Sociobiological Control of Plasmid Copy Number in Bacteria

All genes critical for plasmid replication regulation are located on the plasmid rather than on the host chromosome. It is possible therefore that there can be copy-up “cheater” mutants. In spite of this possibility, low copy number plasmids appear to exist stably in host populations. We examined this paradox using a multilevel selection model. Simulations showed that, a slightly higher copy number mutant could out-compete the wild type. Consequently, another mutant with still higher copy number could invade the first invader. However, the realized benefit of increasing intra-host fitness was saturating whereas that of inter-host fitness was exponential. As a result, above a threshold, intra-host selection was overcompensated by inter-host selection and the low copy number wild type plasmid could back invade a very high copy number plasmid. This led to a rock-paper-scissor (RPS) like situation that allowed the coexistence of plasmids with varied copy numbers. Furthermore, another type of cheater that had lost the genes required for conjugation but could hitchhike on a conjugal plasmid, could further reduce the advantage of copy-up mutants. These sociobiological interactions may compliment molecular mechanisms of replication regulation in stabilizing the copy numbers.     

Dataset of potential targets for Mycobacterium tuberculosis H37Rv through comparative genome analysis

Mycobacterium tuberculosis is the causative agent of the disease, tuberculosis and H37Rv is the most studied clinical strain. We use comparative genome analysis of Mycobacterium tuberculosis H37Rv and human for the identification of potential targets dataset. We used DEG (Database of Essential Genes) to identify essential genes in the H37Rv strain. The analysis shows that 628 of the 3989 genes in Mycobacterium tuberculosis H37Rv were found to be essential of which 324 genes lack similarity to the human genome. Subsequently hypothetical proteins were removed through manual curation. This further resulted in a dataset of 135 proteins with essential function and no homology to human.     

Pulmonary drug delivery and retention: A computational study to identify plausible parameters based on a coupled airway-mucus flow model

Pulmonary drug delivery systems rely on inhalation of drug-laden aerosols produced from aerosol generators such as inhalers, nebulizers etc. On deposition, the drug molecules diffuse in the mucus layer and are also subjected to mucociliary advection which transports the drugs away from the initial deposition site. The availability of the drug at a particular region of the lung is, thus, determined by a balance between these two phenomena. A mathematical analysis of drug deposition and retention in the lungs is developed through a coupled mathematical model of aerosol transport in air as well as drug molecule transport in the mucus layer. The mathematical model is solved computationally to identify suitable conditions for the transport of drug-laden aerosols to the deep lungs. This study identifies the conditions conducive for delivering drugs to the deep lungs which is crucial for achieving systemic drug delivery. The effect of different parameters on drug retention is also characterized for various regions of the lungs, which is important in determining the availability of the inhaled drugs at a target location. Our analysis confirms that drug delivery efficacy remains highest for aerosols in the size range of 1-5 μm. Moreover, it is observed that amount of drugs deposited in the deep lung increases by a factor of 2 when the breathing time period is doubled, with respect to normal breathing, suggesting breath control as a means to increase the efficacy of drug delivery to the deep lung. A higher efficacy also reduces the drug load required to be inhaled to produce the same health effects and hence, can help in minimizing the side effects of a drug.     

Transformation of animal genomics by next-generation sequencing technologies: a decade of challenges and their impact on genetic architecture

For more than a quarter of a century, sequencing technologies from Sanger’s method to next-generation high-throughput techniques have provided fascinating opportunities in the life sciences. The continuing upward trajectory of sequencing technologies will improve livestock research and expedite the development of various new genomic and technological studies with farm animals. The use of high-throughput technologies in livestock research has increased interest in metagenomics, epigenetics, genome-wide association studies, and identification of single nucleotide polymorphisms and copy number variations. Such studies are beginning to provide revolutionary insights into biological and evolutionary processes. Farm animals, such as cattle, swine, and horses, have played a dual role as economically and agriculturally important animals as well as biomedical research models. The first part of this study explores the current state of sequencing methods, many of which are already used in animal genomic studies, and the second part summarizes the state of cattle, swine, horse, and chicken genome sequencing and illustrates its achievements during the last few years. Finally, we describe several high-throughput sequencing approaches for the improved detection of known, unknown, and emerging infectious agents, leading to better diagnosis of infectious diseases. The insights from viral metagenomics and the advancement of next-generation sequencing will strongly support specific and efficient vaccine development and provide strategies for controlling infectious disease transmission among animal populations and/or between animals and humans. However, prospective sequencing technologies will require further research and in-field testing before reaching the marketplace.     

Peristaltic regimes in esophageal transport

Biomechanics and Modeling in Mechanobiology - A FLIP device gives cross-sectional area along the length of the esophagus and one pressure measurement, both as a function of time. Deducing...     

