Quantitative contributions of bacteria and of Deinococcus geothermalis to deposits and slimes in paper industry

Deinococcus geothermalis has frequently been isolated from pink colored deposits of paper industry processes. Laboratory studies have shown that D. geothermalis is capable of forming on nonliving surfaces patchy biofilms that are resistant to adverse agents such as extreme pH, desiccation, solubilising detergents and biocides. This study was done to quantitatively assess the role of D. geothermalis as a biofouler in paper industry. Colored deposits were collected from 24 European and North American paper and board machines and the densities of the bacterial 16S rRNA genes and those of the red slime producers D. geothermalis and Meiothermus spp. were measured by QPCR (quantitative real time PCR). D. geothermalis was found at nine machines, usually from splash area deposits, but its contribution was minor, 0.001–1%, to the total bacterial burden of 8.3 to log 10.5 log units per gram wet-weight of the deposits. When D. geothermalis was found in a measurable quantity, Meiothermus spp. also was found, often in bulk quantity (7–100% of the total bacteria). The data are in line with the properties of D. geothermalis known from laboratory biofilm studies, indicating this species is a pioneer coloniser of machine surfaces and may help other bacteria to adhere and grown into biofilms, rather than competing with them.     

Microbe repelling coated stainless steel analysed by field emission scanning electron microscopy and physicochemical methods

Coating of stainless steel with diamond-like carbon or certain fluoropolymers reduced or almost eliminated adhesion and biofilm growth of Staphylococcus epidermidis, Deinococcus geothermalis, Meiothermus silvanus and Pseudoxanthomonas taiwanensis. These species are known to be pertinent biofilm formers on medical implants or in the wet-end of paper machines. Field emission scanning electron microscopic analysis showed that Staph. epidermidis, D. geothermalis and M. silvanus grew on stainless steel using thread-like organelles for adhesion and biofilm formation. The adhesion threads were fewer in number on fluoropolymer-coated steel than on plain steel and absent when the same strains were grown in liquid culture. Psx. taiwanensis adhered to the same surfaces by a mechanism involving cell ghosts on which the biofilm of live cells grew. Hydrophilic (diamond-like carbon) or hydrophobic (fluoropolymer) coatings reduced the adherence of the four test bacteria on different steels. Selected topographic parameters, including root-mean-square roughness (S (q)), skewness (S (sk)) and surface kurtosis (S (ku)), were analysed by atomic force microscopy. The surfaces that best repelled microbial adhesion of the tested bacteria had higher skewness values than those only slightly repelling. Water contact angle, measured (theta (m)) or roughness corrected (theta (y)), affected the tendency for biofilm growth in a different manner for the four test bacteria.     

培养条件对Shewanella oneidensis MR-1电极生物膜及细胞形貌的影响

【目的】Shewanella oneidensis MR-1是电活性模式微生物,但目前仍缺乏对其细胞及生物膜形貌变化的系统研究,本研究旨在完善对其形貌特征的理解,为支持其作为模式微生物提供有力的基础数据。【方法】选取培养基类型、缓冲液浓度、维生素、微量元素、无机盐、电子穿梭体、电子供体、电子受体等培养条件作为变量,采用恒电位培养法获得生物膜,通过扫描电子显微镜对生物膜形貌进行观察。【结果】低浓度缓冲液中(30 mmol/L和100 mmol/L),其细胞多为短杆状,高浓度缓冲液中(200 mmol/L和300 mmol/L)细胞卷曲伸长;缺乏维生素、微量元素、无机盐则可使生物膜紧贴电极生长,变得致密;而穿梭体和电子受体对于S. oneidensis MR-1极为关键,前者的存在可显著促进生物膜的厚度,后者的缺失可迫使生物膜细胞裂解;此外,通过形貌研究发现,S. oneidensis MR-1可首尾相连形成超过100μm的长线状结构。【结论】可通过改变缓冲液浓度、培养基类型、电子穿梭体和电子供受体等变量,实现Shewanella oneidensis MR-1电极生物膜及细胞形貌的调控。     

Deposition of glucuronoxylans on the secondary cell wall of Japanese beech as observed by immuno-scanning electron microscopy

