“伯杰氏系统细菌学手册(第二版)”第5卷与我国的放线菌系统学研究

"伯杰氏系统细菌学手册"(下文简称"伯杰氏手册"),是世界各国分类学家普遍接受的学术观点的汇总,集科学性、统一性和实用性于一身。2012年5月,随着"伯杰氏手册"第二版第5卷(放线菌专刊)分A、B两册出版,这部经典巨著在Michael Goodfellow等的领导下精心组织并顺利完成。"伯杰氏手册"第5卷对放线菌分类系统做出了重大调整,正式建立了放线菌门,包括6个纲、23个目(含一个未确定目)、53个科、222个属、近3000个种,其分类阶元为细菌域、放线菌门,在门下为纲、目、科、属和种。"伯杰氏手册"收录了我国放线菌分类学研究的大量成果,这是我国四代放线菌分类学家们共同努力的结果。但需要指出的是,由于"伯杰氏手册"过于严谨、保守的著书宗旨与漫长的出版周期,对DNA基因多位点序列分析(MLSA)技术、基因芯片技术和基因组技术等在分类学领域中所做出的新研究成果采纳不足,而这部分内容或许在不久的将来会使原核生物分类学发生深刻的改变。     

Targeting Photoreceptors via Intravitreal Delivery Using Novel,Capsid-Mutated AAV Vectors

Development of viral vectors capable of transducing photoreceptors by less invasive methods than subretinal injection would provide a major advancement in retinal gene therapy. We sought to develop novel AAV vectors optimized for photoreceptor transduction following intravitreal delivery and to develop methodology for quantifying this transduction in vivo. Surface exposed tyrosine (Y) and threonine (T) residues on the capsids of AAV2, AAV5 and AAV8 were changed to phenylalanine (F) and valine (V), respectively. Transduction efficiencies of self-complimentary, capsid-mutant and unmodified AAV vectors containing the smCBA promoter and mCherry cDNA were initially scored in vitro using a cone photoreceptor cell line. Capsid mutants exhibiting the highest transduction efficiencies relative to unmodified vectors were then injected intravitreally into transgenic mice constitutively expressing a Rhodopsin-GFP fusion protein in rod photoreceptors (Rho-GFP mice). Photoreceptor transduction was quantified by fluorescent activated cell sorting (FACS) by counting cells positive for both GFP and mCherry. To explore the utility of the capsid mutants, standard, (non-self-complementary) AAV vectors containing the human rhodopsin kinase promoter (hGRK1) were made. Vectors were intravitreally injected in wildtype mice to assess whether efficient expression exclusive to photoreceptors was achievable. To restrict off-target expression in cells of the inner and middle retina, subsequent vectors incorporated multiple target sequences for miR181, an miRNA endogenously expressed in the inner and middle retina. Results showed that AAV2 containing four Y to F mutations combined with a single T to V mutation (quadY−F+T−V) transduced photoreceptors most efficiently. Robust photoreceptor expression was mediated by AAV2(quadY−F+T−V) −hGRK1−GFP. Observed off-target expression was reduced by incorporating target sequence for a miRNA highly expressed in inner/middle retina, miR181c. Thus we have identified a novel AAV vector capable of transducing photoreceptors following intravitreal delivery to mouse. Furthermore, we describe a robust methodology for quantifying photoreceptor transduction from intravitreally delivered AAV vectors.     

The Evidence for Association of ATP2B2 Polymorphisms with Autism in Chinese Han Population

Background

Autism is a neurodevelopmental disorder with a high estimated heritability. ATP2B2, located on human chromosome 3p25.3, encodes the plasma membrane calcium-transporting ATPase 2 which extrudes Ca²⁺ from cytosol into extracellular space. Recent studies reported association between ATP2B2 and autism in samples from Autism Genetic Resource Exchange (AGRE) and Italy. In this study, we investigated whether ATP2B2 polymorphisms were associated with autism in Chinese Han population.

Methods

We performed a family based association study between five SNPs (rs35678 in exon, rs241509, rs3774180, rs3774179, and rs2278556 in introns) in ATP2B2 and autism in 427 autism trios of Han Chinese descent. All SNPs were genotyped using the Sequenom genotyping platform. The family-based association test (FBAT) program was used to perform association test for SNPs and haplotype analyses.

