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171.
172.
Sebastian Schlafer Javier E. Garcia Matilde Greve Merete K. Raarup Bente Nyvad Irene Dige 《Applied and environmental microbiology》2015,81(4):1267-1273
pH in the extracellular matrix of bacterial biofilms is of central importance for microbial metabolism. Biofilms possess a complex three-dimensional architecture characterized by chemically different microenvironments in close proximity. For decades, pH measurements in biofilms have been limited to monitoring bulk pH with electrodes. Although pH microelectrodes with a better spatial resolution have been developed, they do not permit the monitoring of horizontal pH gradients in biofilms in real time. Quantitative fluorescence microscopy can overcome these problems, but none of the hitherto employed methods differentiated accurately between extracellular and intracellular microbial pH and visualized extracellular pH in all areas of the biofilms. Here, we developed a method to reliably monitor extracellular biofilm pH microscopically with the ratiometric pH-sensitive dye C-SNARF-4, choosing dental biofilms as an example. Fluorescent emissions of C-SNARF-4 can be used to calculate extracellular pH irrespective of the dye concentration. We showed that at pH values of <6, C-SNARF-4 stained 15 bacterial species frequently isolated from dental biofilm and visualized the entire bacterial biomass in in vivo-grown dental biofilms with unknown species composition. We then employed digital image analysis to remove the bacterial biomass from the microscopic images and adequately calculate extracellular pH values. As a proof of concept, we monitored the extracellular pH drop in in vivo-grown dental biofilms fermenting glucose. The combination of pH ratiometry with C-SNARF-4 and digital image analysis allows the accurate monitoring of extracellular pH in bacterial biofilms in three dimensions in real time and represents a significant improvement to previously employed methods of biofilm pH measurement. 相似文献
173.
Esteban D. Babot José C. del Río Marina Ca?ellas Ferran Sancho Fátima Lucas Víctor Guallar Lisbeth Kalum Henrik Lund Glenn Gr?be Katrin Scheibner René Ullrich Martin Hofrichter Angel T. Martínez Ana Gutiérrez 《Applied and environmental microbiology》2015,81(12):4130-4142
The goal of this study is the selective oxyfunctionalization of steroids under mild and environmentally friendly conditions using fungal enzymes. With this purpose, peroxygenases from three basidiomycete species were tested for the hydroxylation of a variety of steroidal compounds, using H2O2 as the only cosubstrate. Two of them are wild-type enzymes from Agrocybe aegerita and Marasmius rotula, and the third one is a recombinant enzyme from Coprinopsis cinerea. The enzymatic reactions on free and esterified sterols, steroid hydrocarbons, and ketones were monitored by gas chromatography, and the products were identified by mass spectrometry. Hydroxylation at the side chain over the steroidal rings was preferred, with the 25-hydroxyderivatives predominating. Interestingly, antiviral and other biological activities of 25-hydroxycholesterol have been reported recently (M. Blanc et al., Immunity 38:106–118, 2013, http://dx.doi.org/10.1016/j.immuni.2012.11.004). However, hydroxylation in the ring moiety and terminal hydroxylation at the side chain also was observed in some steroids, the former favored by the absence of oxygenated groups at C-3 and by the presence of conjugated double bonds in the rings. To understand the yield and selectivity differences between the different steroids, a computational study was performed using Protein Energy Landscape Exploration (PELE) software for dynamic ligand diffusion. These simulations showed that the active-site geometry and hydrophobicity favors the entrance of the steroid side chain, while the entrance of the ring is energetically penalized. Also, a direct correlation between the conversion rate and the side chain entrance ratio could be established that explains the various reaction yields observed. 相似文献
174.
