结核分枝杆菌可视化抗体检测蛋白芯片的制备

目的：利用克隆表达的7种结核分枝杆菌优势表位抗原,建立可视化抗体检测蛋白芯片,用于结核病辅助诊断。方法：将7种结核分枝杆菌优势表位抗原,即38kD、ESAT-6、CFP10、MPT64、Mtb8、Mtb8.4和Mtb16.3点于修饰的基片上,制备可检测7种结核抗体的多靶点蛋白微阵列,建立免疫金银染色检测系统;使用该芯片对48例临床结核病患者血液样品进行检测,并与“金标准”痰涂片（48例）和痰培养（其中的29例）检测结果进行比较,分析其敏感性;对30名献血员血液样品进行检测,分析其特异性。结果：可视化抗体检测蛋白芯片的敏感性分别为98.5％和96.6％,而痰涂片和痰培养检测方法的敏感性分别为35.4％和48.3％;可视化抗体检测蛋白芯片的特异性为93.3％。结论：建立的结核分枝杆菌可视化抗体检测蛋白芯片检测敏感性显著高于痰涂片和痰培养方法,可用于结核病的临床辅助诊断,提高痰涂片和痰培养假阴性的检出率。     

Species delineation using Bayesian model-based assignment tests: a case study using Chinese toad-headed agamas (genus <Emphasis Type="Italic">Phrynocephalus</Emphasis>)

Background  

Species are fundamental units in biology, yet much debate exists surrounding how we should delineate species in nature. Species discovery now requires the use of separate, corroborating datasets to quantify independently evolving lineages and test species criteria. However, the complexity of the speciation process has ushered in a need to infuse studies with new tools capable of aiding in species delineation. We suggest that model-based assignment tests are one such tool. This method circumvents constraints with traditional population genetic analyses and provides a novel means of describing cryptic and complex diversity in natural systems. Using toad-headed agamas of the Phrynocephalus vlangalii complex as a case study, we apply model-based assignment tests to microsatellite DNA data to test whether P. putjatia, a controversial species that closely resembles P. vlangalii morphologically, represents a valid species. Mitochondrial DNA and geographic data are also included to corroborate the assignment test results.     

益生菌混合发酵条件及其对发酵胡萝卜汁色泽及风味的影响

目的 研究3株益生菌混合发酵胡萝卜汁的发酵条件对色泽、风味的影响及其贮藏特性等.方法 通过菌种配比、单因素试验及正交试验优化了胡萝卜汁的发酵条件,并利用分光测色计和气质联用仪分别研究了发酵前后胡萝卜汁的风味成分和色泽变化.结果 添加30％ (m/m)的新鲜胡萝卜汁(原液),接种3％的以1∶1∶1(v/v/v)混合的嗜热...     

Crystine: fibrous biomolecular material from protein crystals cross-linked in a specific geometry

Cysteine substitutions were engineered on the surface of maltose binding protein to produce crystine fibers, linear polymers of folded protein formed within a crystal. Disulfide bond formation between adjacent protein molecules within the lattice was monitored by X-ray crystallography. The cross-linked crystals were resistant to dissolution in water or neutral buffer solutions, even though the cross-linking was one-dimensional. However, crystine fibers were observed by transmission electron microscopy to dissociate from the crystals in acidic solutions. Some fibers remained associated as two-dimensional bundles or sheets, with a repeat unit along the fibers consistent with the packing of the individual protein molecules in the crystal. Neutralization of the acidic solutions caused the fibers to re-associate as a solid. Crystine threads were drawn out of this solution. In scanning electron microscopy images, many individual fibers could be seen unwinding from the ends of some threads. Crystine fibers are a new type of biomolecular material with potential applications wherever the use of proteins in a fibrous form is desirable, for example, the incorporation of enzymes into cloth or filtration material.     

Tomographic reconstruction of treponemal cytoplasmic filaments reveals novel bridging and anchoring components

An understanding of the involvement of bacterial cytoplasmic filaments in cell division requires the elucidation of the structural organization of those filamentous structures. Treponemal cytoplasmic filaments are composed of one protein, CfpA, and have been demonstrated to be involved in cell division. In this study, we used electron tomography to show that the filaments are part of a complex with a novel molecular organization that includes at least two distinct features decorating the filaments. One set of components appears to anchor the filaments to the cytoplasmic membrane. The other set of components appears to bridge the cytoplasmic filaments on the cytoplasmic side, and to be involved in the interfilament spacing within the cell. The filaments occupy between 3 and 18% of the inner surface of the cytoplasmic membrane. These results reveal a novel filamentous molecular organization of independent filaments linked by bridges and continuously anchored to the membrane.     

