Metagenomics analysis of the fecal microbiota in Ring-necked pheasants (Phasianus colchicus) and Green pheasants (Phasianus versicolor) using next generation sequencing

Pheasant reintroduction and conservation efforts have been in place in Pakistan since the 1980 s, yet there is still a scarcity of data on pheasant microbiome and zoonosis. Instead of growing vast numbers of bacteria in the laboratory, to investigate the fecal microbiome, pheasants (green and ring neck pheasant) were analyzed using 16S rRNA metagenomics and using IonS5TMXL sequencing from two flocks more than 10 birds. Operational taxonomic unit (OTU) cluster analysis and phylogenetic tree analysis was performed using Mothur software against the SSUrRNA database of SILVA and the MUSCLE (Version 3.8.31) software. Results of the analysis showed that firmicutes were the most abundant phylum among the top ten phyla, in both pheasant species, followed by other phyla such as actinobacteria and proteobacteria in ring necked pheasant and bacteroidetes in green necked pheasant. Bacillus was the most relatively abundant genus in both pheasants followed by Oceanobacillus and Teribacillus for ring necked pheasant and Lactobacillus for green necked pheasant. Because of their well-known beneficial characteristics, these genus warrants special attention. Bird droppings comprise germs from the urinary system, gut, and reproductive sites, making it difficult to research each anatomical site at the same time. We conclude that metagenomic analysis and classification provides baseline information of the pheasant fecal microbiome that plays a role in disease and health.     

A new model for the anaerobic fermentation of glycerol in enteric bacteria: trunk and auxiliary pathways in Escherichia coli

Anaerobic fermentation of glycerol in the Enterobacteriaceae family has long been considered a unique property of species that synthesize 1,3-propanediol (1,3-PDO). However, we have discovered that Escherichia coli can ferment glycerol in a 1,3-PDO-independent manner. We identified 1,2-propanediol (1,2-PDO) as a fermentation product and established the pathway that mediates its synthesis as well as its role in the metabolism of glycerol. We also showed that the trunk pathway responsible for the conversion of glycerol into glycolytic intermediates is composed of two enzymes: a type II glycerol dehydrogenase (glyDH-II) and a dihydroxyacetone kinase (DHAK), the former of previously unknown physiological role. Based on our findings, we propose a new model for glycerol fermentation in enteric bacteria in which: (i) the production of 1,2-PDO provides a means to consume reducing equivalents generated in the synthesis of cell mass, thus facilitating redox balance, and (ii) the conversion of glycerol to ethanol, through a redox-balanced pathway, fulfills energy requirements by generating ATP via substrate-level phosphorylation. The activity of the formate hydrogen-lyase and F(0)F(1)-ATPase systems were also found to facilitate the fermentative metabolism of glycerol, and along with the ethanol and 1,2-PDO pathways, were considered auxiliary or enabling. We demonstrated that glycerol fermentation in E. coli was not previously observed due to the use of medium formulations and culture conditions that impair the aforementioned pathways. These include high concentrations of potassium and phosphate, low concentrations of glycerol, alkaline pH, and closed cultivation systems that promote the accumulation of hydrogen gas.     

Molecular cloning of enantioselective ester hydrolase from Bacillus pumilus DBRL-191

A gene from Bacillus pumilus expressed under its native promoter was cloned in Escherichia coli. Recombinant B. pumilus esterase (BPE) affects the kinetic resolution of racemic mixtures such as unsubstituted and substituted 1-(phenyl)ethanols (E approximately 33-103), ethyl 3-hydroxy-3-phenylpropanoate (E approximately 45-71), trans-4-fluorophenyl-3-hydroxymethyl-N-methylpiperidine (E approximately 10-13) and ethyl 2-hydroxy-4-phenylbutyrate (E approximately 7). The enzyme is composed of a 34-amino acid signal peptide and a 181-amino acid mature protein corresponding to a molecular weight of approximately 19.2kD and pI approximately 9.4. 3-D the structural model of the enzyme built by homology modelling using the atomic coordinates from the crystal structure of B. subtilis lipase (LipA) showed a compact minimal alpha/beta hydrolase fold.     