Abundance,diversity and geographic distribution of cassava mosaic disease pandemic‐associated Bemisia tabaci in Tanzania 下载免费PDF全文
L. S. Tajebe S. B. Boni D. Guastella V. Cavalieri O. S. Lund C. P. Rugumamu C. Rapisarda J. P. Legg 《Journal of Applied Entomology》2015,139(8):627-637
Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae), one of the most economically important agricultural pests worldwide, is the vector of cassava mosaic geminiviruses that cause cassava mosaic disease (CMD). In East and Central Africa, a severe CMD pandemic that spread from Uganda in the late 1980s still continues to devastate cassava crops. To assess the association of distinct B. tabaci genetic groups with the CMD pandemic, mitochondrial cytochrome oxidase I gene sequences were analysed from whiteflies collected during surveys conducted from 2010 to 2013 in Tanzania. Four genetic groups – Sub‐Saharan Africa 1 (SSA1), Mediterranean, Indian Ocean and East Africa 1, and a group of unknown whitefly species were identified. SSA1 comprised four subgroups: SSA1‐SG1, SSA1‐SG2, SSA1‐SG1/2 and SSA1‐SG3. SSA1‐SG1 was confined to the pandemic‐affected north‐western parts of Tanzania whilst SSA1‐SG2 and SSA1‐SG3 were found in the central and eastern parts not yet affected by the pandemic. The CMD pandemic front was estimated to lie in Geita Region, north‐western Tanzania, and to be spreading south‐east at a rate of ca 26 km/year. The pandemic‐associated B. tabaci SSA1‐SG1 predominated up to 180 km ahead of the CMD front indicating that changes in whitefly population characteristics precede changes in disease characteristics. 相似文献
175.
Diversity of symbiotic bacteria associated with Bemisia tabaci (Homoptera: Aleyrodidae) in cassava mosaic disease pandemic areas of Tanzania 下载免费PDF全文
L.S. Tajebe D. Guastella V. Cavalieri S.E. Kelly M.S. Hunter O.S. Lund J.P. Legg C. Rapisarda 《The Annals of applied biology》2015,166(2):297-310
All Bemisia tabaci individuals harbour an obligate bacterial symbiont (Portiera aleyrodidarum), and many also harbour non‐essential facultative symbionts. The association of symbiotic bacteria with the various genetic groups of B. tabaci remains unknown for East Africa. This study aimed to assess any association between the various whitefly genetic groups and the endosymbionts they harbour; to investigate if a unique endosymbiont is associated with super‐abundant whiteflies, and to provide baseline information on endosymbionts of whiteflies for a part of East Africa. Whiteflies collected during surveys in Tanzania were genotyped and screened for the presence of the obligate and six secondary symbionts (SS): Rickettsia (R), Hamiltonella (H), Arsenophonus (A), Wolbachia (W), Cardinium (C) and Fritschea (F). The results revealed the presence of Mediterranean (MED), East Africa 1 (EA1), Indian Ocean (IO) and Sub‐Saharan Africa 1 (SSA1) genetic groups of Bemisia tabaci, with SSA1 further clustered into four sub‐groups: SSA1‐SG1, SSA1‐SG2, SSA1‐SG1/2 and SSA1‐SG3. F was completely absent from all of the whiteflies tested while R was always found in double or multiple infections. In general, no particular symbiont appeared to be associated with the super‐abundant SSA1‐SG1 B. tabaci, although A or AC infections were common among infected individuals. The most striking feature of these super‐abundant whiteflies, dominating cassava mosaic disease pandemic areas, was the high prevalence of individuals uninfected by any of the six SS tested. This study of the endosymbionts of B. tabaci in East Africa showed contrasting patterns of infection in crop and weed hosts. 相似文献
176.
