植物相关细菌生物膜研究进展

介绍了不同类型的植物相关细菌生物膜。综述了有关生物膜结构、形成和微生物与陆生植物结合所形成生物膜的特性等方面的研究进展。阐述了植物表面环境、细菌在植物表面主动和被动的沉积作用等对生物膜形成的影响。     

Salmonella SipA polymerizes actin by stapling filaments with nonglobular protein arms

Like many bacterial pathogens, Salmonella spp. use a type III secretion system to inject virulence proteins into host cells. The Salmonella invasion protein A (SipA) binds host actin, enhances its polymerization near adherent extracellular bacteria, and contributes to cytoskeletal rearrangements that internalize the pathogen. By combining x-ray crystallography of SipA with electron microscopy and image analysis of SipA-actin filaments, we show that SipA functions as a "molecular staple," in which a globular domain and two nonglobular "arms" mechanically stabilize the filament by tethering actin subunits in opposing strands. Deletion analysis of the tethering arms provides strong support for this model.     

Intracellular bacterial biofilm-like pods in urinary tract infections

Escherichia coli entry into the bladder is met with potent innate defenses, including neutrophil influx and epithelial exfoliation. Bacterial subversion of innate responses involves invasion into bladder superficial cells. We discovered that the intracellular bacteria matured into biofilms, creating pod-like bulges on the bladder surface. Pods contained bacteria encased in a polysaccharide-rich matrix surrounded by a protective shell of uroplakin. Within the biofilm, bacterial structures interacted extensively with the surrounding matrix, and biofilm associated factors had regional variation in expression. The discovery of intracellular biofilm-like pods explains how bladder infections can persist in the face of robust host defenses.     

奶牛乳腺炎葡萄球菌生物被膜形成及相关基因分析

 【目的】研究临床分离的奶牛乳腺炎葡萄球菌生物被膜形成能力,相关基因的分布及二者之间的关系;【方法】利用通用硅胶片法培养葡萄球菌生物被膜,经充分洗涤后对生物被膜进行银染定性判断菌株形成生物被膜的能力,通过生物被膜菌落结晶紫染色法定量检测菌株生物被膜的形成能力,并用扫描电镜观察被膜结构,对葡萄球菌bap、icaAD、icaBC、sar、agr、sigB、clfaA、clfaB、fnbpA和fnbpB进行PCR扩增检测。【结果】 银染法定性结果显示137株葡萄球菌中有120株形成肉眼可见的生物被膜,生物被膜形成率为87.6%;结晶紫染色定量检测与硅胶粘附的有132株,不粘附的有5株。试验所用的137株葡萄球菌中,57株能扩增出bap基因;有43株和54株分别能扩增出icaAD和icaBC;有73株、49株和38株分别扩增出sigB、sar和agr;分别有76株和50株染色体中存在clfaA和clfaB;fnbpA和fnbpB分别在52株和26株菌株中扩增出。【结论】推测bap、sigB、sar、icaAD和icaBC基因是生物被膜形成的重要相关基因;agr及粘附素基因clfaA、clfaB、fnbpA和fnbpB对生物被膜形成的作用不明显。     

溶藻弧菌生物膜研究进展

溶藻弧菌是一种海洋和海洋生物中普遍存在的致病菌,能够感染人类和多种水产品.细菌粘附在介质表面,分泌胞外基质将自身包裹其中从而形成生物膜,具有极高的抗药性和免疫逃逸能力,人类许多细菌感染均与生物膜有关.就溶藻弧菌生物膜进行综述,为相关研究提供理论参考.     

ISOLATION OF TRICHOMONAS VAGINALIS WITH PENICILLIN

Exposure of Trichomonas vaginalis in vaginal discharge for 60 hours to 5,000-10,000 units of penicillin in 10.0 ml of a medium containing cysteine (0.15 per cent.), peptone, liver infusion, maltose and human serum was adequate to destroy the associated bacteria. Seven strains were isolated from seven women without a failure. This technic widens the field of investigation by offering a means of obtaining additional strains of bacteria-free Trichomonas vaginalis for comparative study. Efforts to isolate Trichomonas vaginalis by such laborious methods as washing, micropipetting and migration have all failed to yield bacteria-free cultures at this and other laboratories. The adherence of bacteria to the trichomonads, which appear to have sticky surfaces, and the relatively slow speed with which these protozoa swim are probably in large measure responsible for failure. Further trials by these methods now seem unnecessary. In the cases reported above the success of the penicillin method may be attributed to several factors. Resistance of Trichomonas vaginalis to penicillin in the concentrations reported above, the absence from the specimens of vaginal discharge of bacteria resistant to penicillin in these same concentrations, and the destruction of bacteria adherent to the sticky surfaces of the trichomonads perhaps played a large part in assuring successful isolation of the protozoa.     