Sphingosine kinase activity confers resistance to apoptosis by fumonisin B1 in human embryonic kidney (HEK-293) cells

Fumonisin B1 induces cytotoxicity in sensitive cells by inhibiting ceramide synthase due to its structural similarity to the long-chain backbones of sphingolipids. The resulting accumulation of sphingoid bases has been established as a mechanism for fumonisin B1 cytotoxicity. We found that despite the accumulation of sphinganine, human embryonic kidney (HEK-293) cells are resistant to fumonisin B1 toxicity; 25 microM fumonisin B1 exposure for 48 h did not increase apoptosis in these cells, while it did so in sensitive porcine kidney epithelial (LLC-PK1) cells. In this study, DL-threo-dihydrosphingosine, the sphingosine kinase inhibitor (SKI), considerably increased the sensitivity of HEK-293 cells to fumonisin B1. Treatment of these cells with 25 microM fumonisin B1 and 2.5 microM SKI increased apoptosis. Sphingoid bases, sphinganine or sphingosine, added to cell cultures induced apoptosis by themselves and their effects were potentiated by SKI or fumonisin B1. Addition of physiological amounts of sphingosine-1-phosphate prevented the toxic effects induced by SKI inhibition and fumonisin B1. Results indicated that HEK-293 cells are resistant to fumonisin B1 due to rapid formation of sphingosine-1-phosphate that imparts survival properties. Taken together, these findings suggest that sphingoid base metabolism by sphingosine kinase may be a critical event in rendering the HEK-293 cells relatively resistant to fumonisin B1-induced apoptosis.     

An Approach to Identify SNPs in the Gene Encoding Acetyl-CoA Acetyltransferase-2 (ACAT-2) and Their Proposed Role in Metabolic Processes in Pig

The novel liver protein acetyl-CoA acetyltransferase-2 (ACAT2) is involved in the beta-oxidation and lipid metabolism. Its comprehensive relative expression, in silico non-synonymous single nucleotide polymorphism (nsSNP) analysis, as well as its annotation in terms of metabolic process with another protein from the same family, namely, acetyl-CoA acyltransferase-2 (ACAA2) was performed in Sus scrofa. This investigation was conducted to understand the most important nsSNPs of ACAT2 in terms of their effects on metabolic activities and protein conformation. The two most deleterious mutations at residues 122 (I to V) and 281 (R to H) were found in ACAT2. Validation of expression of genes in the laboratory also supported the idea of differential expression of ACAT2 and ACAA2 conceived through the in silico analysis. Analysis of the relative expression of ACAT2 and ACAA2 in the liver tissue of Jeju native pig showed that the former expressed significantly higher (P<0.05). Overall, the computational prediction supported by wet laboratory analysis suggests that ACAT2 might contribute more to metabolic processes than ACAA2 in swine. Further associations of SNPs in ACAT2 with production traits might guide efforts to improve growth performance in Jeju native pigs.     

Expression, purification, and characterisation of nesiritide using an E. coli expression system

Nesiritide, the recombinant human brain natriuretic peptide, is involved in the regulation of cardiovascular homeostasis and has been approved for treatment of patients with congestive heart failure. We prepared a synthetic cDNA construct of Nesiritide to generate a fusion protein with an affinity handle and 41 amino acid peptide of β-galactosidase. The fusion protein was expressed mainly in the inclusion bodies and accounted for approximately 20% of total cellular protein. After purification by Ni-IDA affinity chromatography and renaturation, the fusion protein was cleaved with purified recombinant enterokinase. Nesiritide was purified by pH precipitation/ion exchange chromatography followed by source phenyl chromatography to obtain protein with > 99% purity (determined by RPHPLC) and a mass of 3,464 Daltons. The potency (ED₅₀) of the purified protein was equivalent to that of Natrecor (Innovator formulation). Analytical methods were developed to identify oxidised, reduced and other related impurities. The expression strategy described in this work allows the convenient generation of high yield Nesiritide and enabled ease of purification.     

Mechanical properties of a bio-inspired robotic knifefish with an undulatory propulsor

South American electric knifefish are a leading model system within neurobiology. Recent efforts have focused on understanding their biomechanics and relating this to their neural processing strategies. Knifefish swim by means of an undulatory fin that runs most of the length of their body, affixed to the belly. Propelling themselves with this fin enables them to keep their body relatively straight while swimming, enabling straightforward robotic implementation with a rigid hull. In this study, we examined the basic properties of undulatory swimming through use of a robot that was similar in some key respects to the knifefish. As we varied critical fin kinematic variables such as frequency, amplitude, and wavelength of sinusoidal traveling waves, we measured the force generated by the robot when it swam against a stationary sensor, and its velocity while swimming freely within a flow tunnel system. Our results show that there is an optimal operational region in the fin's kinematic parameter space. The optimal actuation parameters found for the robotic knifefish are similar to previously observed parameters for the black ghost knifefish, Apteronotus albifrons. Finally, we used our experimental results to show how the force generated by the robotic fin can be decomposed into thrust and drag terms. Our findings are useful for future bio-inspired underwater vehicles as well as for understanding the mechanics of knifefish swimming.