Gopinath Kulasekaran Nadya Nossova Andrea L. Marat Ingrid Lund Christopher Cremer Maria S. Ioannou Peter S. McPherson 《The Journal of biological chemistry》2015,290(29):17999-18008
Connecdenn 1/2 are DENN (differentially expressed in normal and neoplastic cells) domain-bearing proteins that function as GEFs (guanine nucleotide exchange factors) for the small GTPase Rab35. Disruption of connecdenn/Rab35 function leads to defects in the recycling of multiple cargo proteins from endosomes with altered cell function, yet the regulation of connecdenn GEF activity is unexplored. We now demonstrate that connecdenn 1/2 are autoinhibited such that the purified, full-length proteins have significantly less Rab35 binding and GEF activity than the isolated DENN domain. Both proteins are phosphorylated with prominent phosphorylation sites between residues 500 and 600 of connecdenn 1. A large scale proteomics screen revealed that connecdenn 1 is phosphorylated at residues Ser-536 and Ser-538 in an Akt-dependent manner in response to insulin stimulation of adipocytes. Interestingly, we find that an Akt inhibitor reduces connecdenn 1 interaction with Rab35 after insulin treatment of adipocytes. Remarkably, a peptide flanking Ser-536/Ser-538 binds the DENN domain of connecdenn 1, whereas a phosphomimetic peptide does not. Moreover, connecdenn 1 interacts with 14-3-3 proteins, and this interaction is also disrupted by Akt inhibition and by mutation of Ser-536/Ser-538. We propose that Akt phosphorylation of connecdenn 1 downstream of insulin activation regulates connecdenn 1 function through an intramolecular interaction. 相似文献
177.
Marie C. Nyman Annele O. Sainio Mirka M. Pennanen Riikka J. Lund Sanna Vuorikoski Jari T. T. Sundstr?m Hannu T. J?rvel?inen 《The journal of histochemistry and cytochemistry》2015,63(9):710-720
Decorin is generally recognized as a tumor suppressing molecule. Nevertheless, although decorin has been shown to be differentially expressed in malignant tissues, it has often remained unclear whether, in addition to non-malignant stromal cells, cancer cells also express it. Here, we first used two publicly available databases to analyze the current information about decorin expression and immunoreactivity in normal and malignant human colorectal tissue samples. The analyses demonstrated that decorin expression and immunoreactivity may vary in cancer cells of human colorectal tissues. Therefore, we next examined decorin expression in normal, premalignant and malignant human colorectal tissues in more detail using both in situ hybridization and immunohistochemistry for decorin. Our results invariably demonstrate that malignant cells within human colorectal cancer tissues are devoid of both decorin mRNA and immunoreactivity. Identical results were obtained for cells of neuroendocrine tumors of human colon. Using RT-qPCR, we showed that human colon cancer cell lines are also decorin negative, in accordance with the above in vivo results. Finally, we demonstrate that decorin transduction of human colon cancer cell lines causes a significant reduction in their colony forming capability. Thus, strategies to develop decorin-based adjuvant therapies for human colorectal malignancies are highly rational. 相似文献
178.
Xiaoping Wu Mogens S Lund Goutam Sahana Bernt Guldbrandtsen Dongxiao Sun Qin Zhang Guosheng Su 《遗传、选种与进化》2015,47(1)
Background
The sensitivity of genome-wide association studies for the detection of quantitative trait loci (QTL) depends on the density of markers examined and the statistical models used. This study compares the performance of three marker densities to refine six previously detected QTL regions for mastitis traits: 54 k markers of a medium-density SNP (single nucleotide polymorphism) chip (MD), imputed 777 k markers of a high-density SNP chip (HD), and imputed whole-genome sequencing data (SEQ). Each dataset contained data for 4496 Danish Holstein cattle. Comparisons were performed using a linear mixed model (LM) and a Bayesian variable selection model (BVS).Results
After quality control, 587, 7825, and 78 856 SNPs in the six targeted regions remained for MD, HD, and SEQ data, respectively. In general, the association patterns between SNPs and traits were similar for the three marker densities when tested using the same statistical model. With the LM model, 120 (MD), 967 (HD), and 7209 (SEQ) SNPs were significantly associated with mastitis, whereas with the BVS model, 43 (MD), 131 (HD), and 1052 (SEQ) significant SNPs (Bayes factor > 3.2) were observed. A total of 26 (MD), 75 (HD), and 465 (SEQ) significant SNPs were identified by both models. In addition, one, 16, and 33 QTL peaks for MD, HD, and SEQ data were detected according to the QTL intensity profile of SNP bins by post-analysis of the BVS model.Conclusions
The power to detect significant associations increased with increasing marker density. The BVS model resulted in clearer boundaries between linked QTL than the LM model. Using SEQ data, the six targeted regions were refined to 33 candidate QTL regions for udder health. The comparison between these candidate QTL regions and known genes suggested that NPFFR2, SLC4A4, DCK, LIFR, and EDN3 may be considered as candidate genes for mastitis susceptibility.Electronic supplementary material
The online version of this article (doi:10.1186/s12711-015-0129-1) contains supplementary material, which is available to authorized users. 相似文献179.