Bacteria use type IV pili to walk upright and detach from surfaces

Bacterial biofilms are structured multicellular communities involved in a broad range of infections. Knowing how free-swimming bacteria adapt their motility mechanisms near surfaces is crucial for understanding the transition between planktonic and biofilm phenotypes. By translating microscopy movies into searchable databases of bacterial behavior, we identified fundamental type IV pili-driven mechanisms for Pseudomonas aeruginosa surface motility involved in distinct foraging strategies. Bacteria stood upright and "walked" with trajectories optimized for two-dimensional surface exploration. Vertical orientation facilitated surface detachment and could influence biofilm morphology.     

废水处理系统生物膜中细菌的共凝集研究

从不同废水处理系统生物膜中分离得到22株细菌,测定它们的共凝集能力,并通过16SrRNA序列分析对未知细菌进行鉴定。共凝集结果显示,2h和20h时Bacillus cereus G5与其他21株细菌的共凝集率分别达到30%和45%以上,Bacillus megaterium T1与其他17株细菌的共凝集率分别达到34%和45%以上;同属于Shewanella菌属的H2和H3菌株与其它16株细菌配对的共凝集率在2h时达到30%～75%,20h时达到40%～82%;G5和T1与其他21株细菌共培养的生物膜形成能力进一步表明G5和T1的参与增加了单一细菌的生物膜形成量。研究结果表明,上述3种细菌具有将多种细菌固定于生物膜的功能。     

大连海区潮间带海藻附生细菌的抗真菌活性

以分离自石莼、浒苔的16株附生真菌作指示菌，采用双层琼脂扩散法，对从石莼Ulva lactuca、孔石莼Ulvap ertusa等10种海藻表面分离的79株细菌进行拮抗试验，获得有拮抗活性的菌株47株，占总测试菌株的59．5％。不同海藻表面拮抗细菌的分布比例不同。研究结果表明，海藻表面的附生细菌可能帮助宿主阻止真菌等表面污损生物的附着。     

响应面优化大肠杆菌生物膜的培养条件

利用响应面法对大肠杆菌生物膜的培养条件进行优化。在单因素试验基础上,以细菌浓度、pH和培养时间为自变量,生物膜形成量为响应值,根据中心组合(Box-Benhnken)试验设计原理,研究各自变量及其交互作用对生物膜形成的影响,依据回归分析确定各培养条件的最优值,并通过激光共聚焦显微镜观察大肠杆菌生物膜的形成。结果表明,大肠杆菌生物膜的最佳培养条件为:LB培养基pH7.5,细菌接种量为1.2×10~7 cfu/mL,培养时间11.5 h。在该优化条件下,大肠杆菌生物膜的形成量OD_(600nm)可达0.60左右;通过激光共聚焦显微镜观察可见,此时形成大量的细菌聚集体,80%的视野已经被细菌覆盖,细菌活性很高,生物膜形成量达到高峰。     

Visualization and functional analysis of RNA-dependent RNA polymerase lattices

Positive-strand RNA viruses such as poliovirus replicate their genomes on intracellular membranes of their eukaryotic hosts. Electron microscopy has revealed that purified poliovirus RNA-dependent RNA polymerase forms planar and tubular oligomeric arrays. The structural integrity of these arrays correlates with cooperative RNA binding and RNA elongation and is sensitive to mutations that disrupt intermolecular contacts predicted by the polymerase structure. Membranous vesicles isolated from poliovirus-infected cells contain structures consistent with the presence of two-dimensional polymerase arrays on their surfaces during infection. Therefore, host cytoplasmic membranes may function as physical foundations for two-dimensional polymerase arrays, conferring the advantages of surface catalysis to viral RNA replication.     