Results

This study demonstrated a preferential transmission of T allele of rs3774179 to affected offsprings under an additive model (T>C, Z = 2.482, p = 0.013). While C allele of rs3774179 showed an undertransmission from parents to affected children under an additive and a dominant model, respectively (Z = −2.482, p = 0.013; Z = −2.591, p = 0.0096). Haplotype analyses revealed that three haplotypes were significantly associated with autism. The haplotype C-C (rs3774180–rs3774179) showed a significant undertransmission from parents to affected offsprings both in specific and global haplotype FBAT (Z = −2.037, p = 0.042; Global p = 0.03). As for the haplotype constructed by rs3774179 and rs2278556, C-A might be a protective haplotype (Z = −2.206, p = 0.027; Global p = 0.04), while T-A demonstrated an excess transmission from parents to affected offsprings (Z = 2.143, p = 0.032). These results were still significant after using the permutation method to obtain empirical p values.

Conclusions

Our research suggested that ATP2B2 might play a role in the etiology of autism in Chinese Han population.     

Wengen,the Sole Tumour Necrosis Factor Receptor in Drosophila,Collaborates with Moesin to Control Photoreceptor Axon Targeting during Development

Photoreceptor neurons (R cells) in the Drosophila eye define a map of visual space by connecting to targets in distinct layers of the optic lobe, with R1-6 cells connecting to the lamina (the first optic ganglion) and R7 and R8 cells connecting to the medulla (the second optic ganglion). Here, we show that Wengen (Wgn) directly binds Moesin (Moe) through a cytosolic membrane proximal domain and this interaction is important for mediating two distinct aspects of axonal targeting. First, we show that loss of wgn or moe function disrupts cell autonomous R8 axon targeting. Second, we report that wgn or moe mutants show defects in R2–R5 targeting that result from disruption of non-cell autonomous effects, which are secondary to the cell autonomous R8 phenotype. Thus, these studies reveal that the Wgn-Moe signaling cascade plays a key role in photoreceptor target field innervations through cell autonomous and non-cell autonomous mechanisms.     

Bacillus thuringiensis Crystal Protein Cry6Aa Triggers Caenorhabditis elegans Necrosis Pathway Mediated by Aspartic Protease (ASP-1)

Cell death plays an important role in host-pathogen interactions. Crystal proteins (toxins) are essential components of Bacillus thuringiensis (Bt) biological pesticides because of their specific toxicity against insects and nematodes. However, the mode of action by which crystal toxins to induce cell death is not completely understood. Here we show that crystal toxin triggers cell death by necrosis signaling pathway using crystal toxin Cry6Aa-Caenorhabditis elegans toxin-host interaction system, which involves an increase in concentrations of cytoplasmic calcium, lysosomal lyses, uptake of propidium iodide, and burst of death fluorescence. We find that a deficiency in the necrosis pathway confers tolerance to Cry6Aa toxin. Intriguingly, the necrosis pathway is specifically triggered by Cry6Aa, not by Cry5Ba, whose amino acid sequence is different from that of Cry6Aa. Furthermore, Cry6Aa-induced necrosis pathway requires aspartic protease (ASP-1). In addition, ASP-1 protects Cry6Aa from over-degradation in C. elegans. This is the first demonstration that deficiency in necrosis pathway confers tolerance to Bt crystal protein, and that Cry6A triggers necrosis represents a newly added necrosis paradigm in the C. elegans. Understanding this model could lead to new strategies for nematode control.     

Incidence of Co-Infections of HIV,Herpes Simplex Virus Type 2 and Syphilis in a Large Cohort of Men Who Have Sex with Men in Beijing,China

Background

The HIV-epidemic among MSM in China has worsened. In this key population, prevalence of HSV-2 and syphilis infection and co-infection with HIV is high.

Methods

A longitudinal study was conducted (n = 962) in Beijing, China, with three overlapping cohorts (n = 857, 757 and 760) consisting of MSM that were free from pairs of infections of concern (i.e. HIV-HSV-2, HIV-syphilis, HSV-2-syphilis) at baseline to estimate incidence of HIV, HSV-2, syphilis, and those of co-infection.

