Mixed-ligand nickel(II) thiosemicarbazone complexes: Synthesis, characterization and biological evaluation

The syntheses and characterization of some new mixed-ligand nickel(II) complexes {Ni(L-1)(PPh3)] (1), Ni(L-1)(Py)] (2), Ni(L-2)(PPh3)]center dot DMSO (3), Ni(L-2)(Imz)] (4), Ni(L-3)(4-pic)] (5) and RNi(L-3))(2)(mu-4,4'-byp)]center dot 2DMSO (6)1 of three selected thiosemicarbazones the 4-(p-X-phenyl)thiosemicarbazones of salicylaldehyde) (H2L1-3) (A, Scheme 1) are described in the present study, differing in the inductive effect of the substituent X (X = F, Br and OCH3), in order to observe its influence, if any, on the redox potentials and biological activity of the complexes. All the synthesized ligands and the metal complexes were successfully characterized by elemental analysis, IR, UV-Vis, NMR spectroscopy and cyclic voltammetry. The molecular structures of four mononuclear (1-3 and 5) and one dinuclear (6) Ni(II) complex have been determined by X-ray crystallography. The complexes have been screened for their antibacterial activity against Escherichia coli and Bacillus. The minimum inhibitory concentrations of these complexes and their antibacterial activities indicate that compound 4 is the potential lead molecule for drug designing. (C) 2012 Elsevier Ltd. All rights reserved.

Evidence for the role of Mycobacteriumtuberculosis RecG helicase in DNA repair and recombination

In order to survive and replicate in a variety of stressful conditions during its life cycle, Mycobacteriumtuberculosis must possess mechanisms to safeguard the integrity of the genome. Although DNA repair and recombination related genes are thought to play key roles in the repair of damaged DNA in all organisms, so far only a few of them have been functionally characterized in the tubercle bacillus. In this study, we show that M.tuberculosis RecG (MtRecG) expression was induced in response to different genotoxic agents. Strikingly, expression of MtRecG in Escherichiacoli recG mutant strain provided protection against mitomycin C, methyl methane sulfonate and UV induced cell death. Purified MtRecG exhibited higher binding affinity for the Holliday junction (HJ) compared with a number of canonical recombinational DNA repair intermediates. Notably, although MtRecG binds at the core of the mobile and immobile HJs, and with higher binding affinity for the immobile HJ, branch migration was evident only in the case of the mobile HJ. Furthermore, immobile HJs stimulate MtRecG ATPase activity less efficiently than mobile HJs. In addition to HJ substrates, MtRecG exhibited binding affinity for a variety of branched DNA structures including three-way junctions, replication forks, flap structures, forked duplex and a D-loop structure, but demonstrated strong unwinding activity on replication fork and flap DNA structures. Together, these results support that MtRecG plays an important role in processes related to DNA metabolism under normal as well as stress conditions.

``On-the-fly'' kinetics of enzymatic racemization using deuterium NMR in DNA-based chiral oriented media

We report the in situ and real-time monitoring of the interconversion of L- and D-alanine-d(3) by alanine racemase from Bacillus stearothermophilus directly observed by H-2 NMR spectroscopy in anisotropic phase. The enantiomers are distinguished by the difference of their H-2 quadrupolar splittings in a chiral liquid crystal containing short DNA fragments. The proof-of-principle, the reliability, and the robustness of this new method is demonstrated by the determination of the turnover rates of the enzyme using the Michaelis Menten model.

