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Stop codon readthrough (SCR) is the process where translation continues beyond a stop codon on an mRNA. Here, we describe a strategy to enhance or induce SCR in a transcript-selective manner using CRISPR-dCas13 system. Using specific guide RNAs, we targeted dCas13 to the downstream region of the canonical stop codons of mammalianAGO1andVEGFA,which are known to exhibit natural SCR. Results of readthrough assays revealed the enhancement of SCR of these mRNAs (both exogenous and endogenous) caused by dCas13. This effect was associated with ribosomal pausing, which has been reported in several SCR events. Furthermore, our results show that CRISPR-dCas13 can induce SCR across premature termination codons (PTC) in the mRNAs of green fluorescent protein andTP53. Finally, we demonstrate the utility of this strategy in the induction of readthrough across the thalassemia-causing PTC inHBBmRNA. Thus, CRISPR-dCas13 can be programmed to enhance or induce SCR in a transcript-selective and stop codon-specific manner.

Recognition of a stop codon by translation machinery as a sense codon results in translational readthrough instead of termination. This recoding process, termed stop codon readthrough (SCR) or translational readthrough, is found in all domains of life including mammals. The context of the stop codon, local mRNA topology, and molecules that interact with the mRNA region downstream of the stop codon determine SCR. The products of SCR can have localization, stability, and function different from those of the canonical isoforms. In this review, we discuss how recent technological and computational advances have increased our understanding of the SCR process in the mammalian system. Based on the known molecular events that occur during SCR of multiple mRNAs, we propose transient molecular roadblocks on an mRNA downstream of the stop codon as a possible mechanism for the induction of SCR. We argue, with examples, that the insights gained from the natural SCR events can guide us to develop novel strategies for the treatment of diseases caused by premature stop codons. This article is categorized under: TranslationRegulation.

Stop codon read-through (SCR) is a process of continuation of translation beyond a stop codon. This phenomenon, which occurs only in certain mRNAs under specific conditions, leads to a longer isoform with properties different from that of the canonical isoform. MTCH2, which encodes a mitochondrial protein that regulates mitochondrial metabolism, was selected as a potential read-through candidate based on evolutionary conservation observed in the proximal region of its 3′ UTR. Here, we demonstrate translational read-through across two evolutionarily conserved, in-frame stop codons of MTCH2 using luminescence- and fluorescence-based assays, and by analyzing ribosome-profiling and mass spectrometry (MS) data. This phenomenon generates two isoforms, MTCH2x and MTCH2xx (single- and double-SCR products, respectively), in addition to the canonical isoform MTCH2, from the same mRNA. Our experiments revealed that a cis-acting 12-nucleotide sequence in the proximal 3′ UTR of MTCH2 is the necessary signal for SCR. Functional characterization showed that MTCH2 and MTCH2x were localized to mitochondria with a long t1/2 (>36 h). However, MTCH2xx was found predominantly in the cytoplasm. This mislocalization and its unique C terminus led to increased degradation, as shown by greatly reduced t1/2 (

Poultry is the fastest growing livestock sector in the developing countries. The global poultry sector is expected to continue to grow, as demand for poultry meat is driven by growing needs, population, rising incomes and urbanization. The present experiment was conducted to investigate the effect of inorganic and organic trace minerals supplementation on expression of ChTLR2b gene in broilers. A total of 216 broilers randomly divided into twelve groups and each group consisting of 18 broilers in 3 replicates. T1 group was kept as control. T2, T3 and T4 group was supplemented with zinc (40 mg/kg of feed) inorganic, organic and 50% organic form respectively. T5, T6 and T7 groups was supplemented with selenium (0.3 mg/kg of feed) from inorganic, organic and 50%organic form respectively. T8, T9 and T10 groups was supplemented with chromium (2 mg/kg of feed) from inorganic, organic form and 50% organic form respectively. T11 and T12 group was supplemented with combination of all 3 minerals from inorganic and organic form respectively. RT-PCR expression analysis of ChTLR2b gene in spleen revealed that maximum up regulation (3.8413 fold) was found in T3 group (supplemented with organic Zn @ 40 mg/Kg of feed), followed by T4 (3.5325 fold) whereas in bursa of fabricius the maximum up regulation (2.8921 fold) was found in T6 group followed by T12 (2.5310 fold ) as compared to control group. Up-regulation of gene expression ChTLR2b in bursa of fabricius and spleen indicates beneficial effect of organic trace minerals in potentiation of immune system in broilers.