The effective population size of Anopheles gambiae in Kenya: implications for population structure

We estimated current and long-term effective population size (Ne) of two Anopheles gambiae (savanna cytotype) populations in Kenya. Temporal variation at nine microsatellite loci in each population sampled 7 and 9 years apart and genetic diversity in each sample were analyzed to answer the following questions. (1) Do bottlenecks occur in Kenyan populations of A. gambiae? (2) How variable are different populations with respect to their current and long-term Ne values? (3) What are the implications of these results on population structure and history? The estimates of Ne of Asembo and Jego were 6,359 and 4,258, respectively, and the lower 95% limits were 2,455 and 1,669, respectively. Thus, despite the typical observation of low density at the village level during the dry season, large populations are maintained annually. Large current Ne is consistent with previous studies showing low differentiation across the continent, especially under Wright's isolation-by-distance model. Current Ne in Asembo was 1.5-fold higher than in Jego, but this difference was not significant. Long-term Ne in Asembo (22,667) was 2.9-fold higher than that in Jego (7,855) based on the stepwise mutation model. The difference between populations was significant at both time points regardless of whether long-term Ne values were calculated based on the stepwise mutation model or the infinite-alleles model. Heterozygosity in Jego declined significantly between 1987 (59%) and 1996 (54%), whereas heterozygosity in Asembo was stable (66%-65%). Despite the relatively high and significant differentiation between Asembo and Jego (FST = 0.072-0.10, RST = 0.037- 0.038), all alleles in Jego were found in Asembo but not vice versa. All of these findings suggest that lower Ne in Jego magnifies differentiation between the two populations. The long-term Ne was biased downward, because its calculation was based on an upper bound estimate of microsatellite mutation rate. Ne values based on mtDNA and allozymes were an order of magnitude higher. Long-term Ne therefore, is probably measured in hundreds of thousands and hence does not support a recent expansion of this species from a small population.      

Scanning electron microscopy of the conidia produced by the mycelial form of Paracoccidioides brasiliensis

The conidia produced by the mycelial form of Paracoccidioides brasiliensis were examined by scanning electron microscopy for the first time. Several different conidial types were characterized. These included intercalary arthroconidia, several types of septate conidia that are formed from other conidia, pedunculate conidia, and terminal hyphal conidia. In addition, the ultrastructure of the supporting pedestal of the pedunculate conidium was found to be separated from the mother conidium by a septum in some instances, and at other times it was not.     

Peripheral hyaline blebs (podosomes) of macrophages

The plasmalemma and hyaline ectoplasm together constitute the sensory and motor organ of macrophages. The purpose of this study was to isolate this cell fraction in order to analyze it biochemically and functionally. Brief sonification of warmed rabbit lung macrophages caused release of heterodisperse hyaline blebs and filopodia, which were easily collected by differential centrifugation. Viewed in the electron microscope, these structures consisted of membrane-bounded sacs principally containing actin filaments. Some contained secondary lysosomes. They were enriched threefold over whole cell homogenates in specific adenylate cyclase activity and in trichloroacetic-acid-precipitable (125)I when derived from cells labeled with 125(I) by means of a lactoperoxidase-catalyzed reaction. These markers were found to have identical isopycnic densitites when macrophage homogenates were subjected to sedimentation in a focusing sucrose density gradient system, and these markers had densities distinct from those of other cytoplasmic organelles. These markers were therefore assumed to be associated with macrophage plasma membranes. The specific β- glucuronidase activity of the bleb fraction was similar to that of homogenates, but the blebs had considerably lower specific succinic dehydrogenase activity and RNA content, and DNA was undetectable. Electrophoresis of blebs solubilized in sodium dodecyl sulfate on polyacrylamide gels revealed polypeptides co-migrating with macrophage actin-binding protein, myosin, and actin; blebs also had EDTA-activated adenosine triphosphatase activity characteristic of myosin. The concentrations of actin-binding protein and myosin were higher in blebs than in cells or cytoplasmic extracts, whereas actin concentrations were similar (relative to extracts) or only slightly greater (than in cells). Blebs and intact cells had high lactate dehydrogenase activities in the presence but not the absence of Triton X-100. Blebs and cells oxidased 1-[(14)C]glucose, and the rate of glucose oxidation was increased substantially in the presence of latex beads. We conclude that intact sacs of plasmalemma encasing contractile proteins and cytoplasmic enzymes can be isolated from macrophages. They are enriched in myosin and actin-binding protein, indicating that the contractile apparatus is regulated in the cell periphery. These structures have the capacity to respond to environmental signals. We suggest the name "podosomes" for them because of their resemblance to macrophage pseudopodia. We propose that podosome formation results from rapid dissolution of the cortical gel when the membrane is in an actively extended configuration.     

Ultrastructural Changes Associated with Germination and Outgrowth of an Appendage-Bearing Clostridial Spore

The sequence of events occurring during the germination and outgrowth of appendage-bearing spores of Clostridium bifermentans was studied by phase-contrast and electron microscopy. The mature spore was characterized ultrastructurally as having the normal spore components as well as long tubular appendages which orginated from the surface of the spore coat. Spores were incompletely enclosed by a distinctly laminated exosporium which possessed hairlike projections on its outermost layer. During germination, structural changes were observed in the core, core wall, cortex, and spore coat layers. Cortical material was extruded from the spore during outgrowth, which usually occurred from the pole opposite the appendages. The subunits comprising the structure of the appendages and the morphology of the mature appendages were observed. No discernible changes could be observed in the spore appendages during germination and outgrowth.