Cyclic AMP-dependent protein lysine acylation in mycobacteria regulates fatty acid and propionate metabolism

Acetylation of lysine residues is a posttranslational modification that is used by both eukaryotes and prokaryotes to regulate a variety of biological processes. Here we identify multiple substrates for the cAMP-dependent protein lysine acetyltransferase from Mycobacterium tuberculosis (KATmt). We demonstrate that a catalytically important lysine residue in a number of FadD (fatty acyl CoA synthetase) enzymes is acetylated by KATmt in a cAMP-dependent manner and that acetylation inhibits the activity of FadD enzymes. A sirtuin-like enzyme can deacetylate multiple FadDs, thus completing the regulatory cycle. Using a strain deleted for the KATmt ortholog in Mycobacterium bovis Bacillus Calmette-Guerin (BCG), we show for the first time that acetylation is dependent on intracellular cAMP levels. KATmt can utilize propionyl CoA as a substrate and, therefore, plays a critical role in alleviating propionyl CoA toxicity in mycobacteria by inactivating acyl CoA synthetase (ACS). The precision by which mycobacteria can regulate the metabolism of fatty acids in a cAMP-dependent manner appears to be unparalleled in other biological organisms and is ideally suited to adapt to the complex environment that pathogenic mycobacteria experience in the host.

Mycobacterium bovis BCG promotes tumor cell survival from tumor necrosis factor-alpha-induced apoptosis

Background: Increased incidence of lung cancer among pulmonary tuberculosis patients suggests mycobacteria-induced tumorigenic response in the host. The alveolar epithelial cells, candidate cells that form lung adenocarcinoma, constitute a niche for mycobacterial replication and infection. We thus explored the possible mechanism of M. bovis Bacillus Calmette-Guerin (BCG)-assisted tumorigenicity in type II epithelial cells, human lung adenocarcinoma A549 and other cancer cells. Methods: Cancer cell lines originating from lung, colon, bladder, liver, breast, skin and cervix were treated with tumor necrosis factor (TNF)-alpha in presence or absence of BCG infection. p53, COP1 and sonic hedgehog (SHH) signaling markers were determined by immunoblotting and luciferase assays, and quantitative real time PCR was done for p53-responsive pro-apoptotic genes and SHH signaling markers. MTT assays and Annexin V staining were utilized to study apoptosis. Gain-and loss-of-function approaches were used to investigate the role for SHH and COP1 signaling during apoptosis. A549 xenografted mice were used to validate the contribution of BCG during TNF-alpha treatment. Results: Here, we show that BCG inhibits TNF-alpha-mediated apoptosis in A549 cells via downregulation of p53 expression. Substantiating this observation, BCG rescued A549 xenografts from TNF-alpha-mediated tumor clearance in nude mice. Furthermore, activation of SHH signaling by BCG induced the expression of an E3 ubiquitin ligase, COP1. SHH-driven COP1 targeted p53, thereby facilitating downregulation of p53-responsive pro-apoptotic genes and inhibition of apoptosis. Similar effects of BCG could be shown for HCT116, T24, MNT-1, HepG2 and HELA cells but not for HCT116 p53(-/-) and MDA-MB-231 cells. Conclusion: Our results not only highlight possible explanations for the coexistence of pulmonary tuberculosis and lung cancer but also address probable reasons for failure of BCG immunotherapy of cancers.

The adenylyl cyclase Rv2212 modifies the proteome and infectivity of Mycobacterium bovis BCG

All organisms have the capacity to sense and respond to environmental changes. These signals often involve the use of second messengers such as cyclic adenosine monophosphate (cAMP). This second messenger is widely distributed among organisms and coordinates gene expression related with pathogenesis, virulence, and environmental adaptation. Genomic analysis in Mycobacterium tuberculosis has identified 16 adenylyl cyclases (AC) and one phosphodiesterase, which produce and degrade cAMP, respectively. To date, ten AC have been biochemically characterized and only one (Rv0386) has been found to be important during murine infection with M. tuberculosis. Here, we investigated the impact of hsp60-driven Rv2212 gene expression in Mycobacterium bovis Bacillus Calmette-Guerin (BCG) during growth in vitro, and during macrophage and mice infection. We found that hsp60-driven expression of Rv2212 resulted in an increased capacity of replication in murine macrophages but an attenuated phenotype in lungs and spleen when administered intravenously in mice. Furthermore, this strain displayed an altered proteome mainly affecting proteins associated with stress conditions (bfrB, groEL-2, DnaK) that could contribute to the attenuated phenotype observed in mice.