A stop codon ensures termination of translation at a specific position on an mRNA. Sometimes, termination fails as translation machinery recognizes a stop codon as a sense codon. This leads to stop codon readthrough (SCR) resulting in the continuation of translation beyond the stop codon, generating protein isoforms with C-terminal extension. SCR has been observed in viruses, fungi, and multicellular organisms including mammals. However, SCR is largely unexplored in plants. In this study, we have analyzed ribosome profiling datasets to identify mRNAs that undergo SCR in Arabidopsis thaliana. Analyses of the ribosome density, ribosome coverage and three-nucleotide periodicity of the ribosome profiling reads, in the mRNA region downstream of the stop codon, provided strong evidence for SCR in mRNAs of 144 genes. This process generates putative peroxisomal targeting signal, nuclear localization signal, prenylation signal, transmembrane helix and intrinsically disordered regions in the C-terminal extension of several of these proteins. Gene ontology (GO) functional enrichment analysis revealed that these 144 genes belong to three major functional groups - translation, photosynthesis and abiotic stress tolerance. Finally, using a luminescence-based assay, we experimentally demonstrate SCR in representative mRNAs belonging to these functional classes. Based on these observations, we propose that SCR plays an important role in plant physiology by regulating the protein localization and function.AUTHOR SUMMARYProtein synthesis executed by macromolecular complexes, termed ribosomes, starts and stops at specific locations on a messenger RNA (mRNA). This fidelity is critical for the normal functioning of cells. However, sometimes ribosomes don’t stop translation at the stop signal (termed stop codon) on an mRNA resulting in longer proteins with properties different from those of the canonical shorter protein. This process called stop codon readthrough (SCR) has been observed in viruses, fungi, and multicellular organisms including mammals. However, it remains largely unexplored in plants. In this study, we report evidence of SCR in 144 genes of Arabidopsis thaliana, a small flowering weed widely used as a model system to study plant biology. These genes are involved in protein synthesis, photosynthesis and stress tolerance in plants. We have also experimentally demonstrated SCR in a few genes that represent these functional classes. Our analysis shows that SCR can change the localization and functional properties of these proteins. We propose that SCR plays an important role in plant physiology.

Stop codon readthrough (SCR) is the process of continuation of translation beyond the stop codon, generating protein isoforms with C-terminal extensions. SCR has been observed in viruses, fungi, and multicellular organisms, including mammals. However, SCR is largely unexplored in plants. In this study, we have analyzed ribosome profiling datasets to identify mRNAs that exhibit SCR in Arabidopsis thaliana. Analyses of the ribosome density, ribosome coverage, and three-nucleotide periodicity of the ribosome profiling reads in the mRNA region downstream of the stop codon provided strong evidence for SCR in mRNAs of 144 genes. We show that SCR generated putative evolutionarily conserved nuclear localization signals, transmembrane helices, and intrinsically disordered regions in the C-terminal extensions of several of these proteins. Furthermore, gene ontology functional enrichment analysis revealed that these 144 genes belong to three major functional groups-translation, photosynthesis, and abiotic stress tolerance. Using a luminescence-based readthrough assay, we experimentally demonstrated SCR in representative mRNAs belonging to each of these functional classes. Finally, using microscopy, we show that the SCR product of one gene that contains a nuclear localization signal at the C-terminal extension, CURT1B, localizes to the nucleus as predicted. Based on these observations, we propose that SCR plays an important role in plant physiology by regulating protein localization and function.