Summary Glucuronoxylans (GXs), the main hemicellulosic component of hardwoods, are localized exclusively in the secondary wall of Japanese beech and gradually increase during the course of fiber differentiation. To reveal where GXs deposit within secondary wall and how they affect cell wall ultrastructure, immuno-scanning electron microscopy using anti-GXs antiserum was applied in this study. In fibers forming the outer layer of the secondary wall (S₁), cellulose fibrils were small in diameter and deposited sparsely on the inner surface of the cell wall. Fine fibrils with approximately 5 nm width aggregated and formed thick fibrils with 12 nm width. Some of these thick fibrils further aggregated to form bundles which labelled positively for GXs. In fibers forming the middle layer of the secondary wall (S₂), fibrils were thicker than those found in S₁ forming fibers and were densely deposited. The S₂ layer labelled intensely for GXs with no preferential distribution recognized. Compared with newly formed secondary walls, previously formed secondary walls were composed of thick and highly packed microfibrils. Labels against GXs were much more prevalent on mature secondary walls than on newly deposited secondary walls. This result implies that the deposition of GXs into the cell wall may occur continuously after cellulose microfibril deposition and may be responsible for the increase in diameter of the microfibrils.Abbreviations GXs glucuronoxylans - PBS phosphate-buffered saline - RFDE rapid-freeze and deep-etching technique - FE-SEM field emission scanning electron microscope - TEM transmission electron microscope     

Enzymatic synthesis of salicin glycosides through transglycosylation catalyzed by amylosucrases from Deinococcus geothermalis and Neisseria polysaccharea

Amylosucrase (ASase, EC 2.4.1.4) is a member of family 13 of the glycoside hydrolases that catalyze the synthesis of an α-(1→4)-linked glucan polymer from sucrose instead of an expensive activated sugar, such as ADP- or UDP-glucose. Transglycosylation reactions mediated by the ASases of Deinococcus geothermalis (DGAS) and Neisseria polysaccharea (NPAS) were applied to the synthesis of salicin glycosides with sucrose serving as the glucopyranosyl donor and salicin as the acceptor molecule. Two salicin glycoside transfer products were detected by TLC and HPLC analyses. The synthesis of salicin glycosides was very efficient with NPAS with a yield of over 90%. In contrast, DGAS specifically synthesized only one salicin transglycosylation product. The transglycosylation products were identified as α-d-glucopyranosyl-(1→4)-salicin (glucosyl salicin) and α-d-glucopyranosyl-(1→4)-α-d-glucopyranosyl-(1→4)-salicin (maltosyl salicin) by NMR analysis. The ratio between donor and acceptor had a significant effect on the type of product that resulted from the transglycosylation reaction. With more acceptors present in the reaction, more glucosyl salicin and less maltosyl salicin were synthesized.     

Characterization of the Mn2+-stimulated (di)adenosine polyphosphate hydrolase encoded by the Deinococcus radiodurans DR2356 nudix gene

The DR2356 nudix hydrolase gene from Deinococcus radiodurans has been cloned and the product expressed as an 18 kDa histidine-tagged protein. The enzyme hydrolysed adenosine and diadenosine polyphosphates, always generating ATP as one of the initial products. ATP and other (deoxy)nucleoside triphosphates were also substrates, yielding (d)NDP and Pi as products. The DR2356 protein was most active at pH 8.6–9.0 and showed a strong preference for Mn²⁺ as activating cation. Mg²⁺ ions at 15 mM supported only 5% of the activity achieved with 2 mM Mn²⁺. K _m and k _cat values for diadenosine tetra-, penta- and hexaphosphates were 2.0, 2.4 and 1.1 μM and 11.4, 28.6 and 12.0 s⁻¹, respectively, while for GTP they were 20.3 μM and 1.8 s⁻¹, respectively. The K _m for adenosine 5′-pentaphosphate was <1 μM. Expression analysis showed the DR2356 gene to be induced eight- to ninefold in stationary phase and in cells subjected to slow dehydration plus rehydration. Superoxide (but not peroxide) treatment and rapid dehydration caused a two-to threefold induction. The Mn-requirement and induction in stationary phase suggest that DR2356 may have a specific role in maintenance mode metabolism in stationary phase as Mn²⁺ accumulates.     

