C16S - a Hidden Markov Model based algorithm for taxonomic classification of 16S rRNA gene sequences

Recent advances in high throughput sequencing technologies and concurrent refinements in 16S rDNA isolation techniques have facilitated the rapid extraction and sequencing of 16S rDNA content of microbial communities. The taxonomic affiliation of these 16S rDNA fragments is subsequently obtained using either BLAST-based or word frequency based approaches. However, the classification accuracy of such methods is observed to be limited in typical metagenomic scenarios, wherein a majority of organisms are hitherto unknown. In this study, we present a 16S rDNA classification algorithm, called C16S, that uses genus-specific Hidden Markov Models for taxonomic classification of 16S rDNA sequences. Results obtained using C16S have been compared with the widely used RDP classifier. The performance of C16S algorithm was observed to be consistently higher than the RDP classifier. In some scenarios, this increase in accuracy is as high as 34%. A web-server for the C16S algorithm is available at http://metagenomics.atc.tcs.com/C16S/.     

The N-terminal β-sheet of the hyperthermophilic endoglucanase from Pyrococcus horikoshii is critical for thermostability

The β-1,4-endoglucanase (EC 3.2.1.4) from the hyperthermophilic archaeon Pyrococcus horikoshii (EGPh) has strong hydrolyzing activity toward crystalline cellulose. When EGPh is used in combination with β-glucosidase (EC 3.2.1.21), cellulose is completely hydrolyzed to glucose at high temperature, suggesting great potential for EGPh in bioethanol industrial applications. The crystal structure of EGPh shows a triosephosphate isomerase (TIM) (β/α)(8)-barrel fold with an N-terminal antiparallel β-sheet at the opposite side of the active site and a very short C-terminal sequence outside of the barrel structure. We describe here the function of the peripheral sequences outside of the TIM barrel core structure. Sequential deletions were performed from both N and C termini. The activity, thermostability, and pH stability of the expressed mutants were assessed and compared to the wild-type EGPh enzyme. Our results demonstrate that the TIM barrel core is essential for enzyme activity and that the N-terminal β-sheet is critical for enzyme thermostability. Bioinformatics analyses identified potential key residues which may contribute to enzyme hyperthermostability.     

TWARIT: an extremely rapid and efficient approach for phylogenetic classification of metagenomic sequences

Phylogenetic assignment of individual sequence reads to their respective taxa, referred to as 'taxonomic binning', constitutes a key step of metagenomic analysis. Existing binning methods have limitations either with respect to time or accuracy/specificity of binning. Given these limitations, development of a method that can bin vast amounts of metagenomic sequence data in a rapid, efficient and computationally inexpensive manner can profoundly influence metagenomic analysis in computational resource poor settings. We introduce TWARIT, a hybrid binning algorithm, that employs a combination of short-read alignment and composition-based signature sorting approaches to achieve rapid binning rates without compromising on binning accuracy and specificity. TWARIT is validated with simulated and real-world metagenomes and the results demonstrate significantly lower overall binning times compared to that of existing methods. Furthermore, the binning accuracy and specificity of TWARIT are observed to be comparable/superior to them. A web server implementing TWARIT algorithm is available at http://metagenomics.atc.tcs.com/Twarit/     

Targeting the intrinsic apoptosis pathway as a strategy for melanoma therapy

Melanoma drug resistance is often attributed to abrogation of the intrinsic apoptosis pathway. Targeting regulators of apoptosis is thus considered a promising approach to sensitizing melanomas to treatment. The development of small‐molecule inhibitors that mimic natural antagonists of either antiapoptotic members of the BCL‐2 family or the inhibitor of apoptosis proteins (IAPs), known as BH3‐ or SMAC‐mimetics, respectively, are helping us to understand the mechanisms behind apoptotic resistance. Studies using BH3‐mimetics indicate that the antiapoptotic BCL‐2 protein MCL‐1 and its antagonist NOXA are particularly important regulators of BCL‐2 family signaling, while SMAC‐mimetic studies show that both XIAP and the cIAPs must be targeted to effectively induce apoptosis of cancer cells. Although most solid tumors, including melanoma, are insensitive to these mimetic drugs as single agents, combinations with other therapeutics have yielded promising results, and tests combining them with BRAF‐inhibitors, which have already revolutionized melanoma treatment, are a clear priority.     