Results

The incidence of HIV, HSV-2 and syphilis in the overall cohort was 3.90 (95% CI = 2.37, 5.43), 7.87 (95% CI = 5.74, 10.00) and 6.06 (95% CI = 4.18, 7.94) cases per 100 person-years (PYs), respectively. The incidence of HIV-HSV-2, HIV-Syphilis and HSV-2-Syphilis co-infections was 0.30 (95% CI = 0.29, 0.88), 1.02 (95% CI = 0.13, 2.17) and 1.41 (95% CI: 0.04, 2.78) cases per 100 PYs, respectively, in the three sub-cohorts constructed for this study.

Conclusions

The incidence of HIV, HSV-2 and syphilis was very high and those of their co-infections were relatively high. Such co-infections have negative impacts on the HIV/STI epidemics. Prevention practices need to take such co-infections into account.     

RY10-4 Inhibits the Proliferation of Human Hepatocellular Cancer HepG2 Cells by Inducing Apoptosis In Vitro and In Vivo

This study aimed to investigate the anti-tumor activity of RY10-4, a small molecular that was designed and synthesized based on the structure of protoapigenone. A previous screening study showed that RY10-4 possessed anti-proliferative effects against HepG2 human hepatocellular carcinoma cells. However, the full range of RY10-4 anti-cancer effects on liver tumors and the underlying mechanisms have not been identified. Herein, employing flow cytometry, and Western blot analysis, we demonstrate that RY10-4 can induce cell cycle arrest, intracellular reactive oxygen species (ROS) production and apoptosis in HepG2 cells. In HepG2 cell xenograft tumor model, RY10-4 significantly inhibited the growth of tumors and induced apoptosis in tumor cells, with little side effects. Moreover, RY10-4 caused the suppression of STAT3 activation, which may be involved the apoptosis induction. In addition, RY10-4 inhibited the proliferation of Hep3B and HuH-7 human hepatocellular carcinoma cells in a concentration-dependent manner. Taken together, our results suggest that RY10-4 has a great potential to develop as chemotherapeutic agent for liver cancer.     

Identification and Severity Determination of Wheat Stripe Rust and Wheat Leaf Rust Based on Hyperspectral Data Acquired Using a Black-Paper-Based Measuring Method

It is important to implement detection and assessment of plant diseases based on remotely sensed data for disease monitoring and control. Hyperspectral data of healthy leaves, leaves in incubation period and leaves in diseased period of wheat stripe rust and wheat leaf rust were collected under in-field conditions using a black-paper-based measuring method developed in this study. After data preprocessing, the models to identify the diseases were built using distinguished partial least squares (DPLS) and support vector machine (SVM), and the disease severity inversion models of stripe rust and the disease severity inversion models of leaf rust were built using quantitative partial least squares (QPLS) and support vector regression (SVR). All the models were validated by using leave-one-out cross validation and external validation. The diseases could be discriminated using both distinguished partial least squares and support vector machine with the accuracies of more than 99%. For each wheat rust, disease severity levels were accurately retrieved using both the optimal QPLS models and the optimal SVR models with the coefficients of determination (R²) of more than 0.90 and the root mean square errors (RMSE) of less than 0.15. The results demonstrated that identification and severity evaluation of stripe rust and leaf rust at the leaf level could be implemented based on the hyperspectral data acquired using the developed method. A scientific basis was provided for implementing disease monitoring by using aerial and space remote sensing technologies.     

Mono-sulfonated tetrazolium salt based NAD(P)H detection reagents suitable for dehydrogenase and real-time cell viability assays

Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and is important for several biological processes. For GDH inhibitor screening, we developed a novel mono-sulfonated tetrazolium salt (EZMTT), which can be synthesized using H₂O₂ oxidation and purified easily on silica gel in large quantities. The EZMTT detection method showed linear dose responses to NAD(P)H, dehydrogenase concentration and cell numbers. In E. coli GDH assay, the EZMTT method showed excellent assay reproducibility with a Z factor of 0.9 and caused no false positives in the presence of antioxidants (such as BME). Using the EZMTT-formazan-NAD(P)H system, we showed that EGCG is a potent E. coli GDH inhibitor (IC₅₀ 45 nM) and identified that Ebselen, a multifunctional thioredoxin reductase inhibitor, inactivated E. coli GDH (IC₅₀ 213 nM). In cell-based assays at 0.5 mM tetrazolium concentration, EZMTT showed essentially no toxicity after a 3-day incubation, whereas 40% of inhibition was observed for WST-8. In conclusion, EZMTT is a novel tetrazolium salt which provides improved features that are suitable for dehydrogenases and real-time cell-based high-throughput screening (HTS).