly lower than that at 35 °C for 8 h. The developmental duration of egg (4.1 d) was shortest when treated at 38 °C for 8 h. The egg couldn't hatch when treated at 45 °C for 2 h. The survival rate of larvae which was treated at 45 °C for 4 h was significantly lower than. that at 35 °C for 4 h, and the larvae wouldn' t survive when treated at 45 °C for 8 h. The spawning period and total eggs of female adults increased firstly and then decreased. The egg-laying amount of a female N. californicus was 38.9, 36.7 and 14.5 at 35, 38 and 45 °C, respectively. High temperature exposure had significant effects on the egg hatching rate, survival rate and development duration of N. californicus, but had little effect on the pre-oviposition and survival rate of the adults.

Characterization and virus susceptibility of a skin cell line from red-spotted grouper (Epinephelus akaara).

A red-spotted grouper Epinephelus akaara skin (RGS) cell line was established and characterized. RGS cells had a normal diploid chromosome number of 2n = 48, the morphology of which was fibroblastic-like in 3 days and epithelial-like over 5 after 16 passages. The cells multiplied well in Dulbecco's modified Eagle's medium supplemented with 10% of fetal bovine serum at 25°C. Susceptibilities of RGS and grass carp ovary (GCO) cells to two viruses were tested, and the results showed that the titer of an iridovirus Rana grylio virus (RGV) in RGS cells was 10³·5 TCID50 ml⁻¹, which was much higher than a rhabdovirus spring viremia of carp virus (SVCV) in the cells (10°·5 TCID50 ml⁻¹). The titers of RGV and SVCV in GCO were 106·° TCID50 ml⁻¹ and 108·° TCID50 ml⁻¹, respectively, which were higher than those in RGS cells. The data may imply that RGS cells could be selectively resistible to some viruses during infection. RT-PCR analysis of RGV-infected RGS cells showed that RGV could replicate in RGS cells. Further study of virus replications in RGS cells was conducted by electron microscopy and immunofluorescence microscopy has shown that virus particles scattered in the cytoplasm and virus protein appeared in both the cytoplasm and nucleus. The results suggested that RGS cells could be used as a potential in vitro model to study the cutaneous barrier function against virus infection.

Herpes-like virus infection in Yangtze finless porpoise (Neophocaena phocaenoides): pathology, ultrastructure and molecular analysis.

A moribund juvenile Yangtze finless porpoise (Neophocaena phocaenoides) with skin lesions and ulceration was found in Dongting Lake, China. Pathologic examination, electron microscopy, and polymerase chain reaction of liver tissue revealed widely distributed necrotic lesions, sinusoidal dilatation, congestion and herpes-like virus particles.

Genome of turbot rhabdovirus exhibits unusual non-coding regions and an additional ORF that could be expressed in fish cell.

Genomic sequence of Scophthalmus maximus rhabdovirus (SMRV) isolated from diseased turbot has been characterized. The complete genome of SMRV comprises 11,492 nucleotides and encodes five typical rhabdovirus genes N, P, M, G and L. In addition, two open reading frames (ORF) are predicted overlapping with P gene, one upstream of P and smaller than P (temporarily called Ps), and another in P gene which may encodes a protein similar to the vesicular stomatitis virus C protein. The C ORF is contained within the P ORF. The five typical proteins share the highest sequence identities (48.9%) with the corresponding proteins of rhabdoviruses in genus Vesiculovirus. Phylogenetic analysis of partial L protein sequence indicates that SMRV is close to genus Vesiculovirus. The first 13 nucleotides at the ends of the SMRV genome are absolutely inverse complementarity. The gene junctions between the five genes show conserved polyadenylation signal (CATGA(7)) and intergenic dinucleotide (CT) followed by putative transcription initiation sequence A(A/G)(C/G)A(A/G/T), which are different from known rhabdoviruses. The entire Ps ORF was cloned and expressed, and used to generate polyclonal antibody in mice. One obvious band could be detected in SMRV-infected carp leucocyte cells (CLCs) by anti-Ps/C serum via Western blot, and the subcellular localization of Ps-GFP fusion protein exhibited cytoplasm distribution as multiple punctuate or doughnut shaped foci of uneven size.

