bluuebunny/crossref_metadata_2025 huggingface dataset:
Hepatic stimulator substance mitigates hepatic cell injury through suppression of the mitochondrial permeability transition
Authors: Wu Yuan , Zhang Jing , Dong Lingyue , Li Wen , Jia Jidong , An Wei
Published: February 2010

Hepatic stimulator substance (HSS) has been shown to protect liver cells from various toxins. However, the mechanism by which HSS protects hepatocytes remains unclear. In this study, we established BEL‐7402 cells that stably express HSS and analyzed the protective ability of HSS on cells through mitochondrial permeability (MP). After administration of carbonyl cyanidem‐chlorophenylhydrazone (CCCP), a specific agent that leads to depolarization of the mitochondrial transmembrane potential, the apoptosis rate of HSS‐expressing cells was significantly reduced, as measured using Hoechst staining and flow cytometry. The mitochondrial membrane transition and cytochrome cleakage were significantly inhibited in the HSS‐expressing cells as compared with the untransfected cells, and, as a consequence, the cellular ATP content in the HSS‐expressing cells was relatively preserved. Additionally, decreased caspase‐3 activity was observed in the HSS‐expressing cells treated with CCCP as compared with the vector‐transfected cells and cells expressing mutant HSS. Furthermore, silencing of HSS expression using small interfering RNA accelerated CCCP‐induced apoptosis. In isolated mitochondria, recombinant HSS reduced the release of cytochrome cinduced by CCCP, indicating a possible role for HSS in regulation of mitochondrial permeability transition (MPT). HSS‐expressing BEL‐7402 cells are resistant to CCCP injury, and HSS protection is identical to that observed with cyclosporin A, an inhibitor of MPT. Therefore, we propose that the protective effect of HSS may be associated with blockade of MPT.
The plasminogen activator inhibitor 2 transcript is destabilized via a multi‐component 3′ UTR localized adenylate and uridylate‐rich instability element in an analogous manner to cytokines and oncogenes
Authors: Stasinopoulos Stan , Mariasegaram Mythily , Gafforini Chris , Nagamine Yoshikuni , Medcalf Robert L.
Published: February 2010

Plasminogen activator inhibitor type 2 (PAI‐2; SERPINB2) is a highly‐regulated gene that is subject to both transcriptional and post‐transcriptional control. For the latter case, inherent PAI‐2 mRNA instability was previously shown to require a nonameric adenylate‐uridylate element in the 3′ UTR. However, mutation of this site was only partially effective at restoring complete mRNA stabilization. In the present study, we have identified additional regulatory motifs within the 3′ UTR that cooperate with the nonameric adenylate‐uridylate element to promote mRNA destabilization. These elements are located within a 74 nucleotide U‐rich stretch (58%) of the 3′ UTR that flanks the nonameric motif; deletion or substitution of this entire region results in complete mRNA stabilization. These new elements are conserved between species and optimize the destabilizing capacity with the nonameric element to ensure complete mRNA instability in a manner analogous to some class I and II adenylate‐uridylate elements present in transcripts encoding oncogenes and cytokines. Hence, post‐transcriptional regulation of the PAI‐2 mRNA transcript involves an interaction between closely spaced adenylate‐uridylate elements in a manner analogous to the post‐transcriptional regulation of oncogenes and cytokines.
Sensitivity to Abscisic Acid Modulates Positive Interactions between Arabidopsis thaliana Individuals
Authors: Zhang Hao , Shen Zhuxia , Wang Genxuan , Dai Xinfeng , Huang Qiaoqiao , Zheng Kefeng
Published: February 2010

The ability of abscisic acid (ABA) to modulate positive interactions between Arabidopsis thaliana individuals under salinity stress was investigated using abi1‐1 (insensitive to ABA), era1‐2 (hypersensitive to ABA) mutant and wild type plants. The results showed that sensitivity to ABA affects relative interaction intensity (RII) between Arabidopsis thaliana individuals. The neighbor removal experiments also confirmed the role of phenotypic responses in linking plant‐plant interactions and sensitivity to ABA. For abi1‐1 mutants, the absolute value differences between neighbor removal and control of stem length, root length, leaf area, leaf thickness, flower density, above biomass/belowground biomass (A/U), photosynthetic rate, stomatal conductance, leaf water content and water‐use efficiency were smaller than those of the wild type, while for era1‐2 mutants, these absolute value differences were larger than those of the wild type. Thus, it is suggested that positive interactions between Arabidopsis thaliana individuals are at least partly modulated by different sensitivity to ABA through different physiological and phenotypic plasticity.
Characterization of a Novel Plantain Asr Gene, MpAsr, that is Regulated in Response to Infection of Fusarium oxysporum f. sp. cubense and Abiotic Stresses
Authors: Liu Hai‐Yan , Dai Jin‐Ran , Feng Dong‐Ru , Liu Bing , Wang Hong‐Bin , Wang Jin‐Fa
Published: February 2010