CrxV对水稻白叶枯病菌致病力的调控

为分析GGDEF和EAL结构域的蛋白CrxV对水稻黄单胞菌（Xanthomonas oryzaepv.oryzae,Xoo）致病性及相关致病因子的影响,通过同源重组法构建了crxV的突变体及其互补菌株;采用剪叶法比较了突变体、野生型和互补菌株对水稻叶片的侵染能力;分析了这3个菌株产生相关致病因子（运动性、胞外多糖、生物被膜和胞外酶）的能力。结果显示,crxV突变体的致病力比野生型和互补菌株的低。crxV突变导致病原菌生物被膜含量显著升高,但对运动性、胞外多糖和胞外酶均无明显影响。结果表明,CrxV可能通过负调控生物被膜影响Xoo的致病性。     

Hemocyte-mediated shell mineralization in the eastern oyster

The growth of molluscan shell crystals is usually thought to be initiated from solution by extracellular organic matrix. We report a class of granulocytic hemocytes that may be directly involved in shell crystal production for oysters. On the basis of scanning electron microscopy (SEM) and x-ray microanalysis, these granulocytes contain calcium carbonate crystals, and they increase in abundance relative to other hemocytes following experimentally induced shell regeneration. Hemocytes are observed at the mineralization front using vital fluorescent staining and SEM. Some cells are observed releasing crystals that are subsequently remodeled, thereby at least augmenting matrix-mediated crystal-forming processes in this system.     

Molecular architecture and assembly principles of Vibrio cholerae biofilms

In their natural environment, microbes organize into communities held together by an extracellular matrix composed of polysaccharides and proteins. We developed an in vivo labeling strategy to allow the extracellular matrix of developing biofilms to be visualized with conventional and superresolution light microscopy. Vibrio cholerae biofilms displayed three distinct levels of spatial organization: cells, clusters of cells, and collections of clusters. Multiresolution imaging of living V. cholerae biofilms revealed the complementary architectural roles of the four essential matrix constituents: RbmA provided cell-cell adhesion; Bap1 allowed the developing biofilm to adhere to surfaces; and heterogeneous mixtures of Vibrio polysaccharide, RbmC, and Bap1 formed dynamic, flexible, and ordered envelopes that encased the cell clusters.     

Mussel growth supported by methane as sole carbon and energy source

Symbioses between chemoautotrophic bacteria and several specialized marine invertebrates are well documented. However, none of these symbioses have been demonstrated to provide sufficient energy and carbon to the host to enable it to grow. Growth rates of seep mussels collected from hydrocarbon seeps off the coast of Louisiana were measured in a controlled environment where methane was the sole carbon and energy source. The growth rates increased to a maximum of 17.2 micrometers per day in response to methane and approached zero in the absence of methane. These mussels contain methanotrophic symbiotic bacteria in their gills, which suggests that these bacteria provide their hosts with a net carbon flux originating from methane.     

A molecular clutch disables flagella in the Bacillus subtilis biofilm

Biofilms are multicellular aggregates of sessile bacteria encased by an extracellular matrix and are important medically as a source of drug-resistant microbes. In Bacillus subtilis, we found that an operon required for biofilm matrix biosynthesis also encoded an inhibitor of motility, EpsE. EpsE arrested flagellar rotation in a manner similar to that of a clutch, by disengaging motor force-generating elements in cells embedded in the biofilm matrix. The clutch is a simple, rapid, and potentially reversible form of motility control.     

