Strain-specific salt tolerance and osmoregulatory mechanisms in Azospirillum brasilense

Salinity stress inhibits the growth and nitrogen fixation ability of the plant growth-promoting rhizobacterium Azospirillum brasilense. Five strains of A. brasilense were isolated from the rhizosphere of Indian cereals and grasses and identified on the basis of their phenotypic features and 16S rRNA gene sequence. The five Indian isolates and two standard strains of A. brasilense, Sp7 and Cd, showed notable differences in growth, acetylene-reducing activity under salt stress, and ability to take up and use glycine betaine for the restoration of growth and acetylene-reducing activity under salt stress. Salt stress also enhanced the production of exopolysaccharides and cell aggregates, the extent of which varied in different strains of A. brasilense at different carbon to nitrogen ratios in the culture medium. It can be concluded that the production of exopolysaccharides and cell aggregates is a more consistent physiological response of A. brasilense to salt stress than is the uptake and osmoprotection by glycine betaine.     

Novel approaches on identification of conserved miRNAs for broad-spectrum Potyvirus control measures

Molecular Biology Reports - Potyviridae comprises more than 200 ssRNA viruses, many of which have a broad host range and geographical distributions. Potyvirids (members of Potyviridae) infect...     

Data on vildagliptin and vildagliptin plus metformin combination in type-2 diabetes mellitus management

It is of interest to evaluate the clinical effectiveness and safety of vildagliptin as monotherapy and combination therapy of vildagliptin and metformin for the management of type 2 diabetes mellitus (T2DM) patients in Indian settings. The study included patients with T2DM (aged >18 years) receiving vildagliptin monotherapy and vildagliptin in combination with metformin therapy of various strengths. Data related to demographics, risk factors, medical history, glycated hemoglobin (HbA1c) levels, and medical therapies were retrieved from medical records. Out of 9678 patients (median age, 52.0 years), 59.1% were men. A combination of vildagliptin and metformin (50/500 mg) was the most commonly used therapy (54.8%), and the median duration of therapy was 24.0 months. The predominant reason for selecting vildagliptin therapy was to improve HbA1c levels (87.8%). A total of 87.5% of patients required dosage up-titration. Vildagliptin therapy was used in patients with T2DM and associated complications (peripheral neuropathy, CAD, nephropathy, retinopathy, autonomous neuropathy, stroke/TIA, and peripheral artery disease). Among 5175 patients who experienced body weight changes, a majority of patients showed a loss of weight (68.6%). The target glycemic control was achieved in 95.3% of patients. The mean HbA1c levels were significantly decreased post-treatment (mean change: 1.34%; p<0.001). Adverse events were reported in 0.4% of patients. Physicians rated the majority of patients as good to excellent on the global evaluation of efficacy and tolerability scale (98.9%, each). Vildagliptin as monotherapy and combination therapy of vildagliptin and metformin was an effective therapy in reducing HbA1c helps in achieving target glycemic control, and was well tolerated in Indian patients with T2DM continuum.     

High Resolution Magnetic Resonance Imaging for Characterization of the Neuroligin-3 Knock-in Mouse Model Associated with Autism Spectrum Disorder

Autism spectrum disorders (ASD) comprise an etiologically heterogeneous set of neurodevelopmental disorders. Neuroligin-3 (NL-3) is a cell adhesion protein that mediates synapse development and has been implicated in ASD. We performed ex-vivo high resolution magnetic resonance imaging (MRI), including diffusion tensor imaging (DTI) and behavioral (social approach and zero maze) tests at 3 different time points (30, 50 and 70 days-of-age) on NL-3 and wild-type littermates to assess developmental brain abnormalities in NL-3 mice. MRI data were segmented in 39 different gray and white matter regions. Volumetric measurements, along with DTI indices from these segmented regions were also performed. After controlling for age and gender, the NL-3 knock-in animals demonstrated significantly reduced sociability and lower anxiety-related behavior in comparison to their wild type littermates. Significantly reduced volume of several white and gray matter regions in the NL-3 knock-in mice were also observed after considering age, gender and time point as covariates. These findings suggest that structural changes in the brain of NL-3 mice are induced by the mutation in the NL-3 gene. No significant differences in DTI indices were observed, which suggests that the NL-3 mutation may not have a profound effect on water diffusion as detected by DTI. The volumetric and DTI studies aid in understanding the biology of disrupting function on an ASD risk model and may assist in the development of imaging biomarkers for ASD.     

Synthesis of new 2,5-disubstituted-1,3,4-thiadiazole derivatives and their in vivo anticonvulsant activity

A series of 2,5-disubstituted-1,3,4-thiadiazole derivatives were synthesized by the reaction of 3-(2-cyanopropan-2-yl)-N-(5-(piperazine-1-yl)-1,3,4-thiadiazol-2-yl)benzamide with various sulfonyl chlorides and evaluated for their anticonvulsant activity in MES test. Rotorod method was employed to determine the neurotoxicity. The purity of the compounds is confirmed on the basis of their elemental analysis. The structures of all the new compounds are established on the basis of ¹H NMR and mass spectral data. Out of fifteen compounds, three were found to be potent anticolvunstants. The same compounds showed no neurotoxicity at the maximum dose administered (100 mg/kg).     

