Synthesis and biological evaluation of trans 6-methoxy-1,1-dimethyl-2-phenyl-3-aryl-2,3-dihydro-1H-inden-4-yloxyalkylamine derivatives against drug susceptible,non-replicating M. tuberculosis H37Rv and clinical multidrug resistant strains

Synthesis of a library of novel trans 6-methoxy-1,1-dimethyl-2-phenyl-3-aryl-2,3-dihydro-1H-inden-4-yloxy alkyl amines and their antimycobacterial activity against drug sensitive and multidrug resistant strains of Mycobacterium tuberculosis have been reported. All the new compounds in the series exhibited MIC between 1.56 and 6.25 μg/ml. Two compounds 1i and 1j with low MIC and low cytotoxicity showed significant reduction in CFU in infected mouse macrophages at 1× MIC concentration. The compound 1i inhibited the growth of M. tuberculosis in mice at 100 mg/kg dose with 1.35 log₁₀ reduction of CFU in lungs tissue and was active against non-replicating Mycobacterium tuberculosis under anaerobic condition.     

Neto2 Interacts with the Scaffolding Protein GRIP and Regulates Synaptic Abundance of Kainate Receptors

Kainate receptors (KARs) are a class of ionotropic glutamate receptors that are expressed throughout the central nervous system. The function and subcellular localization of KARs are tightly regulated by accessory proteins. We have previously identified the single-pass transmembrane proteins, Neto1 and Neto2, to be associated with native KARs. In the hippocampus, Neto1, but not Neto2, controls the abundance and modulates the kinetics of postsynaptic KARs. Here we evaluated whether Neto2 regulates synaptic KAR levels in the cerebellum where Neto1 expression is limited to the deep cerebellar nuclei. In the cerebellum, where Neto2 is present abundantly, we found a ∼40% decrease in GluK2-KARs at the postsynaptic density (PSD) of Neto2-null mice. No change, however, was observed in total level of GluK2-KARs, thereby suggesting a critical role of Neto2 on the synaptic localization of cerebellar KARs. The presence of a putative class II PDZ binding motif on Neto2 led us to also investigate whether it interacts with PDZ domain-containing proteins previously implicated in regulating synaptic abundance of KARs. We identified a PDZ-dependent interaction between Neto2 and the scaffolding protein GRIP. Furthermore, coexpression of Neto2 significantly increased the amount of GRIP associated with GluK2, suggesting that Neto2 may promote and/or stabilize GluK2:GRIP interactions. Our results demonstrate that Neto2, like Neto1, is an important auxiliary protein for modulating the synaptic levels of KARs. Moreover, we propose that the interactions of Neto1/2 with various scaffolding proteins is a critical mechanism by which KARs are stabilized at diverse synapses.     

Assessment of bacterial diversity in agricultural by-product compost by sequencing of cultivated isolates and amplified rDNA restriction analysis

An investigation of bacterial diversity in compost was performed using molecular chronometer in order to reveal its phylogeny. Thirty-three bacterial isolates isolated from compost were analyzed by 16S rRNA gene sequencing which revealed phylogenetic lineage of class Bacilli, γ, β-Proteobacteria, and Actinobacteria. Among these lineages, isolates belonging to class Bacilli consisted of species from genera Staphylococcus, Bacillus, Terribacillus, and Lysinibacillus. From phylum Actinobacteria: Microbacterium barkeri and Kocuria sp. were identified. Other bacterial groups had phylogenetic linkage with genera Comamonas and Acidovorax (class β-Proteobacteria); Serratia, Klebsiella, and Enterobacter (class γ-Proteobacteria). Similar isolates were analyzed through ARDRA. Amplified product of 16S rRNA gene from each isolates was subjected to cleavage by enzymes HpaII, HinfI, and MspI in separate reaction tubes. HpaII generated 2–6 bands ranging from 90–688 bp, HinfI generated 2–5 bands of 71–1,038 bp, and MspI 2–7 bands of 69–793 bp. The restriction patterns from HpaII, HinfI, and MspI were normalized separately and combined by means of pattern recognition software “Diversity Database.” HpaII had highest discrimination index (0.72) than HinfI (0.68) and MspI (0.65), and the combination of all three showed discrimination index (0.69). Numerical analysis of ARDRA patterns demonstrated sufficient phylogenetic information for characterizing bacterial diversity. Phylogenetic relationship obtained among isolates through ARDRA was compared with 16S rRNA gene sequence and ARDRA results showed sufficiently similar 16S rRNA gene sequence analysis, but not an overlapping. It has been observed that ARDRA technique facilitates the identification of bacteria in less than 36 h as compared to traditional 16S rRNA gene sequencing.     