Generation and characterization of monoclonal antibodies against the flounder Paralichthys olivaceus rhabdovirus.

Two MAbs (3C7 and 3C9) against flounder Paralichthys olivaceus rhabdovirus (PORV) were generated with hybridoma cell fusion technology and characterized by an indirect enzyme-linked immunosorbent assay, isotype test, Western blot and immunodot analysis and immunofluorescence assay. Isotyping tests demonstrated that both of the two MAbs belonged to IgM subclass. Western blot analysis showed the MAbs reacted with 42, 30, and 22 kDa viral proteins, which were localized within the cytoplasm of PORV-infected grass carp ovary (GCO) cells analyzed by indirect immunofluorescences tests. The MAb 3C7 was also selected at random for detecting virus antigens in the inoculated grass carp tissues by immunohistochemistry assay. Flow cytometry tests showed that at the 36 h postinfection (0.25 PFU/cell), the 23% PORV-infected GCO cells could be distinguished from the uninfected cells with the MAb 3C7. Such MAbs could be useful for diagnosis and potential treatment of viral infection.

Establishment, characterization, and virus susceptibility of a new marine cell line from red spotted grouper (Epinephelus akaara).

A marine fish cell line from the snout of red spotted grouper Epinephelus akaara, a protogynous hermaphrodite, was established, characterized, and subcultured with more than 60 passages. The grouper snout cell line (GSC) cells multiplied well in Dulbecco's modified Eagle's medium (DMEM) medium supplemented with 10% fetal bovine serum. The optimal growth temperature was 25 degrees C, and morphologically the cells were fibroblastic. Chromosome analysis revealed that the GSC cell line has a normal diploid karyotype with 2n = 8st + 40t. A virus titration study indicated that the cells were susceptible to turbot Scophthalmus Maximus rhabdovirus (SMRV) (10(8.5) TCID(50) ml(-1)), while the viral titer of frog Rana grylio virus 9807 (RGV(9807)) reached 10(3.5) TCID(50) ml(-1). The infection was confirmed by cytopathic effect (CPE), immunofluorescence, and electron microscopy experiments, which detected the viral particles in the cytoplasm of virus-infected cells, respectively. Further, significant fluorescent signals were observed when the GSC cells were transfected with pEGFP vector DNA, indicating their potential utility for transgenic and genetic manipulation studies.

Infection and propagation of lymphocystis virus isolated from the cultured flounder Paralichthys olivaceus in grass carp cell lines.

The causative agent of lymphocystis disease that frequently occurs in cultured flounder Paralichthys olivaceus in China is lymphocystis virus (LV). In this study, 13 fish cell lines were tested for their susceptibility to LV. Of these, 2 cell lines derived from the freshwater grass carp Ctenopharyngodon idellus proved susceptible to the LV, and 1 cell line, GCO (grass carp ovary), was therefore used to replicate and propagate the virus. An obvious cytopathic effect (CPE) was first observed in cell monolayers at 1 d post-inoculation, and at 3 d this had extended to about 75% of the cell monolayer. However, no further CPE extension was observed after 4 d. Cytopathic characteristics induced by the LV were detected by Giemsa staining and fluorescence microscopic observation with Hoechst 33258 staining. The propagated virus particles were also observed by electron microscopy. Ultrastructure analysis revealed several distinct cellular changes, such as chromatin compaction and margination, vesicle formation, cell-surface convolution, nuclear fragmentation and the occurrence of characteristic 'blebs' and cell fusion. This study provides a detailed report of LV infection and propagation in a freshwater fish cell line, and presents direct electron microscopy evidence for propagation of the virus in infected cells. A possible process by which the CPEs are controlled is suggested.