Microbial biosensors: a review

A microbial biosensor is an analytical device which integrates microorganism(s) with a physical transducer to generate a measurable signal proportional to the concentration of analytes. In recent years, a large number of microbial biosensors have been developed for environmental, food, and biomedical applications. Starting with the discussion of various sensing techniques commonly used in microbial biosensing, this review article concentrates on the summarization of the recent progress in the fabrication and application of microbial biosensors based on amperometry, potentiometry, conductometry, voltammetry, microbial fuel cell, fluorescence, bioluminescence, and colorimetry, respectively. Prospective strategies for the design of future microbial biosensors will also be discussed.     

Novel Bacillus thuringiensis δ-endotoxin active against Locusta migratoria manilensis

A novel δ-endotoxin gene was cloned from a Bacillus thuringiensis strain with activity against Locusta migratoria manilensis by PCR-based genome walking. The sequence of the cry gene was 3,432 bp long, and it encoded a Cry protein of 1,144 amino acid residues with a molecular mass of 129,196.5 kDa, which exhibited 62% homology with Cry7Ba1 in the amino acid sequence. The δ-endotoxin with five conserved sequence blocks in the amino-terminal region was designated Cry7Ca1 (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"EF486523","term_id":"1121787749","term_text":"EF486523"}}EF486523). Protein structure analysis suggested that the activated toxin of Cry7Ca1 has three domains: 227 residues forming 7 α-helices (domain I); 213 residues forming three antiparallel β-sheets (domain II); and 134 residues forming a β-sandwich (domain III). The three domains, respectively, exhibited 47, 44, and 34% sequence identity with corresponding domains of known Cry toxins. SDS-PAGE and Western blot analysis showed that Cry7Ca1, encoded by the full-length open reading frame of the cry gene, the activated toxin 1, which included three domains but without the N-terminal 54 amino acid residues and the C terminus, and the activated toxin 2, which included three domains and N-terminal 54 amino acid residues but without the C terminus, could be expressed in Escherichia coli. Bioassay results indicated that the expressed proteins of Cry7Ca1 and the activated toxins (toxins 1 and 2) showed significant activity against 2nd instar locusts, and after 7 days of infection, the estimated 50% lethal concentrations (LC₅₀s) were 8.98 μg/ml for the expressed Cry7Ca1, 0.87 μg/ml for the activated toxin 1, and 4.43 μg/ml for the activated toxin 2. The δ-endotoxin also induced histopathological changes in midgut epithelial cells of adult L. migratoria manilensis.     

Alpha-melanocyte stimulating hormone protects retinal pigment epithelium cells from oxidative stress through activation of melanocortin 1 receptor–Akt–mTOR signaling

Patients with age related macular degeneration (AMD) will develop vision loss in the center of the visual field. Reactive oxygen species (ROS)-mediated retinal pigment epithelium (RPE) cell apoptosis is an important contributor of AMD. In this study, we explored the pro-survival effect of α-melanocyte stimulating hormone (α-MSH) on oxidative stressed RPE cells. We found that α-MSH receptor melanocortin 1 receptor (MC1R) was functionally expressed in primary and transformed RPE cells. RPE cells were response to α-MSH stimulation. α-MSH activated Akt/mammalian target of rapamycin (mTOR) and Erk1/2 signalings in RPE cells, which were inhibited by MC1R siRNA knockdown. α-MSH protected RPE cells from hydrogen peroxide (H₂O₂)-induced apoptosis, an effect that was almost abolished when MC1R was depleted by siRNA. α-MSH-mediated S6K1 activation and pro-survival effect against H₂O₂ was inhibited by Akt inhibitors (perifosine, MK-2206 and LY294002). Further, mTOR inhibition by rapamycin, or by mTOR siRNA knockdown, diminished α-MSH’s pro-survival effect in RPE cells. Thus, Akt and its downstream mTOR signaling mediates α-MSH-induced survival in RPE cells. In summary, we have identified a new α-MSH–MC1R physiologic pathway that reduces H₂O₂-induced RPE cell damage, and might minimize the risk of developing AMD.     