Functional Characterization of Recombinant Bromelain of Ananas comosus Expressed in a Prokaryotic System

Bromelain (BRM) is a defense protein present in the fruit and stem of pineapple (Ananas comosus) and it is grouped as a cysteine protease enzyme with diversified medicinal uses. Based on its therapeutic applications, bromelain has got sufficient attention in pharmaceutical industries. In the present study, the full coding gene of bromelain in pineapple stem (1,093 bp) was amplified by RT-PCR. The PCR product was cloned, sequenced, and characterized. The sequence analysis of the gene revealed the single nucleotide polymorphism and its phylogenetic relatedness. The peptide sequence deduced from the gene showed the amino acid variations, physicochemical properties and secondary and tertiary structural features of the protein. The full BRM gene was transformed to prokaryotic vector pET32b and expressed in Escherichia coli BL21 DE3pLysS host cells successfully. The identity of the recombinant bromelain (rBRM) protein was confirmed by Western blot analysis using anti-BRM-rabbit IgG antibody. The activity of recombinant bromelain compared with purified native bromelain was determined by protease assay. The inhibitory effect of rBRM compared with native BRM in the growth of Gram-positive and Gram-negative strains of Streptococcus agalactiae and Escherichia coli O111 was evident from the antibacterial sensitivity test. To the best of our knowledge, this is the first report showing the bactericidal property of rBRM expressed in a prokaryotic system.     

Identification of small molecules inhibiting diguanylate cyclases to control bacterial biofilm development

Biofilm formation by pathogenic bacteria is an important virulence factor in the development of numerous chronic infections, thereby causing a severe health burden. Many of these infections cannot be resolved, as bacteria in biofilms are resistant to the host’s immune defenses and antibiotic therapy. An urgent need for new strategies to treat biofilm-based infections is critically needed. Cyclic di-GMP (c-di-GMP) is a widely conserved second-messenger signal essential for biofilm formation. The absence of this signalling system in higher eukaryotes makes it an attractive target for the development of new anti-biofilm agents. In this study, the results of an in silico pharmacophore-based screen to identify small-molecule inhibitors of diguanylate cyclase (DGC) enzymes that synthesize c-di-GMP are described. Four small molecules, LP 3134, LP 3145, LP 4010 and LP 1062 that antagonize these enzymes and inhibit biofilm formation by Pseudomonas aeruginosa and Acinetobacter baumannii in a continuous-flow system are reported. All four molecules dispersed P. aeruginosa biofilms and inhibited biofilm development on urinary catheters. One molecule dispersed A. baumannii biofilms. Two molecules displayed no toxic effects on eukaryotic cells. These molecules represent the first compounds identified from an in silico screen that are able to inhibit DGC activity to prevent biofilm formation.     

Protein Kinase B (PknB) of Mycobacterium tuberculosis Is Essential for Growth of the Pathogen in Vitro as well as for Survival within the Host

The Mycobacterium tuberculosis protein kinase B (PknB) comprises an intracellular kinase domain, connected through a transmembrane domain to an extracellular region that contains four PASTA domains. The present study describes the comprehensive analysis of different domains of PknB in the context of viability in avirulent and virulent mycobacteria. We find stringent regulation of PknB expression necessary for cell survival, with depletion or overexpression of PknB leading to cell death. Although PknB-mediated kinase activity is essential for cell survival, active kinase lacking the transmembrane or extracellular domain fails to complement conditional mutants not expressing PknB. By creating chimeric kinases, we find that the intracellular kinase domain has unique functions in the virulent strain, which cannot be substituted by other kinases. Interestingly, we find that although the presence of the C-terminal PASTA domain is dispensable in the avirulent M. smegmatis, all four PASTA domains are essential in M. tuberculosis. The differential behavior of PknB vis-à-vis the number of essential PASTA domains and the specificity of kinase domain functions suggest that PknB-mediated growth and signaling events differ in virulent compared with avirulent mycobacteria. Mouse infection studies performed to determine the role of PknB in mediating pathogen survival in the host demonstrate that PknB is not only critical for growth of the pathogen in vitro but is also essential for the survival of the pathogen in the host.     

Skp2B attenuates p53 function by inhibiting prohibitin

The F‐box protein Skp2 and its isoform Skp2B are both overexpressed in breast cancers. Skp2 alters the activity of p53 by inhibiting its interaction with p300 and by promoting p300 degradation. Here, we report that Skp2B also attenuates the activity of p53; however, this effect is independent of p300, suggesting that another mechanism might be involved. Prohibitin, a protein reported to activate p53, was isolated in a two‐hybrid screen with the carboxy‐terminal domain unique to Skp2B. We observed that prohibitin is a new substrate of Skp2B and that the degradation of prohibitin is responsible for the attenuated activity of p53 in cells overexpressing Skp2B. Furthermore, we show that the activity of p53 is reduced in the mammary glands of Skp2B transgenic mice. This study indicates that both Skp2 and Skp2B attenuate p53 activity through different pathways, suggesting that amplification of the Skp2 locus represents a powerful mechanism to attenuate p53 function in cancer.