In the present investigation we explore the use of organic and inorganic zinc, selenium and chromium to modulate antioxidant defense system, biochemical status and enhance production response of Broiler. Total 312 day old Cobb broiler chicks were used in the experiment. The results indicate that mean plasma superoxide dismutase, glutathione peroxidise and glucose concentration were non-significantly different in all groups. Significantly (p less than 0.05) higher mean plasma catalase concentration and TBA value was found in T13 and control group respectively. Mean plasma total protein, albumin and globulin were significantly (p less than 0.01) higher in organic Zn, Se and Cr (T13) followed by organic Cr (T10) supplemented group. Mean body weight gain, feed intake, FER and PI were significantly (p less than 0.01) higher in organic Zn, Se and Cr (T13) followed by organic Se (T7) supplemented group. In the present investigation organic Zn, Se and Cr supplementation showed systemic effects with a better biochemical status, which could be translated into better production performance and lower mortality.

Translational readthrough generates proteins with extended C‐termini, which often possess distinct properties. Here, we have used various reporter assays to demonstrate translational readthrough of AGO1 mRNA. Analysis of ribosome profiling data and mass spectrometry data provided additional evidence for translational readthrough of AGO1. The endogenous readthrough product, Ago1x, could be detected by a specific antibody both in vitro and in vivo. This readthrough process is directed by a cis sequence downstream of the canonical AGO1 stop codon, which is sufficient to drive readthrough even in a heterologous context. This cis sequence has a let‐7a miRNA‐binding site, and readthrough is promoted by let‐7a miRNA. Interestingly, Ago1x can load miRNAs on target mRNAs without causing post‐transcriptional gene silencing, due to its inability to interact with GW182. Because of these properties, Ago1x can serve as a competitive inhibitor of miRNA pathway. In support of this, we observed increased global translation in cells overexpressing Ago1x. Overall, our results reveal a negative feedback loop in the miRNA pathway mediated by the translational readthrough product of AGO1.

The present investigation was carried out with the objective to study the productive and reproductive performances of dairy cattle under organic farming systems. Total 24 medium to high yielding cattle has been selected for the study, out of which 12 were reared under organic management system and 12 under conventional management system. For cows maintained under organic management system, the animals were fed ad lib quality green fodder grown organically and limited quantity (1.5 to 2.0 kg/cows/day) of concentrate mixture. Production and reproduction performance of animals of both the groups were compared. The results revealed that the reproductive performance was better in cows maintained under organic production management system than their conventional counterparts. The study on milk quality in terms of composition revealed significantly (P less than 0.05) higher fat % of milk, while lower (P less than 0.05) protein % of milk for the cows maintained under organic system as compared to those under conventional system. It can be concluded that the organic management system of dairy cows can be beneficial as depicted by better reproductive efficiency of cows with higher fat % of the milk.

In cyanobacterium Anabaena 7120, iron deficiency leads to oxidative stress with unavoidable consequences. Nitric oxide reduces pigment damage and supported the growth of Anabaena 7120 in iron-deficient conditions. Elevation in nitric oxide accumulation and reduced superoxide radical production justified the role of nitric oxide in alleviating oxidative stress in iron deficiency. Increased activities of antioxidative enzymes and higher levels of ROS scavengers (ascorbate, glutathione and thiol) in iron deficiency were also observed in the presence of nitric oxide. Nitric oxide also supported the membrane integrity of Anabaena cells and reduces protein and DNA damage caused by oxidative stress induced by iron deficiency. Results suggested that nitric oxide alleviates the damaging effects of oxidative stress induced by iron deficiency in cyanobacterium Anabaena 7120.

The efficacy of dihydroxamate siderophore in attenuation/mitigation of cadmium toxicity, an optimum level of cadmium under which cyanobacterium can grow and produce siderophore, complexation behaviour of cadmium and siderophore and kinetics of complexation along with in silico analysis have been investigated in paddy field cyanobacterium Anabaena oryzae under iron replete and deplete conditions. Results showed that cadmium concentration as 1.0 μM is optimum for growth as well as siderophore production in iron replete and deplete culture conditions. The complexation between cadmium–siderophore was evident in spectral scan. Kinetic studies as well as in silico docking analysis indicate that like iron, cadmium also forms a thermodynamically stable complex with siderophore. Thus our results suggest that the siderophore produced by Anabaena oryzae can act as a biological sequestering agent for the mitigation of cadmium metal ions from the paddy fields to improve crop productivity.