Short homologies efficiently generate detectable homologous recombination events

When recombineering bacterial artificial chromosomes (BACs), it is common practice to design the ends of the donor molecule with 50 bp of homology specifying its insertion site. We demonstrate that desired recombinants can be produced using intermolecular homologies as short as 15 bp. Although the use of shorter donor end regions decreases total recombinants by several fold, the frequency of recombinants with correctly inserted donor molecules was high enough for easy detection by simple polymerase chain reaction (PCR) screening. This observation may have important implications for the design of oligonucleotides for recombineering, including significant cost savings, especially for high-throughput projects that use large quantities of primers.     

Predicting response to endovascular therapies: Dissecting the roles of local lesion complexity,systemic comorbidity,and clinical uncertainty

Through decades of use and refinement, endovascular stents have become part and parcel of the management of obstructive atherosclerotic lesions. Upon stent placement, a variety of biophysical reactions ensue, governed not only by the mechanical and material properties of the device, but also the impact these properties have on the local vascular biology. Anatomic changes and vascular deformations give rise to solid mechanical and fluid forces that are the proximate, functional drivers of the induced reparative response. Powerful computational tools and advanced imaging techniques allow us to define these forces with high precision and increasingly, at a patient-specific level. We have also gained fundamental insights into how these forces influence subcellular and cellular processes, and, through application of a variety of model systems, how they subsequently drive an integrated tissue response. Clinical studies extend understanding to actual patients and pathophysiologic scenarios. These tools and insights take on added weight given the real risks that accompany the many substantial benefits of stenting. Complex lesions remain difficult to manage and continue to be associated with worse outcomes. While many patients respond well to treatment, others suffer treatment failures and recurrent events – sometimes catastrophic. Overcoming such variability requires that we move towards individualized treatment plans. Doing so necessitates that we develop not just a qualitative understanding of involved phenomena, but a quantitative ability to predict integrated outcomes. Given the multi-scale nature of the vascular response to stenting, it is critical that models, be they computational, bench-top, animal, or clinical, can be verified, validated, and made interrelated. This review provides an overview of the biophysics governing endovascular stenting, their integration in real-world endovascular settings, and how simulation and statistical approaches are helping to bridge the gap between qualitative model understanding and quantitative clinical prediction.     

Phylogenetic analysis of different breeds of domestic chickens in selected area of Peninsular Malaysia inferred from partial cytochrome b gene information and RAPD markers

The present investigation was carried out in an attempt to study the phylogenetic analysis of different breeds of domestic chickens in Peninsular Malaysia inferred from partial cytochrome b gene information and random amplified polymorphic DNA (RAPD) markers. Phylogenetic analysis using both neighbor-joining (NJ) and maximum parsimony (MP) methods produced three clusters that encompassed Type-I village chickens, the red jungle fowl subspecies and the Japanese Chunky broilers. The phylogenetic analysis also revealed that majority of the Malaysian commercial chickens were randomly assembled with the Type-II village chickens. In RAPD assay, phylogenetic analysis using neighbor-joining produced six clusters that were completely distinguished based on the locality of chickens. High levels of genetic variations were observed among the village chickens, the commercial broilers, and between the commercial broilers and layer chickens. In this study, it was found that Type-I village chickens could be distinguished from the commercial chickens and Type-II village chickens at the position of the 27th nucleotide of the 351?bp cytochrome b gene. This study also revealed that RAPD markers were unable to differentiate the type of chickens, but it showed the effectiveness of RAPD in evaluating the genetic variation and the genetic relationships between chicken lines and populations.