Cholesterol binding reserve of hyperlipemic rat serum lipoproteins in chronic ethanol administration

Chronic administration of ethanol in rats caused the reduction of serum cholesterol binding reserve. The very low density and high density lipoproteins, main serum cholesterol binding reserves, were slightly increased with corresponding increases in their lipid and protein components during initial stage of alcohol consumption. However, these capacities get deminished during reversal of hyperlipemia induced by prolonged action of ethanol. This situation may be an early indicator for the initiation of hepatic damage and a variety of secondary effects of ethanol.     

Synthesis of phosphatidylcholine by rat lung during choline deficiency

The effect of choline deficiency on the de novo pathway for phosphatidylcholine (PC) synthesis in the lung was investigated in rats fed a washed soy protein (lipotrophic) diet deficient in choline and methionine for 2-3 wk. Lungs from lipotrophic rats showed a decreased content of choline and choline-phosphate (P less than 0.05) compared with control but no change in content of cytidine 5'-diphosphocholine or PC. Isolated perfused lungs from lipotrophic rats were evaluated for choline and fatty acid utilization for PC synthesis. Lipotrophic lungs perfused with 5 microM [14C-methyl]-choline chloride showed increased incorporation into PC while there was no significant effect at saturating levels of choline (100 microM). There was increased incorporation of [1-14C]-palmitic acid into PC and diglyceride and increased incorporation of D-[U-14C]glucose into fatty acids of PC. Increased choline and glucose incorporation was not due to alteration of intracellular specific activity of these substrates. This study indicates the utilization of choline and fatty acid for PC synthesis is stimulated as a result of choline deficiency while lung CDP-choline concentration is maintained, possibly through regulation of choline phosphate cytidyl transferase activity. These mechanisms compensate for decreased choline availability to maintain the PC content of lungs.     

Uptake hydrogenase in fast-growing strains of Rhizobium sp. (Sesbania) in relation to nitrogen fixation

Eight of 104 fast-growing strains of Rhizobium sp. ( Sesbania ) tested possessed a hydrogen recycling system. Depression of uptake hydrogenase occurred in poor as well as in rich carbon media. A statistically significant increase (> 22%) in total plant nitrogen content and dry matter yield was observed by the inoculation of hup + strains over hup ^- strains.     

Genetic and phylogenetic analyses of human T-cell lymphotropic virus type I variants from Melanesians with an without spastic myelopathy

Molecular variants of human T-cell lymphotropic virus type I (HTLV-I) have been isolated recently from lifelong residents of remote Melanesian populations, including a Solomon Islander with tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM) or HTLV-I myeloneuropathy. To clarify the genetic heterogeneity and molecular epidemiology of disease-associated strains of HTLV-I, we enzymatically amplified, then directly sequenced representative regions of thegag, pol, env, andpX genes of HTLV-I strains from Melanesians with and without TSP/HAM, and aligned and compared these sequences with those of HTLV-I strains from patients with TSP/HAM or adult T-cell leukemia/lymphoma and from asymptomatic carriers from widely separated and culturally disparate populations. Overall, the HTLV-I variant from the Solomon Islander with TSP/HAM, like HTLV-I strains from asymptomatically infected Melanesians, diverged by approx 7% from cosmopolitan HTLV-I strain. No disease-specific viral sequences were found. Gene phylogenies, as determined by the unweighted pair-goup method of assortment and by the maximum parsimony method, indicated that the Melanesian and cosmopolitan strains of HTLV-I have evolved along separate geographically dependent lineages, one comprised of HTLV-I strains from Papua New Guinea and the Solomon Islands, and the other composed of virus strains from Japan, India, the Caribbean, Polynesia, the Americas, and Africa. The total absence of nonhuman primates in Papua New Guinea and the Solomon Islands precludes any possibility that the Melanesian HTLV-I strains have evolved recently from the simian homolog of HTLV-I.     

Topcats and underdogs: intraguild interactions among three apex carnivores across Asia's forestscapes