The flexibility of UV-inducible mutation in Deinococcus ficus as evidenced by the existence of the imuB-dnaE2 gene cassette and generation of superior feather degrading bacteria

The lexA–imuB–dnaE2 gene cassette contributing to the TLS (translesion synthesis) polymerase activity and can easily cause mutation after DNA damage in many bacteria. But it was previously thought that TLS polymerase activity was unlikely to exist in the radio-resistant genus Deinococcus. In our preliminary studies, the lexA–imuB–dnaE2 gene cassette was found in a newly isolated feather-degrading Deinococcus ficus. Here we have attempted to determine the imuB gene sequence from another Deinococcus species namely D. grandis, by using the newly designed primers. The destroying of either imuB or dnaE2 gene in D. ficus leads to the increase in UV sensitivity and decrease in UV-induced mutations, which demonstrated the existence of TLS polymerase activity in D. ficus. In the presence of lexA–imuB–dnaE2, it is possible to obtain mutants with various keratinolytic activities after UV exposure. The keratinolytic activity of mutant strain CC-ZG207 increased by approximately twofold during growth in liquid feather medium. In contrast, the mutant strain CC-ZG227 showed only half of the keratinolytic activity compared with the wild type strain. By utilizing SDS–PAGE and zymogram profile analysis, the change in the protease activity was observed. We have proposed that the superior mutants of D. ficus can be created under UV stress, which is mediated by the lexA–imuB–dnaE2 gene cassette.     

国产18种鳞盖蕨属植物叶表皮微形态特征及其系统学意义

鳞盖蕨属是一个自然类群,由于各类群之间的形态差异以及种内变异比较大,因此其分类一直存在争议。该研究利用光学显微镜(LM)和扫描电子显微镜(SEM)对鳞盖蕨属18种植物(含3变种)的叶表皮微形态进行了观察和比较,并结合形态学、孢粉学和分子系统学探讨其系统学意义。结果表明:在光学显微镜下,18种鳞盖蕨属植物叶上、下表皮的脉上均有毛,叶表皮细胞都为不规则形,垂周壁为深波状或波状; 气孔都分布在下表皮,气孔器类型主要有极细胞型和腋下细胞型两种,叶表皮微形态特征支持鳞盖蕨属是碗蕨科中的一个单系类群; 叶上、下表皮脉间毛的变异特征支持光叶鳞盖蕨、毛叶边缘鳞盖蕨和二回边缘鳞盖蕨处理为边缘鳞盖蕨变种; 在扫描电镜下,叶上表皮角质层多为脊状凸起,多数还具丝状纹饰,角质膜特征与叶片回数有一定的相关性; 共环极细胞型、聚腋下细胞型、不等细胞型和不规则四细胞型只在少数种出现,气孔外拱盖多数凹陷,光滑或有颗粒; 气孔外拱盖内缘平滑、浅波状或齿状; 保卫细胞两极大多数有“T”型加厚,不同种间叶表皮微形态特征表现出一定差异。该研究结果为其分类、演化、系统位置等进一步研究提供了参考。     

吉林省豚草瘦果形态及微结构特征

【目的】研究吉林省不同地区豚草瘦果形态及微结构特征,为豚草的识别及防控提供参考。【方法】采集吉林省吉林市磨盘山、吉林市龙潭山、桦甸市、蛟河市、长春市、四平市、公主岭市、通化市、白山市、白城市、松原市和乾安县12个地区不同豚草种群的瘦果进行扫描电镜观察、描述和拍照,分析豚草瘦果表型性状与地理-气候因子的相关性,并对瘦果形态及微结构特征进行聚类分析。【结果】豚草果皮纹饰有条状纹饰、网状纹饰和穴状纹饰;瘦果的喙长度随年均降水量增加而增加、随纬度升高而减少;果形系数随经度升高及年均降水量的增加而增加。根据形态和微结构性状聚类分析结果,将豚草瘦果分为3类（Ⅰ、Ⅱ、Ⅲ）。其中,Ⅰ类瘦果果皮纹饰网状、中果形、短喙类,种源主要来自蛟河市、通化市、公主岭市、松原市和乾安县;Ⅱ类瘦果果皮纹饰条状、小果形、中长喙类,种源主要来自吉林市磨盘山、吉林市龙潭山、桦甸市、长春市和四平市;Ⅲ类瘦果果皮纹饰穴状、大果形、长喙类,种源主要来自白山市和白城市。【结论】吉林省豚草瘦果形态及微结构存在差异,低纬度、高降水量有利于豚草瘦果生长发育。     

