一株解磷细菌的筛选、鉴定及其溶磷培养条件的优化

从土壤作物根际筛选分离出的一株解磷能力较强的溶磷菌P0417,对其进行16S r DNA基因水平上的初步鉴定,测定其溶解磷的能力,并对该菌的溶磷培养基条件进行优化。结果表明,经序列分析,确定该菌株P0417为洋葱伯克霍尔德氏菌。且其溶磷能力与培养液p H呈显著相关性,当培养基条件为葡萄糖10 g/L、草酸铵0.5 g/L、Na Cl 1.0 g/L时,菌株P0417对Ca3(PO4)2盐培养基具有较好的解磷能力,其解磷能力可达791.84μg/m L。     

Design and application of highly responsive fluorescence resonance energy transfer biosensors for detection of sugar in living Saccharomyces cerevisiae cells

A protein sensor with a highly responsive fluorescence resonance energy transfer (FRET) signal for sensing sugars in living Saccharomyces cerevisiae cells was developed by combinatorial engineering of the domain linker and the binding protein moiety. Although FRET sensors based on microbial binding proteins have previously been created for visualizing various sugars in vivo, such sensors are limited due to a weak signal intensity and a narrow dynamic range. In the present study, the length and composition of the linker moiety of a FRET-based sensor consisting of CFP-linker(1)-maltose-binding protein-linker(2)-YFP were redesigned, which resulted in a 10-fold-higher signal intensity. Molecular modeling of the composite linker moieties, including the connecting peptide and terminal regions of the flanking proteins, suggested that an ordered helical structure was preferable for tighter coupling of the conformational change of the binding proteins to the FRET response. When the binding site residue Trp62 of the maltose-binding protein was diversified by saturation mutagenesis, the Leu mutant exhibited an increased binding constant (82 microM) accompanied by further improvement in the signal intensity. Finally, the maltose sensor with optimized linkers was redesigned to create a sugar sensor with a new specificity and a wide dynamic range. When the optimized maltose sensors were employed as in vivo sensors, highly responsive FRET images were generated from real-time analysis of maltose uptake of Saccharomyces cerevisiae (baker's yeast).     

Role of Nucleotide Binding and GTPase Domain Dimerization in Dynamin-like Myxovirus Resistance Protein A for GTPase Activation and Antiviral Activity

Myxovirus resistance (Mx) GTPases are induced by interferon and inhibit multiple viruses, including influenza and human immunodeficiency viruses. They have the characteristic domain architecture of dynamin-related proteins with an N-terminal GTPase (G) domain, a bundle signaling element, and a C-terminal stalk responsible for self-assembly and effector functions. Human MxA (also called MX1) is expressed in the cytoplasm and is partly associated with membranes of the smooth endoplasmic reticulum. It shows a protein concentration-dependent increase in GTPase activity, indicating regulation of GTP hydrolysis via G domain dimerization. Here, we characterized a panel of G domain mutants in MxA to clarify the role of GTP binding and the importance of the G domain interface for the catalytic and antiviral function of MxA. Residues in the catalytic center of MxA and the nucleotide itself were essential for G domain dimerization and catalytic activation. In pulldown experiments, MxA recognized Thogoto virus nucleocapsid proteins independently of nucleotide binding. However, both nucleotide binding and hydrolysis were required for the antiviral activity against Thogoto, influenza, and La Crosse viruses. We further demonstrate that GTP binding facilitates formation of stable MxA assemblies associated with endoplasmic reticulum membranes, whereas nucleotide hydrolysis promotes dynamic redistribution of MxA from cellular membranes to viral targets. Our study highlights the role of nucleotide binding and hydrolysis for the intracellular dynamics of MxA during its antiviral action.     

Mycobacterium kansasii-induced death of murine macrophages involves endoplasmic reticulum stress responses mediated by reactive oxygen species generation or calpain activation

Although pathogenic mechanisms of tuberculosis have been extensively studied, little is known about the pathogenic mechanisms of Mycobacterium kansasii. In this work the influence of virulence and ER-stress mediated apoptosis of macrophages during two different strains of M. kansasii infection was investigated. We show that M. kansasii infection is associated with ER stress-mediated apoptosis in the murine macrophage cell line RAW 264.7. Infection of RAW 264.7 cells in vitro with apoptosis-inducing a clinical isolate of M. kansasii SM-1 (SM-1) resulted in strong induction of ER stress responses compared with M. kansasii type strain (ATCC 12478)-infected RAW 264.7 cells. Interestingly, inhibition of calpain prevented the induction of CHOP and Bip in ATCC 12478-infected RAW 264.7 cells but not in RAW 264.7 cells infected with SM-1. In contrast, reactive oxygen species (ROS) were significantly increased only in RAW 264.7 cells infected with SM-1. We propose that ROS generation is important for triggering ER stress-mediated apoptosis during SM-1 infection, whereas ATCC 12478-induced, ER stress-mediated apoptosis is associated with calpain activation. Our results demonstrate that the ER stress pathway plays important roles in the pathogenesis of M. kansasii infections, and that different strains of M. kansasii induce different patterns of ER stress-mediated apoptosis.     

三种几丁质合成抑制剂对意大利蝗的防治研究

本文用3种几丁质合成抑制剂卡死克、噻嗪酮和灭幼脲对意大利蝗Calliptatmnus italicus(L.)卵和3龄蝗蝻进行药剂试验.实验结果显示卡死克对意大利蝗药效最高:LC50、LC90分别为1.34、14.17 mg/L.灭幼脲次之LC50、LC90分别为2.09、45.22 mg/L.灭幼脲和卡死克在50 m...     

Intentionally introduced species: more easily invited than removed

Alien species are brought into countries world wide on a massive scale for agricultural production, ex situ conservation, landscape aesthetics, gardens, and ecosystem restoration. Unfortunately, some of these species have escaped and adversely impacted on regional as well as global biodiversity conservation and agricultural production. To reduce such risks, it is necessary to implement specific and effective measures. Since various government departments and institutions are involved in the management of alien species, it is difficult to prevent native and agroecosystems from being invaded by invited species. We propose the establishment of a supervision and inspection continuum over intentional species introduction, similar to that which exists in some countries over unintentional species introductions. Namely, a justification of the necessity to import, a risk assessment, assurances as to provision of an adequate containment facility assessment, and a damage-limitation protocol should that need to be invoked. These requirements should be satisfied before an alien species is knowingly imported, and the necessary follow-up supervision is important post- importation.