Anita Skafjeld Marjolein. M. Iversen Ingar Holme Lis Ribu Kjetil Hvaal Bente. K. Kilhovd 《BMC endocrine disorders》2015,15(1):55
Background
Although monitoring foot skin temperatures has been associated with diabetic foot ulcer recurrence, no studies have been carried out to test the feasibility among European Caucasians. Moreover, the educational and/or motivational models that promote cognitive or psychosocial processes in these studies are lacking. Thus, we conducted a pilot randomized controlled trial to test the feasibility of monitoring foot skin temperatures in combination with theory-based counselling to standard foot care to reduce diabetic foot ulcer recurrence.Methods
In a single-blinded nurse-led 1-year controlled trial, conducted at a hospital setting in Norway, 41 patients with diabetic neuropathy and previous foot ulcer were randomized to the intervention (n?=?21) or control groups (n?=?20). All participants were instructed in foot care and recording observations daily. Additionally, the intervention group was taught how to monitor and record skin temperature at baseline, and received counselling every third month supporting them to use the new treatment. Subjects observing temperature differences >2.0 °C between corresponding sites on the left and right foot on two consecutive days were asked to contact the study nurse and reduce physical activity. Fisher exact test was used to evaluate the effect of the intervention on the proportion of subjects with a foot ulcer. Kaplan-Meier survival analysis was performed to compare the two groups in regard to the time to development of a foot ulcer.Results
In the intervention group, 67 % (n?=?14/21) monitored and recorded skin temperatures ≥80 % of the time while 70 % (n?=?14/20) of the controls recorded foot inspections. Foot ulcer incidence was 39 % (7/21) vs. 50 % (10/20) in the intervention and control groups, respectively (ns).Conclusions
This feasibility study showed that the addition of counselling to promote self-monitoring of skin temperature to standard care to prevent recurrence of foot ulcer is feasible in patients with diabetes in Norway. Home skin temperature monitoring was performed as frequently by the intervention group as usual foot observations in the controls despite the extra effort required. We did not detect a difference in foot ulcer recurrence between groups, but our study may inform future full scale studies.Trial registration
Clinicaltrials.gov NCT01269502180.
Thorn K Nielsen CU Jakobsen P Steffansen B Zercher CK Begtrup M 《Bioorganic & medicinal chemistry letters》2011,21(15):4597-4601
The rationale for targeting the human di-/tripeptide transporter hPEPT1 for oral drug delivery has been well established by several drug and prodrug cases. The aim of this study was to synthesize novel ketomethylene modified tripeptidomimetics and to investigate their binding affinity for hPEPT1. Three related tripeptidomimetics of the structure H-Phe-ψ[COCH2]-Ser(Bz)-Xaa-OH were synthesized applying the tandem chain extension aldol reaction, where amino acid derived β-keto imides were stereoselectively converted to α-substituted γ-keto imides. In addition, three corresponding tripeptides, composed of amide bonds, were synthesized for comparison of binding affinities. The six investigated compounds were all defined as high affinity ligands (Ki-values <0.5 mM) for hPEPT1 by measuring the concentration dependent inhibition of apical [14C]Gly-Sar uptake in Caco-2 cells. Consequently, the ketomethylene replacement for the natural amide bond and α-side chain modifications appears to offer a promising strategy to modify tripeptidic structures while maintaining a high affinity for hPEPT1. 相似文献