Localization of aggregation substances of Enterococcus faecalis after induction by sex pheromones

The distribution of sex pheromone induced aggregation substance was studied on the cell surface of various Enterococcus faecalis strains. In the accompanying paper we have shown that the aggregation substance appears as a layer of hairlike structures. Using direct and indirect immunogold technique, transmission electron microscopy and high resolution scanning electron microscopy we investigated the appearance and distribution of the aggregation substance. The hairs increase in number with increasing exposure to sex pheromones (maximum density: 1300/m²). We show that these structures are unequally distributed over the cell surface, even if the cells were induced by sex pheromones for a long period of time. Statistical analysis of the unequal distribution indicates that aggregation substance is incorporated into pre-existing old cell-walls and that this incorporation shows a saturation ca. 40 min after addition of sex pheromones.Abbreviations cAD1 sex pheromone specific for plasmid pAD1 - cPD1 sex pheromone specific for plasmid pPD1 - FESEM field emission scanning electron microscope - pAD1 conjugative plasmid specifically transferred in the presence of cAD1 - pPD1 conjugative plasmid specifically transferred in the presence of cPD1 - TEM transmission electron microscope     

香薷叶表皮腺毛及其分泌黄酮类物质的组织化学研究

利用光学显微技术和扫描电镜技术,研究了香薷营养生长期和生殖生长期的枝条上部、中部、下部叶片的腺毛种类、分布及数量;采用组织化学染色和荧光显微技术,观察香薷腺毛分泌黄酮类物质的过程,探讨香薷叶表皮腺毛的泌香机理,为生产中确定提取香薷黄酮类物质的最佳取材时期提供依据。结果表明:(1)香薷叶表皮有头状腺毛和盾状腺毛,成熟的头状腺毛包括基细胞、柄细胞和头部,头部较小,呈半圆球型,直径为(20±2)μm;发育近成熟的盾状腺毛头部较大,呈盾状,直径为(60±5)μm。(2)香薷叶片远轴面和近轴面均有腺毛的分布,远轴面分布较多,近轴面分布较少;盾状腺毛主要分布在叶的远轴面,头状腺毛在叶的两面均有分布;腺毛密度随节位的降低而减少,节位相同时,营养生长期叶片上的腺毛密度(283.9个/mm2)高于生殖生长期(194.4个/mm2)。(3)香薷头状腺毛和盾状腺毛均能分泌黄酮类物质,且随着腺毛的发育成熟,黄酮类物质逐渐积累于腺毛的头部。     

Protection of catheter surfaces from adhesion of Pseudomonas aeruginosa by a combination of silver ions and lectins

A catheter surface was modified by coating a cellulose acetate polymer. Adhesion of Pseudomonas aeruginosa ATCC 27853 to the surface was investigated by exposing bacterial cultures to three treatments: polymer impregnated with silver ions (Ag⁺), polymer surfaces coated with lectins and a combination of Ag⁺ and a lectin coating. The effective concentration of Ag⁺ providing protection against bacterial biofilm development was 100g/ml and higher. Lectins alone at 10% also showed inhibition of bacterial attachment. However, the best result was achieved against bacterial adhesion and growth on surfaces using a combination of 100 g Ag⁺/ml and a lectin coating as a surface treatment. This surface treatment was also effective against both fresh culture and a two-week-old culture containing P. aeruginosa producing exopolymers. Our results suggest that Ag⁺impregnation combined with a lectin coating warrants further investigation as a potential means of protecting catheters.     