To investigate the physiology and taxonomic composition of the key players of nitrification and denitrification processes in paddy fields, culture dependent and independent studies have been carried out. A total of 28 bacterial strains have been screened in which six were capable of reducing nitrate and nitrite as well as having significant ammonium removal potential. 16S rRNA-PCR-DGGE-based molecular typing of enriched batch culture was done with time duration to explore and identify dominant and stable soil denitrifiers. Notably, three isolates namely PDN3, PDN19, PDN14 were found to be efficiently involved in the removal of 70.32, 71.46, and 81.50% of NH4 (+) and showed closest similarity (>98%) with Bacillus cereus, Bacillus subtilis, and Pseudomonas aeruginosa strains, respectively. The bacterial strain PDN14 showed maximum growth with highest ammonium removal rate (2.78 gN/(m(3) ·h) has also been characterized based on nosZ gene which showed similarity to uncultured γ- Proteobacteria, P. aeruginosa sp. B3. Median joining (MJ) network and rRNA secondary structure have been analyzed for their detailed taxonomic diversity and derived haplotype-based co-occurrence. Results demonstrated that such strains can serve as good candidate for in situ nitrogen transformation in paddy soils and improvingly characterized by physiological and detailed phylogenetic approaches.

16S rRNA and nifD-nifK sequences were used to study the molecular phylogeny and evolutionary genetics of Frankia strains isolated from Hippophae salicifolia D. Don growing at different altitudes (ecologically classified as riverside and hillside isolates) of the Eastern Himalayan region of North Sikkim, India. Genetic information for the small subunit rRNA (16S rRNA) revealed that the riverside Frankia isolates markedly differed from the hillside isolates suggesting that the riverside isolates are genetically compact. Further, for enhanced resolutions, the partial sequence of nifD (3' end), nifK (5' end) and nifD-K IGS region have been investigated. The sequences obtained, failed to separate riverside isolates and hillside isolates, thus suggesting a possible role of genetic transfer events either from hillside to riverside or vice versa. The evolutionary genetic analyses using evogenomic extrapolations of gene sequence data obtained from 16S rRNA and nifD-K provided differing equations with the pace of evolution being more appropriately, intermediate. Values of recombination frequency (R), nucleotide diversity per site (Pi), and DNA divergence estimates supported the existence of an intermixed zone where spatial isolations occurred in sync with the temporal estimates. J. Basic Microbiol. 2015, 54, 1-9.

Siderophore production in Anabaena oryzae was investigated under the influence of various levels of iron and other abiotic factors such as pH, temperature, light and different nitrogen sources. Optimization of culture conditions under controlled mechanisms of these abiotic factors lead to the siderophore production in significant amount. Under iron-starved condition, A. oryzae extracellularly releases 89.17 % hydroxymate-type siderophore. Slightly alkaline pH and 30 °C temperature was found stimulatory for the cyanobacterial growth and siderophorogenesis (88.52 % SU and 83.87 % SU, respectively). Excess iron loading had a negative impact on siderophore production along with the alterations in the morphology and growth. Further, scanning electron microphotographs signified that higher concentrations of iron lead to complete damage of the cells and alterations in membrane proteins possibly transporters responsible for exchange of siderophore complex from environment to the cell. SDS-PAGE analysis of whole cell proteins showed overexpression of low molecular weight proteins ranges between 20.1 to 29.0 kDa up to 100-μM iron concentrations. These polypeptides/proteins might be involved in maintaining iron homeostasis by regulating siderophore production. Results suggest that lower concentrations of iron ≤50 μM along with other abiotic factors are stimulatory, whereas higher concentrations (>50 μM) are toxic. Data further suggested that cyanobacterium A. oryzae can serve as a potential biofertilizer especially in iron-rich soil through sequestration by the power of natural Fe(III)-siderophore complex formation.