Asr (abscisic acid, stress, ripening induced) genes are typically upregulated by a wide range of factors, including drought, cold, salt, abscisic acid (ABA) and injury; in addition to plant responses to developmental and environmental signals. We isolated an Asr gene, MpAsr, from a suppression subtractive hybridization (SSH) cDNA library of cold induced plantain (Musa paradisiaca) leaves. MpAsr expression was upregulated in Fusarium oxysporum f. sp. cubense infected plantain leaves, peels and roots, suggesting that MpAsr plays a role in plantain pathogen response. In addition, a 581‐bp putative promoter region of MpAsr was isolated via genome walking and cis‐elements involved in abiotic stress and pathogen‐related responses were detected in this same region. Furthermore, the MpAsr promoter demonstrated positive activity and inducibility in tobacco under F. oxysporum f. sp. cubense infection and ABA, cold, dehydration and high salt concentration treatments. Interestingly, transgenic Arabidopsis plants overexpressing MpAsr exhibited higher drought tolerance, but showed no significant decreased sensitivity to F. oxysporum f. sp. cubense. These results suggest that MpAsr might be involved in plant responses to both abiotic stress and pathogen attack.
Inducible Positive Mutant Screening System to Unveil the Signaling Pathway of Late Blight Resistance
Authors: Jia Zhiqi , Cui Yanhong , Li Ying , Wang Xiaoxuan , Du Yongchen , Huang Sanwen
Published: February 2010

Late blight is the most devastating potato disease and it also causes serious yield loss in tomato. Several disease resistance genes (R genes) to late blight have been cloned from potato in the past decade. However, the resistance mechanisms remain elusive. Tomato and potato belong to the botanical family Solanaceace and share remarkably conserved genome structure. Since tomato is a model system in genetic and plant pathology research, we used tomato to develop a powerful mutant screening system that will greatly facilitate the analysis of the signaling pathway of resistance to Phytophthora infestans. First we proved that the R3a transgenic tomatoes developed specific hypersensitive cell death response (HR) to P. infestans strains carrying the corresponding avirulence gene Avr3a, indicating that the signaling pathway from the R3a‐Avr3a recognition to HR is conserved between potato and tomato. Second, we generated transgenic tomatoes carrying both R3a and Avr3a genes, with the latter under the control of a glucocorticiod‐inducible promoter. Dexamethasone induced expression of Avr3a and resulted in localized HR. This versatile system can be used to construct a mutant library to screen surviving mutants whose resistance signal transduction was interrupted, providing the basis to identify key genes involved in the resistance to late blight in Solanaceae.
Explaining the reception of the Code Napoleon in Germany: a fuzzy-set qualitative comparative analysis
Authors: Arvind TT , Stirton Lindsay
Published: February 2010

This paper examines the diverse responses of the German states to the Code Napoleon at the beginning of the nineteenth century. These states differed both in the extent to which they adopted the Code, and the extent to which they retained the Code after Napoleon's influence waned. In order to identify the causes of adoption and retention of the Code, we use fuzzy-set qualitative comparative analysis (fsQCA). This method is now well established in comparative research in the social sciences but has been little used in comparative legal analysis. We find the following to be among the conditions relevant to the reception of the Code: territorial diversity, control by Napoleon, central state institutions, a feudal economy and society, liberal (enlightented absolutist) rule, nativism among the governing elites and popular anti-French sentiment. The paper also serves to demonstrate the potential of fsQCA as a method for comparative lawyers.
Addressing the imbalance: empowering older people in disaster response and preparedness
Authors: Deeny Pat , Vitale Catherine T. , Spelman Ruth , Duggan Seana
Published: February 2010

Deeny P., Vitale C.T., Spelman R. & Duggan S. (2010) Addressing the imbalance: empowering older people in disaster response and preparedness. International Journal of Older People Nursing 5, 77–80 doi: 10.1111/j.1748‐3743.2009.00204.x

This paper explores the role of the nurse in empowering older people at all stages in the disaster cycle. Evidence points to the need to increase the level of consultation and inclusivity of older people on this topic. Caution is called for in relation to classifying older people as a vulnerable group in disasters without first recognising that older people are probably the richest resource within any culture when it comes to emergency planning and\or coping with disasters. Practical advice on how nurses can best assist older people is presented.
Palestinian Water I: Resources, Allocation and Perception
Authors: Hassan Marwan A. , McIntyre Graham , Klinkenberg Brian , Al‐Rahman Tamimi Abed , Paisely Richard K. , Diabat Mousa , Shahin Khaled
Published: February 2010

Transboundary water issues are not new; however, the Palestinian case represents a unique situation that is atypical from most transboundary water conflicts. This difference is marked most importantly by the issue of lack of domestic control over water resources within the Palestinian Territories. Since the Israeli occupation of the West Bank in 1967, Palestinian water policy has been dictated by Israeli control, which has resulted in an allocational shift in water distribution in the Palestinian Territories. In this article we review transboundary water practices and conflicts, water resources, allocation and consumption, water perception, and water reuse and conservation in the Palestinian West Bank in order to assess contemporary trends in water practices as well as recommend strategies for improving regional water management. Water use and perception are assessed based on an extensive unpublished research survey conducted by the Palestinian Hydrology Group in 2002. The final report from this survey focused on developing criteria and linking initial results from the survey to the design of a cost effective and equitable water delivery system that would provide all Palestinians with basic water needs. In this article, through a reassessment and statistical analysis of their data, we support the recommendations of this initial survey, including the implementation of an increasing block tariff system as a means of sustainably delivering water throughout the West Bank. However, any future water management cannot proceed in an effective manner unless some level of water control is relinquished by Israel.
Physical Stability Assessment and Sensory Optimization of a Dairy‐Free Emulsion Using Response Surface Methodology
Authors: Granato Daniel , De Castro I. Alves , Ellendersen L. Souza Neves , Masson M. Lucia
Published: February 2010