微生物被膜形成因子及其对厚壳贻贝附着的影响

为调查环境因子对微生物被膜形成能力的影响以及在此条件下形成的微生物被膜对厚壳贻贝Mytilus coruscus稚贝(壳长为1.81 mm±0.13 mm)附着的影响,通过显微共聚焦技术和荧光染色技术,探讨了在18℃条件下盐度为10、20、30时和盐度为30的条件下温度为8、18、28℃时,环境因子对希瓦氏细菌Shewanella loihica ECSMB14101形成微生物被膜产生的影响,调查了上述环境下形成微生物被膜的细菌密度、分布、膜厚等生物学特性及其对厚壳贻贝稚贝附着的影响。结果表明:S.loihica ECSMB14101形成微生物被膜的最终密度随盐度的升高而增加;不同密度的微生物被膜诱导活性不同,且与盐度呈线性关系;微生物被膜上细菌密度随温度的升高而增多,在8℃和18℃时对稚贝的附着诱导活性有显著性差异(P0.05),附着率分别为31%和68%,18℃与28℃时的微生物被膜诱导活性无显著性差异(P0.05);随盐度和温度的上升,微生物被膜上的细菌分布逐渐密集,膜厚显著增加,且在盐度为30、温度为28℃条件下微生物被膜的膜厚达到最大值(4.4μm±0.1μm);在温度为18℃、盐度为30条件下,膜厚的最大值为(4.3±0.1)μm。研究表明,盐度和温度改变微生物被膜的生物学特性,并导致贻贝附着率的显著差异。     

小麦条锈菌与寄主交界面的超微结构和细胞化学

用电镜技术和细胞化学方法研究结果发现,小麦条锈菌吸器颈部和吸器体壁均由两层构成,即染色较深的外层和染色较浅的内层,但是,具邻位羟基多糖物质的分布模式和糖基的种类在颈部壁和吸器体壁并非一致。在颈部处凹陷的寄主细胞质膜紧贴于其外壁,而在吸器体部位,它与吸器体始终保持一定距离。吸器外间质中存在有多糖物质,α-葡萄糖或α-甘露糖,乙酸半乳糖胺,半乳糖等糖基。此外,在吸器外围的寄主原生质中存在有呈管状的复合体。     

果壳种类对生物膜特性及废水处理效果的影响

为促进植物果壳的资源化利用,选择花生壳、核桃壳和夏威夷果壳3种果壳,使用SEM和FTIR表征其表面微观特征与化学基团,探索植物果壳作为SBBR反应器填料的可行性,考察果壳类型对生物膜特性和废水处理效果的影响。结果表明：3种果壳表面粗糙多孔且富有OH、COOH等亲水性基团,有利于微生物附着生长。花生壳上生物膜量呈现“先增后降”变化,试验前期从6.80 mg·cm^-3上升至24.66 mg·cm^-3。后期花生壳软化分解,生物膜量不断降低,与试验前期基本持平,生物膜中夹杂过多腐烂的代谢产物,导致EPS含量最低（49.90 mg·g^-1）,DHA低,生物膜活性差,对COD、NH₄⁺-N长期去除效果不理想,仅能达到约65%;核桃壳、夏威夷果壳上生物膜量含量高,试验前期分别从7.98 mg·cm^-3和8.45 mg·cm^-3快速上升至26.75 mg·cm^-3和25.96 mg·cm^-3,之后缓慢上升。悬浮于反应器中的核桃壳受到强烈水力剪切作用,污染物传质效果的增强促进微生物EPS分泌,EPS含量最高达66.44 mg·g^-1;2种果壳生物膜DHA均呈现“先增后稳”变化,最高达到81.72 mg·g^-1·h^-1,生物膜活性高,对COD、NH₄⁺-N去除率不断增长,最高均达到90%左右。因此,除花生壳外,核桃壳和夏威夷果壳结构稳定,生物膜特性良好、污染物去除效果较好,可作为填料长期使用。     

生物膜磁场流化床应用于硫酸盐废水脱硫处理研究

将磁场稳态流化床（MSFB）技术应用于高硫酸盐废水的处理。具体做法是,将驯化后的硫酸盐还原菌（SRB）菌群接入已填充有磁性聚苯乙烯多孔微球（MPB）的厌氧磁性流化床（anMSFB）,采用载体浸泡挂膜法进行启动运行,然后通入模拟废水进入稳定负荷运行。通过显微、扫描电镜等方法研究了系统anMSFB的生物膜组成及生物膜量;考察了pH、温度对SO42-去除率的影响;确定了anMSFB反应器中SO42-的降解动力学方程;理论推导出在特定进液废水浓度和脱硫率下anMSFB反应器中MPB颗粒的填充高度。该研究为固定化生物膜磁场流化床反应系统应用于含硫废水处理奠定了坚实的基础。