Iron deficiency ends up into several unavoidable consequences including damaging oxidative stress in cyanobacteria. NtcA is a global nitrogen regulator controls wide range of metabolisms in addition to regulation of nitrogen metabolism. In present communication, NtcA based regulation of iron homeostasis, ROS production and cellular phenotype under iron deficiency in Anabaena 7120 has been investigated. NtcA regulates the concentration dependent iron uptake by controlling the expression of furA gene. NtcA also regulated pigment synthesis and phenotypic alterations in Anabaena 7120. A significant increase in ROS production and corresponding reduction in the activities of antioxidative enzymes (SOD, CAT, APX and GR) in CSE2 mutant strain in contrast to wild type Anabaena 7120 also suggested the possible involvement of NtcA in protection against oxidative stress in iron deficiency. NtcA has no impact on the expression of furB and furC in spite of presence of consensus NtcA binding site (NBS) and -10 boxes in their promoter. NtcA also regulates the thylakoid arrangement as well as related photosynthetic and respiration rates under iron deficiency in Anabaena 7120. Overall results suggested that NtcA regulates iron acquisition and in turn protect Anabaena cells from the damaging effects of oxidative stress induced under iron deficiency.

An appreciation of comparative microbial survival is most easily done while evaluating their adaptive strategies during stress. In the present experiment, antioxidative and whole cell proteome variations based on spectrophotometric analysis and SDS-PAGE and 2-dimensional gel electrophoresis have been analysed among salt-tolerant and salt-sensitive Frankia strains. This is the first report of proteomic basis underlying salt tolerance in these newly isolated Frankia strains from Hippophae salicifolia D. Don. Salt-tolerant strain HsIi10 shows higher increment in the contents of superoxide dismutase, catalase and ascorbate peroxidase as compared to salt-sensitive strain HsIi8. Differential 2-DGE profile has revealed differential profiles for salt-tolerant and salt-sensitive strains. Proteomic confirmation of salt tolerance in the strains with inbuilt efficiency of thriving in nitrogen-deficient locales is a definite advantage for these microbes. This would be equally beneficial for improvement of soil nitrogen status. Efficient protein regulation in HsIi10 suggests further exploration for its potential use as biofertilizer in saline soils.

To decipher an evolutionary lineage between two different but important bacterial groups, i.e., Pseudomonas strain (γ-Proteobacteria) and Frankia strain (actinobacteria) growing in the same ecological niche in and around of an actinorhizal plant Hippophae salicifolia D. Don, genetic diversity and comparative molecular phylogeny have been investigated using 16S rRNA gene sequences and computer-simulated and virtually directed restriction fragment length polymorphism (RFLP) through 10 restriction enzymes. Bayesian and coalescent analyses on the basis of 16S rRNA gene sequences suggested three major groups with close proximity between Pseudomonas and Frankia isolates. This result has been further validated based on the data observed through similarity coefficient value and computational RFLP. Principal component analysis and Mandel h and k statistical analysis also confirmed and strengthen the findings. Approximately 458 aligned sequence of all the taxa were used to decipher nucleotide diversity, polymorphism and gene flow between these taxa. Thus, our results suggest for a possible co-evolution or a heterologous gene transfer of distantly related microbial forms. Further, our study also advocate for the use of computer aided, virtual RFLP analysis as a cost effective and rapid identification tool. Electronic supplementary material The online version of this article (doi:10.1007/s13205-016-0488-5) contains supplementary material, which is available to authorized users.

In order to investigate the role of frankial siderophores in minimizing metal toxicity, Frankia strains were grown at different concentrations of Mg2+, Cu2+, and Zn2+. Growth was observed up to 500 µmol L−1 Mg2+, 10 µmol L−1 Cu2+, and 10 µmol L−1 Zn2+ while the maxima were at 200 µmol L−1 Mg2+, 1 µmol L−1 Cu2+, and 1 µmol L−1 Zn2+. The siderophore production was increased up to 500 µmol L−1 Mg2+, 10 µmol L−1 Cu2+, 100 µmol L−1 (hydroxamate type), and 200 µmol L−1 (catecholate type) Zn2+ while maximum production was found at Mg2+ (200 µmol L−1), Cu2+ (10 µmol L−1), and Zn2+ (10 µmol L−1). The results suggested that the growth was protected at higher concentrations of Mg2+ (up to 500 µmol L−1), Cu2+ (10 µmol L−1), and Zn2+ (10 µmol L−1), possibly due to enhanced siderophore production. Thus, siderophores minimize the metal-induced inhibition of growth in Frankia, likely due to regulation of nutritional imbalances and metabolic processes during adaptation towards metal stress and/or metal toxicity.