ABSTRACT:  Desserts made with soy cream, which are oil‐in‐water emulsions, are widely consumed by lactose‐intolerant individuals in Brazil. In this regard, this study aimed at using response surface methodology (RSM) to optimize the sensory attributes of a soy‐based emulsion over a range of pink guava juice (GJ: 22% to 32%) and soy protein (SP: 1% to 3%). WHC and backscattering were analyzed after 72 h of storage at 7 °C. Furthermore, a rating test was performed to determine the degree of liking of color, taste, creaminess, appearance, and overall acceptability. The data showed that the samples were stable against gravity and storage. The models developed by RSM adequately described the creaminess, taste, and appearance of the emulsions. The response surface of the desirability function was used successfully in the optimization of the sensory properties of dairy‐free emulsions, suggesting that a product with 30.35% GJ and 3% SP was the best combination of these components. The optimized sample presented suitable sensory properties, in addition to being a source of dietary fiber, iron, copper, and ascorbic acid.
Trypsin Inhibitor from 3 Legume Seeds: Fractionation and Proteolytic Inhibition Study
Authors: Wati Richa Kusuma , Theppakorn Theerapong , Benjakul Soottawat , Rawdkuen Saroat
Published: February 2010

ABSTRACT:  The trypsin inhibitor from navy beans (Phaseoulus vulgaris), red kidney beans (Phaseoulus vulgaris L.), and adzuki beans (Vigna angularis) provided by the Royal Project Foundation in Thailand was isolated by heat and ammonium sulfate (AS) precipitation. Incubation at 70 °C for 10 min produced the highest trypsin inhibitor recovery for all legumes. The AS precipitation with 60% to 80% saturation (precipitate IV) resulted in 41‐, 88‐, and 34‐fold of the purity and (−)26%, 126%, and (−)47% of percentage of activity increase for navy beans, red kidney beans, and adzuki beans, respectively. The trypsin inhibitors had a molecular weight of 132 kDa for navy beans, 118 kDa for red kidney beans, and 13 kDa for adzuki beans under nonreducing conditions. The obtained precipitate IV fraction from each legume effectively prevented the degradation of the tilapia muscle with concentration dependent. The myosin heavy chain increased as the concentration of the inhibitor fraction increased, especially at the highest level of addition. The result indicated that the precipitate IV from these legumes have potential for use as a protease inhibitor in fishery related products.
Rofecoxib augments anticancer effects by reversing intrinsic multidrug resistance gene expression in BGC‐823 gastric cancer cells
Authors: ZHU Feng Shang , CHEN Xi Mei , HUANG Zhi Gang , WANG Zhi Rong , ZHANG Dong Wei , ZHANG Xia
Published: February 2010

OBJECTIVE:  To investigate combined chemotherapeutic effects of rofecoxib in combination with 5‐fluorouracil (5‐FU), cisplatin (DDP) and etoposide (VP‐16) in vitro, and to explore the potential mechanisms in modulating multidrug resistance (MDR) expression.

MEHTODS:  The BGC‐823 gastric cancer cell line was incubated for 48 h with 0.1 µmol/L rofecoxib, 5‐FU, DDP and VP‐16 (1 µg/mL, 10 µg/mL and 100 µg/mL) alone, and combined with rofecoxib, respectively. Methyl‐thiazolyl‐tetrazolium and the terminal deoxynucleotidyl transferase–mediated 2′‐deoxyuridine 5′‐yriphosphate nick‐end labeling assays were performed to calculate inhibitory rates and apoptotic index. Middle effects principles (CI values) were used to determine the interaction between rofecoxib and chemotherapeutic agents. Reverse transcription‐polymerase chain reaction (RT‐PCR) and Western blot analysis were employed to determine expression of MDR1, multidrug resistance‐associated protein 1 (MRP1), glutathione S‐tranferase‐π (GST‐π) mRNA and protein in gastric cancer cells administered by rofecoxib, respectively.

RESULTS:  Both anticancer drugs such as 5‐FU, DDP and VP‐16 and rofecoxib inhibited the cells' proliferation and induced apoptosis in a dose‐dependent manner, and a more significant inhibition was achieved when the cells were co‐treated with anticancer drugs and rofecoxib. There was a synergetic role when different concentrations of chemotherapeutic agents were combined with rofecoxib (all CI < 1, P < 0.01 or 0.05). RT‐PCR analyses of MDR gene families in BGC‐823 gastric cancer cells revealed a strong expression in MRP1 and GST‐π mRNA, but MDR1 mRNA was undetectable. After administration with different concentrations of rofecoxib (0.1, 1.0, 10 µmol/L), significant downregulation of MRP1 and GST‐π mRNA was observed (MRP1: from 0.984 ± 0.093–0.513 ± 0.098; GST‐π: from 1.078 ± 0.201–0.472 ± 0.084, P < 0.01 or 0.05). In addition, MRP1 and GST‐π protein expression induced by rofecoxib were also reduced (P < 0.01 or 0.05).

CONCLUSION:  Rofecoxib, a specific cyclooxygenase‐2 inhibitor, plays a chemotherapeutic sensitizer role in various anticancer agents on the BGC‐823 gastric cancer cell line, which could be partly explained by its ability to reverse the intrinsic MRP1 and GST‐πin vitro.
ORIGINAL ARTICLE: Evaluation of the CellaVision™ DM automated microscope in pediatrics
Authors: BILLARD M. , LAINEY E. , ARMOOGUM P. , ALBERTI C. , FENNETEAU O. , DA COSTA L.
Published: February 2010

