Cloning and deletion analysis of a genomic segment of Bacillus subtilis coding for the sdhA, B, C (succinate dehydrogenase) and gerE (spore germination) loci

From a Bacillus subtilis gene bank constructed in Escherichia coli and based on a low copy number cloning vector we have isolated a hybrid plasmid, pSH1047, containing an 8.0 kb segment of B. subtilis DNA coding for the sdhA, B and C genes, which code for the component polypeptides of succinate dehydrogenase, and the gerE gene, which may code for or regulate a protease involved in producing spores which germinate normally. We report the restriction map of this segment and the analysis of deletion derivatives which allow us to correlate the physical and genetic maps of these chromosomal segments.     

Structural Evidence for a Copper-Bound Carbonate Intermediate in the Peroxidase and Dismutase Activities of Superoxide Dismutase

Copper-zinc superoxide dismutase (SOD) is of fundamental importance to our understanding of oxidative damage. Its primary function is catalysing the dismutation of superoxide to O₂ and H₂O₂. SOD also reacts with H₂O₂, leading to the formation of a strong copper-bound oxidant species that can either inactivate the enzyme or oxidise other substrates. In the presence of bicarbonate (or CO₂) and H₂O₂, this peroxidase activity is enhanced and produces the carbonate radical. This freely diffusible reactive oxygen species is proposed as the agent for oxidation of large substrates that are too bulky to enter the active site. Here, we provide direct structural evidence, from a 2.15 Å resolution crystal structure, of (bi)carbonate captured at the active site of reduced SOD, consistent with the view that a bound carbonate intermediate could be formed, producing a diffusible carbonate radical upon reoxidation of copper. The bound carbonate blocks direct access of substrates to Cu(I), suggesting that an adjunct to the accepted mechanism of SOD catalysed dismutation of superoxide operates, with Cu(I) oxidation by superoxide being driven via a proton-coupled electron transfer mechanism involving the bound carbonate rather than the solvent. Carbonate is captured in a different site when SOD is oxidised, being located in the active site channel adjacent to the catalytically important Arg143. This is the probable route of diffusion from the active site following reoxidation of the copper. In this position, the carbonate is poised for re-entry into the active site and binding to the reduced copper.     

Identifying Elements of Patient-Centered Care in Underserved Populations: A Qualitative Study of Patient Perspectives

Patient-centered care is an important goal in the delivery of healthcare. However, many patients do not engage in preventive medical care. In this pilot study, we conducted twenty in depth, semi-structured qualitative interviews at the University of Illinois at Chicago Health Sciences campus in a four month time frame. Many patients were underserved and underinsured, and we wanted to understand their experiences in the healthcare system. Using content analysis, several themes emerged from the interview data. Participants discussed the need for empathy and rapport with their providers. They identified provider behaviors that fostered a positive clinical relationship, including step-by step explanations of procedures, attention to body language and clinic atmosphere, and appropriate time management. Participants identified cost as the most common barrier to engaging in preventive care and discussed children and social support as motivating factors. A long-term relationship with a provider was an important motivator for preventive care, suggesting that the therapeutic alliance was essential to many patients. Conversely, many participants discussed a sense of dehumanization in the healthcare system, reporting that their life circumstances were overlooked, or that they were judged based on insurance status or ethnicity. We discuss implications for provider training and healthcare delivery, including the importance of patient-centered medical homes.     

Effects of Porosity and Mixed Convection on MHD Two Phase Fluid Flow in an Inclined Channel

The present study deals with the flow and heat transfer analysis of two immiscible fluids in an inclined channel embedded in a porous medium. The channel is divided in two phases such that a third grade fluid occupies the phase I and a viscous fluid occupies the phase II. Both viscous and third grade fluids are electrically conducting. A constant magnetic field is imposed perpendicular to the channel walls. The mathematical model is developed by using Darcy''s and modified Darcy''s laws for viscous and third grade fluids respectively. The transformed ordinary differential equations are solved numerically using a shooting method. The obtained results are presented graphically and influence of emerging parameters is discussed in detail.     

Antifungal and antibacterial activities of indigenous <Emphasis Type="Italic">Streptomyces</Emphasis> isolates from saline farmlands: prescreening,ribotyping and metabolic diversity

A culture collection of 110 indigenous Streptomyces strains originally isolated from saline farmlands (Punjab, Pakistan) using stringent methods was screened biologically and chemically to investigate their potential for the production of bioactive secondary metabolites. In a biological screening the crude extracts obtained from the culture broth of selected strains were analysed for their activity against a set of test organisms, including Gram-positive, Gram-negative bacteria, fungi and microalgae using the disk diffusion bioassay method. Additionally a cytotoxicity test was performed by means of the brine shrimp microwell cytotoxicity assay. In a chemical screening each of the crude extracts was analysed by TLC using various staining reagents and by HPLC-MS/MS measurements. The results depicted an impressive chemical diversity of crude extracts produced by these strains. The taxonomic status of the selected strains was confirmed by preliminary physiological testing and 16S rRNA gene sequencing.     

Operculina turpethum attenuates N-nitrosodimethylamine induced toxic liver injury and clastogenicity in rats

The root extract of Operculina turpethum (OTE) has been used as an anti-inflammatory, purgative, and hepato-protective agent. N-Nitrosodimethylamine (NDMA) is a potent hepatotoxin that induces fibrosis of the liver. In the present study, we examined the therapeutic effects of OTE root extract against NDMA-induced hepatotoxicity and clastogenicity in rats. Hepatic fibrosis was induced in adult male albino rats through serial intraperitoneal administrations of NDMA at a concentration of 10 mg/kg body weight on three consecutive days of each week over a period of three weeks. A group of rats received OTE orally in doses of 75, 150 and 200 mg/kg body weight at 5 h after the administration of NDMA. The controls and treated animals were sacrificed on days-7, 14 and 21 after the start of the administration of NDMA. The progression of hepatic fibrosis as well as the amelioration effect of OTE was evaluated through histopathologically as well as by immunohistochemical staining for the activation of hepatic stellate cells. Alterations in serum and liver biochemical parameters and LDH isoenzymes were also studied. Serial administration of NDMA resulted in well formed fibrosis in the liver and induction of micronuclei in the bone marrow cells. Staining of α-SMA demonstrated activated stellate cells from day-7 onwards which was dramatically increased on day-21. An elevation of micronuclei count, liver function enzymes, serum hydroxyproline levels and LDH isoenzymes 4 and 5 were also observed. All these changes were remarkably reduced in OTE administered animals and fibrogenesis was completely absent. Our results suggest that OTE has hepatoprotective and anti-clastogenic effects against NDMA-induced hepatic fibrosis. Therefore OTE may be used as a hepatoprotective agent against various liver diseases including toxic liver injury.