Spatiotemporally controlled active manipulation of external micro-/nanoprobes inside living cells can lead to development of innovative biomedical technologies and inspire fundamental studies of various biophysical phenomena. Examples include gene silencing applications, real-time mechanical mapping of the intracellular environment, studying cellular response to local stress, and many more. Here, for the first time, cellular internalization and subsequent intracellular manipulation of a system of helical nanomotors driven by small rotating magnetic fields with no adverse effect on the cellular viability are demonstrated. This remote method of fuelling and guidance limits the effect of mechanical transduction to cells containing external probes, in contrast to ultrasonically or chemically powered techniques that perturb the entire experimental volume. The investigation comprises three cell types, containing both cancerous and noncancerous types, and is aimed toward analyzing and engineering the motion of helical propellers through the crowded intracellular space. The studies provide evidence for the strong anisotropy, heterogeneity, and spatiotemporal variability of the cellular interior, and confirm the suitability of helical magnetic nanoprobes as a promising tool for future cellular investigations and applications.

Different Frankia strains (HsIi2, HsIi4, HsIi5, HsIi8, HsIi9, HsIi10, HsIi11, HsIi12, HsIi13, HsIi14) nodulating Hippophae salicifolia D. Don, were characterized on the basis of physiological, biochemical and molecular attributes. Results suggest that the physiological approaches i.e., nitrogenase activity, glutamine synthetase (GS) activity and ammonia excretion are strain specific. The highest rate of nitrogen fixation and maximum production of ammonia with low GS makes the strain HsIi11, a suitable biofertilizer as compared to other strains. Analysis of total protein pattern (SDS–PAGE) revealed that the most closely related strains HsIi10 and HsIi4 were found to be most distantly related to the most similar strains HsIi14, HsIi5, HsIi13, HsIi11 and HsIi12. RAPD PCR analyses with an arbitrary primer 1253 produced distinct, unique and specific DNA fingerprints for each of the Frankia strain and 100% polymorphism was observed which uncovers the genetic diversity. These approaches might be helpful in rapid identification, in designing the marker for the specific strains as well as in improving nitrogen fixation in agroforestry.

To investigate the physiology and taxonomic composition of the key players of nitrification and denitrification processes in paddy fields, culture dependent and independent studies have been carried out. A total of 28 bacterial strains have been screened in which six were capable of reducing nitrate and nitrite as well as having significant ammonium removal potential. 16S rRNA-PCR-DGGE-based molecular typing of enriched batch culture was done with time duration to explore and identify dominant and stable soil denitrifiers. Notably, three isolates namely PDN3, PDN19, PDN14 were found to be efficiently involved in the removal of 70.32, 71.46, and 81.50% of NH4 (+) and showed closest similarity (98%) with Bacillus cereus, Bacillus subtilis, and Pseudomonas aeruginosa strains, respectively. The bacterial strain PDN14 showed maximum growth with highest ammonium removal rate (2.78 gN/(m(3) ·h) has also been characterized based on nosZ gene which showed similarity to uncultured γ- Proteobacteria, P. aeruginosa sp. B3. Median joining (MJ) network and rRNA secondary structure have been analyzed for their detailed taxonomic diversity and derived haplotype-based co-occurrence. Results demonstrated that such strains can serve as good candidate for in situ nitrogen transformation in paddy soils and improvingly characterized by physiological and detailed phylogenetic approaches.

Three cyanobacterial strains originating from different habitats were subjected to temperature shift exposures and monitored for levels of proline, thiol and activity of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Thermophile Mastigocladus laminosus (growth optimum, 40 °C), raised the proline level 4.2-fold at low temperature (20 °C), for the psychrophile Nostoc 593 (growth optimum, 20 °C), it was raised 8-fold at 40 °C while in the mesophile Nostoc muscorum (growth optimum, 30 °C), the imino acid level increased 2.3-fold during temperature ‘shiftdown’ to 20 °C or 3.5-fold in sets facing ‘shiftup’ (40 °C). Alterations in thiol levels in the above strains were in line with proline. It is suggested that such fluctuations reflect metabolic shifts as a response to stress. Interestingly, GAPDH activity was maximum at the respective growth temperature optimum of M. laminosus (122 nmol NADPH oxidized min −1 mg −1 protein) and Nostoc 593 (141 nmol NADPH oxidized min −1 mg −1 protein) while in N. muscorum, it increased at 40 °C (101 nmol NADPH oxidized min −1 mg −1 protein) and to 93.3 nmol NADPH oxidized min −1 mg −1 protein (20 °C) relative to 86 nmol NADPH oxidized min −1 mg −1 protein at 30 °C. It seems that extremophiles maintain the GAPDH activity/level during growth at their respective temperatures optimal while the mesophile increases it in order to cope up with temperature-stress.