Effect of solvent; enhancing the wettability and engineering the porous structure of a calcium phosphate/agarose composite for drug delivery

Tissue engineering deals with the regeneration of tissues for bone repair, wound healing, drug delivery, etc., and a highly porous 3D artificial scaffold is required to accommodate the cells and direct their growth. We prepared 3D porous calcium phosphate ((hydroxyapatite/beta-tricalcium phosphate)/agarose, (HAp/beta-TCP)/agarose) composite scaffolds by sol-gel technique with water (WBS) and ethanol (EBS) as solvents. The crystalline phases of HAp and beta-TCP in the scaffolds were confirmed by X-ray diffraction (XRD) analysis. The EBS had reduced crystallinity and crystallite size compared to WBS. WBS and EBS revealed interconnected pores of 1 mu m and 100 nm, respectively. The swelling ratio was higher for EBS in water and phosphate buffered saline (PBS). An in vitro drug loading/release experiment was carried out on the scaffolds using gentamicin sulphate (GS) and amoxicillin (AMX). We observed initial burst release followed by sustained release from WBS and EBS. In addition, GS showed more extended release than AMX from both the scaffolds. GS and AMX loaded scaffolds showed greater efficacy against Pseudomonas than Bacillus species. WBS exhibited enhanced mechanical properties, wettability, drug loading and haemocompatibility compared to EBS. In vitro cell studies showed that over the scaffolds, MC3T3 cells attached and proliferated and there was a significant increase in live MC3T3 cells. Both scaffolds supported MC3T3 proliferation and mineralization in the absence of osteogenic differentiation supplements in media which proves the scaffolds are osteoconducive. Microporous scaffolds (WBS) could assist the bone in-growth, whereas the presence of nanopores (EBS) could enhance the degradation process. Hence, WBS and EBS could be used as scaffolds for tissue engineering and drug delivery. This is a cost effective technique to produce scaffolds of degradable 3D ceramic-polymer composites.

Production and characterization of bioflocculants for mineral processing applications

A comparative study of two bacterial strains namely, Bacillus licheniformis and Bacillus firmus in the production of bioflocculants was made. The highest bioflocculant yield of 16.55 g/L was obtained from B. licheniformis (L) and 10 g/L from B. firmus (F). The bioflocculants obtained from the bacterial species were water soluble and insoluble in organic solvents. FTIR spectral analysis revealed the presence of hydroxyl, carboxyl and sugar derivatives in the bioflocculants. Thermal characterization by differential scanning calorimetry (DSC) showed the crystalline transition and the melting point (T-m) at 90-100 degrees C. Effects of bioflocculant dosage and pH on the flocculation of clay fines were evaluated. Highest bioflocculation efficiency on kaolin clay suspensions was observed at an optimum bioflocculant dosage of 5 g/L. The optimum pH range for the maximum bioflocculation was at pH 7-9. Bioflocculants exhibited high efficiency in dye decolorization. The maximum Cr (VI) removal was found to be 85 % for L (bioflocculant dosage at 2 g/L). This study demonstrates that microbial bioflocculants find potential applications in mineral processing such as selective flocculation of mineral fines, decolorization of dye solutions and in the remediation of toxic metal solutions. (C) 2015 Elsevier B.V. All rights reserved.

