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971.
972.
Jason R. Andrews Krishna G. Prajapati Elizabeth Eypper Poojan Shrestha Mila Shakya Kamal R. Pathak Niva Joshi Priyanka Tiwari Manisha Risal Samir Koirala Abhilasha Karkey Sabina Dongol Shawn Wen Amy B. Smith Duncan Maru Buddha Basnyat Stephen Baker Jeremy Farrar Edward T. Ryan Elizabeth Hohmann Amit Arjyal 《PLoS neglected tropical diseases》2013,7(6)
Background
In many rural areas at risk for enteric fever, there are few data on Salmonella enterica serotypes Typhi (S. Typhi) and Paratyphi (S. Paratyphi) incidence, due to limited laboratory capacity for microbiologic culture. Here, we describe an approach that permits recovery of the causative agents of enteric fever in such settings. This approach involves the use of an electricity-free incubator based upon use of phase-change materials. We compared this against conventional blood culture for detection of typhoidal Salmonella.Methodology/Principal Findings
Three hundred and four patients with undifferentiated fever attending the outpatient and emergency departments of a public hospital in the Kathmandu Valley of Nepal were recruited. Conventional blood culture was compared against an electricity-free culture approach. Blood from 66 (21.7%) patients tested positive for a Gram-negative bacterium by at least one of the two methods. Sixty-five (21.4%) patients tested blood culture positive for S. Typhi (30; 9.9%) or S. Paratyphi A (35; 11.5%). From the 65 individuals with culture-confirmed enteric fever, 55 (84.6%) were identified by the conventional blood culture and 60 (92.3%) were identified by the experimental method. Median time-to-positivity was 2 days for both procedures. The experimental approach was falsely positive due to probable skin contaminants in 2 of 239 individuals (0.8%). The percentages of positive and negative agreement for diagnosis of enteric fever were 90.9% (95% CI: 80.0%–97.0%) and 96.0% (92.7%–98.1%), respectively. After initial incubation, Salmonella isolates could be readily recovered from blood culture bottles maintained at room temperature for six months.Conclusions/Significance
A simple culture approach based upon a phase-change incubator can be used to isolate agents of enteric fever. This approach could be used as a surveillance tool to assess incidence and drug resistance of the etiologic agents of enteric fever in settings without reliable local access to electricity or local diagnostic microbiology laboratories. 相似文献973.
Rama Krishna Gudiminchi Martina Geier Anton Glieder Dr. Andrea Camattari 《Biotechnology journal》2013,8(1):146-152
Cytochrome P450 (CYP) enzymes are useful biocatalysts for the pharmaceutical and biotechnological industries. A high-throughput method for quantification of CYP expression in yeast is needed in order to fully exploit the yeast expression system. Carbon monoxide (CO) difference spectra of whole cells have been successfully used for the quantification of heterologous CYP expressed in Escherichia coli in the 96-well format; however, very few researchers have shown whole-cell CO difference spectra with yeast cells using 1-cm path length. Spectral interference from the native hemoproteins often obscures the P450 peak, challenging functional CYP quantification in whole yeast cells. For the first time, we describe the high-throughput determination of CO difference spectra using whole cells in the 96-well format for the quantification of CYP genes expressed in Pichia pastoris. Very little interference from the hemoproteins of P. pastoris enabled CYP quantification even at relatively low expression levels. P. pastoris strains carrying a single copy or three copies of both hCPR and CYP2D6 integrated into the chromosomal DNA were used to establish the method in 96-well format, allowing to detect quantities of CYP2D6 as low as 6 nmol gCDW–1 and 12 pmol per well. Finally, the established method was successfully demonstrated and used to screen P. pastoris clones expressing Candida CYP52A13. 相似文献
974.
975.
976.
Gonghao Wang Wenbin Mao Rebecca Byler Krishna Patel Caitlin Henegar Alexander Alexeev Todd Sulchek 《PloS one》2013,8(10)
Abnormal cell mechanical stiffness can point to the development of various diseases including cancers and infections. We report a new microfluidic technique for continuous cell separation utilizing variation in cell stiffness. We use a microfluidic channel decorated by periodic diagonal ridges that compress the flowing cells in rapid succession. The compression in combination with secondary flows in the ridged microfluidic channel translates each cell perpendicular to the channel axis in proportion to its stiffness. We demonstrate the physical principle of the cell sorting mechanism and show that our microfluidic approach can be effectively used to separate a variety of cell types which are similar in size but of different stiffnesses, spanning a range from 210 Pa to 23 kPa. Atomic force microscopy is used to directly measure the stiffness of the separated cells and we found that the trajectories in the microchannel correlated to stiffness. We have demonstrated that the current processing throughput is 250 cells per second. This microfluidic separation technique opens new ways for conducting rapid and low-cost cell analysis and disease diagnostics through biophysical markers. 相似文献
977.