The DM is an automated microscope, which performs WBC differential counts and monitors red cell morphology. The user either validates the cell recognition if the DM has correctly identified the WBCs or reclassifies the WBCs in the good category in case of a DM mis‐assignment. Morphological anomalies of leukocytes, red blood cells or platelets are analyzed and registered. We studied 521 newborns and infants sorted by age and pathology. The results correlated well with those using conventional microscopy except for samples containing blasts, in which the percentage of malignant cells was underestimated. Newborns had the lowest rates of overall accuracy and postclassification agreement. For red cell analysis, 10% of the selected areas were considered unreadable. However, the DM diagnosed faithfully all studied red cell pathologies. The DM was also very useful in analyzing samples of storage diseases. Timesavings ranging from 1 up to 10 min (for vacuolated lymphocyte screening) were observed when performing analysis with the DM. The DM represents a useful diagnostic and training tool. However, conventional microscopy remains essential, in particular when the image quality is poor, such as in the case of lymphoblasts, and in the screening of platelet clusters.
Metabolic Syndrome Risk Factors Among a Sample of Overweight and Obese Mexican Children
Authors: Elizondo‐Montemayor Leticia , Serrano‐González Mónica , Ugalde‐Casas Patricia A. , Cuello‐García Carlos , Borbolla‐Escoboza José R.
Published: February 2010

J Clin Hypertens (Greenwich).

The objective of this study was to estimate the prevalence and correlations of components of the metabolic syndrome (MetS) using the International Diabetes Federation (IDF) pediatric definition in a cross‐sectional study of 215 overweight/obese Mexican children aged 6 to 12. There are no previous studies of this kind in Mexican children. Clinical, anthropometric, and laboratory measurements were performed. The prevalence of MetS using the pediatric IDF criteria was 6.7% (95% confidence interval, 4.0–11.1). A higher proportion of children in the younger age group had waist circumference above the cutoff, while a higher proportion in the older age group had hyperglycemia. Children with MetS had higher percentages of body fat, body mass index, total cholesterol, and low‐density lipoprotein cholesterol. Increased triglycerides, decreased high‐density lipoprotein cholesterol, and waist circumference were most highly associated with MetS. This has significant implications for public health.
Variability and divergence in Pongamia pinnata (L.) Pierre germplasm – a candidate tree for biodiesel
Authors: SUNIL N. , KUMAR VINOD , SIVARAJ N. , LAVANYA C. , PRASAD R. B. N. , RAO B. V. S. K. , VARAPRASAD K. S.
Published: February 2010

Three explorations were undertaken in South East Coastal zone of India covering parts of Andhra Pradesh (AP) and Orissa states to collect Pongamia pinnata (L.) Pierre germplasm during March–June 2007. A total of 123 accessions were collected and seed data recorded were analyzed for morphometric traits viz., seed length, seed width, seed thickness, 100‐seed weight and oil content. Variation in the collected germplasm was analyzed using anova, simple measures of variation and D2 statistics. Significant genetic variability between seed traits and oil content and association among the seed traits was recorded. Phenotypic variance was higher than genotypic variance for all the characters indicating dominant role of environment. High heritability (broad sense) for 100‐seed weight (97.6%) and oil content (86.7%) indicated the reliability of these characters as selection criteria for plus trees. Genetic gain was maximum for 100‐seed weight (62.6%) followed by oil content (30.5%). D2 analysis grouped the accessions into 12 clusters. Cluster XII and cluster IX were the most diverse based on the intercluster distance. Based on the observed diversity, Chittoor, Srikakulam and Adilabad districts of AP are most suitable for collecting diverse germplasm lines and also for in situ conservation.
The impact of co‐occurring tree and grassland species on carbon sequestration and potential biofuel production
Authors: LAUNGANI RAMESH , KNOPS JOHANNES M. H.
Published: February 2010

We evaluated how three co‐occurring tree and four grassland species influence potentially harvestable biofuel stocks and above‐ and belowground carbon pools. After 5 years, the tree Pinus strobus had 6.5 times the amount of aboveground harvestable biomass as another tree Quercus ellipsoidalis and 10 times that of the grassland species. P. strobus accrued the largest total plant carbon pool (1375 g C m−2 or 394 g C m−2 yr), while Schizachyrium scoparium accrued the largest total plant carbon pool among the grassland species (421 g C m−2 or 137 g C m−2 yr). Quercus ellipsoidalis accrued 850 g C m−2, Q. macrocarpa 370 g C m−2, Poa pratensis 390 g C m−2, Solidago canadensis 132 g C m−2, and Lespedeza capitata 283 g C m−2. Only P. strobus and Q. ellipsoidalis significantly sequestered carbon during the experiment. Species differed in total ecosystem carbon accumulation from −21.3 to +169.8 g C m−2 yr compared with the original soil carbon pool. Plant carbon gains with P. strobus were paralleled by a decrease of 16% in soil carbon and a nonsignificant decline of 9% for Q. ellipsoidalis. However, carbon allocation differed among species, with P. strobus allocating most aboveground in a disturbance prone aboveground pool, whereas Q. ellipsoidalis, allocated most carbon in less disturbance sensitive belowground biomass. These differences have strong implications for terrestrial carbon sequestration and potential biofuel production. For P. strobus, aboveground plant carbon harvest for biofuel would result in no net carbon sequestration as declines in soil carbon offset plant carbon gains. Conversely the harvest of Q. ellipsoidalis aboveground biomass would result in net sequestration of carbon belowground due to its high allocation belowground, but would yield lower amounts of aboveground biomass. Our results demonstrate that plant species can differentially impact ecosystem carbon pools and the distribution of carbon above and belowground.
The potential of Miscanthus to sequester carbon in soils: comparing field measurements in Carlow, Ireland to model predictions
Authors: DONDINI MARTA , HASTINGS ASTLEY , SAIZ GUSTAVO , JONES MICHAEL B. , SMITH PETE
Published: February 2010