A novel peptide derived from human apolipoprotein E is an inhibitor of tumor growth and ocular angiogenesis

Angiogenesis is a hallmark of tumor development and metastasis and now a validated target for cancer treatment. We previously reported that a novel dimer peptide (apoEdp) derived from the receptor binding region of human apolipoprotein E (apoE) inhibits virus-induced angiogenesis. However, its role in tumor anti-angiogenesis is unknown. This study demonstrates that apoEdp has anti-angiogenic property in vivo through reduction of tumor growth in a mouse model and ocular angiogenesis in a rabbit eye model. Our in vitro studies show that apoEdp inhibits human umbilical vein endothelial cell proliferation, migration, invasion and capillary tube formation. We document that apoEdp inhibits vascular endothelial growth factor-induced Flk-1 activation as well as downstream signaling pathways that involve c-Src, Akt, eNOS, FAK, and ERK1/2. These in vitro data suggest potential sites of the apoE dipeptide inhibition that could occur in vivo.This is the first evidence that a synthetic dimer peptide mimicking human apoE has anti-angiogenesis functions and could be an anti-tumor drug candidate.     

In vivo andin vitro methylation of lysine residues ofEuglena gracilis histone H1

We have earlier identified and purified two protein-lysine N-methyltransferases (Protein methylase III) fromEuglena gracilis [J. Biol. Chem.,260, 7114 (1985)]. The enzymes were highly specific toward histone H1 (lysine-rich), and the enzymatic products were identified as -N-mono-, di- and trimethyllysines. These earlier studies, however, were carried out with rat liver histone H1 as thein vitro substrate. Presently, histone H1 has been purified fromEuglena gracilis through Bio-Rex 70 and Bio-Gel P-100 column chromatography. TheEuglena histone H1 showed a single band on SDS-polyacrylamide gel electrophoresis and behaved like other histone H1 of higher animals, whereas it had a much higherR _f value than the other histones H1 in acid/urea gel electrophoresis. When theEuglena histone H1 was [methyl-³H]-labeledin vitro by a homologous enzyme (one of the twoEuglena protein methylase III) and analyzed on two-dimensional gel electrophoresis, three distinctive subtypes of histone H1 were shown to be radiolabeled, whereas five subtypes of rat liver histone H1 were found to be labeled. Finally, by the combined use of a strong cation exchange and reversed-phase Resolve C₁₈ columns on HPLC, we demonstrated thatEuglena histone H1 contains approximately 9 mol% of -N-methyllysines (1.40, 1.66, and 5.62 mol% for -N-mono-, di- and trimethyllysines, respectively). This is the first demonstration of the natural occurrence of -N-methyllysines in histone H1.     

Liver glycogen phosphorylase is upregulated in glioblastoma and provides a metabolic vulnerability to high dose radiation

Channelling of glucose via glycogen, known as the glycogen shunt, may play an important role in the metabolism of brain tumours, especially in hypoxic conditions. We aimed to dissect the role of glycogen degradation in glioblastoma (GBM) response to ionising radiation (IR). Knockdown of the glycogen phosphorylase liver isoform (PYGL), but not the brain isoform (PYGB), decreased clonogenic growth and survival of GBM cell lines and sensitised them to IR doses of 10–12 Gy. Two to five days after IR exposure of PYGL knockdown GBM cells, mitotic catastrophy and a giant multinucleated cell morphology with senescence-like phenotype developed. The basal levels of the lysosomal enzyme alpha-acid glucosidase (GAA), essential for autolysosomal glycogen degradation, and the lipidated forms of gamma-aminobutyric acid receptor-associated protein-like (GABARAPL1 and GABARAPL2) increased in shPYGL U87MG cells, suggesting a compensatory mechanism of glycogen degradation. In response to IR, dysregulation of autophagy was shown by accumulation of the p62 and the lipidated form of GABARAPL1 and GABARAPL2 in shPYGL U87MG cells. IR increased the mitochondrial mass and the colocalisation of mitochondria with lysosomes in shPYGL cells, thereby indicating reduced mitophagy. These changes coincided with increased phosphorylation of AMP-activated protein kinase and acetyl-CoA carboxylase 2, slower ATP generation in response to glucose loading and progressive loss of oxidative phosphorylation. The resulting metabolic deficiencies affected the availability of ATP required for mitosis, resulting in the mitotic catastrophy observed in shPYGL cells following IR. PYGL mRNA and protein levels were higher in human GBM than in normal human brain tissues and high PYGL mRNA expression in GBM correlated with poor patient survival. In conclusion, we show a major new role for glycogen metabolism in GBM cancer. Inhibition of glycogen degradation sensitises GBM cells to high-dose IR indicating that PYGL is a potential novel target for the treatment of GBMs.Subject terms: Cancer metabolism, CNS cancer     