Zhifu Sun Yan W. Asmann Asha Nair Yuji Zhang Liguo Wang Krishna R. Kalari Aditya V. Bhagwate Tiffany R. Baker Jennifer M. Carr Jean-Pierre A. Kocher Edith A. Perez E. Aubrey Thompson 《PloS one》2013,8(8)
Objectives
The sequencing by the PolyA selection is the most common approach for library preparation. With limited amount or degraded RNA, alternative protocols such as the NuGEN have been developed. However, it is not yet clear how the different library preparations affect the downstream analyses of the broad applications of RNA sequencing.Methods and Materials
Eight human mammary epithelial cell (HMEC) lines with high quality RNA were sequenced by Illumina’s mRNA-Seq PolyA selection and NuGEN ENCORE library preparation. The following analyses and comparisons were conducted: 1) the numbers of genes captured by each protocol; 2) the impact of protocols on differentially expressed gene detection between biological replicates; 3) expressed single nucleotide variant (SNV) detection; 4) non-coding RNAs, particularly lincRNA detection; and 5) intragenic gene expression.Results
Sequences from the NuGEN protocol had lower (75%) alignment rate than the PolyA (over 90%). The NuGEN protocol detected fewer genes (12–20% less) with a significant portion of reads mapped to non-coding regions. A large number of genes were differentially detected between the two protocols. About 17–20% of the differentially expressed genes between biological replicates were commonly detected between the two protocols. Significantly higher numbers of SNVs (5–6 times) were detected in the NuGEN samples, which were largely from intragenic and intergenic regions. The NuGEN captured fewer exons (25% less) and had higher base level coverage variance. While 6.3% of reads were mapped to intragenic regions in the PolyA samples, the percentages were much higher (20–25%) for the NuGEN samples. The NuGEN protocol did not detect more known non-coding RNAs such as lincRNAs, but targeted small and “novel” lincRNAs.Conclusion
Different library preparations can have significant impacts on downstream analysis and interpretation of RNA-seq data. The NuGEN provides an alternative for limited or degraded RNA but it has limitations for some RNA-seq applications. 相似文献978.
Tatiana Lemos Bisi Paulo Renato Dorneles José Lailson-Brito Gilles Lepoint Alexandre de Freitas Azevedo Leonardo Flach Olaf Malm Krishna Das 《PloS one》2013,8(12)
To investigate the foraging habitats of delphinids in southeastern Brazil, we analyzed stable carbon (δ13C) and nitrogen (δ15N) isotopes in muscle samples of the following 10 delphinid species: Sotalia guianensis, Stenella frontalis, Tursiops truncatus, Steno bredanensis, Pseudorca crassidens, Delphinus sp., Lagenodelphis hosei, Stenella attenuata, Stenella longirostris and Grampus griseus. We also compared the δ13C and δ15N values among four populations of S. guianensis. Variation in carbon isotope results from coast to ocean indicated that there was a significant decrease in δ13C values from estuarine dolphins to oceanic species. S. guianensis from Guanabara Bay had the highest mean δ13C value, while oceanic species showed significantly lower δ13C values. The highest δ15N values were observed for P. crassidens and T. truncatus, suggesting that these species occupy the highest trophic position among the delphinids studied here. The oceanic species S. attenuata, G. griseus and L. hosei had the lowest δ15N values. Stable isotope analysis showed that the three populations of S. guianensis in coastal bays had different δ13C values, but similar δ15N results. Guiana dolphins from Sepetiba and Ilha Grande bays had different foraging habitat, with specimens from Ilha Grande showing more negative δ13C values. This study provides further information on the feeding ecology of delphinids occurring in southeastern Brazil, with evidence of distinctive foraging habitats and the occupation of different ecological niches by these species in the study area. 相似文献
979.
Panwen Wang Wing-Fu Lai Mulin Jun Li Feng Xu Hari Krishna Yalamanchili Robin Lovell-Badge Junwen Wang 《PloS one》2013,8(10)
One of the fundamental goals of genetics is to understand gene functions and their associated phenotypes. To achieve this goal, in this study we developed a computational algorithm that uses orthology and protein-protein interaction information to infer gene-phenotype associations for multiple species. Furthermore, we developed a web server that provides genome-wide phenotype inference for six species: fly, human, mouse, worm, yeast, and zebrafish. We evaluated our inference method by comparing the inferred results with known gene-phenotype associations. The high Area Under the Curve values suggest a significant performance of our method. By applying our method to two human representative diseases, Type 2 Diabetes and Breast Cancer, we demonstrated that our method is able to identify related Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes pathways. The web server can be used to infer functions and putative phenotypes of a gene along with the candidate genes of a phenotype, and thus aids in disease candidate gene discovery. Our web server is available at http://jjwanglab.org/PhenoPPIOrth. 相似文献
980.
Shashi Bhushan Lal Pankaj Kumar Pandey Punit K Rai Anil Rai Anu Sharma Krishna Kumar Chaturvedi 《Bioinformation》2013,9(11):588-598
The application of novel and modern techniques in genetic engineering and genomics has resulted in information explosion in
genomics. Three major genome databases under International Nucleotide Sequence Database collaboration NCBI, DDBJ and EMBL
have been providing a convenient platform for submission of sequences which they share among themselves. Many institutes in
India under Indian Council of Agricultural Research have scientists working on biotechnology and bioinformatics research. The
various studies conducted by them, generate massive data related to biological information of plants, animals, insects, microbes
and fisheries. These scientists are dependent on NCBI, EMBL, DDBJ and other portals for their sequence submissions, analysis and
other data mining tasks. Due to various limitations imposed on these sites and the poor connectivity problem prevents them to
conduct their studies on these open domain databases. The valued information generated by them needs to be shared by the
scientific communities to eliminate the duplication of efforts and expedite their knowledge extended towards new findings. A
secured common submission portal system with user-friendly interfaces, integrated help and error checking facilities has been
developed in such a way that the database at the backend consists of a union of the items available on the above mentioned
databases. Standard database management concepts have been employed for their systematic storage management. Extensive
hardware resources in the form of high performance computing facility are being installed for deployment of this portal.