Growing bioenergy crops such as Miscanthus has the potential to mitigate atmospheric carbon dioxide emissions by the replacement of fossil fuels and by storing carbon (C) in the soil due to land use change. Here we compare direct measurements of soil organic C fractions made in Carlow (Ireland) to model predictions made by RothC and a cohort model. Our results show that when Miscanthus is grown on land previously under arable agriculture, the soil organic C will increase to a level above that of native pasture, as Miscanthus organic material is shown to have a slow decomposition rate. In addition we demonstrate that for measured organic C, fractions of different lability are similar to the C pools used in RothC. Using the model predictions from RothC and Miscanthus yields from MISCANFOR, we predict that in Ireland, changing the land use from arable to Miscanthus plantations has the potential to store between 2 and 3 Mg C ha−1 y−1 depending on the crop yield and the initial soil organic C level.
Etching studies of silica glasses in SF6/Ar inductively coupled plasmas: Implications for microfluidic devices fabrication
Authors: Lallement L. , Gosse C. , Cardinaud C. , Peignon-Fernandez M.-C. , Rhallabi A.
Published: February 2010

To fabricate microlaboratories, commercially available silica glasses represent a good alternative to the expensive quartz or fused silica substrates. Therefore, the authors have here investigated the behavior of four of them—Vycor, Pyrex, D263, and AF45—in SF6 and SF6/Ar inductively coupled plasmas. Using Vycor, a material close to pure SiO2, as a reference, they demonstrated that the etch rate negatively correlates with the global content in metallic oxides. However, no such clear trend was found for the surface roughness and they hypothesize that the large asperities (>500 nm) sometimes observed might be due to local variation in the glass surface composition. Furthermore, investigations on the influence of the plasma conditions (i.e., source power, dc self-bias, gas mixture, and pressure) on the etch rate, surface chemistry, and surface morphology, as well as positive ion current and fluorine concentration measurements, enable them to unravel an ion enhanced chemical etching mechanism, where stronger ion assistance is needed when more metallic oxides are present. By increasing the ion to neutral flux ratio, they consequently could, for all the materials, reduce the surface roughness to less than 5 nm while maintaining etch rates around 150 nm/min. These conditions have further been used to optimize pattern transfer experiments.
Double Trouble
Authors: Wong Wei
Published: February 2010

Neighboring cells with distinct mutations can cooperate to form invasive tumors.
Promoting Axon Formation
Authors: Stern Peter R.
Published: February 2010

Cyclic adenosine monophosphate and cyclic guanosine monophosphate regulate axon formation and exert opposite actions on dendrite formation.
Dueling Kinases Finesse the Fat
Authors: Ray L. Bryan
Published: February 2010

The energy-sensing kinase AMPK is coupled to hormonal control of metabolism through PKA.
Science Signaling
Podcast: 2 February 2010
Authors: Kino Tomoshige , VanHook Annalisa M.
Published: February 2010

A noncoding RNA competes with DNA for binding to the glucocorticoid receptor.
Transformation of Acinetobacter calcoaceticus RAG-1 by electroporation
Authors: Leahy Joseph G. , Jones-Meehan Joanne M. , Colwell Rita R.
Published: February 2010

A method is described for transforming Acinetobacter calcoaceticus RAG-1 with a transposon-delivery plasmid, pLOFPttKm, and the broad-host-range cloning vector pKT230. Kanamycin-resistant transformants isolated following electroporation with pLOFPttKm were shown to have lost the plasmid, and contained the transposon mini-Tn10PttKm in at least five unique, single sites. Transformation with pKT230 was confirmed by plasmid screening and acquisition of the kanamycin-resistant, streptomycin-resistant phenotype. By optimizing conditions of electric field strength and pulse length for electroporation, we obtained a maximum efficiency of 4.36 × 102/(μg DNA∙109 cells) for insertion of the mini-Tn10PttKm transposon, and 9.88 × 104/(μg DNA∙109 cells) for transformation with plasmid pKT230. Our results indicate that electroporation is a suitable method for introducing transposon and cloning vectors into A. calcoaceticus RAG-1, and possibly other Acinetobacter spp. The availability of a transformation system should enhance the molecular genetic and physiological analysis of these bacteria, particularly the vast majority of strains that are not competent for transformation with DNA.Key words: electroporation, Acinetobacter, transformation, plasmid, transposon.
Numerical taxonomy of Vibrionaceae from Newfoundland coastal waters
Authors: Martin-Kearley Jennifer , Gow John A.
Published: February 2010

Numerical analysis was used to characterize 111 Vibrionaceae strains. These included 31 reference cultures belonging to the genera Aeromonas, Listonella, and Vibrio and 80 strains isolated from the seasonally cold coastal waters of Newfoundland. The sources of the regional strains were the brown alga Alaria esculenta and the giant scallop Placopecten magellanicus. Most of the regional strains grew at temperatures associated with psychrotrophs, while a small proportion may have been psychrophilic. All of the regional strains grew at 4 °C. A routine incubation temperature of 20 °C was chosen and in tests for utilization of organic components as sole sources of carbon and energy the strains were incubated for 3 weeks rather than the more common 6-day period. The treatment of weak positive results as weak positive, positive, or negative was investigated and it was decided that the general conclusions reached in the study would not be significantly altered by the interpretation of weak positive results. Using numerical analysis it was shown that most of the strains clustered according to source. Most reference cultures were more closely related to each other than they were to the regional strains. Some strains were phenotypically similar to Vibrio splendidus biovar I, which is arginine dihydrolase positive. Although there were differences, some strains were similar to the fish pathogen Vibrio ordalii, which is negative for arginine dihydrolase. Both species are reported to grow at 4 °C. It was shown that most of the regional Vibrionaceae strains studied were different from previously described species belonging to the family Vibrionaceae.Key words: numerical taxonomy, Vibrionaceae, Vibrio, marine bacteria, psychrotroph.
The optimization and application of two direct viable count methods for bacteria in distributed drinking water
Authors: Coallier Josée , Prévost Michèle , Rompré Annie , Duchesne Daniel
Published: February 2010