小瓜虫在澳洲龙纹斑鳃器官上的分布及其影响

【目的】小瓜虫病是澳洲龙纹斑苗种阶段危害巨大的寄生虫病。探究小瓜虫在澳洲龙纹斑鳃器官上的分布及其影响可以丰富小瓜虫的致病性及病理学方面的研究内容,也可以为渔业生产中小瓜虫病的防治提供参考。【方法】采用光镜及扫描电镜技术确定病原,并观察小瓜虫在澳洲龙纹斑鳃丝、鳃小片及鳃盖上的分布情况及这些器官的变化情况。【结果】小瓜虫侵染澳洲龙纹斑的鳃器官后,分布在鳃丝、鳃小片及鳃盖的表面,上皮细胞之下及鳃小片之间,或是包裹在黏液细胞里,但侵染后期在鳃丝及鳃小片上的数量明显少于鳃盖。侵染后期,鳃丝、鳃小片及鳃盖出现一定程度的膨胀变形,黏液细胞分泌增多,鳃小片末端膨大变形甚至黏连融合,顶端充血呈球状或棒状。【结论】对于鳃部而言,侵染后期小瓜虫主要分布在澳洲龙纹斑的鳃盖上。小瓜虫主要通过侵染引发澳洲龙纹斑鳃部器官的变形膨大,使其丧失正常的功能,造成血液循环受阻、渗透压调节失衡,最终导致鱼体缺氧死亡。     

10种金花茶组植物的花粉形态及分类学意义

金花茶组(Sect.Chrysantha)植物具有很高的观赏价值,但其分类存在争议.为探讨金花茶组植物花粉的形态特征和分类学意义,该研究用扫描电镜观察了10种金花茶的花粉形状、外壁纹饰以及穿孔特征,统计了极轴长、赤道轴长和萌发沟长,并进行聚类分析.结果表明:(1)10种供试金花茶植物花粉为中粒花粉,大部分花粉形状呈近球...     

Photocatalytic antimicrobial activity of thin surface films of TiO(2), CuO and TiO (2)/CuO dual layers on Escherichia coli and bacteriophage T4

TiO₂-coated surfaces are increasingly studied for their ability to inactivate microorganisms. The activity of glass coated with thin films of TiO₂, CuO and hybrid CuO/TiO₂ prepared by atmospheric Chemical Vapour Deposition (Ap-CVD) and TiO₂ prepared by a sol–gel process was investigated using the inactivation of bacteriophage T4 as a model for inactivation of viruses. The chemical oxidising activity was also determined by measuring stearic acid oxidation. The results showed that the rate of inactivation of bacteriophage T4 increased with increasing chemical oxidising activity with the maximum rate obtained on highly active sol–gel preparations. However, these were delicate and easily damaged unlike the Ap-CVD coatings. Inactivation rates were highest on CuO and CuO/TiO₂ which had the lowest chemical oxidising activities. The inactivation of T4 was higher than that of Escherichia coli on low activity surfaces. The combination of photocatalysis and toxicity of copper acted synergistically to inactivate bacteriophage T4 and retained some self-cleaning activity. The presence of phosphate ions slowed inactivation but NaCl had no effect. The results show that TiO₂/CuO coated surfaces are highly antiviral and may have applications in the food and healthcare industries.     

Action of antimicrobial substances produced by different oil reservoir Bacillus strains against biofilm formation

Microbial colonization of petroleum industry systems takes place through the formation of biofilms, and can result in biodeterioration of the metal surfaces. In a previous study, two oil reservoir Bacillus strains (Bacillus licheniformis T6-5 and Bacillus firmus H₂O-1) were shown to produce antimicrobial substances (AMS) active against different Bacillus strains and a consortium of sulfate-reducing bacteria (SRB) on solid medium. However, neither their ability to form biofilms nor the effect of the AMS on biofilm formation was adequately addressed. Therefore, here, we report that three Bacillus strains (Bacillus pumilus LF4—used as an indicator strain, B. licheniformis T6-5, and B. firmus H₂O-1), and an oil reservoir SRB consortium (T6lab) were grown as biofilms on glass surfaces. The AMS produced by strains T6-5 and H₂O-1 prevented the formation of B. pumilus LF4 biofilm and also eliminated pre-established LF4 biofilm. In addition, the presence of AMS produced by H₂O-1 reduced the viability and attachment of the SRB consortium biofilm by an order of magnitude. Our results suggest that the AMS produced by Bacillus strains T6-5 and H₂O-1 may have a potential for pipeline-cleaning technologies to inhibit biofilm formation and consequently reduce biocorrosion.     