Simultaneous presence of fhs and purT genes is disadvantageous for the fitness of Escherichia coli growth

In bacteria, alternate mechanisms are known to synthesize N-10-formyltetrahydrofolate (N10-formyl-THF) and formyl glycinamide ribotide (fGAR), which are important in purine biosynthesis. In one of the mechanisms, a direct transfer of one carbon unit from formate allows Fhs to convert tetrahydrofolate to N-10-formyl-THF, and PurT to convert glycinamide ribotide (GAR) to fGAR. Our bioinformatics analysis of fhs and purT genes (encoding Fhs and PurT) showed that in a majority of bacteria (similar to 94%), their presence was mutually exclusive. A large number of organisms possessing fhs lacked purT and vice versa. The phenomenon is so penetrating that even within a genus (Bacillus) if a species possessed fhs it lacked purT and vice versa. To investigate physiological importance of this phenomenon, we used Escherichia coli, which naturally lacks fhs (and possesses purT) as model. We generated strains, which possessed fhs and purT genes in singles or together. Deletion of purT from E. coli in the presence or absence of fhs did not confer a detectable growth disadvantage in pure cultures. However, growth competition assays revealed that the strains possessing either of the single genes outcompeted those possessing both the genes suggesting that mutual exclusion of purT and fhs in organisms confers fitness advantage in mixed cultures.

Green Nanosilver as Reinforcing Eco-Friendly Additive to Epoxy Coating for Augmented Anticorrosive and Antimicrobial Behavior

Epoxy resin GY250 representing diglycidyl ethers of bisphenol-A (DGEBA) was reinforced with 1, 3 and 5 wt % of surface functionalized silver nanoparticles (F-AgNPs) which were synthesized using Couroupita guianensis leaves extract with a view of augmenting the corrosion control property of the epoxy resin and also imparting antimicrobial activity to epoxy coatings on mild steel. Corrosion resistance of the coatings was evaluated by EIS, potentiodynamic polarization studies and cross scratch tests. AFM, SEM, HRTEM and EDX were utilized to investigate the surface topography, morphology and elemental composition of the coatings on MS specimens. Results showed that the corrosion resistance, hardness and T-g of the DGEBA/F-AgNPs coatings increased at 1 wt % of F-AgNPs. The DGEBA/F-AgNPs coatings also offered manifold antimicrobial protection to the MS surfaces by inhibiting the growth of biofilm forming bacteria like P. aeruginosa, B. subtilis, the most common human pathogen E. coli and the most virulent human pathogenic yeast C. albicans.

Efecto de policultivos repollo-arroz, repollo-zanahoria, repollo-frijol, repollo-cebolla sobre la entomofauna en el cultivo del repollo (Brassica oleraceae L.) híbrido Izalco

El experimento de campo se llevó a cabo en San Ignacio, municipio de la Concepción, departamento de Masaya con el objetivo de evaluar el efecto de policultivos (repollo-arroz, repollo-zanahoria, repollo-frijol y repollo-cebolla) sobre la palomilla de la col (Plutella xylostella L.) en el cultivo de repollo (Brassica oleracea L.) híbrido Izalco, además se evaluó un tratamiento en monocultivo de repollo y otro con aplicación de insecticida biológico dipel (Bacillus thuringiensis) para ser usados como testigo absoluto y relativo respectivamente. Las plagas predominantes durante el ensayo fueron Plutella Xylostella, Spodoptera sp., Ascia monuste y Afidos. En la etapa de crecimiento vegetativo (0-32 días después del trasplante, DDT),las población de Plutella xylostella fue baja, pero en la etapa de preformación de cabeza (32-64 DDT) la población se incrementó, los tratamientos que presentaron menores poblaciones fueron repollo-arroz, repollo + dipel seguido de repollo-cebolla. En la etapa de formación de cabeza (64-88 DDT) la incidencia de la plaga fue siempre alta en el monocultivo de repollo y repollo-frijol a excepción de los demás tratamientos, siendo esta la etapa de mayor importancia del cultivo. El uso de cultivos en asocio no ejerció ningún efecto sobre Ascia monuste, Spodoptera sp. y Afidos. En la incidencia de insectos benéficos sólo se encontró & ralas y sus poblaciones se mantuvieron a niveles bajos.