Intraguild interactions among carnivores have long held the fascination of ecologists. Ranging from competition to facilitation and coexistence, these interactions and their complex interplay influence everything from species persistence to ecosystem functioning. Yet, the patterns and pathways of such interactions are far from understood in tropical forest systems, particularly across countries in the Global South. Here, we examined the determinants and consequences of competitive interactions between dholes Cuon alpinus and the two large felids (leopards Panthera pardus and tigers Panthera tigris) with which they most commonly co-occur across Asia. Using a combination of traditional and novel data sources (N = 118), we integrate information from spatial, temporal, and dietary niche dimensions. These three species have faced catastrophic declines in their extent of co-occurrence over the past century; most of their source populations are now confined to Protected Areas. Analysis of dyadic interactions between species pairs showed a clear social hierarchy. Tigers were dominant over dholes, although pack strength in dholes helped ameliorate some of these effects; leopards were subordinate to dholes. Population-level spatio-temporal interactions assessed at 25 locations across Asia did not show a clear pattern of overlap or avoidance between species pairs. Diet-profile assessments indicated that wild ungulate biomass consumption by tigers was highest, while leopards consumed more primate and livestock prey as compared to their co-predators. In terms of prey offtake (ratio of wild prey biomass consumed to biomass available), the three species together harvested 0.4–30.2% of available prey, with the highest offtake recorded from the location where the carnivores reach very high densities. When re-examined in the context of prey availability and offtake, locations with low wild prey availability showed spatial avoidance and temporal overlap among the carnivore pairs, and locations with high wild prey availability showed spatial overlap and temporal segregation. Based on these observations, we make predictions for 40 Protected Areas in India where temporally synchronous estimates of predator and prey densities are available. We expect that low prey availability will lead to higher competition, and in extreme cases, to the complete exclusion of one or more species. In Protected Areas with high prey availability, we expect intraguild coexistence and conspecific competition among carnivores, with spill-over to forest-edge habitats and subsequent prey-switching to livestock. We stress that dhole–leopard–tiger co-occurrence across their range is facilitated through an intricate yet fragile balance between prey availability, and intraguild and conspecific competition. Data gaps and limitations notwithstanding, our study shows how insights from fundamental ecology can be of immense utility for applied aspects like large predator conservation and management of human–carnivore interactions. Our findings also highlight potential avenues for future research on tropical carnivores that can broaden current understanding of intraguild competition in forest systems of Asia and beyond.     

Interaction of heavy metal toxicants with brain constitutive nitric oxide synthase

This study was designed to evaluate thein vitro effects of transition heavy metal cations on activity of constitutive isoform of nitric oxide synthase (cNOS) in rat brain. NOS activity was determined in the cytosolic fractions of rat cerebral hemispheres by conversion of³H-L-arginine to³H-L-citrulline. Different concentrations of mercury (Hg²⁺), nickel (Ni²⁺), manganese (Mn²⁺), zinc (Zn²⁺), cadmium (Cd²⁺), lead (Pb²⁺) and calcium (Ca²⁺) were tested on NOS activity. While all the cations caused inhibition, there were differences in the apparent inhibition constants (Ki) among the cations. With the exception of calcium ion no other cation required preincubation with the enzyme preparation. These results indicate that while calcium ion modulate cNOS activity at regulatory site(s), inhibitory influence of toxic heavy metal cations may be exerted on the catalytic site(s) either by direct binding to it or by interfering with the electron transfer during catalysis.     

Role of NAD+ and ADP-Ribosylation in the Maintenance of the Golgi Structure

We have investigated the role of the ADP- ribosylation induced by brefeldin A (BFA) in the mechanisms controlling the architecture of the Golgi complex. BFA causes the rapid disassembly of this organelle into a network of tubules, prevents the association of coatomer and other proteins to Golgi membranes, and stimulates the ADP-ribosylation of two cytosolic proteins of 38 and 50 kD (GAPDH and BARS-50; De Matteis, M.A., M. DiGirolamo, A. Colanzi, M. Pallas, G. Di Tullio, L.J. McDonald, J. Moss, G. Santini, S. Bannykh, D. Corda, and A. Luini. 1994. Proc. Natl. Acad. Sci. USA. 91:1114–1118; Di Girolamo, M., M.G. Silletta, M.A. De Matteis, A. Braca, A. Colanzi, D. Pawlak, M.M. Rasenick, A. Luini, and D. Corda. 1995. Proc. Natl. Acad. Sci. USA. 92:7065–7069). To study the role of ADP-ribosylation, this reaction was inhibited by depletion of NAD⁺ (the ADP-ribose donor) or by using selective pharmacological blockers in permeabilized cells. In NAD⁺-depleted cells and in the presence of dialized cytosol, BFA detached coat proteins from Golgi membranes with normal potency but failed to alter the organelle's structure. Readdition of NAD⁺ triggered Golgi disassembly by BFA. This effect of NAD⁺ was mimicked by the use of pre–ADP- ribosylated cytosol. The further addition of extracts enriched in native BARS-50 abolished the ability of ADP-ribosylated cytosol to support the effect of BFA. Pharmacological blockers of the BFA-dependent ADP-ribosylation (Weigert, R., A. Colanzi, A. Mironov, R. Buccione, C. Cericola, M.G. Sciulli, G. Santini, S. Flati, A. Fusella, J. Donaldson, M. DiGirolamo, D. Corda, M.A. De Matteis, and A. Luini. 1997. J. Biol. Chem. 272:14200–14207) prevented Golgi disassembly by BFA in permeabilized cells. These inhibitors became inactive in the presence of pre–ADP-ribosylated cytosol, and their activity was rescued by supplementing the cytosol with a native BARS-50–enriched fraction. These results indicate that ADP-ribosylation plays a role in the Golgi disassembling activity of BFA, and suggest that the ADP-ribosylated substrates are components of the machinery controlling the structure of the Golgi apparatus.