Ultrastructure of the tentacles of the apodous holothurian Leptosynapta spp (Holothurioidea: Echinodermata) with special reference to the epidermis and surface coats

Summary The tentacles of the apodous holothurian Genus Leptosynapta have been studied by use of transmission and scanning electron microscopy. The gross anatomy, water vascular system, fibre systems and ectoneural nerve ring are described. A fuzzy coat of attenuated filaments covers the surface of the tentacle, broken only by secretory ducts. A cuticle underlies the fuzzy coat. Bacteria are common in the subcuticular space. Fixation without osmium gives poor preservation of the surface coats. The epidermis consists of a single layer of columnar cells consisting of Type-1, Type-2, support, goblet and uniciliated cells. Type-1 cells secrete electron-dense material and appear to be homologous to adhesive cells of the tentacles of other holothurians. The support cells contain large, granular vesicles not found in other holothurians. Goblet cells contain flocculent mucus and have an apical cilium. Goblet cells are not found in other holothurian tentacles and may function to lubricate and wrap adhering particles to aid their ingestion. The uniciliated cells are rare, poorly developed and the cilium does not extend past the cuticle. The ultrastructure of the tentacles is discussed in relation to those of other holothurians.     

Diurnal differences in the supply of glucomannans and xylans to innermost surface of cell walls at various developmental stages from cambium to mature xylem in Cryptomeria japonica

Summary. We investigated the diurnal differences in the innermost surface of tracheid cell walls at various developmental stages from cambium to mature xylem. Cryptomeria japonica saplings were cultivated in a growth chamber with a light cycle set at 14 h of light and 10 h of darkness. Samples were collected from the saplings during both the light and dark periods. The innermost surface of cell walls was immunogold-labeled with anti-glucomannan or anti-xylan antiserum and was observed by field emission scanning electron microscopy. Diurnal differences in the aspect of the innermost surface of cell walls were seen only in S₂-layer-forming tracheids; cellulose microfibrils were clearly evident during the light period, and amorphous material containing glucomannans and xylans was prevalent during the dark period. Cellulose microfibrils were present at the primary-wall formation and S₁-layer-forming stages, and many warts were observed in the mature tracheids, regardless of the time of sampling. The densities of labeled glucomannans on the innermost surface of cell walls in S₁- and S₂-forming tracheids and of labeled xylans in S₂-forming tracheids during the dark period were significantly higher than those during the light period. These results suggest a diurnal periodicity in the supply of cell wall matrix containing hemicellulose to the innermost surface of developing secondary walls. Correspondence and reprints: Laboratory of Bio-material Physics, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan. Present address: Chair of Climate Change Science for Forestry and Water Resources, Graduate School of Science and Technology, Niigata University, Niigata, Japan.     

Integrated biosorption and photocatalytic oxidation treatment of di(2-ethylhexyl)phthalate

Di(2-ethylhexyl)phthalate (DEHP), a toxic phthalate ester, is a ubiquitous contaminant due to its extensive use and persistence. No available treatment method can cost-effectively remove it from industrial wastewater. In a previous study, DEHP was effectively removed from aqueous solution by adsorption onto the biomass of selected seaweed, i.e., beached seaweed and Sargassum siliquastrum. Since biosorption cannot detoxify DEHP, the degradation (and detoxification) of desorbed DEHP from seaweed biomass by photocatalytic oxidation (PCO) was attempted. The first part of this study was to optimize the conditions for the degradation of desorbed DEHP in aqueous solution by PCO. Under optimized conditions, a total degradation of 20 mg/L of DEHP was achieved within 45 min. The degradation intermediates/products such as phthalic anhydride and 2-ethyhexanol were identified by GC-MS analysis. Total organic carbon analysis was also used to ensure the complete mineralization of DEHP. The Microtox^® test was used to assess the toxicities of the parent and degraded compounds. In the second part of this study, DEHP was first removed and concentrated by adsorption onto seaweed biomass under the conditions optimized in the previous study. It was then desorbed from seaweed biomass and degraded by PCO. Results indicate that the treatment for DEHP by integrating biosorption and PCO is feasible.