The optimal incubation conditions for the direct viable count method with nalidixic acid were determined. They do not differ from those proposed in the literature for a laboratory strain and a mixed bacterial population isolated from drinking water. The direct viable count method with 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) was performed under in situ conditions. The bacteria were incubated with CTC at a concentration of 1 mM for 4–6 h at the temperature of the water in the pipes and without the addition of an exogenous substrate. The results obtained for a laboratory strain using the two direct count methods were similar. However, for a mixed bacterial population, the counts were always higher with the CTC method than with the nalidixic acid method.Key words: drinking water, CTC, nalidixic acid, direct viable count.
Characterization of the diversity of sulfate-reducing bacteria in soil and mining waste water environments by nucleic acid hybridization techniques
Authors: Telang Anita J. , Voordouw Gerrit , Ebert Sara , Sifeldeen Neili , Foght Julia M. , Fedorak Phillip M. , Westlake D. W. S.
Published: February 2010

Nucleic acid hybridization techniques were used to characterize the sulfate-reducing bacterial communities at seven waste water and two soil sites in Canada. Genomic DNA was obtained from liquid enrichment cultures of samples taken from these nine sites. The liquid enrichment protocol favored growth of the sulfate-reducing bacterial component of the communities at these sites. The genomic DNA preparations were analyzed with (i) a specific gene probe aimed at a single genus (Desulfovibrio), (ii) a general 16S rRNA gene probe aimed at all genera of sulfate-reducing bacteria and other bacteria, and (iii) whole genome probes aimed at specific bacteria. This three-pronged approach provided information on the sulfate-reducing bacterial community structures for the nine sites. These were compared with each other and with the sulfate-reducing bacterial communities of western Canadian oil field production waters, studied previously. It was found that there is considerable diversity in the sulfate-reducing bacterial community at each site. Most sulfate-reducing bacteria isolated from distinct sites are genomically different and differ also from sulfate-reducing bacteria found in oil field production waters.Key words: sulfate-reducing bacteria, genomic diversity, nucleic acid hybridization, microbial community.
Production and partial characterization of an endopolygalacturonase from Saccharomyces cerevisiae
Authors: Blanco P. , Sieiro C. , Diaz A. , Villa T. G.
Published: February 2010

Saccharomyces cerevisiae CECT1389 secreted an extracellular endopolygalacturonase (EC 3.2.1.15) when grown in shake flasks in medium containing galactose alone, or either galactose and polygalacturonic acid or galactose and galacturonic acid as the carbon sources. The synthesis of the enzyme was repressed by glucose and by high oxygen tensions. The enzyme was partially purified by gel exclusion chromatography over Sephacryl S-200, where it showed an apparent molecular mass of 39 kDa; the value determined by high-performance liquid chromatography (HPLC) was 65 kDa. The optimal temperature and pH for enzyme activity were45 °C and 5.5, respectively. The Km and Vmax values for polygalacturonic acid were 4.7 mg∙mL−1 and 6.4 nmol∙mL−1∙min−1. The Ki for HgCl2 was 6.8 × 10−5 M. The enzyme exhibited an endo-splitting mechanism as deduced from viscosimetry experiments as well as from an HPLC study of the end products.Key words: characterization, endopolygalacturonase, Saccharomyces cerevisiae.
Genetic variability of the commonnodgene in soybean bradyrhizobia isolated in Thailand and Japan
Authors: Yokoyama Tadashi , Ando Shotaro , Murakami Toshifumi , Imai Hideo
Published: February 2010

To determine the taxonomic relationship between Thai soybean bradyrhizobia and soybean bradyrhizobia from other regions, a total of 62 Bradyrhizobium strains were isolated in Thailand. The genetic diversity of the strains was examined with reference to 46 Japanese and 15 USDA strains. The degree of sequence divergence in and around common nod gene regions of the 123 strains was estimated by restriction fragment length polymorphism analysis using the Bradyrhizobium japonicum USDA 110 common nodDYABC gene probe. The phylogenetic grouping of the strains resulted in four major clusters. Cluster 1 comprised the Japanese and USDA strains, which originated in temperate regions, whereas clusters 3 and 4 comprised the tropical Thai strains. Cluster 1 strains comprised the DNA homology groups I and Ia, and hence, were classified as B. japonicum. Cluster 2 strains were in the DNA homology group II, and hence, were classified as Bradyrhizobium elkanii. Clusters 3 and 4 strains, however, did not correspond to any known DNA homology groups. These results indicate that Thai soybean bradyrhizobia are distantly related to B. japonicum and B. elkanii.Key words: Bradyrhizobium japonicum, Bradyrhizobium elkanii, common nod gene, RFLP, genetic diversity.
Comparative study of five polycyclic aromatic hydrocarbon degrading bacterial strains isolated from contaminated soils
Authors: Dagher Fadi , Déziel Eric , Lirette Patricia , Paquette Gilles , Bisaillon Jean-Guy , Villemur Richard
Published: February 2010