青海藏区B型沙眼衣原体的分离培养与鉴定

【目的】对青海藏区沙眼患者标本进行沙眼衣原体分离培养与鉴定。【方法】分别采集患者的单眼结膜和结膜囊拭子标本至1 mL样本保护培养基中。取50μL样本采用离心法感染BGM细胞,37°C培养72 h,连续传代3次,相差显微镜观察衣原体包涵体。对临床样本和分离株分别进行主要外膜蛋白基因ompA序列分析。【结果】共采集了45例活动性沙眼患者的115份临床样本,其中54份样本为ompA PCR阳性,15份样本为沙眼衣原体培养阳性。ompA分析发现,青海藏区沙眼衣原体有3个不同的同源ompA变异株,均为基因B型,都包含有一个泌尿生殖道型沙眼衣原体特有密码子。分离株QH111L和QH111R分别来自编号111患者的左眼和右眼样本,它们ompA基因的可变区有一个非同义碱基差异。该碱基变异仅存在于111号患者的左眼样本中,说明QH111L可能是新出现的ompA突变体。【结论】青海藏区的眼型沙眼衣原体流行株为基因B型,至少存在3个不同的ompA变异株。从青海藏区分离培养了15株眼型沙眼衣原体,发现同一患者的左右眼样本中的沙眼衣原体有不同ompA。本研究为研制沙眼疫苗和诊断试剂奠定了基础,也将有助于理解沙眼的进化和传播。     

Soluble expression, purification and functional identification of a disulfide-rich conotoxin derived from Conus litteratus

Conotoxins are a diverse array of small peptides mostly with multiple disulfide bridges. These peptides become an increasing significant source of neuro-pharmacological probes and drugs as a result of the high selectivity for ion channels and receptors. Usually, the analogue of natural conotoxins is produced by means of chemical synthesis. Here, we present a simple and fast strategy of producing disulfide-rich conotoxins via recombinant expression. By fused with thioredoxin and His tag, a novel O-superfamily conotoxin lt7a was successfully expressed in Escherichia coli and purified, resulting in a high yield of recombinant lt7a about 6 mg/l. The purity of target protein is up to 95% as identified by HPLC results. Whole cell patch-clamp recording revealed that the new conotoxin blocked voltage-sensitive sodium channels in rat dorsal root ganglion neurons, indicating it might be a novel microO-conotoxin.     

Three lysine residues in the common β chain of the interleukin-5 receptor are required for Janus kinase (JAK)-dependent receptor ubiquitination, endocytosis, and signaling

Eosinophils are multifunctional leukocytes implicated in the pathogenesis of numerous inflammatory diseases including allergic asthma and hypereosinophilic syndrome. Eosinophil physiology is critically dependent on IL-5 and the IL-5 receptor (IL-5R), composed of a ligand binding α chain (IL-5Rα), and a common β chain, βc. Previously, we demonstrated that the βc cytoplasmic tail is ubiquitinated and degraded by proteasomes following IL-5 stimulation. However, a complete understanding of the role of βc ubiquitination in IL-5R biology is currently lacking. By using a well established, stably transduced HEK293 cell model system, we show here that in the absence of ubiquitination, βc subcellular localization, IL-5-induced endocytosis, turnover, and IL-5R signaling were significantly impaired. Whereas ubiquitinated IL-5Rs internalized into trafficking endosomes for their degradation, ubiquitination-deficient IL-5Rs accumulated on the cell surface and displayed blunted signaling even after IL-5 stimulation. Importantly, we identified a cluster of three membrane-proximal βc lysine residues (Lys(457), Lys(461), and Lys(467)) whose presence was required for both JAK1/2 binding to βc and receptor ubiquitination. These findings establish that JAK kinase binding to βc requires the presence of three critical βc lysine residues, and this binding event is essential for receptor ubiquitination, endocytosis, and signaling.     