16S rRNA and nifD-nifK sequences were used to study the molecular phylogeny and evolutionary genetics of Frankia strains isolated from Hippöphae salicifolia D. Don growing at different altitudes (ecologically classified as riverside and hillside isolates) of the Eastern Himalayan region of North Sikkim, India. Genetic information for the small subunit rRNA (16S rRNA) revealed that the riverside Frankia isolates markedly differed from the hillside isolates suggesting that the riverside isolates are genetically compact. Further, for enhanced resolutions, the partial sequence of nifD (3' end), nifK (5' end) and nifD-K IGS region have been investigated. The sequences obtained, failed to separate riverside isolates and hillside isolates, thus suggesting a possible role of genetic transfer events either from hillside to riverside or vice versa. The evolutionary genetic analyses using evogenomic extrapolations of gene sequence data obtained from 16S rRNA and nifD-K provided differing equations with the pace of evolution being more appropriately, intermediate. Values of recombination frequency (R), nucleotide diversity per site (Pi), and DNA divergence estimates supported the existence of an intermixed zone where spatial isolations occurred in sync with the temporal estimates. J. Basic Microbiol. 2015, 54, 1-9.

Studies have been conducted on the enzymic activity of Baker's yeast and also of Brewer's yeast entrapped into the reversed micelles formed by cetyl pyridinium chloride (CPC1) in n-hexane. The activities of alpha-amylase and invertase enzymes in the entrapped cells have been estimated and compared with those in the control experiments where there was no entrapment. The following significant observations have been made: 1. except for invertase, enzymes in Brewer's yeast, the entrapped yeast cells showed enhanced enzymic activities; 2. when the yeast cells were entrapped inside the reversed micelles along with substrates of the two enzymes, alpha-amylase, and invertase, the activity of each of these enzymes showed a further enhancement in comparison to that showed in the experiments in which substrates of the individual enzymes alone were entrapped-the phenomenon of synergism; 3. when the yeast cells and the respective substrates were entrapped inside separate reversed micelles and the solutions containing entrapped cells and entrapped substrates were mixed, the activities of the individual enzymes, alpha-amylase and invertase, showed further enhancement in comparison to the case in which the cells and the substrates were entrapped inside the same reversed micelle (in this case also the phenomenon of synergism was observed); and (4) In the case of experiments in which there was no entrapment, it was observed that the presence of substrates induced more release of enzymes from the yeast cells. These observations on yeast cells, which to the best of our knowledge have not been reported before, should be biotechnologically relevant.

Takotsubo cardiomyopathy ("broken heart") exhibits a highly possible link between acute emotional stress and the onset of left ventricular dysfunction. This article describes a case report of takotsubo cardiomyopathy in an 89-year-old female; the patient is significantly older than the median age for this condition, which ranges from 63 to 76 years of age. The exact mechanism of this condition is unclear and there are several hypotheses under investigation. Several studies have shown a link between takotsubo cardiomyopathy and elevated catecholamines. Similarly, other studies have documented emotional and physical stress to be the underlying pathophysiology of this condition. Another case study has proposed a novel hypothesis of a link between septal thickening in cases of takotsubo cardiomyopathy. This article is an interesting case report of an elderly patient with takotsubo cardiomyopathy.

In this paper, we propose a novel scheme to generate sinusoidal pulse width modulated (SPWM) switching pulses using Direct Modulation Strategy and its implementation has been done through software developed using VHDL code. In the given proposed scheme, the periodic PWM signals are generated, separated by positive and negative group of polarity control switching pulse and evoked at the output as a control signal for FPGA interfaced H bridge inverter circuit.