Five polycyclic aromatic hydrocarbon (PAH) degrading bacterial strains, Pseudomonas putida 34, Pseudomonas fluorescens 62, Pseudomonas aeruginosa 57, Sphingomonas sp. strain 107, and the unidentified strain PL1, were isolated from two contaminated soils and characterized for specific features regarding PAH degradation. Degradation efficiency was determined by the rapidity to form clearing zones around colonies when sprayed with different PAH solutions and the growth in liquid medium with different PAHs as sole source of carbon and energy. The presence of plasmids, the production of biosurfactants, the effect of salicylate on PAH degradation, the transformation of indole to indigo indicating the presence of an aromatic ring dioxygenase activity, and the hybridization with the SphAb probe representing a sequence highly homologous to the naphthalene dioxygenase ferredoxin gene nahAb were examined. The most efficient strain in terms of substrate specificity and rapidity to degrade different PAHs was Sphingomonas sp. strain 107, followed by strain PL1 and P. aeruginosa 57. The less efficient strains were P. putida 34 and P. fluorescens 62. Each strain transformed indole to indigo, except strain PL1. Biosurfactants were produced by P. aeruginosa 57 and P. putida 34, and a bioemulsifier was produced by Sphingomonas sp. strain 107. The presence of salicylate in solid medium has accelerated the formation of clearing zones and the transformation of indole by Sphingomonas sp. strain 107 and P. aeruginosa 57 colonies. Plasmids were found in Sphingomonas sp. strain 107 and strain PL1. The SphAb probe hybridized with DNA extracted from each strain. However, hybridization signals were detected only in the plasmidic fraction of Sphingomonas sp. strain 107 and strain PL1. Using a polymerase chain reaction (PCR) approach, we determined that several genes encoding enzymes involved in the upper catabolic pathway of naphthalene were present in each strain. Sequencing of PCR DNA fragments revealed that, for all the five strains, these genes are highly homologous with respective genes found in the pah, dox, and nah opérons, and are arranged in a polycistronic operon. Results suggest that these genes are ordered in the five selected strains like the pah, nah, and dox opérons.Key words: polycyclic aromatic hydrocarbons, biodegradation, polymerase chain reaction, naphthalene catabolic genes.
Molecular analysis of lytic genes of bacteriophage 80α ofStaphylococcus aureus
Authors: Bon Jon , Mani Nagraj , Jayaswal R. K.
Published: February 2010

Nucleotide sequencing of a 3779-bp fragment of the Staphylococcus aureus bacteriophage 80α revealed two open reading frames: ORF1, designated as lytA, which encodes a polypeptide of 481 amino acids with an apparent Mrof 53.81 kDa; and ORF2, designated as holin, which encodes for a hydrophobic polypeptide of 145 amino acids with an apparent Mrof 15.58 kDa and exhibits two putative transmembrane helices. Both genes showed 100% sequence homology to that of the peptidoglycan hydrolase and holin genes of the S. aureus phage [Formula: see text] reported earlier. In addition, the downstream sequences of the lytA gene were homologous to the phage attachment site (attP) of the phage [Formula: see text]. Based on our data we propose that the lytic system of the phage 80α evolved from that of phage [Formula: see text].Key words: attachment site, bacteriophage 80α, holin, peptidoglycan hydrolase, Staphylococcus aureus.
Intracytoplasmic membrane formation inMethylomicrobium albumBG8 is stimulated by copper in the growth medium
Authors: Brantner Christine A. , Buchholz Lorie A. , McSwain Claudia L. , Newcomb Laura L. , Remsen Charles C. , Collins Mary Lynne Perille
Published: February 2010

Methylomicrobium album BG8 uses methane as its sole source of carbon and energy. The oxidation of methane to methanol is catalyzed by the enzyme methane monooxygenase. Methane monooxygenase activity, intracytoplasmic membrane abundance, and cell mass increased with increasing copper concentration in the medium. When copper was added to copper-deficient cultures, cell mass and intracytoplasmic membrane structure increased. These findings are consistent with the presence of copper in the particulate methane monooxygenase. Methane monooxygenase activity and intracytoplasmic membrane abundance were correlated, suggesting that the methane monooxygenase may be involved in intracytoplasmic membrane proliferation.Key words: Methylomicrobium album BG8, copper, intracytoplasmic membrane, methane monooxygenase.
Monitoring the insecticidal toxins fromBacillus thuringiensisin soil with flow cytometry
Authors: Tapp H. , Stotzky G.
Published: February 2010

The accumulation and persistance in soil and other natural habitats of the insecticidal toxins from Bacillus thuringiensis may result in environmental hazards, such as toxicity to nontarget species and the selection of toxin-resistant target species. We describe the use of flow cytometry as a method for detecting and tracking the fate of these insecticidal toxins in soil that does not require their extraction and purification. The toxins from B. thuringiensis subspp. tenebrionis and kurstaki were bound on clay- or silt-sized particles separated from Kitchawan soil that was unamended (naturally contains predominantly kaolinite) or amended to 6% v/v with the clay minerals montmorillonite or kaolinite (as an internal control). The particle–toxin mixtures were suspended in 0.1 M phosphate buffer (pH 7) containing 3% nonfat milk powder to block nonspecific binding of antibody, resuspended in a solution of antibody to the toxin from B. thuringiensis subsp. tenebrionis, and then resuspended in a solution of anti-rabbit antibody conjugated with fluorescein isothiocyanate (FITC–Ab). Controls consisted of the particles alone and bound complexes of the particles with the toxin from B. thuringiensis subsp. kurstaki. All particles that bound the toxin from B. thuringiensis subsp. tenebrionis showed a significant shift in the peak of fluorescence to the right on the x axis as compared with the nonspecific fluorescence from the control FITC–Ab complexes with particles in the absence of the toxin. There was also a slight shift in the peak to the right for some particles that bound the toxin from B. thuringiensis subsp. tenebrionis, as there is some cross-reactivity between the toxins from B. thuringiensis subspp. tenebrionis and kurstaki and the antibodies that they induce. This method is more sensitive and rapid than the dot-blot ELISA, and processing of many samples is easily accomplished.Key words: flow cytometry, soil, insecticidal toxins, Bacillus thuringiensis, clay, silt.
Production of cercosporin toxin by the phytopathogenicCercosporafungi is affected by diverse environmental signals
Authors: You Bang-Jau , Lee Miin-Hui , Chung Kuang-Ren
Published: February 2010