Validación de diferentes opciones de manejo para el control de la roya del café (Hemileia vastatrix B y Br.) en la finca Santa Ana, El Mombacho, Nicaragua

El presente trabajo tuvo como objetivo buscar nuevas alternativas de manejo para el control de la roya del café (Hemileia vastatrix B & BR.) con el uso de Bacillus thuringiensis Berliner (Bt) y oxicloruro de cobre, como complemento al manejo de la sombra, para buscar como disminuir la incidencia de la roya y determinar el mejor momento para la aplicación tomando en cuenta los porcentajes de ésta enfermedad. Para ello se escogió la finca Santa Ana ubicada en el cerro Mombacho (Nicaragua) a 750 metros sobre el nivel del mar. De junio de 1994 a febrero de 1995 se realizaron quincenalmente muestreos aleatorios por conglomerados para roya, mancha de hierro (Cercospora coffeicola B & CK.) y antracnosis (Colletotrichum sp), así como se registraron todas las labores de manejo que realizó el productor. Se establecieron 4 opciones: 2 con cobre: una en forma calendarizado y otra con 10 por ciento de incidencia acumulada, una tercera opc1on usando Bt y por último una parcela testigo. Con los datos obtenidos se elaboraron curvas de incidencia simple y acumulada. Las proporciones de incidencia fueron transformadas a monolitos y lógitos para la realización de regresión simple y poder calcular la velocidad de crecimiento de la epidemia, se calculó el área bajo la curva de la epidemia y las X máximas alcanzadas, se realizaron análisis de varianza para establecer diferencias estadísticas entre los tratamientos y las repeticiones. De manera general la epidemia de roya se hace evidente en agosto y septiembre manteniéndose siempre en aumento, alcanzando los máximos valores en enero y febrero. De los tratamientos en estudio podemos decir que las aplicaciones de cobre calendarizado y con 10 por ciento de incidencia acumulada, presentaron la menor incidencia para la epidemia de roya, sin embargo en el cobre de aplicación calendarizada el efecto de cobre se observa mejor por lo que presenta el mejor comportamiento. En el caso de Bt no se logró determinar su efecto a nivel de campo, a la dosis empleada y en el momento aplicado, su comportamiento fue similar al testigo, quien presentó los mayores porcentajes de acumulados. Para el caso de mancha de hierro se observó una menor incidencia bajo el efecto de aplicaciones calendarizadas. La epidemia de antracnosis no se consideró un problema grave, debido a que los acumulados finales no sobre pasaron el nivel considerado riesgoso (15 por ciento).

Validación del umbral de acción para el manejo del gusano del fruto Helicoverpa spp (Boddie) Lepidoptera: noctuidae: en el cultivo del tomate Tisma, Nicaragua

Con el objetivo de validar un método determinado por ICTA/CATIE Guatemala, utilizando un umbral de acción para el manejo de Helicoverpa spp también conocido como gusano del fruto, en el cultivo del tomate se estableció un experimento de campo en la época de riego durante el período comprendido (Diciembre 1991 - Abril 1992) en Tisma, Masaya. El estudio se realizó con la colaboración de dos productores de tomate de la zona, en la finca de los productores se establecieron dos parcelas de validación, bajo dos sistemas de manejo: 1. Los productores realizaron aplicaciones periódicas semanales utilizando insecticidas químicos. 2. Aplicación de la mezcla de DIPEL (Bacillus thurinqiensis: 800 gr/mz) y LANNATE (Metomil: 75 gr/mz) según el umbral de acción, cuando se encontró cuatro huevos y/o larvas, o un 8% de frutos dañados por Helicoverpa spp en treinta plantas. El análisis económico de los tratamientos permite afirmar que las parcelas manejadas con umbral son más rentables, se obtiene mayores ingresos y menos costos a las parcelas manejadas por productor. Bajo las condiciones de este ensayo es factible económicamente, la implementación del umbral de acción, y la aplicación de la mezcla, para el combate de Helicoverpa spp en el cultivo del tomate.