Improvement of xylose utilization in Clostridium acetobutylicum via expression of the talA gene encoding transaldolase from Escherichia coli

Clostridium acetobutylicum ATCC 824 was metabolically engineered for improved xylose utilization. The gene talA, which encodes transaldolase from Escherichia coli K-12, was cloned and overexpressed in C. acetobutylicum ATCC 824. Compared with C. acetobutylicum ATCC 824 (824-WT), the transformant bearing the E. coli talA gene (824-TAL) showed improved ability on xylose utilization and solvents production using xylose as the sole carbon source. During the fermentation of xylose and glucose mixtures with three xylose/glucose ratios (approximately 1:2, 1:1 and 2:1), the rate of xylose consumption and final solvents titers of 824-TAL were all higher than those of 824-WT, despite glucose repression on xylose uptake still existing. These results suggest that the insufficiency of transaldolase in the pentose phosphate pathway (PPP) of C. acetobutylicum is one of the bottlenecks for xylose metabolism and therefore, overexpressing the gene encoding transaldolase is able to improve xylose utilization and solvent production.     

Antitumor mechanism of Se-containing polysaccharide, a novel organic selenium compound

Recent studies on the inhibition of tumor growth by Se-containing polysaccharide were reviewed. Meanwhile, the possible molecular mechanisms of the inhibition of tumor cell growth through antioxidation, induction of tumor cell apoptosis, blockade of cell cycle, and enhancement of immunity by Se-containing polysaccharide were proposed. In the end, the potential application of Se-containing polysaccharide in the prevention and treatment of tumor was elucidated.     

鹿衔草化学成分的研究：羟基肾叶鹿蹄草甙的结构鉴定

从鹿衔草（Ｐｙｒｏｌａ ｃａｌｌｉａｎｔｈａ Ｈ．Ａｎｄｒｅｓ）分离得到两个化合物。经光谱分析,确定其中１个化合物为新化合物,命名为羟基肾叶鹿蹄草甙（１, ｈｙｄｒｏｘｙｌｒｅｎｉｆｏｌｉｎ）,另一化合物为儿茶素（３）。     

库布齐沙漠东部不同生物结皮发育阶段土壤温室气体通量

以流动沙地为对照,采用时空替代法分析库布齐沙漠东部固定沙地上不同发育阶段生物结皮藻类结皮和地衣结皮土壤温室气体通量特征及其与环境因子之间的关系,研究生物结皮发育对荒漠土壤温室气体通量的影响.结果表明: 荒漠土壤CO₂排放通量大小为地衣结皮(128.5 mg·m^-2·h^-1)>藻结皮(70.2 mg·m^-2·h^-1)>流动沙地(48.2 mg·m^-2·h^-1),CH₄吸收通量大小为地衣结皮(30.4 μg·m^-2·h^-1)>藻结皮(21.2 μg·m^-2·h^-1)>流动沙地(18.2 μg·m^-2·h^-1),N₂O排放通量大小为地衣结皮(6.6 μg·m^-2·h^-1)>藻结皮(5.4 μg·m^-2·h^-1)>流动沙地(2.5 μg·m^-2·h^-1).CO₂排放具有明显的季节变化,生长季显著大于非生长季;CH₄和N₂O季节变化差异不显著,前者生长季吸收大于非生长季,后者非生长季排放大于生长季.土壤有机碳和全氮含量、土壤微生物数量均是影响温室气体通量的重要因素,环境水热因子是影响土壤CO₂排放的关键因子,但CH₄和N₂O通量对水热因子的变化不敏感.随着植被恢复和生物结皮发育,荒漠土壤温室气体累积通量的不断增大导致其百年尺度的全球增温潜势亦显著提高,依次为地衣结皮(1135.7 g CO₂-e·m^-2·a^-1)>藻结皮(626.5 g CO₂-e·m^-2·a^-1) >流动沙地(422.7 g CO₂-e·m^-2·a^-1).