Cercosporin is a polyketide phytotoxin produced by many phytopathogenic Cercospora spp. We investigated environmental signals that have elaborate control of cercosporin production. Light is the most critical factor for cercosporin production. Cercospora nicotianae accumulated substantial quantities of cercosporin only when grown on a particular potato dextrose agar under light but produced little cercosporin on other brands of potato dextrose agar or media with defined ingredients. In addition to light regulation, numerous factors including salts, buffers, and ions markedly affected cercosporin production. By contrast, pH had little effect on cercosporin production. Depletion or alteration of the carbon or nitrogen sources also affected cercosporin production. Production of cercosporin was elevated to varying levels by metal ions, such as cobalt, ferric, manganese, and zinc. Significant differences in cercosporin production were observed among various Cercospora species. Further, regulation of cercosporin production by phosphate buffer, ammonium, LiCl, but not metal ions appeared to occur at transcriptional levels. Expression of the genes involved in cercosporin biosynthesis and regulation decreased markedly and was closely concomitant with the amounts of cercosporin reduced as the fungus was grown on medium containing phosphate, LiCl, ammonium, or dimethyl sulfoxide. The results reveal the complexity of cercosporin production at the physiological and genetic levels. A model delineating regulatory controls of cercosporin biosynthesis is proposed and discussed.
Pretreatment of lignocellulosic material with fungi capable of higher lignin degradation and lower carbohydrate degradation improves substrate acid hydrolysis and the eventual conversion to ethanol
Authors: Kuhar Sarika , Nair Lavanya M. , Kuhad Ramesh Chander
Published: February 2010

Phanerochaete chrysosporium , Pycnoporus cinnabarinus ,and fungal isolates RCK-1 and RCK-3 were tested for their lignin degradation abilities when grown on wheat straw (WS) and Prosopis juliflora (PJ) under solid-state cultivation conditions. Fungal isolate RCK-1 degraded more lignin in WS (12.26% and 22.64%) and PJ (19.30% and 21.97%) and less holocellulose in WS (6.27% and 9.39%) and PJ (3.01% and 4.58%) after 10 and 20 days, respectively, than other fungi tested. Phanerochaete chrysosporium caused higher substrate mass loss and degraded more of holocellulosic content (WS: 55.67%; PJ: 48.89%) than lignin (WS: 18.89%; PJ: 20.20%) after 20 days. The fungal pretreatment of WS and PJ with a high-lignin-degrading and low-holocellulose-degrading fungus (fungal isolate RCK-1) for 10 days resulted in (i) reduction in acid load for hydrolysis of structural polysaccharides (from 3.5% to 2.5% in WS and from 4.5% to 2.5% in PJ), (ii) an increase in the release of fermentable sugars (from 30.27 to 40.82 g·L–1in WS and from 18.18 to 26.00 g·L–1in PJ), and (iii) a reduction in fermentation inhibitors (total phenolics) in acid hydrolysate of WS (from 1.31 to 0.63 g·L–1) and PJ (from 2.05 to 0.80 g·L–1). Ethanol yield and volumetric productivity from RCK-1-treated WS (0.48 g·g–1and 0.54 g·L–1·h–1, respectively) and PJ (0.46 g·g–1and 0.33 g·L–1·h–1, respectively) were higher than untreated WS (0.36 g·g–1and 0.30 g·L–1·h–1, respectively) and untreated PJ (0.42 g·g–1and 0.21 g·L–1·h–1, respectively).
Occurrence of herpes simplex virus 1 and three periodontal bacteria in patients with chronic periodontitis and necrotic pulp
Authors: Nishiyama Sheila Alexandra Belini , Nakano Viviane , Velásquez-Melendez Gustavo , Avila-Campos Mario Julio
Published: February 2010

Viral and bacterial associations appear to be implicated in the development of periodontal infections. Little information is available describing the periodontopathic agents in root canals with necrotic pulp. In this study, the occurrence and the combinations among herpes simplex virus type 1 (HSV-1) and Dialister pneumosintes , Tannerella forsythia , and Treponema denticola in patients with chronic periodontitis and necrotic pulp were evaluated. Clinical samples from healthy subjects and patients with periodontal or pulp infections were analyzed using a nested polymerase chain reaction PCR to detect HSV and PCR to detect the 3 periodontal bacteria. The presence of Tannerella forsythia and Treponema denticola was observed in healthy, periodontitis, and necrotic pulp patients. HSV was observed in periodontitis and necrotic pulp patients, and no healthy subject harbored D. pneumosintes or HSV. The occurrence of Tannerella forsythia was not statistically significant in patients with necrotic pulp (P = 0.704). Periodontal bacteria were observed varying from 10.3% to 20.7% in periodontitis and necrotic pulp patients. The presence of Treponema denticola – HSV association was predominant in patients showing necrotic pulp (24.1%); however, HSV alone was observed in one patient with periodontitis and in another patient with necrotic pulp. The presence of double association among bacteria or bacteria – HSV could indicate a role in both periodontitis and necrotic pulp, and Tannerella forsythia – Treponema denticola – HSV and Tannerella forsythia – D. pneumosintes – Treponema denticola – HSV associations might be important in periodontitis.