Evaluación de productos botánicos y biológicos para el control de Plutella xylostella en el cultivo del repollo (Brassica oleraceae L.) hibrido Izalco en época de apante

Con el fin de encontrar el mejor producto para el control de Plutella xylostella en el cultivo del repollo, se llevó a cabo un ensayo a nivel de campo en época seca (Die 91-Mar 92) en el valle de Sébaco-Matagalpa donde se evaluaron los tratamientos Bacillus thuringiensis nacional, Dipel comercial, Javelin, Nim, Mamey y Júpiter, con un criterio de aplicación de 0.5 larvas por plantas. Las poblaciones de Plutella xylostella fueron relativamente bajas en la etapa de crecimiento vegetativo disminuyendo en las últimas dos etapas formación y llenado de cabeza. Durante las tres fases del cultivo las poblaciones de Plutella xylostella fueron menores en los tratamientos Javelin Mamey y Júpiter. Los insecticidas Dipel. Javelin y Mamey fueron diferentes estadísticamente del testigo (sin aplicación), según el análisis económico los primeros dos insecticidas son rentables económicamente sucediendo lo contrario con el insecticida Mamey por lo tanto su uso no es recomendado para el manejo de la plaga. El insecticida Júpiter puede ser utilizado contra la plaga ya que muestra una tasa de retorno marginal superior a la tasa comparativa. La presencia de enemigos naturales (Araña, Polybia sppy el parasitoide Diadegma insularis) fue notoria en el cultivo, cabe señalar su importancia ya que intervienen en el equilibrio del ecosistema dentro del cultivo (plaga-repollo). Estos predadores no se vieron afectados por las aplicaciones de los insecticidas botánicos y biológicos evaluados en el cultivo. El uso de estos productos parecen ser una alternativa de manejo de las plagas del cultivo a largo plazo ya que el establecimiento de la fauna benéfica en el sistema y el no uso de productos sintéticos evitan los problemas de resistencia en las plagas y la contaminación ambiental.

Evaluación económica de diferentes criterios de aplicación para el control de Plutella xylostella L. en el cultivo de repollo (Brassica oleracea L.) en época de primera

Con el objetivo de encontrar el mejor criterio de aplicación para el manejo de P. xylostella en el cultivo del repollo, se rea1izó un experimento a nivel de campo en época de primera, Mayo-Agosto ,1991 en la Concepción­ Masaya (IV Región de Nicaragua). Los tratamientos evaluados fueron: 0.1; O.3: O.5; O.7: O.9; y 1.2larvas por planta, sin control y calendarizado. Se mantuvieron los diferentes criterios de esta plaga utilizando aplicaciones semanales del insecticida Javelin WG NRD-12 Bacillus thuringiensis sub spp Kurstaki una dosis de 783 grlha. La incidencia de P. Xylosella no fue significativa durante la fase de establecimiento del cultivo del repollo (0-30 días después del trasplante), oscilando entre 0.01-007 larvas por planta. No así en las etapas posteriores comprendidas entre 30-90 días después del trasplante, se registraron poblaciones altas hasta 2. 30 larvas por planta encontrándose diferencias significativas. Esta incidencia en la última etapa afecta a las variables: daño foliar, precio e ingreso bruto. Se tomaron datos sobré los costos agrícolas de cada uno de los tratamientos y sus respectivos rendimientos. Se analizaron mediante el análisis de presupuestos parciales incluyendo un análisis de retorno mínimo y un análisis de sensibilidad de los precios del producto final. El análisis económico mostró que para una producción de repollo rentable, es necesario el uso de plaguicidas utilizando umbrales de aplicación que permitan mantener la rentabilidad del cultivo y los niveles poblacionales de la plaga. Tal es el caso del umbral O.S larvas por planta que demostró presentar estas características aún en las peores condiciones de producción, al igual que cuando existen cambios en el precio del insecticida y el precio del repollo, es decir que cuando existe un aumento de 5,300% en el precio del insecticida y una disminución de 95.8% en el precio del repollo este umbral mantiene una tasa de retorno marginal similar a la tasa mínima comparativa de 125%.