Synthesis and biological evaluation of 4β-sulphonamido and 4β-[(4'-sulphonamido)benzamide]podophyllotoxins as DNA topoisomerase-IIα and apoptosis inducing agents

A series of new 4β-sulphonamido and 4β-[(4'-sulphonamido)benzamide] conjugates of podophyllotoxin (11a-j and 15a-g) were synthesized and evaluated for anticancer activity against six human cancer cell lines and found to be more potent than etoposide. Some of the compounds 11b, 11d and 11e that showed significant antiproliferative activity in Colo-205 cells, were superior to etoposide. The flow cytometric analysis indicates that these compounds (11b, 11d and 11e) showed G2/M cell cycle arrest and among them 11e is the most effective. It is observed that this compound (11e) caused both single-strand DNA breaks as observed by comet assay as well as double-strand DNA breaks as indicated by γ-H2AX. Further 11e showed inhibition of topo-IIα as observed from Western blot analysis and related studies. Compounds caused activation of ATM as well as Chk1 protein indicating that the compound caused effective DNA damage. Moreover activation of caspase-3, p21, p16, NF-kB and down regulation of Bcl-2 protein suggests that this compound (11e) has apoptotic cell death inducing ability, apart from acting as a topo-IIα inhibitor.     

The importance of leaf surface wax as short-range attractant and oviposition stimulant in a generalist Lepidoptera

Green gram, Vigna radiata (L.) Wilczek, is an important pulse crop of Asia. Severe attack by the larvae of Spilosoma obliqua Walker (Lepidoptera: Arctiidae) causes defoliation of green gram and reduces seed yield. Females lay eggs on the leaf surface, and therefore, surface wax plays an important role as short-range attractant and oviposition stimulant. So, we have attempted to find out whether leaf surface wax compounds (alkanes and free fatty acids) from three green gram cultivars (PDM 54, PUSA BAISAKHI and SAMRAT) could act as short-range attractant and oviposition stimulant in females. The TLC, GC-MS and GC-FID analyses of n-hexane extracts revealed 20 n-alkanes from n-C₁₅ to n-C₃₆ and 13 free fatty acids from C12:0 to C21:0, whilst linoleic acid was unique in SAMRAT. Pentacosane was the predominant amongst n-alkanes in the leaf surface waxes of three cultivars. Heneicosanoic acid and palmitoleic acid were the predominant free fatty acids in the leaf surface waxes of PDM 54, and PUSA BAISAKHI and SAMRAT, respectively. Females were attracted towards one leaf equivalent surface wax of three green gram cultivars against solvent controls (n-hexane) in Y-tube olfactometer bioassays. A synthetic blend of pentacosane, heptacosane, nonacosane, hexatriacontane, palmitoleic acid, linolenic acid and stearic acid, a synthetic blend of pentacosane, hexatriacontane and stearic acid, and a synthetic blend of hexatriacontane, linolenic acid and stearic acid resembling in amounts present in one leaf equivalent surface wax of PDM 54, PUSA BAISAKHI and SAMRAT, respectively, served as short-range attractant and oviposition stimulant in females. Females showed equal preference for egg laying towards the above three synthetic blends when these blends were tested against each other, and hence, these blends could be employed in development of baited traps in pest management strategies.     

Fullerenol [60] Nano-cages for Protection of Crops Against Oxidative Stress: A Critical Review

Journal of Plant Growth Regulation - Fullerenols are carbon nanoparticles that have been declared as free radical sponges. There is a need to take a prudent path toward its applications in various...